AU2015227398B2 - Method for using gene expression to determine prognosis of prostate cancer - Google Patents

Abstract

Molecular assays that involve measurement of expression levels of prognostic biomarkers, or co-expressed biomarkers, from a biological sample obtained from a prostate cancer patient, and analysis of the measured expression levels to provide information concerning the likely prognosis for said patient, and likelihood that said patient will have a recurrence of prostate cancer, or to classify the tumor by likelihood of clinical outcome or TMPRSS2 fusion status, are provided herein. The prognostic markers comprise gene or microRNA expression levels.

Description

2015227398 15 Sep 2015

METHOD FOR USING GENE EXPRESSION TO DETERMINE PROGNOSIS

OF PROSTATE CANCER

[0001] The present application is a divisional of AU2011282892, the entire contents of which are incorporated herein by reference.

TECHNICAL FIELD

[0002] The present disclosure relates to molecular diagnostic assays that provide information concerning methods to use gene expression profiles to determine prognostic information for cancer patients. Specifically, the present disclosure provides genes and microRNAs, the expression levels of which may be used to determine the likelihood that a prostate cancer patient will experience a local or distant cancer recurrence.

INTRODUCTION

[0003] Prostate cancer is the most common solid malignancy in men and the second most common cause of cancer-related death in men in North America and the European Union (EU).

In 2008, over 180,000 patients will be diagnosed with prostate cancer in the United States alone and nearly 30,000 will die of this disease. Age is the single most important risk factor for the development of prostate cancer, and applies across all racial groups that have been studied. With the aging of the U.S. population, it is projected that the annual incidence of prostate cancer will double by 2025 to nearly 400,000 cases per year.

[0004] Since the introduction of prostate-specific antigen (PSA) screening in the 1990’s, the proportion of patients presenting with clinically evident disease has fallen dramatically such that patients categorized as “low risk” now constitute half of new diagnoses today. PSA is used as a tumor marker to determine the presence of prostate cancer as high PSA levels are associated with prostate cancer. Despite a growing proportion of localized prostate cancer patients presenting with low-risk features such as low stage (Tl) disease, greater than 90% of patients in the US still undergo definitive therapy, including prostatectomy or radiation. Only about 15% of these patients would develop metastatic disease and die from their prostate cancer, even in the absence of definitive therapy. A. Bill-Axelson, et al., J Nat’l Cancer Inst. 100(16):1144-1154 (2008). Therefore, the majority of prostate cancer patients are being over-treated. 1 2015227398 15 Sep 2015 [0005] Estimates of recurrence risk and treatment decisions in prostate cancer are currently based primarily on PSA levels and/or tumor stage. Although tumor stage has been demonstrated to have significant association with outcome sufficient to be included in pathology reports, the College of American Pathologists Consensus Statement noted that variations in approach to the acquisition, interpretation, reporting, and analysis of this information exist. C. Compton, et al., Arch Pathol Lab Med 124:979-992 (2000). As a consequence, existing pathologic staging methods have been criticized as lacking reproducibility and therefore may provide imprecise estimates of individual patient risk.

SUMMARY

[0006] This application discloses molecular assays that involve measurement of expression level(s) of one or more genes, gene subsets, microRNAs, or one or more microRNAs in combination with one or more genes or gene subsets, from a biological sample obtained from a prostate cancer patient, and analysis of the measured expression levels to provide information concerning the likelihood of cancer recurrence. For example, the likelihood of cancer recurrence could be described in terms of a score based on clinical or biochemical recurrence-free interval.

[0007] In addition, this application discloses molecular assays that involve measurement of expression level(s) of one or more genes, gene subsets, microRNAs, or one or more microRNAs in combination with one or more genes or gene subsets, from a biological sample obtained to identify a risk classification for a prostate cancer patient. For example, patients may be stratified using expression level(s) of one or more genes or microRNAs associated, positively or negatively, with cancer recurrence or death from cancer, or with a prognostic factor. In an exemplary embodiment, the prognostic factor is Gleason pattern.

[0008] The biological sample may be obtained from standard methods, including surgery, biopsy, or bodily fluids. It may comprise tumor tissue or cancer cells, and, in some cases, histologically normal tissue, e.g., histologically normal tissue adjacent the tumor tissue. In exemplary embodiments, the biological sample is positive or negative for a TMPRSS2 fusion.

[0009] In exemplary embodiments, expression level(s) of one or more genes and/or microRNAs that are associated, positively or negatively, with a particular clinical outcome in prostate cancer are used to determine prognosis and appropriate therapy. The genes disclosed herein may be used alone or arranged in functional gene subsets, such as cell adhesion/migration, immediate-early stress response, and extracellular matrix-associated. Each gene subset comprises 2 2015227398 15 Sep 2015 the genes disclosed herein, as well as genes that are co-expressed with one or more of the disclosed genes. The calculation may be performed on a computer, programmed to execute the gene expression analysis. The microRNAs disclosed herein may also be used alone or in combination with any one or more of the microRNAs and/or genes disclosed.

[0010] In exemplary embodiments, the molecular assay may involve expression levels for at least two genes. The genes, or gene subsets, may be weighted according to strength of association with prognosis or tumor microenvironment. In another exemplary embodiment, the molecular assay may involve expression levels of at least one gene and at least one microRNA. The gene-microRNA combination may be selected based on the likelihood that the gene-microRNA combination functionally interact.

BRIEF DESCRIPTION OF THE DRAWING

[0011] Figure 1 shows the distribution of clinical and pathology assessments of biopsy Gleason score, baseline PSA level, and clinical T-stage.

DEFINITIONS

[0012] Unless defined otherwise, technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Singleton et al., Dictionary of Microbiology and Molecular Biology 2nd ed., J. Wiley & Sons (New York, NY 1994), and March, Advanced Organic Chemistry Reactions, Mechanisms and Structure 4th ed., John Wiley & Sons (New York, NY 1992), provide one skilled in the art with a general guide to many of the terms used in the present application.

[0013] One skilled in the art will recognize many methods and materials similar or equivalent to those described herein, which could be used in the practice of the present invention. Indeed, the present invention is in no way limited to the methods and materials described herein. For purposes of the invention, the following terms are defined below.

[0014] The terms “tumor” and “lesion” as used herein, refer to all neoplastic cell growth and proliferation, whether malignant or benign, and all pre-cancerous and cancerous cells and tissues. Those skilled in the art will realize that a tumor tissue sample may comprise multiple biological elements, such as one or more cancer cells, partial or fragmented cells, tumors in 3 2015227398 15 Sep 2015 various stages, surrounding histologically normal-appearing tissue, and/or macro or micro-dissected tissue.

[0015] The terms “cancer” and “cancerous” refer to or describe the physiological condition in mammals that is typically characterized by unregulated cell growth. Examples of cancer in the present disclosure include cancer of the urogenital tract, such as prostate cancer.

[0016] The “pathology” of cancer includes all phenomena that compromise the wellbeing of the patient. This includes, without limitation, abnormal or uncontrollable cell growth, metastasis, interference with the normal functioning of neighboring cells, release of cytokines or other secretory products at abnormal levels, suppression or aggravation of inflammatory or immunological response, neoplasia, premalignancy, malignancy, invasion of surrounding or distant tissues or organs, such as lymph nodes, etc.

[0017] As used herein, the term “prostate cancer” is used interchangeably and in the broadest sense refers to all stages and all forms of cancer arising from the tissue of the prostate gland.

[0018] According to the tumor, node, metastasis (TNM) staging system of the American Joint Committee on Cancer (AJCC), AJCC Cancer Staging Manual (7th Ed., 2010), the various stages of prostate cancer are defined as follows: Tumor: Tl: clinically inapparent tumor not palpable or visible by imaging, Tla: tumor incidental histological finding in 5% or less of tissue resected, Tib: tumor incidental histological finding in more than 5% of tissue resected, Tic: tumor identified by needle biopsy; T2: tumor confined within prostate, T2a: tumor involves one half of one lobe or less, T2b: tumor involves more than half of one lobe, but not both lobes, T2c: tumor involves both lobes; T3: tumor extends through the prostatic capsule, T3a: extracapsular extension (unilateral or bilateral), T3b: tumor invades seminal vesicle(s); T4: tumor is fixed or invades adjacent structures other than seminal vesicles (bladder neck, external sphincter, rectum, levator muscles, or pelvic wall). Node: NO: no regional lymph node metastasis; Nl: metastasis in regional lymph nodes. Metastasis: M0: no distant metastasis; Ml: distant metastasis present.

[0019] The Gleason Grading system is used to help evaluate the prognosis of men with prostate cancer. Together with other parameters, it is incorporated into a strategy of prostate cancer staging, which predicts prognosis and helps guide therapy. A Gleason “score” or “grade” is given to prostate cancer based upon its microscopic appearance. Tumors with a low Gleason score typically grow slowly enough that they may not pose a significant threat to the patients in 4 2015227398 15 Sep 2015 their lifetimes. These patients are monitored (“watchful waiting” or “active surveillance”) over time. Cancers with a higher Gleason score are more aggressive and have a worse prognosis, and these patients are generally treated with surgery (e.g., radical prostectomy) and, in some cases, therapy (e.g., radiation, hormone, ultrasound, chemotherapy). Gleason scores (or sums) comprise grades of the two most common tumor patterns. These patterns are referred to as Gleason patterns 1-5, with pattern 1 being the most well-differentiated. Most have a mixture of patterns. To obtain a Gleason score or grade, the dominant pattern is added to the second most prevalent pattern to obtain a number between 2 andlO. The Gleason Grades include: Gl: well differentiated (slight anaplasia) (Gleason 2-4); G2: moderately differentiated (moderate anaplasia) (Gleason 5-6); G3-4: poorly differentiated/undifferentiated (marked anaplasia) (Gleason 7-10).

[0020] Stage groupings: Stage I: Tla NO M0 Gl; Stage II: (Tla NO M0 G2-4) or (Tib, c, ΤΙ, T2, NO M0 Any G); Stage III: T3 NO M0 Any G; Stage IV: (T4 NO M0 Any G) or (Any T N1 M0 Any G) or (Any T Any N Ml Any G).

[0021] As used herein, the term “tumor tissue” refers to a biological sample containing one or more cancer cells, or a fraction of one or more cancer cells. Those skilled in the art will recognize that such biological sample may additionally comprise other biological components, such as histologically appearing normal cells (e.g., adjacent the tumor), depending upon the method used to obtain the tumor tissue, such as surgical resection, biopsy, or bodily fluids.

[0022] As used herein, the term “AUA risk group” refers to the 2007 updated American Urological Association (AUA) guidelines for management of clinically localized prostate cancer, which clinicians use to determine whether a patient is at low, intermediate, or high risk of biochemical (PSA) relapse after local therapy.

[0023] As used herein, the term “adjacent tissue (AT)” refers to histologically “normal” cells that are adjacent a tumor. For example, the AT expression profile may be associated with disease recurrence and survival.

[0024] As used herein “non-tumor prostate tissue” refers to histologically normalappearing tissue adjacent a prostate tumor.

[0025] Prognostic factors are those variables related to the natural history of cancer, which influence the recurrence rates and outcome of patients once they have developed cancer. Clinical parameters that have been associated with a worse prognosis include, for example, 5 2015227398 15 Sep 2015 increased tumor stage, PSA level at presentation, and Gleason grade or pattern. Prognostic factors are frequently used to categorize patients into subgroups with different baseline relapse risks.

[0026] The term “prognosis” is used herein to refer to the likelihood that a cancer patient will have a cancer-attributable death or progression, including recurrence, metastatic spread, and drug resistance, of a neoplastic disease, such as prostate cancer. For example, a “good prognosis” would include long term survival without recurrence and a “bad prognosis” would include cancer recurrence.

[0027] As used herein, the term “expression level” as applied to a gene refers to the normalized level of a gene product, e.g. the normalized value determined for the RNA expression level of a gene or for the polypeptide expression level of a gene.

[0028] The term “gene product” or “expression product” are used herein to refer to the RNA (ribonucleic acid) transcription products (transcripts) of the gene, including mRNA, and the polypeptide translation products of such RNA transcripts. A gene product can be, for example, an unspliced RNA, an mRNA, a splice variant mRNA, a microRNA, a fragmented RNA, a polypeptide, a post-translationally modified polypeptide, a splice variant polypeptide, etc.

[0029] The term “RNA transcript” as used herein refers to the RNA transcription products of a gene, including, for example, mRNA, an unspliced RNA, a splice variant mRNA, a microRNA, and a fragmented RNA.

[0030] The term “microRNA” is used herein to refer to a small, non-coding, single-stranded RNA of -18 - 25 nucleotides that may regulate gene expression. For example, when associated with the RNA-induced silencing complex (RISC), the complex binds to specific mRNA targets and causes translation repression or cleavage of these mRNA sequences.

[0031] Unless indicated otherwise, each gene name used herein corresponds to the Official Symbol assigned to the gene and provided by Entrez Gene (URL: www.ncbi.nlm.nih.gov/sites/entrez) as of the filing date of this application.

[0032] The terms “correlated” and “associated” are used interchangeably herein to refer to the association between two measurements (or measured entities). The disclosure provides genes,gene subsets, microRNAs, or microRNAs in combination with genes or gene subsets, the expression levels of which are associated with tumor stage. For example, the increased 6 2015227398 15 Sep 2015 expression level of a gene or microRNA may be positively correlated (positively associated) with a good or positive prognosis. Such a positive correlation may be demonstrated statistically in various ways, e.g. by a cancer recurrence hazard ratio less than one. In another example, the increased expression level of a gene or microRNA may be negatively correlated (negatively associated) with a good or positive prognosis. In that case, for example, the patient may experience a cancer recurrence.

[0033] The terms “good prognosis” or “positive prognosis” as used herein refer to a beneficial clinical outcome, such as long-term survival without recurrence. The terms “bad prognosis” or “negative prognosis” as used herein refer to a negative clinical outcome, such as cancer recurrence.

[0034] The term “risk classification” means a grouping of subjects by the level of risk (or likelihood) that the subject will experience a particular clinical outcome. A subject may be classified into a risk group or classified at a level of risk based on the methods of the present disclosure, e.g. high, medium, or low risk. A “risk group” is a group of subjects or individuals with a similar level of risk for a particular clinical outcome.

[0035] The term “long-term” survival is used herein to refer to survival for a particular time period, e.g., for at least 5 years, or for at least 10 years.

[0036] The term “recurrence” is used herein to refer to local or distant recurrence (i.e., metastasis) of cancer. For example, prostate cancer can recur locally in the tissue next to the prostate or in the seminal vesicles. The cancer may also affect the surrounding lymph nodes in the pelvis or lymph nodes outside this area. Prostate cancer can also spread to tissues next to the prostate, such as pelvic muscles, bones, or other organs. Recurrence can be determined by clinical recurrence detected by, for example, imaging study or biopsy, or biochemical recurrence detected by, for example, sustained follow-up prostate-specific antigen (PSA) levels > 0.4 ng/mL or the initiation of salvage therapy as a result of a rising PSA level.

[0037] The term “clinical recurrence-free interval (cRFI)” is used herein as time (in months) from surgery to first clinical recurrence or death due to clinical recurrence of prostate cancer. Losses due to incomplete follow-up, other primary cancers or death prior to clinical recurrence are considered censoring events; when these occur, the only information known is that up through the censoring time, clinical recurrence has not occurred in this subject. Biochemical recurrences are ignored for the purposes of calculating cRFI. 7 2015227398 15 Sep 2015 [0038] The term “biochemical recurrence-free interval (bRFI)” is used herein to mean the time (in months) from surgery to first biochemical recurrence of prostate cancer. Clinical recurrences, losses due to incomplete follow-up, other primary cancers, or death prior to biochemical recurrence are considered censoring events.

[0039] The term “Overall Survival (OS)” is used herein to refer to the time (in months) from surgery to death from any cause. Losses due to incomplete follow-up are considered censoring events. Biochemical recurrence and clinical recurrence are ignored for the purposes of calculating OS.

[0040] The term “Prostate Cancer-Specific Survival (PCSS)” is used herein to describe the time (in years) from surgery to death from prostate cancer. Losses due to incomplete followup or deaths from other causes are considered censoring events. Clinical recurrence and biochemical recurrence are ignored for the purposes of calculating PCSS.

[0041] The term “upgrading” or “upstaging” as used herein refers to a change in Gleason grade from 3+3 at the time of biopsy to 3+4 or greater at the time of radical prostatectomy (RP), or Gleason grade 3+4 at the time of biopsy to 4+3 or greater at the time of RP, or seminal vessical involvement (SVI), or extracapsular involvement (ECE) at the time of RP.

[0042] In practice, the calculation of the measures listed above may vary from study to study depending on the definition of events to be considered censored.

[0043] The term “microarray” refers to an ordered arrangement of hybridizable array elements, e.g. oligonucleotide or polynucleotide probes, on a substrate.

[0044] The term “polynucleotide” generally refers to any polyribonucleotide or polydeoxribonucleotide, which may be unmodified RNA or DNA or modified RNA or DNA. Thus, for instance, polynucleotides as defined herein include, without limitation, single- and double-stranded DNA, DNA including single- and double-stranded regions, single- and double-stranded RNA, and RNA including single- and double-stranded regions, hybrid molecules comprising DNA and RNA that may be single-stranded or, more typically, double-stranded or include single- and double-stranded regions. In addition, the term “polynucleotide” as used herein refers to triple-stranded regions comprising RNA or DNA or both RNA and DNA. The strands in such regions may be from the same molecule or from different molecules. The regions may include all of one or more of the molecules, but more typically involve only a region of some of the molecules. One of the molecules of a triple-helical region often is an 8 2015227398 15 Sep 2015 oligonucleotide. The term “polynucleotide” specifically includes cDNAs. The term includes DNAs (including cDNAs) and RNAs that contain one or more modified bases. Thus, DNAs or RNAs with backbones modified for stability or for other reasons, are “polynucleotides” as that term is intended herein. Moreover, DNAs or RNAs comprising unusual bases, such as inosine, or modified bases, such as tritiated bases, are included within the term “polynucleotides” as defined herein. In general, the term “polynucleotide” embraces all chemically, enzymatically and/or metabolically modified forms of unmodified polynucleotides, as well as the chemical forms of DNA and RNA characteristic of viruses and cells, including simple and complex cells.

[0045] The term “oligonucleotide” refers to a relatively short polynucleotide, including, without limitation, single-stranded deoxyribonucleotides, single- or double-stranded ribonucleotides, RNArDNA hybrids and double-stranded DNAs. Oligonucleotides, such as single-stranded DNA probe oligonucleotides, are often synthesized by chemical methods, for example using automated oligonucleotide synthesizers that are commercially available.

However, oligonucleotides can be made by a variety of other methods, including in vitro recombinant DNA-mediated techniques and by expression of DNAs in cells and organisms.

[0046] The term “Ct” as used herein refers to threshold cycle, the cycle number in quantitative polymerase chain reaction (qPCR) at which the fluorescence generated within a reaction well exceeds the defined threshold, i.e. the point during the reaction at which a sufficient number of amplicons have accumulated to meet the defined threshold.

[0047] The term “Cp” as used herein refers to “crossing point.” The Cp value is calculated by determining the second derivatives of entire qPCR amplification curves and their maximum value. The Cp value represents the cycle at which the increase of fluorescence is highest and where the logarithmic phase of a PCR begins.

[0048] The terms “threshold” or “thresholding” refer to a procedure used to account for non-linear relationships between gene expression measurements and clinical response as well as to further reduce variation in reported patient scores. When thresholding is applied, all measurements below or above a threshold are set to that threshold value. Non-linear relationship between gene expression and outcome could be examined using smoothers or cubic splines to model gene expression in Cox PH regression on recurrence free interval or logistic regression on recurrence status. D. Cox, Journal of the Royal Statistical Society, Series B 34:187-220 (1972). 9 2015227398 15 Sep 2015

Variation in reported patient scores could be examined as a function of variability in gene expression at the limit of quantitation and/or detection for a particular gene.

[0049] As used herein, the term “amplicon,” refers to pieces of DNA that have been synthesized using amplification techniques, such as polymerase chain reactions (PCR) and ligase chain reactions.

[0050] “Stringency” of hybridization reactions is readily determinable by one of ordinary skill in the art, and generally is an empirical calculation dependent upon probe length, washing temperature, and salt concentration. In general, longer probes require higher temperatures for proper annealing, while shorter probes need lower temperatures. Hybridization generally depends on the ability of denatured DNA to re-anneal when complementary strands are present in an environment below their melting temperature. The higher the degree of desired homology between the probe and hybridizable sequence, the higher the relative temperature which can be used. As a result, it follows that higher relative temperatures would tend to make the reaction conditions more stringent, while lower temperatures less so. For additional details and explanation of stringency of hybridization reactions, see Ausubel et al., Current Protocols in Molecular Biology (Wiley Interscience Publishers, 1995).

[0051] “Stringent conditions” or “high stringency conditions”, as defined herein, typically: (1) employ low ionic strength and high temperature for washing, for example 0.015 M sodium chloride/0.0015 M sodium citrate/0.1% sodium dodecyl sulfate at 50°C; (2) employ during hybridization a denaturing agent, such as formamide, for example, 50% (v/v) formamide with 0.1% bovine serum albumin/0.1% Ficoll/0.1% polyvinylpyrrolidone/50mM sodium phosphate buffer at pH 6.5 with 750 mM sodium chloride, 75 mM sodium citrate at 42°C; or (3) employ 50% formamide, 5 x SSC (0.75 M NaCl, 0.075 M sodium citrate), 50 mM sodium phosphate (pH 6.8), 0.1% sodium pyrophosphate, 5 x Denhardt's solution, sonicated salmon sperm DNA (50 pg/ml), 0.1% SDS, and 10% dextran sulfate at 42°C, with washes at 42°C in 0.2 x SSC (sodium chloride/sodium citrate) and 50% formamide, followed by a high-stringency wash consisting of 0.1 x SSC containing EDTA at 55°C.

[0052] “Moderately stringent conditions” may be identified as described by Sambrook et al., Molecular Cloning: A Laboratory Manual, New York: Cold Spring Harbor Press, 1989, and include the use of washing solution and hybridization conditions (e.g., temperature, ionic strength and %SDS) less stringent that those described above. An example of moderately 10 2015227398 15 Sep 2015 stringent conditions is overnight incubation at 37°C in a solution comprising: 20% formamide, 5 x SSC (150 mM NaCl, 15 mM trisodium citrate), 50 mM sodium phosphate (pH 7.6), 5 x Denhardt's solution, 10% dextran sulfate, and 20 mg/ml denatured sheared salmon sperm DNA, followed by washing the filters in 1 x SSC at about 37-500C. The skilled artisan will recognize how to adjust the temperature, ionic strength, etc. as necessary to accommodate factors such as probe length and the like.

[0053] The terms “splicing” and “RNA splicing” are used interchangeably and refer to RNA processing that removes introns and joins exons to produce mature mRNA with continuous coding sequence that moves into the cytoplasm of an eukaryotic cell.

[0054] The terms “co-express” and “co-expressed”, as used herein, refer to a statistical correlation between the amounts of different transcript sequences across a population of different patients. Pairwise co-expression may be calculated by various methods known in the art, e.g., by calculating Pearson correlation coefficients or Spearman correlation coefficients. Co-expressed gene cliques may also be identified using graph theory. An analysis of co-expression may be calculated using normalized expression data. A gene is said to be co-expressed with a particular disclosed gene when the expression level of the gene exhibits a Pearson correlation coefficient greater than or equal to 0.6.

[0055] A “computer-based system” refers to a system of hardware, software, and data storage medium used to analyze information. The minimum hardware of a patient computer-based system comprises a central processing unit (CPU), and hardware for data input, data output (e.g., display), and data storage. An ordinarily skilled artisan can readily appreciate that any currently available computer-based systems and/or components thereof are suitable for use in connection with the methods of the present disclosure. The data storage medium may comprise any manufacture comprising a recording of the present information as described above, or a memory access device that can access such a manufacture.

[0056] To “record” data, programming or other information on a computer readable medium refers to a process for storing information, using any such methods as known in the art. Any convenient data storage structure may be chosen, based on the means used to access the stored information. A variety of data processor programs and formats can be used for storage, e.g. word processing text file, database format, etc. 11 2015227398 15 Sep 2015 [0057] A ‘ ‘processor” or “computing means” references any hardware and/or software combination that will perform the functions required of it. For example, a suitable processor may be a programmable digital microprocessor such as available in the form of an electronic controller, mainframe, server or personal computer (desktop or portable). Where the processor is programmable, suitable programming can be communicated from a remote location to the processor, or previously saved in a computer program product (such as a portable or fixed computer readable storage medium, whether magnetic, optical or solid state device based). For example, a magnetic medium or optical disk may carry the programming, and can be read by a suitable reader communicating with each processor at its corresponding station.

[0058] As used herein, the terms “active surveillance” and “watchful waiting” mean closely monitoring a patient’s condition without giving any treatment until symptoms appear or change. For example, in prostate cancer, watchful waiting is usually used in older men with other medical problems and early-stage disease.

[0059] As used herein, the term “surgery” applies to surgical methods undertaken for removal of cancerous tissue, including pelvic lymphadenectomy, radical prostatectomy, transurethral resection of the prostate (TURP), excision, dissection, and tumor biopsy/removal. The tumor tissue or sections used for gene expression analysis may have been obtained from any of these methods.

[0060] As used herein, the term “therapy” includes radiation, hormonal therapy, cryosurgery, chemotherapy, biologic therapy, and high-intensity focused ultrasound.

[0061] As used herein, the term “TMPRSS fusion” and “TMPRSS2 fusion” are used interchangeably and refer to a fusion of the androgen-driven TMPRSS2 gene with the ERG oncogene, which has been demonstrated to have a significant association with prostate cancer. S. Perner, et al., Urologe A. 46(7):754-760 (2007); S.A. Narod, et al., Br J Cancer 99(6):847-851 (2008). As used herein, positive TMPRSS fusion status indicates that the TMPRSS fusion is present in a tissue sample, whereas negative TMPRSS fusion status indicates that the TMPRSS fusion is not present in a tissue sample. Experts skilled in the art will recognize that there are numerous ways to determine TMPRSS fusion status, such as real-time, quantitative PCR or high-throughput sequencing. See, e.g., K. Mertz, et al., Neoplasis 9(3):200-206 (2007); C. Maher, Nature 458(7234):97-101 (2009). 12 2015227398 15 Sep 2015

Gene Expression Methods Using Genes, Gene Subsets, and microRNAs [0062] The present disclosure provides molecular assays that involve measurement of expression level(s) of one or more genes, gene subsets, microRNAs, or one or more microRNAs in combination with one or more genes or gene subsets, from a biological sample obtained from a prostate cancer patient, and analysis of the measured expression levels to provide information concerning the likelihood of cancer recurrence.

[0063] The present disclosure further provides methods to classify a prostate tumor based on expression level(s) of one or more genes and/or microRNAs. The disclosure further provides genes and/or microRNAs that are associated, positively or negatively, with a particular prognostic outcome. In exemplary embodiments, the clinical outcomes include cRFI and bRFI.

In another embodiment, patients may be classified in risk groups based on the expression level(s) of one or more genes and/or microRNAs that are associated, positively or negatively, with a prognostic factor. In an exemplary embodiment, that prognostic factor is Gleason pattern.

[0064] Various technological approaches for determination of expression levels of the disclosed genes and microRNAs are set forth in this specification, including, without limitation, RT-PCR, microarrays, high-throughput sequencing, serial analysis of gene expression (SAGE) and Digital Gene Expression (DGE), which will be discussed in detail below. In particular aspects, the expression level of each gene or microRNA may be determined in relation to various features of the expression products of the gene including exons, introns, protein epitopes and protein activity.

[0065] The expression level(s) of one or more genes and/or microRNAs may be measured in tumor tissue. For example, the tumor tissue may obtained upon surgical removal or resection of the tumor, or by tumor biopsy. The tumor tissue may be or include histologically “normal” tissue, for example histologically “normal” tissue adjacent to a tumor. The expression level of genes and/or microRNAs may also be measured in tumor cells recovered from sites distant from the tumor, for example circulating tumor cells, body fluid (e.g., urine, blood, blood fraction, etc.).

[0066] The expression product that is assayed can be, for example, RNA or a polypeptide. The expression product may be fragmented. For example, the assay may use primers that are complementary to target sequences of an expression product and could thus 13 2015227398 15 Sep 2015 measure full transcripts as well as those fragmented expression products containing the target sequence. Further information is provided in Table A (inserted in specification prior to claims).

[0067] The RNA expression product may be assayed directly or by detection of a cDNA product resulting from a PCR-based amplification method, e.g., quantitative reverse transcription polymerase chain reaction (qRT-PCR). (See e.g., U.S. Patent No. 7,587,279). Polypeptide expression product may be assayed using immunohistochemistry (IHC). Further, both RNA and polypeptide expression products may also be is assayed using microarrays.

Clinical Utility [0068] Prostate cancer is currently diagnosed using a digital rectal exam (DRE) and Prostate-specific antigen (PSA) test. If PSA results are high, patients will generally undergo a prostate tissue biopsy. The pathologist will review the biopsy samples to check for cancer cells and determine a Gleason score. Based on the Gleason score, PSA, clinical stage, and other factors, the physician must make a decision whether to monitor the patient, or treat the patient with surgery and therapy.

[0069] At present, clinical decision-making in early stage prostate cancer is governed by certain histopathologic and clinical factors. These include: (1) tumor factors, such as clinical stage (e.g. ΤΙ, T2), PSA level at presentation, and Gleason grade, that are very strong prognostic factors in determining outcome; and (2) host factors, such as age at diagnosis and co-morbidity. Because of these factors, the most clinically useful means of stratifying patients with localized disease according to prognosis has been through multifactorial staging, using the clinical stage, the serum PSA level, and tumor grade (Gleason grade) together. In the 2007 updated American Urological Association (AUA) guidelines for management of clinically localized prostate cancer, these parameters have been grouped to determine whether a patient is at low, intermediate, or high risk of biochemical (PSA) relapse after local therapy. I. Thompson, et al., Guideline for the management of clinically localized prostate cancer, J Urol. 177(6):2106-31 (2007).

[0070] Although such classifications have proven to be helpful in distinguishing patients with localized disease who may need adjuvant therapy after surgery/radiation, they have less ability to discriminate between indolent cancers, which do not need to be treated with local therapy, and aggressive tumors, which require local therapy. In fact, these algorithms are of increasingly limited use for deciding between conservative management and definitive therapy 14 2015227398 15 Sep 2015 because the bulk of prostate cancers diagnosed in the PSA screening era now present with clinical stage Tic and PSA <10 ng/mL.

[0071] Patients with T1 prostate cancer have disease that is not clinically apparent but is discovered either at transurethral resection of the prostate (TURP, Tla, Tib) or at biopsy performed because of an elevated PSA (> 4 ng/mL, Tic). Approximately 80% of the cases presenting in 2007 are clinical T1 at diagnosis. In a Scandinavian trial, OS at 10 years was 85% for patients with early stage prostate cancer (T1/T2) and Gleason score < 7, after radical prostatectomy.

[0072] Patients with T2 prostate cancer have disease that is clinically evident and is organ confined; patients with T3 tumors have disease that has penetrated the prostatic capsule and/or has invaded the seminal vesicles. It is known from surgical series that clinical staging underestimates pathological stage, so that about 20% of patients who are clinically T2 will be pT3 after prostatectomy. Most of patients with T2 or T3 prostate cancer are treated with local therapy, either prostatectomy or radiation. The data from the Scandinavian trial suggest that for T2 patients with Gleason grade <7, the effect of prostatectomy on survival is at most 5% at 10 years; the majority of patients do not benefit from surgical treatment at the time of diagnosis. For T2 patients with Gleason > 7 or for T3 patients, the treatment effect of prostatectomy is assumed to be significant but has not been determined in randomized trials. It is known that these patients have a significant risk (10-30%) of recurrence at 10 years after local treatment, however, there are no prospective randomized trials that define the optimal local treatment (radical prostatectomy, radiation) at diagnosis, which patients are likely to benefit from neo-adjuvant/adjuvant androgen deprivation therapy, and whether treatment (androgen deprivation, chemotherapy) at the time of biochemical failure (elevated PSA) has any clinical benefit.

[0073] Accurately determining Gleason scores from needle biopsies presents several technical challenges. First, interpreting histology that is "borderline" between Gleason pattern is highly subjective, even for urologic pathologists. Second, incomplete biopsy sampling is yet another reason why the “predicted” Gleason score on biopsy does not always correlate with the actual “observed” Gleason score of the prostate cancer in the gland itself. Hence, the accuracy of Gleason scoring is dependent upon not only the expertise of the pathologist reading the slides, but also on the completeness and adequacy of the prostate biopsy sampling strategy. T. Stamey, Urology 45:2-12 (1995). The gene/microRNA expression assay and associated information 15 2015227398 15 Sep 2015 provided by the practice of the methods disclosed herein provide a molecular assay method to facilitate optimal treatment decision-making in early stage prostate cancer. An exemplary embodiment provides genes and microRNAs, the expression levels of which are associated (positively or negatively) with prostate cancer recurrence. For example, such a clinical tool would enable physicians to identify T2/T3 patients who are likely to recur following definitive therapy and need adjuvant treatment.

[0074] In addition, the methods disclosed herein may allow physicians to classify tumors, at a molecular level, based on expression level(s) of one or more genes and/or microRNAs that are significantly associated with prognostic factors, such as Gleason pattern and TMPRSS fusion status. These methods would not be impacted by the technical difficulties of intra-patient variability, histologically determining Gleason pattern in biopsy samples, or inclusion of histologically normal appearing tissue adjacent to tumor tissue. Multi-analyte gene/microRNA expression tests can be used to measure the expression level of one or more genes and/or microRNAs involved in each of several relevant physiologic processes or component cellular characteristics. The methods disclosed herein may group the genes and/or microRNAs. The grouping of genes and microRNAs may be performed at least in part based on knowledge of the contribution of those genes and/or microRNAs according to physiologic functions or component cellular characteristics, such as in the groups discussed above. Furthermore, one or more microRNAs may be combined with one or moregenes. The gene-microRNA combination may be selected based on the likelihood that the gene-microRNA combination functionally interact. The formation of groups (or gene subsets), in addition, can facilitate the mathematical weighting of the contribution of various expression levels to cancer recurrence. The weighting of a gene/microRNA group representing a physiological process or component cellular characteristic can reflect the contribution of that process or characteristic to the pathology of the cancer and clinical outcome.

[0075] Optionally, the methods disclosed may be used to classify patients by risk, for example risk of recurrence. Patients can be partitioned into subgroups (e.g., tertiles or quartiles) and the values chosen will define subgroups of patients with respectively greater or lesser risk.

[0076] The utility of a disclosed gene marker in predicting prognosis may not be unique to that marker. An alternative marker having an expression pattern that is parallel to that of a disclosed gene may be substituted for, or used in addition to, that co-expressed gene or 16 2015227398 15 Sep 2015 microRNA. Due to the co-expression of such genes or microRNAs, substitution of expression level values should have little impact on the overall utility of the test. The closely similar expression patterns of two genes or microRNAs may result from involvement of both genes or microRNAs in the same process and/or being under common regulatory control in prostate tumor cells. The present disclosure thus contemplates the use of such co-expressed genes,gene subsets, or microRNAs as substitutes for, or in addition to, genes of the present disclosure.

Methods of Assaying Expression Levels of a Gene Product [0077] The methods and compositions of the present disclosure will employ, unless otherwise indicated, conventional techniques of molecular biology (including recombinant techniques), microbiology, cell biology, and biochemistry, which are within the skill of the art. Exemplary techniques are explained fully in the literature, such as, “Molecular Cloning: A Laboratory Manual”, 2nd edition (Sambrook et al., 1989); “Oligonucleotide Synthesis” (M.J. Gait, ed., 1984); “Animal Cell Culture” (R.I. Freshney, ed., 1987); “Methods in Enzymology” (Academic Press, Inc.); “Handbook of Experimental Immunology”, 4th edition (D.M. Weir &amp; C.C. Blackwell, eds., Blackwell Science Inc., 1987); “Gene Transfer Vectors for Mammalian Cells” (J.M. Miller &amp; M.P. Calos, eds., 1987); “Current Protocols in Molecular Biology” (F.M. Ausubel et al., eds., 1987); and “PCR: The Polymerase Chain Reaction”, (Mullis et al., eds., 1994).

[0078] Methods of gene expression profiling include methods based on hybridization analysis of polynucleotides, methods based on sequencing of polynucleotides, and proteomics-based methods. Exemplary methods known in the art for the quantification of RNA expression in a sample include northern blotting and in situ hybridization (Parker &amp; Barnes, Methods in Molecular Biology 106:247-283 (1999)); RNAse protection assays (Hod, Biotechniques 13:852-854 (1992)); and PCR-based methods, such as reverse transcription PCT (RT-PCR) (Weis et al., Trends in Genetics 8:263-264 (1992)). Antibodies may be employed that can recognize sequence-specific duplexes, including DNA duplexes, RNA duplexes, and DNA-RNA hybrid duplexes or DNA-protein duplexes. Representative methods for sequencing-based gene expression analysis include Serial Analysis of Gene Expression (SAGE), and gene expression analysis by massively parallel signature sequencing (MPSS). 17 2015227398 15 Sep 2015

Reverse Transcriptase PCR (RT-PCR) [0079] Typically, mRNA or microRNA is isolated from a test sample. The starting material is typically total RNA isolated from a human tumor, usually from a primary tumor. Optionally, normal tissues from the same patient can be used as an internal control. Such normal tissue can be histologically-appearing normal tissue adjacent a tumor. mRNA or microRNA can be extracted from a tissue sample, e.g., from a sample that is fresh, frozen (e.g. fresh frozen), or paraffin-embedded and fixed (e.g. formalin-fixed).

[0080] General methods for mRNA and microRNA extraction are well known in the art and are disclosed in standard textbooks of molecular biology, including Ausubel et al., Current Protocols of Molecular Biology, John Wiley and Sons (1997). Methods for RNA extraction from paraffin embedded tissues are disclosed, for example, in Rupp and Locker, Lab Invest. 56:A67 (1987), and De Andres et al., BioTechniques 18:42044 (1995). In particular, RNA isolation can be performed using a purification kit, buffer set and protease from commercial manufacturers, such as Qiagen, according to the manufacturer’s instructions. For example, total RNA from cells in culture can be isolated using Qiagen RNeasy mini-columns. Other commercially available RNA isolation kits include MasterPure™ Complete DNA and RNA Purification Kit (EPICENTRE®, Madison, WI), and Paraffin Block RNA Isolation Kit (Ambion, Inc.). Total RNA from tissue samples can be isolated using RNA Stat-60 (Tel-Test). RNA prepared from tumor can be isolated, for example, by cesium chloride density gradient centrifugation.

[0081] The sample containing the RNA is then subjected to reverse transcription to produce cDNA from the RNA template, followed by exponential amplification in a PCR reaction. The two most commonly used reverse transcriptases are avilo myeloblastosis virus reverse transcriptase (AMV-RT) and Moloney murine leukemia virus reverse transcriptase (MMLV-RT). The reverse transcription step is typically primed using specific primers, random hexamers, or oligo-dT primers, depending on the circumstances and the goal of expression profiling. For example, extracted RNA can be reverse-transcribed using a GeneAmp RNA PCR kit (Perkin Elmer, CA, USA), following the manufacturer’s instructions. The derived cDNA can then be used as a template in the subsequent PCR reaction.

[0082] PCR-based methods use a thermostable DNA-dependent DNA polymerase, such as a Taq DNA polymerase. For example, TaqMan® PCR typically utilizes the 5’-nuclease activity of Taq or Tth polymerase to hydrolyze a hybridization probe bound to its target 18 2015227398 15 Sep 2015 amplicon, but any enzyme with equivalent 5’ nuclease activity can be used. Two oligonucleotide primers are used to generate an amplicon typical of a PCR reaction product. A third oligonucleotide, or probe, can be designed to facilitate detection of a nucleotide sequence of the amplicon located between the hybridization sites the two PCR primers. The probe can be detectably labeled, e.g., with a reporter dye, and can further be provided with both a fluorescent dye, and a quencher fluorescent dye, as in a Taqman® probe configuration. Where a Taqman® probe is used, during the amplification reaction, the Taq DNA polymerase enzyme cleaves the probe in a template-dependent manner. The resultant probe fragments disassociate in solution, and signal from the released reporter dye is free from the quenching effect of the second fluorophore. One molecule of reporter dye is liberated for each new molecule synthesized, and detection of the unquenched reporter dye provides the basis for quantitative interpretation of the data.

[0083] TaqMan® RT-PCR can be performed using commercially available equipment, such as, for example, high-throughput platforms such as the ABI PRISM 7700 Sequence Detection System® (Perkin-Elmer-Applied Biosystems, Foster City, CA, USA), or Lightcycler (Roche Molecular Biochemicals, Mannheim, Germany). In a preferred embodiment, the procedure is run on a LightCycler® 480 (Roche Diagnostics) real-time PCR system, which is a microwell plate-based cycler platform.

[0084] 5'-Nuclease assay data are commonly initially expressed as a threshold cycle (“CT”)· Fluorescence values are recorded during every cycle and represent the amount of product amplified to that point in the amplification reaction. The threshold cycle (Ct) is generally described as the point when the fluorescent signal is first recorded as statistically significant. Alternatively, data may be expressed as a crossing point ( “Cp”). The Cp value is calculated by determining the second derivatives of entire qPCR amplification curves and their maximum value. The Cp value represents the cycle at which the increase of fluorescence is highest and where the logarithmic phase of a PCR begins.

[0085] To minimize errors and the effect of sample-to-sample variation, RT-PCR is usually performed using an internal standard. The ideal internal standard gene (also referred to as a reference gene) is expressed at a quite constant level among cancerous and non-cancerous tissue of the same origin (i.e., a level that is not significantly different among normal and cancerous tissues), and is not significantly affected by the experimental treatment (i.e., does not 19 2015227398 15 Sep 2015 exhibit a significant difference in expression level in the relevant tissue as a result of exposure to chemotherapy), and expressed at a quite constant level among the same tissue taken from different patients. For example, reference genes useful in the methods disclosed herein should not exhibit significantly different expression levels in cancerous prostate as compared to normal prostate tissue. RNAs frequently used to normalize patterns of gene expression are mRNAs for the housekeeping genes glyceraldehyde-3-phosphate-dehydrogenase (GAPDH) and β-actin. Exemplary reference genes used for normalization comprise one or more of the following genes: AAMP, ARF1, ATP5E, CLTC, GPS1, and PGK1. Gene expression measurements can be normalized relative to the mean of one or more (e.g., 2, 3, 4, 5, or more) reference genes. Reference-normalized expression measurements can range from 2 to 15, where a one unit increase generally reflects a 2-fold increase in RNA quantity.

[0086] Real time PCR is compatible both with quantitative competitive PCR, where internal competitor for each target sequence is used for normalization, and with quantitative comparative PCR using a normalization gene contained within the sample, or a housekeeping gene for RT-PCR. For further details see, e.g. Held et al., Genome Research 6:986-994 (1996).

[0087] The steps of a representative protocol for use in the methods of the present disclosure use fixed, paraffin-embedded tissues as the RNA source. For example, mRNA isolation, purification, primer extension and amplification can be performed according to methods available in the art. (see, e.g., Godfrey et al. J. Molec. Diagnostics 2: 84-91 (2000); Specht et al., Am. J. Pathol. 158: 419-29 (2001)). Briefly, a representative process starts with cutting about 10 μιη thick sections of paraffin-embedded tumor tissue samples. The RNA is then extracted, and protein and DNA depleted from the RNA-containing sample. After analysis of the RNA concentration, RNA is reverse transcribed using gene specific primers followed by RT-PCR to provide for cDNA amplification products.

Design of Intron-Based PCR Primers and Probes [0088] PCR primers and probes can be designed based upon exon or intron sequences present in the mRNA transcript of the gene of interest. Primer/probe design can be performed using publicly available software, such as the DNA BLAT software developed by Kent, W.J., Genome Res. 12(4):656-64 (2002), or by the BLAST software including its variations.

[0089] Where necessary or desired, repetitive sequences of the target sequence can be masked to mitigate non-specific signals. Exemplary tools to accomplish this include the Repeat 20 2015227398 15 Sep 2015

Masker program available on-line through the Baylor College of Medicine, which screens DNA sequences against a library of repetitive elements and returns a query sequence in which the repetitive elements are masked. The masked intron sequences can then be used to design primer and probe sequences using any commercially or otherwise publicly available primer/probe design packages, such as Primer Express (Applied Biosystems); MGB assay-by-design (Applied Biosystems); Primer3 (Steve Rozen and Helen J. Skaletsky (2000) Primer3 on the WWW for general users and for biologist programmers. See S. Rrawetz, S. Misener, Bioinformatics Methods and Protocols: Methods in Molecular Biology, pp. 365-386 (Humana Press).

[0090] Other factors that can influence PCR primer design include primer length, melting temperature (Tm), and G/C content, specificity, complementary primer sequences, and 3 '-end sequence. In general, optimal PCR primers are generally 17-30 bases in length, and contain about 20-80%, such as, for example, about 50-60% G+C bases, and exhibit Tm's between 50 and 80 0C, e.g. about 50 to 70 0C.

[0091] For further guidelines for PCR primer and probe design see, e.g. Dieffenbach, CW. et al, “General Concepts for PCR Primer Design” in: PCR Primer, A Laboratory Manual, Cold Spring Harbor Laboratory Press,. New York, 1995, pp. 133-155; Innis and Gelfand, “Optimization of PCRs” in: PCR Protocols, A Guide to Methods and Applications, CRC Press, London, 1994, pp. 5-11; and Plasterer, T.N. Primerselect: Primer and probe design. Methods Mol. Biol. 70:520-527 (1997), the entire disclosures of which are hereby expressly incorporated by reference.

[0092] Table A provides further information concerning the primer, probe, and amplicon sequences associated with the Examples disclosed herein.

MassARRAY® System [0093] In Mass ARRAY-based methods, such as the exemplary method developed by Sequenom, Inc. (San Diego, CA) following the isolation of RNA and reverse transcription, the obtained cDNA is spiked with a synthetic DNA molecule (competitor), which matches the targeted cDNA region in all positions, except a single base, and serves as an internal standard. The cDNA/competitor mixture is PCR amplified and is subjected to a post-PCR shrimp alkaline phosphatase (SAP) enzyme treatment, which results in the dephosphorylation of the remaining nucleotides. After inactivarion of the alkaline phosphatase, the PCR products from the competitor and cDNA are subjected to primer extension, which generates distinct mass signals 21 2015227398 15 Sep 2015 for the competitor- and cDNA-derives PCR products. After purification, these products are dispensed on a chip array, which is pre-loaded with components needed for analysis with matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) analysis. The cDNA present in the reaction is then quantified by analyzing the ratios of the peak areas in the mass spectrum generated. For further details see, e.g. Ding and Cantor, Proc. Natl. Acad. Sci. USA 100:3059-3064 (2003).

Other PCR-based Methods [0094] Further PCR-based techniques that can find use in the methods disclosed herein include, for example, BeadArray® technology (Illumina, San Diego, CA; Oliphant et al., Discovery of Markers for Disease (Supplement to Biotechniques), June 2002; Ferguson et al., Analytical Chemistry 72:5618 (2000)); BeadsArray for Detection of Gene Expression® (BADGE), using the commercially available LuminexlOO LabMAP® system and multiple color-coded microspheres (Luminex Corp., Austin, TX) in a rapid assay for gene expression (Yang et al., Genome Res. 11:1888-1898 (2001)); and high coverage expression profiling (HiCEP) analysis (Fukumura et al., Nucl. Acids. Res. 31(16) e94 (2003).

Microarrays [0095] Expression levels of a gene or microArray of interest can also be assessed using the microarray technique. In this method, polynucleotide sequences of interest (including cDNAs and oligonucleotides) are arrayed on a substrate. The arrayed sequences are then contacted under conditions suitable for specific hybridization with detectably labeled cDNA generated from RNA of a test sample. As in the RT-PCR method, the source of RNA typically is total RNA isolated from a tumor sample, and optionally from normal tissue of the same patient as an internal control or cell lines. RNA can be extracted, for example, from frozen or archived paraffin-embedded and fixed (e.g. formalin-fixed) tissue samples.

[0096] For example, PCR amplified inserts of cDNA clones of a gene to be assayed are applied to a substrate in a dense array. Usually at least 10,000 nucleotide sequences are applied to the substrate. For example, the microarrayed genes, immobilized on the microchip at 10,000 elements each, are suitable for hybridization under stringent conditions. Fluorescently labeled cDNA probes may be generated through incorporation of fluorescent nucleotides by reverse transcription of RNA extracted from tissues of interest. Labeled cDNA probes applied to the chip hybridize with specificity to each spot of DNA on the array. After washing under stringent 22 2015227398 15 Sep 2015 conditions to remove non-specifically bound probes, the chip is scanned by confocal laser microscopy or by another detection method, such as a CCD camera. Quantitation of hybridization of each arrayed element allows for assessment of corresponding RNA abundance.

[0097] With dual color fluorescence, separately labeled cDNA probes generated from two sources of RNA are hybridized pair wise to the array. The relative abundance of the transcripts from the two sources corresponding to each specified gene is thus determined simultaneously. The miniaturized scale of the hybridization affords a convenient and rapid evaluation of the expression pattern for large numbers of genes. Such methods have been shown to have the sensitivity required to detect rare transcripts, which are expressed at a few copies per cell, and to reproducibly detect at least approximately two-fold differences in the expression levels (Schena et at, Proc. Natl. Acad. ScL USA 93(2):106-149 (1996)). Microarray analysis can be performed by commercially available equipment, following manufacturer's protocols, such as by using the Affymetrix GenChip® technology, or Incyte's microarray technology.

Serial Analysis of Gene Expression (SAGE) [0098] Serial analysis of gene expression (SAGE) is a method that allows the simultaneous and quantitative analysis of a large number of gene transcripts, without the need of providing an individual hybridization probe for each transcript. First, a short sequence tag (about 10-14 bp) is generated that contains sufficient information to uniquely identify a transcript, provided that the tag is obtained from a unique position within each transcript. Then, many transcripts are linked together to form long serial molecules, that can be sequenced, revealing the identity of the multiple tags simultaneously. The expression pattern of any population of transcripts can be quantitatively evaluated by determining the abundance of individual tags, and identifying the gene corresponding to each tag. For more details see, e.g. Velculescu et al., Science 270:484-487 (1995); and Velculescu et al., Cell 88:243-51 (1997).

Gene Expression Analysis by Nucleic Acid Sequencing [0099] Nucleic acid sequencing technologies are suitable methods for analysis of gene expression. The principle underlying these methods is that the number of times a cDNA sequence is detected in a sample is directly related to the relative expression of the RNA corresponding to that sequence. These methods are sometimes referred to by the term Digital Gene Expression (DGE) to reflect the discrete numeric property of the resulting data. Early methods applying this principle were Serial Analysis of Gene Expression (SAGE) and Massively 23 2015227398 15 Sep 2015

Parallel Signature Sequencing (MPSS). See, e.g., S. Brenner, et al., Nature Biotechnology 18(6):630-634 (2000). More recently, the advent of “next-generation” sequencing technologies has made DGE simpler, higher throughput, and more affordable. As a result, more laboratories are able to utilize DGE to screen the expression of more genes in more individual patient samples than previously possible. See, e.g., J. Marioni, Genome Research 18(9):1509-1517 (2008); R. Morin, Genome Research 18(4):610-621 (2008); A. Mortazavi, Nature Methods 5(7):621-628 (2008); N. Cloonan, Nature Methods 5(7):613-619 (2008).

Isolating RNA from Body Fluids [00100] Methods of isolating RNA for expression analysis from blood, plasma and serum (see, e.g., K. Enders, et al., Clin Chem 48,1647-53 (2002) (and references cited therein) and from urine (see, e.g., R. Boom, et al., J Clin Microbiol. 28, 495-503 (1990) and references cited therein) have been described.

Immunohistochemistrv [00101] Immunohistochemistry methods are also suitable for detecting the expression levels of genes and applied to the method disclosed herein. Antibodies (e.g., monoclonal antibodies) that specifically bind a gene product of a gene of interest can be used in such methods. The antibodies can be detected by direct labeling of the antibodies themselves, for example, with radioactive labels, fluorescent labels, hapten' labels such as, biotin, or an enzyme such as horse radish peroxidase or alkaline phosphatase. Alternatively, unlabeled primary antibody can be used in conjunction with a labeled secondary antibody specific for the primary antibody. Immunohistochemistry protocols and kits are well known in the art and are commercially available.

Proteomics [00102] The term “proteome” is defined as the totality of the proteins present in a sample (e.g. tissue, organism, or cell culture) at a certain point of time. Proteomics includes, among other things, study of the global changes of protein expression in a sample (also referred to as “expression proteomics”). Proteomics typically includes the following steps: (1) separation of individual proteins in a sample by 2-D gel electrophoresis (2-D PAGE); (2) identification of the individual proteins recovered from the gel, e.g. my mass spectrometry or N- terminal sequencing, and (3) analysis of the data using bioinformatics. 24 2015227398 15 Sep 2015

General Description of the mRNA/microRNA Isolation, Purification and Amplification [00103] The steps of a representative protocol for profiling gene expression using fixed, paraffin-embedded tissues as the RNA source, including mRNA or microRNA isolation, purification, primer extension and amplification are provided in various published journal articles. (See, e.g., T.E. Godfrey, et al,. J. Molec. Diagnostics 2: 84-91 (2000); K. Specht et al., Am. J. Pathol. 158: 419-29 (2001), M. Cronin, et al., Am J Pathol 164:35-42 (2004)). Briefly, a representative process starts with cutting a tissue sample section (e.g.about 10 pm thick sections of a paraffin-embedded tumor tissue sample). The RNA is then extracted, and protein and DNA are removed. After analysis of the RNA concentration, RNA repair is performed if desired. The sample can then be subjected to analysis, e.g., by reverse transcribed using gene specific promoters followed by RT-PCR.

Statistical Analysis Of Expression Levels in Identification of Genes and MicroRNAs [00104] One skilled in the art will recognize that there are many statistical methods that may be used to determine whether there is a significant relationship between a parameter of interest (e.g., recurrence) and expression levels of a marker gene/microRNA as described here.

In an exemplary embodiment, the present invention provides a stratified cohort sampling design (a form of case-control sampling) using tissue and data from prostate cancer patients. Selection of specimens was stratified by T stage (ΤΙ, T2), year cohort (<1993, >1993), and prostatectomy Gleason Score (low/intermediate, high). All patients with clinical recurrence were selected and a sample of patients who did not experience a clinical recurrence was selected. For each patient, up to two enriched tumor specimens and one normal-appearing tissue sample was assayed.

[00105] All hypothesis tests were reported using two-sided p-values. To investigate if there is a significant relationship of outcomes (clinical recurrence-free interval (cRFI), biochemical recurrence-free interval (bRFI), prostate cancer-specific survival (PCSS), and overall survival (OS)) with individual genes and/or microRNAs, demographic or clinical covariates Cox Proportional Hazards (PH) models using maximum weighted pseudo partial-likelihood estimators were used and p-values from Wald tests of the null hypothesis that the hazard ratio (HR) is one are reported. To investigate if there is a significant relationship between individual genes and/or microRNAs and Gleason pattern of a particular sample, ordinal logistic 25 2015227398 15 Sep 2015 regression models using maximum weighted likelihood methods were used and p-values from Wald tests of the null hypothesis that the odds ratio (OR) is one are reported.

Coexpression Analysis [00106] The present disclosure provides a method to determine tumor stage based on the expression of staging genes, or genes that co-express with particular staging genes. To perform particular biological processes, genes often work together in a concerted way, i.e. they are coexpressed. Co-expressed gene groups identified for a disease process like cancer can serve as biomarkers for tumor status and disease progression. Such co-expressed genes can be assayed in lieu of, or in addition to, assaying of the staging gene with which they are co-expressed.

[00107] In an exemplary embodiment, the joint correlation of gene expression levels among prostate cancer specimens under study may be assessed. For this purpose, the correlation structures among genes and specimens may be examined through hierarchical cluster methods. This information may be used to confirm that genes that are known to be highly correlated in prostate cancer specimens cluster together as expected. Only genes exhibiting a nominally significant (unadjusted p < 0.05) relationship with cRFI in the univariate Cox PH regression analysis will be included in these analyses.

[00108] One skilled in the art will recognize that many co-expression analysis methods now known or later developed will fall within the scope and spirit of the present invention. These methods may incorporate, for example, correlation coefficients, co-expression network analysis, clique analysis, etc., and may be based on expression data from RT-PCR, microarrays, sequencing, and other similar technologies. For example, gene expression clusters can be identified using pair-wise analysis of correlation based on Pearson or Spearman correlation coefficients. (See, e.g., Pearson K. and Lee A., Biometrika 2, 357 (1902); C. Spearman, Amer. J. Psychol 15:72-101 (1904); J. Myers, A. Well, Research Design and Statistical Analysis, p. 508 (2nd Ed., 2003).)

Normalization of Expression Levels [00109] The expression data used in the methods disclosed herein can be normalized. Normalization refers to a process to correct for (normalize away), for example, differences in the amount of RNA assayed and variability in the quality of the RNA used, to remove unwanted sources of systematic variation in Ct or Cp measurements, and the like. With respect to RT-PCR experiments involving archived fixed paraffin embedded tissue samples, sources of systematic 26 2015227398 15 Sep 2015 variation are known to include the degree of RNA degradation relative to the age of the patient sample and the type of fixative used to store the sample. Other sources of systematic variation are attributable to laboratory processing conditions.

[00110] Assays can provide for normalization by incorporating the expression of certain normalizing genes, which do not significantly differ in expression levels under the relevant conditions. Exemplary normalization genes disclosed herein include housekeeping genes. (See, e.g., E. Eisenberg, et al., Trends in Genetics 19(7):362-365 (2003).) Normalization can be based on the mean or median signal (Ct or Cp) of all of the assayed genes or a large subset thereof (global normalization approach). In general, the normalizing genes, also referred to as reference genes should be genes that are known not to exhibit significantly different expression in prostate cancer as compared to non-cancerous prostate tissue, and are not significantly affected by various sample and process conditions, thus provide for normalizing away extraneous effects.

[00111] In exemplary embodiments, one or more of the following genes are used as references by which the mRNA or microRNA expression data is normalized: AAMP, ARF1, ATP5E, CLTC, GPS1, and PGK1. In another exemplary embodiment, one or more of the following microRNAs are used as references by which the expression data of microRNAs are normalized: hsa-miR-106a; hsa-miR-146b-5p; hsa-miR-191; hsa-miR-19b; and hsa-miR-92a. The calibrated weighted average Ct or Cp measurements for each of the prognostic and predictive genes or microRNAs may be normalized relative to the mean of five or more reference genes or microRNAs.

[00112] Those skilled in the art will recognize that normalization may be achieved in numerous ways, and the techniques described above are intended only to be exemplary, not exhaustive.

Standardization of Expression Levels [00113] The expression data used in the methods disclosed herein can be standardized. Standardization refers to a process to effectively put all the genes or microRNAs on a comparable scale. This is performed because some genes or microRNAs will exhibit more variation (a broader range of expression) than others. Standardization is performed by dividing each expression value by its standard deviation across all samples for that gene or microRNA. Hazard ratios are then interpreted as the relative risk of recurrence per 1 standard deviation increase in expression. 27 2015227398 15 Sep 2015

Kits of the Invention [00114] The materials for use in the methods of the present invention are suited for preparation of kits produced in accordance with well-known procedures. The present disclosure thus provides kits comprising agents, which may include gene (or microRNA)-specific or gene (or microRNA)-selective probes and/or primers, for quantifying the expression of the disclosed genes or microRNAs for predicting prognostic outcome or response to treatment. Such kits may optionally contain reagents for the extraction of RNA from tumor samples, in particular fixed paraffin-embedded tissue samples and/or reagents for RNA amplification. In addition, the kits may optionally comprise the reagent(s) with an identifying description or label or instructions relating to their use in the methods of the present invention. The kits may comprise containers (including microliter plates suitable for use in an automated implementation of the method), each with one or more of the various materials or reagents (typically in concentrated form) utilized in the methods, including, for example, chromatographic columns, pre-fabricated microarrays, buffers, the appropriate nucleotide triphosphates (e.g., dATP, dCTP, dGTP and dTTP; or rATP, rCTP, rGTP and UTP), reverse transcriptase, DNA polymerase, RNA polymerase, and one or more probes and primers of the present invention (e.g., appropriate length poly(T) or random primers linked to a promoter reactive with the RNA polymerase). Mathematical algorithms used to estimate or quantify prognostic or predictive information are also properly potential components of kits.

Reports [00115] The methods of this invention, when practiced for commercial diagnostic purposes, generally produce a report or summary of information obtained from the herein-described methods. For example, a report may include information concerning expression levels of one or more genes and /or microRNAs, classification of the tumor or the patient’s risk of recurrence, the patient’s likely prognosis or risk classification, clinical and pathologic factors, and/or other information. The methods and reports of this invention can further include storing the report in a database. The method can create a record in a database for the subject and populate the record with data. The report may be a paper report, an auditory report, or an electronic record. The report may be displayed and/or stored on a computing device (e.g., handheld device, desktop computer, smart device, website, etc.). It is contemplated that the report is provided to a physician and/or the patient. The receiving of the report can further 28 2015227398 15 Sep 2015 include establishing a network connection to a server computer that includes the data and report and requesting the data and report from the server computer.

Computer program [00116] The values from the assays described above, such as expression data, can be calculated and stored manually. Alternatively, the above-described steps can be completely or partially performed by a computer program product. The present invention thus provides a computer program product including a computer readable storage medium having a computer program stored on it. The program can, when read by a computer, execute relevant calculations based on values obtained from analysis of one or more biological sample from an individual (e.g., gene expression levels, normalization, standardization, thresholding, and conversion of values from assays to a score and/or text or graphical depiction of tumor stage and related information). The computer program product has stored therein a computer program for performing the calculation.

[00117] The present disclosure provides systems for executing the program described above, which system generally includes: a) a central computing environment; b) an input device, operatively connected to the computing environment, to receive patient data, wherein the patient data can include, for example, expression level or other value obtained from an assay using a biological sample from the patient, or microarray data, as described in detail above; c) an output device, connected to the computing environment, to provide information to a user (e.g., medical personnel); and d) an algorithm executed by the central computing environment (e.g., a processor), where the algorithm is executed based on the data received by the input device, and wherein the algorithm calculates an expression score, thresholding, or other functions described herein. The methods provided by the present invention may also be automated in whole or in part.

[00118] All aspects of the present invention may also be practiced such that a limited number of additional genes and/or microRNAs that are co-expressed or functionally related with the disclosed genes, for example as evidenced by statistically meaningful Pearson and/or Spearman correlation coefficients, are included in a test in addition to and/or in place of disclosed genes. 29 2015227398 15 Sep 2015 [00119] Having described the invention, the same will be more readily understood through reference to the following Examples, which are provided by way of illustration, and are not intended to limit the invention in any way.

Examples

Example 1: RNA Yield and Gene Expression Profiles in Prostate Cancer Biopsy Cores [00120] Clinical tools based on prostate needle core biopsies are needed to guide treatment planning at diagnosis for men with localized prostate cancer. Limiting tissue in needle core biopsy specimens poses significant challenges to the development of molecular diagnostic tests. This study examined RNA extraction yields and gene expression profiles using an RT-PCR assay to characterize RNA from manually micro-dissected fixed paraffin embedded (FPE) prostate cancer needle biopsy cores. It also investigated the association of RNA yields and gene expression profiles with Gleason score in these specimens.

Patients and Samples [00121] This study determined the feasibility of gene expression profile analysis in prostate cancer needle core biopsies by evaluating the quantity and quality of RNA extracted from fixed paraffin-embedded (FPE) prostate cancer needle core biopsy specimens. Forty-eight (48) formalin-fixed blocks from prostate needle core biopsy specimens were used for this study. Classification of specimens was based on interpretation of the Gleason score (2005 Int’l Society of Urological Pathology Consensus Conference) and percentage tumor (<33%, 33-66%, >66%) involvement as assessed by pathologists.

Table 1: Distribution of cases

Gleason score Category ~<33% Tumor -33-66% Tumor ~>66% Tumor Low (<6) 5 5 6 Intermediate (7) 5 5 6 High (8, 9, 10) 5 5 6 Total 15 15 18

Assay Methods [00122] Fourteen (14) serial 5 pm unstained sections from each FPE tissue block were included in the study. The first and last sections for each case were H&amp;E stained and histologically reviewed to confirm the presence of tumor and for tumor enrichment by manual micro-dissection. 30 2015227398 15 Sep 2015 [00123] RNA from enriched tumor samples was extracted using a manual RNA extraction process. RNA was quantitated using the RiboGreen® assay and tested for the presence of genomic DNA contamination. Samples with sufficient RNA yield and free of genomic DNA tested for gene expression levels of a 24-gene panel of reference and cancer-related genes using quantitative RT-PCR. The expression was normalized to the average of 6 reference genes (AAMP, ARF1, ATP5E, CLTC, EEF1A1, and GPX1).

Statistical Methods [00124] Descriptive statistics and graphical displays were used to summarize standard pathology metrics and gene expression, with stratification for Gleason Score category and percentage tumor involvement category. Ordinal logistic regression was used to evaluate the relationship between gene expression and Gleason Score category.

Results [00125] The RNA yield per unit surface area ranged from 16 to 2406 ng/mm2. Higher RNA yield was observed in samples with higher percent tumor involvement (p=0.02) and higher Gleason score (p=0.01). RNA yield was sufficient (> 200ng) in 71% of cases to permit 96-well RT-PCR, with 87% of cases having >100ng RNA yield. The study confirmed that gene expression from prostate biopsies, as measured by qRT-PCR, was comparable to FPET samples used in commercial molecular assays for breast cancer. In addition, it was observed that greater biopsy RNA yields are found with higher Gleason score and higher percent tumor involvement. Nine genes were identified as significantly associated with Gleason score (p < 0.05) and there was a large dynamic range observed for many test genes.

Example 2; Gene Expression Analysis for Genes Associated with Prognosis in Prostate Cancer

Patients and Samples [00126] Approximately 2600 patients with clinical stage T1/T2 prostate cancer treated with radical prostatectomy (RP) at the Cleveland Clinic between 1987 and 2004 were identified. Patients were excluded from the study design if they received neo-adjuvant and/or adjuvant therapy, if pre-surgical PSA levels were missing, or if no tumor block was available from initial diagnosis. 127 patients with clinical recurrence and 374 patients without clinical recurrence after radical prostatectomy were randomly selected using a cohort sampling design. The specimens were stratified by T stage (ΤΙ, T2), year cohort (<1993, >1993), and prostatectomy Gleason 31 2015227398 15 Sep 2015 score (low/intermediate, high). Of the 501 sampled patients, 51 were excluded for insufficient tumor; 7 were excluded due to clinical ineligibility; 2 were excluded due to poor quality of gene expression data; and 10 were excluded because primary Gleason pattern was unavailable. Thus, this gene expression study included tissue and data from 111 patients with clinical recurrence and 330 patients without clinical recurrence after radical prostatectomies performed between 1987 and 2004 for treatment of early stage (ΤΙ, T2) prostate cancer.

[00127] Two fixed paraffin embedded (FPE) tissue specimens were obtained from prostate tumor specimens in each patient. The sampling method (sampling method A or B) depended on whether the highest Gleason pattern is also the primary Gleason pattern. For each specimen selected, the invasive cancer cells were at least 5.0 mm in dimension, except in the instances of pattern 5, where 2.2 mm was accepted. Specimens were spatially distinct where possible.

Table 2; Sampling Methods

Sampling Method A Sampling Method B For patients whose prostatectomy primary Gleason pattern is also the highest Gleason pattern For patients whose prostatectomy primary Gleason pattern is not the highest Gleason pattern Specimen 1 (Al) = primary Gleason pattern Select and mark largest focus (greatest cross-sectional area) of primary Gleason pattern tissue. Invasive cancer area >5.0 mm. Specimen 1 (Bl) = highest Gleason pattern Select highest Gleason pattern tissue from spatially distinct area from specimen B2, if possible. Invasive cancer area at least 5.0 mm if selecting secondary pattern, at least 2.2 mm if selecting Gleason pattern 5. Specimen 2 (A2) = secondary Gleason pattern Select and mark secondary Gleason pattern tissue from spatially distinct area from specimen Al. Invasive cancer area > 5.0 mm. Specimen 2 (B2) = primary Gleason pattern Select largest focus (greatest cross-sectional area) of primary Gleason pattern tissue. Invasive cancer area >5.0 mm.

[00128] Histologically normal appearing tissue (NAT) adjacent to the tumor specimen (also referred to in these Examples as “non-tumor tissue”) was also evaluated. Adjacent tissue was collected 3 mm from the tumor to 3 mm from the edge of the FPET block. NAT was preferentially sampled adjacent to the primary Gleason pattern. In cases where there was insufficient NAT adjacent to the primary Gleason pattern, then NAT was sampled adjacent to the secondary or highest Gleason pattern (A2 or Bl) per the method set forth in Table 2. Six (6) 10 pm sections with beginning H&amp;E at 5 pm and ending unstained slide at 5 pm were prepared 32 2015227398 15 Sep 2015 from each fixed paraffin-embedded tumor (FPET) block included in the study. All cases were histologically reviewed and manually micro-dissected to yield two enriched tumor samples and, where possible, one normal tissue sample adjacent to the tumor specimen.

Assay Method [00129] In this study, RT-PCR analysis was used to determine RNA expression levels for 738 genes and chromosomal rearrangements (e.g., TMPRSS2-ERG fusion or other ETS family genes) in prostate cancer tissue and surrounding NAT in patients with early-stage prostate cancer treated with radical prostatectomy.

[00130] The samples were quantified using the RiboGreen assay and a subset tested for presence of genomic DNA contamination. Samples were taken into reverse transcription (RT) and quantitative polymerase chain reaction (qPCR). All analyses were conducted on reference-normalized gene expression levels using the average of the of replicate well crossing point (CP) values for the 6 reference genes (AAMP, ARF1, ATP5E, CLTC, GPS1, PGK1).

Statistical Analysis and Results [00131] Primary statistical analyses involved 111 patients with clinical recurrence and 330 patients without clinical recurrence after radical prostatectomy for early-stage prostate cancer stratified by T-stage (ΤΙ, T2), year cohort (<1993, >1993), and prostatectomy Gleason score (low/intermediate, high). Gleason score categories are defined as follows: low (Gleason score < 6), intermediate (Gleason score = 7), and high (Gleason score > 8). A patient was included in a specified analysis if at least one sample for that patient was evaluable. Unless otherwise stated, all hypothesis tests were reported using two-sided p-values. The method of Storey was applied to the resulting set of p-values to control the false discovery rate (FDR) at 20%. J. Storey, R. Tibshirani, Estimating the Positive False Discovery Rate Under Dependence, with Applications to DNA Microarrays, Dept, of Statistics, Stanford Univ. (2001).

[00132] Analysis of gene expression and recurrence-free interval was based on univariate Cox Proportional Hazards (PH) models using maximum weighted pseudo-partial-likelihood estimators for each evaluable gene in the gene list (727 test genes and 5 reference genes). P-values were generated using Wald tests of the null hypothesis that the hazard ratio (HR) is one. Both unadjusted p-values and the q-value (smallest FDR at which the hypothesis test in question is rejected) were reported. Un-adjusted p-values <0.05 were considered statistically significant. Since two tumor specimens were selected for each patient, this analysis was performed using the 33 2015227398 15 Sep 2015 2 specimens from each patient as follows: (1) analysis using the primary Gleason pattern specimen from each patient (Specimens A1 and B2 as described in Table 2); (2) analysis using the highest Gleason pattern specimen from each patient (Specimens A1 and B1 as described in Table 2).

[00133] Analysis of gene expression and Gleason pattern (3, 4, 5) was based on univariate ordinal logistic regression models using weighted maximum likelihood estimators for each gene in the gene list (727 test genes and 5 reference genes). P-values were generated using a Wald test of the null hypothesis that the odds ratio (OR) is one. Both unadjusted p-values and the q-value (smallest FDR at which the hypothesis test in question is rejected) were reported. Un-adjusted p-values <0.05 were considered statistically significant. Since two tumor specimens were selected for each patient, this analysis was performed using the 2 specimens from each patient as follows: (1) analysis using the primary Gleason pattern specimen from each patient (Specimens A1 and B2 as described in Table 2); (2) analysis using the highest Gleason pattern specimen from each patient (Specimens A1 and B1 as described in Table 2).

[00134] It was determined whether there is a significant relationship between cRFI and selected demographic, clinical, and pathology variables, including age, race, clinical tumor stage, pathologic tumor stage, location of selected tumor specimens within the prostate (peripheral versus transitional zone), PSA at the time of surgery, overall Gleason score from the radical prostatectomy, year of surgery, and specimen Gleason pattern. Separately for each demographic or clinical variable, the relationship between the clinical covariate and cRFI was modeled using univariate Cox PH regression using weighted pseudo partial-likelihood estimators and a p-value was generated using Wald’s test of the null hypothesis that the hazard ratio (HR) is one. Covariates with unadjusted p-values <0.2 may have been included in the covariate-adjusted analyses.

[00135] It was determined whether there was a significant relationship between each of the individual cancer-related genes and cRFI after controlling for important demographic and clinical covariates. Separately for each gene, the relationship between gene expression and cRFI was modeled using multivariate Cox PH regression using weighted pseudo partial-likelihood estimators including important demographic and clinical variables as covariates. The independent contribution of gene expression to the prediction of cRFI was tested by generating a p-value from a Wald test using a model that included clinical covariates for each nodule 34 2015227398 15 Sep 2015 (specimens as defined in Table 2). Un-adjusted p-values <0.05 were considered statistically significant.

[00136] Tables 3A and 3B provide genes significantly associated (p<0.05), positively or negatively, with Gleason pattern in the primary and/or highest Gleason pattern. Increased expression of genes in Table 3A is positively associated with higher Gleason score, while increased expression of genes in Table 3B are negatively associated with higher Gleason score.

Table 3A

Gene significantly (p<0.05) associated with Gleason pattern for all specimens in the primary Gleason pattern or highest Gleason pattern odds ratio (OR) >1.0 (Increased expression is positively associated with higher Gleason Score)

Table 3A Primary Pattern Highest Pattern Official Symbol OR p-value OR p-value ALCAM 1.73 <.001 1.36 0.009 ANLN 1.35 0.027 APOC1 1.47 0.005 1.61 <001 APOE 1.87 <.001 2.15 <001 ASAP2 1.53 0.005 ASPN 2.62 <.001 2.13 <001 ATP5E 1.35 0.035 AURKA 1.44 0.010 AURKB 1.59 <001 1.56 <001 BAX 1.43 0.006 BGN 2.58 <001 2.82 <001 BIRC5 1.45 0.003 1.79 <001 BMP6 2.37 <001 1.68 <001 BMPR1B 1.58 0.002 BRCA2 1.45 0.013 BUB1 1.73 <001 1.57 <001 CACNA1D 1.31 0.045 1.31 0.033 CADPS 1.30 0.023 CCNB1 1.43 0.023 CCNE2 1.52 0.003 1.32 0.035 CD276 2.20 <001 1.83 <001 CD68 1.36 0.022 CDC20 1.69 <001 1.95 <001 CDC6 1.38 0.024 1.46 <001 CDH11 1.30 0.029 CDKN2B 1.55 0.001 1.33 0.023 CDKN2C 1.62 <001 1.52 <001 CDKN3 1.39 0.010 1.50 0.002 CENPF 1.96 <001 1.71 <001 CHRAC1 1.34 0.022 CLDN3 1.37 0.029 35 2015227398 15 Sep 20

Table 3A Primary Pattern Highest Pattern Official Symbol OR p-value OR p-value COL1A1 2.23 <001 2.22 <001 COL1A2 1.42 0.005 COL3A1 1.90 <001 2.13 <001 COL8A1 1.88 <001 2.35 <001 CRISP3 1.33 0.040 1.26 0.050 CTHRC1 2.01 <001 1.61 <001 CTNND2 1.48 0.007 1.37 0.011 DAPK1 1.44 0.014 DIAPH1 1.34 0.032 1.79 <001 DI02 1.56 0.001 DLL4 1.38 0.026 1.53 <001 ECE1 1.54 0.012 1.40 0.012 ENY2 1.35 0.046 1.35 0.012 EZH2 1.39 0.040 F2R 2.37 <001 2.60 <001 FAM49B 1.57 0.002 1.33 0.025 FAP 2.36 <001 1.89 <001 FCGR3A 2.10 <001 1.83 <001 GNPTAB 1.78 <001 1.54 <001 GSK3B 1.39 0.018 HRAS 1.62 0.003 HSD17B4 2.91 <001 1.57 <001 HSPA8 1.48 0.012 1.34 0.023 IFI30 1.64 <001 1.45 0.013 IGFBP3 1.29 0.037 IL11 1.52 0.001 1.31 0.036 INHBA 2.55 <001 2.30 <001 ITGA4 1.35 0.028 JAG1 1.68 <001 1.40 0.005 KCNN2 1.50 0.004 KCTD12 1.38 0.012 KHDRBS3 1.85 <001 1.72 <001 KIF4A 1.50 0.010 1.50 <001 KLK14 1.49 0.001 1.35 <001 KPNA2 1.68 0.004 1.65 0.001 KRT2 1.33 0.022 KRT75 1.27 0.028 LAMC1 1.44 0.029 LAPTM5 1.36 0.025 1.31 0.042 LTBP2 1.42 0.023 1.66 <001 MANF 1.34 0.019 MAOA 1.55 0.003 1.50 <001 MAP3K5 1.55 0.006 1.44 0.001 MDK 1.47 0.013 1.29 0.041 36 2015227398 15 Sep 20

Table 3A Primary Pattern Highest Pattern Official Symbol OR p-value OR p-value MDM2 1.31 0.026 MELK 1.64 <001 1.64 <001 MMP11 2.33 <001 1.66 <001 MYBL2 1.41 0.007 1.54 <001 MY06 1.32 0.017 NET02 1.36 0.018 NOX4 1.84 <001 1.73 <001 NPM1 1.68 0.001 NRIP3 1.36 0.009 NRP1 1.80 0.001 1.36 0.019 OSM 1.33 0.046 PATE1 1.38 0.032 PEC AMI 1.38 0.021 1.31 0.035 PGD 1.56 0.010 PLK1 1.51 0.004 1.49 0.002 PLOD2 1.29 0.027 POSTN 1.70 0.047 1.55 0.006 PPP3CA 1.38 0.037 1.37 0.006 PTK6 1.45 0.007 1.53 <001 PTTG1 1.51 <001 RAB31 1.31 0.030 RAD21 2.05 <001 1.38 0.020 RAD51 1.46 0.002 1.26 0.035 RAF1 1.46 0.017 RALBP1 1.37 0.043 RHOC 1.33 0.021 R0B02 1.52 0.003 1.41 0.006 RRM2 1.77 <001 1.50 <001 SAT1 1.67 0.002 1.61 <001 SDC1 1.66 0.001 1.46 0.014 SEC14L1 1.53 0.003 1.62 <001 SESN3 1.76 <001 1.45 <001 SFRP4 2.69 <001 2.03 <001 SHMT2 1.69 0.007 1.45 0.003 SKIL 1.46 0.005 SOX4 1.42 0.016 1.27 0.031 SPARC 1.40 0.024 1.55 <001 SPINK1 1.29 0.002 SPP1 1.51 0.002 1.80 <001 TFDP1 1.48 0.014 THBS2 1.87 <001 1.65 <001 THY1 1.58 0.003 1.64 <001 TK1 1.79 <001 1.42 0.001 TOP2A 2.30 <001 2.01 <001 37 2015227398 15 Sep 2015

Table 3A Primary Pattern Highest Pattern Official Symbol OR p-value OR p-value TPD52 1.95 <001 1.30 0.037 TPX2 2.12 <001 1.86 <001 TYMP 1.36 0.020 TYMS 1.39 0.012 1.31 0.036 UBE2C 1.66 <001 1.65 <001 UBE2T 1.59 <001 1.33 0.017 UGDH 1.28 0.049 UGT2B15 1.46 0.001 1.25 0.045 UHRF1 1.95 <001 1.62 <001 VDR 1.43 0.010 1.39 0.018 WNT5A 1.54 0.001 1.44 0.013 Table 3B.

Gene significantly (p<0.05) associated with Gleason pattern for all specimens in the primary Gleason pattern or highest Gleason pattern odds ratio (OR) <1.0 (Increased expression is negatively associated with higher Gleason score)

Table 3B Primary Pattern Highest Pattern Official Symbol OR p-value OR p-value ABCA5 0.78 0.041 ABCG2 0.65 0.001 0.72 0.012 ACOX2 0.44 <001 0.53 <001 ADH5 0.45 <001 0.42 <001 AFAP1 0.79 0.038 AIG1 0.77 0.024 AKAP1 0.63 0.002 AKR1C1 0.66 0.003 0.63 <001 AKT3 0.68 0.006 0.77 0.010 ALDH1A2 0.28 <001 0.33 <001 ALKBH3 0.77 0.040 0.77 0.029 AMPD3 0.67 0.007 ANPEP 0.68 0.008 0.59 <001 ANXA2 0.72 0.018 APC 0.69 0.002 AXIN2 0.46 <001 0.54 <001 AZGP1 0.52 <001 0.53 <001 BIK 0.69 0.006 0.73 0.003 BIN1 0.43 <001 0.61 <001 BTG3 0.79 0.030 BTRC 0.48 <001 0.62 <001 C7 0.37 <001 0.55 <001 CADM1 0.56 <001 0.69 0.001 CAV1 0.58 0.002 0.70 0.009 CAV2 0.65 0.029 38 2015227398 15 Sep 20

Table 3B Primary Pattern Highest Pattern Official Symbol OR p-value OR p-value CCNH 0.67 0.006 0.77 0.048 CD 164 0.59 0.003 0.57 <001 CDC25B 0.77 0.035 CDH1 0.66 <001 CDK2 0.71 0.003 CDKN1C 0.58 <001 0.57 <001 CDS2 0.69 0.002 CHN1 0.66 0.002 COL6A1 0.44 <001 0.66 <001 COL6A3 0.66 0.006 CSRP1 0.42 0.006 CTGF 0.74 0.043 CTNNA1 0.70 <001 0.83 0.018 CTNNB1 0.70 0.019 CTNND1 0.75 0.028 CUL1 0.74 0.011 CXCL12 0.54 <001 0.74 0.006 CYP3A5 0.52 <001 0.66 0.003 CYR61 0.64 0.004 0.68 0.005 DDR2 0.57 0.002 0.73 0.004 DES 0.34 <001 0.58 <001 DLGAP1 0.54 <001 0.62 <001 DNM3 0.67 0.004 DPP4 0.41 <001 0.53 <001 DPT 0.28 <001 0.48 <001 DUSP1 0.59 <001 0.63 <001 EDNRA 0.64 0.004 0.74 0.008 EGF 0.71 0.012 EGR1 0.59 <001 0.67 0.009 EGR3 0.72 0.026 0.71 0.025 EIF5 0.76 0.025 ELK4 0.58 0.001 0.70 0.008 ENPP2 0.66 0.002 0.70 0.005 EPHA3 0.65 0.006 EPHB2 0.60 <001 0.78 0.023 EPHB4 0.75 0.046 0.73 0.006 ERBB3 0.76 0.040 0.75 0.013 ERBB4 0.74 0.023 ERCC1 0.63 <001 0.77 0.016 FAAH 0.67 0.003 0.71 0.010 FAM107A 0.35 <001 0.59 <001 FAM13C 0.37 <001 0.48 <001 FAS 0.73 0.019 0.72 0.008 FGF10 0.53 <001 0.58 <001 39 2015227398 15 Sep 20

Table 3B Primary Pattern Highest Pattern Official Symbol OR p-value OR p-value FGF7 0.52 <001 0.59 <001 FGFR2 0.60 <001 0.59 <001 FKBP5 0.70 0.039 0.68 0.003 FLNA 0.39 <001 0.56 <001 FLNC 0.33 <001 0.52 <001 FOS 0.58 <001 0.66 0.005 FOXOl 0.57 <001 0.67 <001 FOXQ1 0.74 0.023 GADD45B 0.62 0.002 0.71 0.010 GHR 0.62 0.002 0.72 0.009 GNRH1 0.74 0.049 0.75 0.026 GPM6B 0.48 <001 0.68 <001 GPS1 0.68 0.003 GSN 0.46 <001 0.77 0.027 GSTM1 0.44 <001 0.62 <001 GSTM2 0.29 <001 0.49 <001 HGD 0.77 0.020 HIRIP3 0.75 0.034 HK1 0.48 <001 0.66 0.001 HLF 0.42 <001 0.55 <001 HNF1B 0.67 0.006 0.74 0.010 HPS1 0.66 0.001 0.65 <001 HSP90AB1 0.75 0.042 HSPA5 0.70 0.011 HSPB2 0.52 <001 0.70 0.004 IGF1 0.35 <001 0.59 <001 IGF2 0.48 <001 0.70 0.005 IGFBP2 0.61 <001 0.77 0.044 IGFBP5 0.63 <001 IGFBP6 0.45 <001 0.64 <001 IL6ST 0.55 0.004 0.63 <001 ILK 0.40 <001 0.57 <001 ING5 0.56 <001 0.78 0.033 ITGA1 0.56 0.004 0.61 <001 ITGA3 0.78 0.035 ITGA5 0.71 0.019 0.75 0.017 ITGA7 0.37 <001 0.52 <001 ITGB3 0.63 0.003 0.70 0.005 ITPR1 0.46 <001 0.64 <001 ITPR3 0.70 0.013 ITSN1 0.62 0.001 JUN 0.48 <001 0.60 <001 JUNB 0.72 0.025 KIT 0.51 <001 0.68 0.007 40 2015227398 15 Sep 20

Table 3B Primary Pattern Highest Pattern Official Symbol OR p-value OR p-value KLC1 0.58 <001 KLK1 0.69 0.028 0.66 0.003 KLK2 0.60 <001 KLK3 0.63 <001 0.69 0.012 KRT15 0.56 <001 0.60 <001 KRT18 0.74 0.034 KRT5 0.64 <001 0.62 <001 LAMA4 0.47 <001 0.73 0.010 LAMB 3 0.73 0.018 0.69 0.003 LGALS3 0.59 0.003 0.54 <001 LIG3 0.75 0.044 MAP3K7 0.66 0.003 0.79 0.031 MCM3 0.73 0.013 0.80 0.034 MGMT 0.61 0.001 0.71 0.007 MGST1 0.75 0.017 MLXIP 0.70 0.013 MMP2 0.57 <001 0.72 0.010 MMP7 0.69 0.009 MPPED2 0.70 0.009 0.59 <001 MSH6 0.78 0.046 MTA1 0.69 0.007 MTSS1 0.55 <001 0.54 <001 MYBPC1 0.45 <001 0.45 <001 NCAM1 0.51 <001 0.65 <001 NCAPD3 0.42 <001 0.53 <001 NCOR2 0.68 0.002 NDUFS5 0.66 0.001 0.70 0.013 NEXN 0.48 <001 0.62 <001 NFAT5 0.55 <001 0.67 0.001 NFKBIA 0.79 0.048 NRG1 0.58 0.001 0.62 0.001 OLFML3 0.42 <001 0.58 <001 OMD 0.67 0.004 0.71 0.004 OR51E2 0.65 <001 0.76 0.007 PAGE4 0.27 <001 0.46 <001 PCA3 0.68 0.004 PCDHGB7 0.70 0.025 0.65 <001 PGF 0.62 0.001 PGR 0.63 0.028 PHTF2 0.69 0.033 PLP2 0.54 <001 0.71 0.003 PPAP2B 0.41 <001 0.54 <001 PPP1R12A 0.48 <001 0.60 <001 PRIM A1 0.62 0.003 0.65 <001 41 2015227398 15 Sep 20

Table 3B Primary Pattern Highest Pattern Official Symbol OR p-value OR p-value PRKAR1B 0.70 0.009 PRKAR2B 0.79 0.038 PRKCA 0.37 <001 0.55 <001 PRKCB 0.47 <001 0.56 <001 PTCH1 0.70 0.021 PTEN 0.66 0.010 0.64 <001 PTGER3 0.76 0.015 PTGS2 0.70 0.013 0.68 0.005 PTH1R 0.48 <001 PTK2B 0.67 0.014 0.69 0.002 PYCARD 0.72 0.023 RAB27A 0.76 0.017 RAGE 0.77 0.040 0.57 <001 RARB 0.66 0.002 0.69 0.002 RECK 0.65 <001 RHOA 0.73 0.043 RHOB 0.61 0.005 0.62 <001 RND3 0.63 0.006 0.66 <001 SDHC 0.69 0.002 SEC23A 0.61 <001 0.74 0.010 SEMA3A 0.49 <001 0.55 <001 SERPINA3 0.70 0.034 0.75 0.020 SH3RF2 0.33 <001 0.42 <001 SLC22A3 0.23 <001 0.37 <001 SMAD4 0.33 <001 0.39 <001 SMARCC2 0.62 0.003 0.74 0.008 SMO 0.53 <001 0.73 0.009 SORBS 1 0.40 <001 0.55 <001 SPARCL1 0.42 <001 0.63 <001 SRD5A2 0.28 <001 0.37 <001 ST5 0.52 <001 0.63 <001 STAT5A 0.60 <001 0.75 0.020 STAT5B 0.54 <001 0.65 <001 STS 0.78 0.035 SUMOl 0.75 0.017 0.71 0.002 SVIL 0.45 <001 0.62 <001 TARP 0.72 0.017 TGFB1I1 0.37 <001 0.53 <001 TGFB2 0.61 0.025 0.59 <001 TGFB3 0.46 <001 0.60 <001 TIMP2 0.62 0.001 TIMP3 0.55 <001 0.76 0.019 TMPRSS2 0.71 0.014 TNF 0.65 0.010 42 2015227398 15 Sep 2015

Table 3B Primary Pattern Highest Pattern Official Symbol OR p-value OR p-value TNFRSF10A 0.71 0.014 0.74 0.010 TNFRSF10B 0.74 0.030 0.73 0.016 TNFSF10 0.69 0.004 TP53 0.73 0.011 TP63 0.62 <001 0.68 0.003 TPM1 0.43 <001 0.47 <001 TPM2 0.30 <001 0.47 <001 TPP2 0.58 <001 0.69 0.001 TRA2A 0.71 0.006 TRAF3IP2 0.50 <001 0.63 <001 TRO 0.40 <001 0.59 <001 TRPC6 0.73 0.030 TRPV6 0.80 0.047 VCL 0.44 <001 0.55 <001 VEGFB 0.73 0.029 VIM 0.72 0.013 VTI1B 0.78 0.046 WDR19 0.65 <001 WFDC1 0.50 <001 0.72 0.010 YY1 0.75 0.045 ZFHX3 0.52 <001 0.54 <001 ZFP36 0.65 0.004 0.69 0.012 ZNF827 0.59 <001 0.69 0.004 [00137] To identify genes associated with recurrence (cRFI, bRFI) in the primary and the highest Gleason pattern, each of 727 genes were analyzed in univariate models using specimens A1 and B2 (see Table 2, above). Tables 4A and 4B provide genes that were associated, positively or negatively, with cRFI and/or bRFI in the primary and/or highest Gleason pattern. Increased expression of genes in Table 4A is negatively associated with good prognosis, while increased expression of genes in Table 4B is positively associated with good prognosis.

Table 4A.

Genes significantly (p<0.05) associated with cRFI or bRFI in the primary Gleason pattern or highest Gleason pattern with hazard ratio (HR) > 1.0 (increased expression is negatively

Table 4A cRFI cRFI bRFI bRFI Primar; Y Pattern Highes Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value AKR1C3 1.304 0.022 1.312 0.013 ANLN 1.379 0.002 1.579 <001 1.465 <001 1.623 <001 AQP2 1.184 0.027 1.276 <001 ASAP2 1.442 0.006 43 associated with good prognosis) 2015227398 15 Sep 20

Table 4A cRFI cRFI bRFI bRFI Primary γ Pattern Highesl Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value ASPN 2.272 <001 2.106 <001 1.861 <001 1.895 <001 ATP5E F414 0.013 1.538 <001 BAG5 1.263 0.044 BAX 1.332 0.026 1.327 0.012 1.438 0.002 BGN F947 <001 2.061 <001 1.339 0.017 BIRC5 F497 <001 1.567 <001 1.478 <001 1.575 <001 BMP6 F705 <001 2.016 <001 1.418 0.004 1.541 <001 BMPR1B F401 0.013 1.325 0.016 BRCA2 1.259 0.007 BUB1 F411 <001 1.435 <001 1.352 <001 1.242 0.002 CADPS 1.387 0.009 1.294 0.027 CCNB1 1.296 0.016 1.376 0.002 CCNE2 F468 <001 1.649 <001 1.729 <001 1.563 <001 CD276 F678 <001 1.832 <001 1.581 <001 1.385 0.002 CDC20 F547 <001 1.671 <001 1.446 <001 1.540 <001 CDC6 F400 0.003 1.290 0.030 1.403 0.002 1.276 0.019 CDH7 F403 0.003 1.413 0.002 CDKN2B F569 <001 1.752 <001 1.333 0.017 1.347 0.006 CDKN2C F612 <001 1.780 <001 1.323 0.005 1.335 0.004 CDKN3 F384 <001 1.255 0.024 1.285 0.003 1.216 0.028 CENPF F578 <001 1.692 <001 1.740 <001 1.705 <001 CKS2 F390 0.007 1.418 0.005 1.291 0.018 CLTC 1.368 0.045 COL1A1 F873 <001 2.103 <001 1.491 <001 1.472 <001 COL1A2 1.462 0.001 COL3A1 F827 <001 2.005 <001 1.302 0.012 1.298 0.018 COL4A1 F490 0.002 1.613 <001 COL8A1 F692 <001 1.926 <001 1.307 0.013 1.317 0.010 CRISP3 F425 0.001 1.467 <001 1.242 0.045 CTHRC1 F505 0.002 2.025 <001 1.425 0.003 1.369 0.005 CTNND2 1.412 0.003 CXCR4 F312 0.023 1.355 0.008 DDIT4 F543 <001 1.763 <001 DYNLL1 F290 0.039 1.201 0.004 EIF3H 1.428 0.012 ENY2 F361 0.014 1.392 0.008 1.371 0.001 EZH2 1.311 0.010 F2R F773 <001 1.695 <001 1.495 <001 1.277 0.018 FADD 1.292 0.018 FAM171B 1.285 0.036 FAP F455 0.004 1.560 0.001 1.298 0.022 1.274 0.038 FASN F263 0.035 44 2015227398 15 Sep 20

Table 4A cRFI cRFI bRFI bRFI Primary γ Pattern Highesl Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value FCGR3A 1.654 <001 1.253 0.033 1.350 0.007 FGF5 E219 0.030 GNPTAB E388 0.007 1.503 0.003 1.355 0.005 1.434 0.002 GPR68 1.361 0.008 GREM1 E470 0.003 1.716 <001 1.421 0.003 1.316 0.017 HDAC1 1.290 0.025 HDAC9 1.395 0.012 HRAS E424 0.006 1.447 0.020 HSD17B4 E342 0.019 1.282 0.026 1.569 <001 1.390 0.002 HSPA8 E290 0.034 IGFBP3 E333 0.022 1.442 0.003 1.253 0.040 1.323 0.005 INHBA 2.368 <001 2.765 <001 1.466 0.002 1.671 <001 JAG1 E359 0.006 1.367 0.005 1.259 0.024 KCNN2 E361 0.011 1.413 0.005 1.312 0.017 1.281 0.030 KHDRBS3 E387 0.006 1.601 <001 1.573 <001 1.353 0.006 KIAA0196 1.249 0.037 KIF4A E212 0.016 1.149 0.040 1.278 0.003 KLK14 E167 0.023 1.180 0.007 KPNA2 1.425 0.009 1.353 0.005 1.305 0.019 KRT75 1.164 0.028 LAMA3 1.327 0.011 LAMB1 1.347 0.019 LAMC1 E555 0.001 1.310 0.030 1.349 0.014 LIMS1 1.275 0.022 LOX 1.358 0.003 1.410 <001 LTBP2 E396 0.009 1.656 <001 1.278 0.022 LUM 1.315 0.021 MANF 1.660 <001 1.323 0.011 MCM2 1.345 0.011 1.387 0.014 MCM6 E307 0.023 1.352 0.008 1.244 0.039 MELK E293 0.014 1.401 <001 1.501 <001 1.256 0.012 MMP11 E680 <001 1.474 <001 1.489 <001 1.257 0.030 MRPL13 1.260 0.025 MSH2 1.295 0.027 MYBL2 E664 <001 1.670 <001 1.399 <001 1.431 <001 MY06 1.301 0.033 NET02 E412 0.004 1.302 0.027 1.298 0.009 NFKB1 1.236 0.050 NOX4 E492 <001 1.507 0.001 1.555 <001 1.262 0.019 NPM1 1.287 0.036 NRIP3 1.219 0.031 1.218 0.018 NRP1 1.482 0.002 1.245 0.041 45 2015227398 15 Sep 20

Table 4A cRFI cRFI bRFI bRFI Primary γ Pattern Highesl Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value OLFML2B 1.362 0.015 OR51E1 1.531 <001 1.488 0.003 PAK6 1.269 0.033 PATE1 E308 <001 1.332 <001 1.164 0.044 PCNA 1.278 0.020 PEX10 E436 0.005 1.393 0.009 PGD E298 0.048 1.579 <001 PGK1 1.274 0.023 1.262 0.009 PLA2G7 1.315 0.011 1.346 0.005 PLAU 1.319 0.010 PLK1 E309 0.021 1.563 <001 1.410 0.002 1.372 0.003 PLOD2 1.284 0.019 1.272 0.014 1.332 0.005 POSTN E599 <001 1.514 0.002 1.391 0.005 PPP3CA 1.402 0.007 1.316 0.018 PSMD13 E278 0.040 1.297 0.033 1.279 0.017 1.373 0.004 PTK6 E640 <001 1.932 <001 1.369 0.001 1.406 <001 PTTG1 E409 <001 1.510 <001 1.347 0.001 1.558 <001 RAD21 E315 0.035 1.402 0.004 1.589 <001 1.439 <001 RAF1 1.503 0.002 RALA E521 0.004 1.403 0.007 1.563 <001 1.229 0.040 RALBP1 1.277 0.033 RGS7 E154 0.015 1.266 0.010 RRM1 E570 0.001 1.602 <001 RRM2 E368 <001 1.289 0.004 1.396 <001 1.230 0.015 SAT1 E482 0.016 1.403 0.030 SDC1 1.340 0.018 1.396 0.018 SEC14L1 1.260 0.048 1.360 0.002 SESN3 E485 <001 1.631 <001 1.232 0.047 1.292 0.014 SFRP4 E800 <001 1.814 <001 1.496 <001 1.289 0.027 SHMT2 E807 <001 1.658 <001 1.673 <001 1.548 <001 SKIL 1.327 0.008 SLC25A21 1.398 0.001 1.285 0.018 SOX4 1.286 0.020 1.280 0.030 SPARC E539 <001 1.842 <001 1.269 0.026 SPP1 1.322 0.022 SQLE 1.359 0.020 1.270 0.036 STMN1 E402 0.007 1.446 0.005 1.279 0.031 SULF1 1.587 <001 TAF2 1.273 0.027 TFDP1 1.328 0.021 1.400 0.005 1.416 0.001 THBS2 E812 <001 1.960 <001 1.320 0.012 1.256 0.038 THY1 E362 0.020 1.662 <001 46 2015227398 15 Sep 2015

Table 4A cRFI cRFI bRFI bRFI Primary γ Pattern Highesl Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value TK1 1.251 0.011 1.377 <001 1.401 <001 TOP2A E670 <001 1.920 <001 1.869 <001 1.927 <001 TPD52 E324 0.011 1.366 0.002 1.351 0.005 TPX2 E884 <001 2.154 <001 1.874 <001 1.794 <001 UAP1 1.244 0.044 UBE2C E403 <001 1.541 <001 1.306 0.002 1.323 <001 UBE2T E667 <001 1.282 0.023 1.502 <001 1.298 0.005 UGT2B15 1.295 0.001 1.275 0.002 UGT2B17 1.294 0.025 UHRF1 E454 <001 1.531 <001 1.257 0.029 VCPIP1 E390 0.009 1.414 0.004 1.294 0.021 1.283 0.021 WNT5A 1.274 0.038 1.298 0.020 XIAP 1.464 0.006 ZMYND8 1.277 0.048 ZWINT E259 0.047 47 2015227398 15 Sep 2015

Table 4B. Genes significantly (p<0.05) associated with cRFI or bRFI in the primary Gleason pattern or highest Gleason pattern with hazard ratio (HR) <1.0 (increased expression is positively associated with good prognosis) Table 4B cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value AAMP 0.564 <001 0.571 <001 0.764 0.037 0.786 0.034 ABCA5 0.755 <001 0.695 <001 0.800 0.006 ABCB1 0.777 0.026 ABCG2 0.788 0.033 0.784 0.040 0.803 0.018 0.750 0.004 ABHD2 0.734 0.011 ACE 0.782 0.048 ACOX2 0.639 <001 0.631 <001 0.713 <001 0.716 0.002 ADH5 0.625 <001 0.637 <001 0.753 0.026 AKAP1 0.764 0.006 0.800 0.005 0.837 0.046 AKR1C1 0.773 0.033 0.802 0.032 AKT1 0.714 0.005 AKT3 0.811 0.015 0.809 0.021 ALDH1A2 0.606 <001 0.498 <001 0.613 <001 0.624 <001 AMPD3 0.793 0.024 ANPEP 0.584 <001 0.493 <001 ANXA2 0.753 0.013 0.781 0.036 0.762 0.008 0.795 0.032 APRT 0.758 0.026 0.780 0.044 0.746 0.008 ATXN1 0.673 0.001 0.776 0.029 0.809 0.031 0.812 0.043 AXIN2 0.674 <001 0.571 <001 0.776 0.005 0.757 0.005 AZGP1 0.585 <001 0.652 <001 0.664 <001 0.746 <001 BAD 0.765 0.023 BCL2 0.788 0.033 0.778 0.036 BDKRB1 0.728 0.039 BIK 0.712 0.005 BIN1 0.607 <001 0.724 0.002 0.726 <001 0.834 0.034 BTG3 0.847 0.034 BTRC 0.688 0.001 0.713 0.003 C7 0.589 <001 0.639 <001 0.629 <001 0.691 <001 CADM1 0.546 <001 0.529 <001 0.743 0.008 0.769 0.015 CAS PI 0.769 0.014 0.799 0.028 0.799 0.010 0.815 0.018 CAV1 0.736 0.011 0.711 0.005 0.675 <001 0.743 0.006 CAV2 0.636 0.010 0.648 0.012 0.685 0.012 CCL2 0.759 0.029 0.764 0.024 CCNH 0.689 <001 0.700 <001 CD 164 0.664 <001 0.651 <001 CD1A 0.687 0.004 CD44 0.545 <001 0.600 <001 0.788 0.018 0.799 0.023 CD82 0.771 0.009 0.748 0.004 48 2015227398 15 Sep 20

Table 4B cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value CDC25B 0.755 0.006 0.817 0.025 CDK14 0.845 0.043 CDK2 0.819 0.032 CDK3 0.733 0.005 0.772 0.006 0.838 0.017 CDKN1A 0.766 0.041 CDKN1C 0.662 <001 0.712 0.002 0.693 <001 0.761 0.009 CHN1 0.788 0.036 COL6A1 0.608 <001 0.767 0.013 0.706 <001 0.775 0.007 CSF1 0.626 <001 0.709 0.003 CSK 0.837 0.029 CSRP1 0.793 0.024 0.782 0.019 CTNNB1 0.898 0.042 0.885 <001 CTSB 0.701 0.004 0.713 0.007 0.715 0.002 0.803 0.038 CTSK 0.815 0.042 CXCL12 0.652 <001 0.802 0.044 0.711 0.001 CYP3A5 0.463 <001 0.436 <001 0.727 0.003 CYR61 0.652 0.002 0.676 0.002 DAP 0.761 0.026 0.775 0.025 0.802 0.048 DARC 0.725 0.005 0.792 0.032 DDR2 0.719 0.001 0.763 0.008 DES 0.619 <001 0.737 0.005 0.638 <001 0.793 0.017 DHRS9 0.642 0.003 DHX9 0.888 <001 DLC1 0.710 0.007 0.715 0.009 DLGAP1 0.613 <001 0.551 <001 0.779 0.049 DNM3 0.679 <001 0.812 0.037 DPP4 0.591 <001 0.613 <001 0.761 0.003 DPT 0.613 <001 0.576 <001 0.647 <001 0.677 <001 DUSP1 0.662 0.001 0.665 0.001 0.785 0.024 DUSP6 0.713 0.005 0.668 0.002 EDNRA 0.702 0.002 0.779 0.036 EGF 0.738 0.028 EGR1 0.569 <001 0.577 <001 0.782 0.022 EGR3 0.601 <001 0.619 <001 0.800 0.038 EIF2S3 0.756 0.015 EIF5 0.776 0.023 0.787 0.028 ELK4 0.628 <001 0.658 <001 EPHA2 0.720 0.011 0.663 0.004 EPHA3 0.727 0.003 0.772 0.005 ERBB2 0.786 0.019 0.738 0.003 0.815 0.041 ERBB3 0.728 0.002 0.711 0.002 0.828 0.043 0.813 0.023 ERCC1 0.771 0.023 0.725 0.007 0.806 0.049 0.704 0.002 49 2015227398 15 Sep 20

Table 4B cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value EREG 0.754 0.016 0.777 0.034 ESR2 0.731 0.026 FAAH 0.708 0.004 0.758 0.012 0.784 0.031 0.774 0.007 FAM107A 0.517 <001 0.576 <001 0.642 <001 0.656 <001 FAM13C 0.568 <001 0.526 <001 0.739 0.002 0.639 <001 FAS 0.755 0.014 FAS EG 0.706 0.021 FGF10 0.653 <001 0.685 <001 0.766 0.022 FGF17 0.746 0.023 0.781 0.015 0.805 0.028 FGF7 0.794 0.030 0.820 0.037 0.811 0.040 FGFR2 0.683 <001 0.686 <001 0.674 <001 0.703 <001 FKBP5 0.676 0.001 FEN A 0.653 <001 0.741 0.010 0.682 <001 0.771 0.016 FFNC 0.751 0.029 0.779 0.047 0.663 <001 0.725 <001 FFT1 0.799 0.044 FOS 0.566 <001 0.543 <001 0.757 0.006 FOXOl 0.816 0.039 0.798 0.023 FOXQ1 0.753 0.017 0.757 0.024 0.804 0.018 FYN 0.779 0.031 GADD45B 0.590 <001 0.619 <001 GDF15 0.759 0.019 0.794 0.048 GHR 0.702 0.005 0.630 <001 0.673 <001 0.590 <001 GNRH1 0.742 0.014 GPM6B 0.653 <001 0.633 <001 0.696 <001 0.768 0.007 GSN 0.570 <001 0.697 0.001 0.697 <001 0.758 0.005 GSTM1 0.612 <001 0.588 <001 0.718 <001 0.801 0.020 GSTM2 0.540 <001 0.630 <001 0.602 <001 0.706 <001 HGD 0.796 0.020 0.736 0.002 HIRIP3 0.753 0.011 0.824 0.050 HK1 0.684 <001 0.683 <001 0.799 0.011 0.804 0.014 HFA-G 0.726 0.022 HFF 0.555 <001 0.582 <001 0.703 <001 0.702 <001 HNF1B 0.690 <001 0.585 <001 HPS1 0.744 0.003 0.784 0.020 0.836 0.047 HSD3B2 0.733 0.016 HSP90AB1 0.801 0.036 HSPA5 0.776 0.034 HSPB1 0.813 0.020 HSPB2 0.762 0.037 0.699 0.002 0.783 0.034 HSPG2 0.794 0.044 ICAM1 0.743 0.024 0.768 0.040 IER3 0.686 0.002 0.663 <001 50 2015227398 15 Sep 20

Table 4B cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value IFIT1 0.649 <001 0.761 0.026 IGF1 0.634 <001 0.537 <001 0.696 <001 0.688 <001 IGF2 0.732 0.004 IGFBP2 0.548 <001 0.620 <001 IGFBP5 0.681 <001 IGFBP6 0.577 <001 0.675 <001 IL1B 0.712 0.005 0.742 0.009 IL6 0.763 0.028 IL6R 0.791 0.039 IL6ST 0.585 <001 0.639 <001 0.730 0.002 0.768 0.006 IL8 0.624 <001 0.662 0.001 ILK 0.712 0.009 0.728 0.012 0.790 0.047 0.790 0.042 ING5 0.625 <001 0.658 <001 0.728 0.002 ITGA5 0.728 0.006 0.803 0.039 ITGA6 0.779 0.007 0.775 0.006 ITGA7 0.584 <001 0.700 0.001 0.656 <001 0.786 0.014 ITGAD 0.657 0.020 ITGB4 0.718 0.007 0.689 <001 0.818 0.041 ITGB5 0.801 0.050 ITPR1 0.707 0.001 JUN 0.556 <001 0.574 <001 0.754 0.008 JUNB 0.730 0.017 0.715 0.010 KIT 0.644 0.004 0.705 0.019 0.605 <001 0.659 0.001 KLC1 0.692 0.003 0.774 0.024 0.747 0.008 KLF6 0.770 0.032 0.776 0.039 KLK1 0.646 <001 0.652 0.001 0.784 0.037 KLK10 0.716 0.006 KLK2 0.647 <001 0.628 <001 0.786 0.009 KLK3 0.706 <001 0.748 <001 0.845 0.018 KRT1 0.734 0.024 KRT15 0.627 <001 0.526 <001 0.704 <001 0.782 0.029 KRT18 0.624 <001 0.617 <001 0.738 0.005 0.760 0.005 KRT5 0.640 <001 0.550 <001 0.740 <001 0.798 0.023 KRT8 0.716 0.006 0.744 0.008 LI CAM 0.738 0.021 0.692 0.009 0.761 0.036 LAG3 0.741 0.013 0.729 0.011 LAMA4 0.686 0.011 0.592 0.003 LAMA5 0.786 0.025 LAMB 3 0.661 <001 0.617 <001 0.734 <001 LGALS3 0.618 <001 0.702 0.001 0.734 0.001 0.793 0.012 LIG3 0.705 0.008 0.615 <001 LRP1 0.786 0.050 0.795 0.023 0.770 0.009 51 2015227398 15 Sep 20

Table 4B cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value MAP3K7 0.789 0.003 MGMT 0.632 <001 0.693 <001 MICA 0.781 0.014 0.653 <001 0.833 0.043 MPPED2 0.655 <001 0.597 <001 0.719 <001 0.759 0.006 MSH6 0.793 0.015 MTSS1 0.613 <001 0.746 0.008 MVP 0.792 0.028 0.795 0.045 0.819 0.023 MYBPC1 0.648 <001 0.496 <001 0.701 <001 0.629 <001 NCAM1 0.773 0.015 NCAPD3 0.574 <001 0.463 <001 0.679 <001 0.640 <001 NEXN 0.701 0.002 0.791 0.035 0.725 0.002 0.781 0.016 NFAT5 0.515 <001 0.586 <001 0.785 0.017 NFATC2 0.753 0.023 NFKBIA 0.778 0.037 NRG1 0.644 0.004 0.696 0.017 0.698 0.012 OAZ1 0.777 0.034 0.775 0.022 OLFML3 0.621 <001 0.720 0.001 0.600 <001 0.626 <001 OMD 0.706 0.003 OR51E2 0.820 0.037 0.798 0.027 PAGE4 0.549 <001 0.613 <001 0.542 <001 0.628 <001 PCA3 0.684 <001 0.635 <001 PCDHGB7 0.790 0.045 0.725 0.002 0.664 <001 PGF 0.753 0.017 PGR 0.740 0.021 0.728 0.018 PIK3CG 0.803 0.024 PLAUR 0.778 0.035 PLG 0.728 0.028 PPAP2B 0.575 <001 0.629 <001 0.643 <001 0.699 <001 PPP1R12A 0.647 <001 0.683 0.002 0.782 0.023 0.784 0.030 PRIM A1 0.626 <001 0.658 <001 0.703 0.002 0.724 0.003 PRKCA 0.642 <001 0.799 0.029 0.677 0.001 0.776 0.006 PRKCB 0.675 0.001 0.648 <001 0.747 0.006 PROM1 0.603 0.018 0.659 0.014 0.493 0.008 PTCH1 0.680 0.001 0.753 0.010 0.789 0.018 PTEN 0.732 0.002 0.747 0.005 0.744 <001 0.765 0.002 PTGS2 0.596 <001 0.610 <001 PTH1R 0.767 0.042 0.775 0.028 0.788 0.047 PTHLH 0.617 0.002 0.726 0.025 0.668 0.002 0.718 0.007 PTK2B 0.744 0.003 0.679 <001 0.766 0.002 0.726 <001 PTPN1 0.760 0.020 0.780 0.042 PYCARD 0.748 0.012 RAB27A 0.708 0.004 52 2015227398 15 Sep 20

Table 4B cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value RAB30 0.755 0.008 RAGE 0.817 0.048 RAP IB 0.818 0.050 RARB 0.757 0.007 0.677 <001 0.789 0.007 0.746 0.003 RASSF1 0.816 0.035 RHOB 0.725 0.009 0.676 0.001 0.793 0.039 RLN1 0.742 0.033 0.762 0.040 RND3 0.636 <001 0.647 <001 RNF114 0.749 0.011 SDC2 0.721 0.004 SDHC 0.725 0.003 0.727 0.006 SEMA3A 0.757 0.024 0.721 0.010 SERPINA3 0.716 0.008 0.660 0.001 SERPINB5 0.747 0.031 0.616 0.002 SH3RF2 0.577 <001 0.458 <001 0.702 <001 0.640 <001 SFC22A3 0.565 <001 0.540 <001 0.747 0.004 0.756 0.007 SMAD4 0.546 <001 0.573 <001 0.636 <001 0.627 <001 SMARCD1 0.718 <001 0.775 0.017 SMO 0.793 0.029 0.754 0.021 0.718 0.003 SOD1 0.757 0.049 0.707 0.006 SORBS 1 0.645 <001 0.716 0.003 0.693 <001 0.784 0.025 SPARCF1 0.821 0.028 0.829 0.014 0.781 0.030 SPDEF 0.778 <001 SPINT1 0.732 0.009 0.842 0.026 SRC 0.647 <001 0.632 <001 SRD5A1 0.813 0.040 SRD5A2 0.489 <001 0.533 <001 0.544 <001 0.611 <001 ST5 0.713 0.002 0.783 0.011 0.725 <001 0.827 0.025 STAT3 0.773 0.037 0.759 0.035 STAT5A 0.695 <001 0.719 0.002 0.806 0.020 0.783 0.008 STAT5B 0.633 <001 0.655 <001 0.814 0.028 SUMOl 0.790 0.015 SVIE 0.659 <001 0.713 0.002 0.711 0.002 0.779 0.010 TARP 0.800 0.040 TBP 0.761 0.010 TFF3 0.734 0.010 0.659 <001 TGFB1I1 0.618 <001 0.693 0.002 0.637 <001 0.719 0.004 TGFB2 0.679 <001 0.747 0.005 0.805 0.030 TGFB3 0.791 0.037 TGFBR2 0.778 0.035 TIMP3 0.751 0.011 TMPRSS2 0.745 0.003 0.708 <001 53 2015227398 15 Sep 2015

Table 4B cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value TNF 0.670 0.013 0.697 0.015 TNFRSF10A 0.780 0.018 0.752 0.006 0.817 0.032 TNFRSF10B 0.576 <.001 0.655 <001 0.766 0.004 0.778 0.002 TNFRSF18 0.648 0.016 0.759 0.034 TNFSF10 0.653 <.001 0.667 0.004 TP53 0.729 0.003 TP63 0.759 0.016 0.636 <001 0.698 <001 0.712 0.001 TPM1 0.778 0.048 0.743 0.012 0.783 0.032 0.811 0.046 TPM2 0.578 <.001 0.634 <001 0.611 <001 0.710 0.001 TPP2 0.775 0.037 TRAF3IP2 0.722 0.002 0.690 <001 0.792 0.021 0.823 0.049 TRO 0.744 0.003 0.725 0.003 0.765 0.002 0.821 0.041 TUBB2A 0.639 <.001 0.625 <001 TYMP 0.786 0.039 VCL 0.594 <.001 0.657 0.001 0.682 <001 VEGFA 0.762 0.024 VEGFB 0.795 0.037 VIM 0.739 0.009 0.791 0.021 WDR19 0.776 0.015 WFDC1 0.746 <001 YY1 0.683 0.001 0.728 0.002 ZFHX3 0.684 <.001 0.661 <001 0.801 0.010 0.762 0.001 ZFP36 0.605 <.001 0.579 <001 0.815 0.043 ZNF827 0.624 <.001 0.730 0.007 0.738 0.004 [00138] Tables 5A and 5B provide genes that were significantly associated (p<0.05), positively or negatively, with recurrence (cRFI, bRFI) after adjusting for AUA risk group in the primary and/or highest Gleason pattern. Increased expression of genes in Table 5A is negatively associated with good prognosis, while increased expression of genes in Table 5B is positively associated with good prognosis.

Table 5A.

Gene significantly (p<0.05) associated with cRFI or bRFI after adjustment for AUA risk group in the primary Gleason pattern or highest Gleason pattern with hazard ratio (HR) > 1.0 (increased expression negatively associated with good prognosis)

Table 5A cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value AKR1C3 1.315 0.018 1.283 0.024 ALOX12 1.198 0.024 54 2015227398 15 Sep 20

Table 5A cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value ANLN E406 <001 1.519 <001 1.485 <001 1.632 <001 AQP2 E209 <001 1.302 <001 ASAP2 1.582 <001 1.333 0.011 1.307 0.019 ASPN E872 <001 1.741 <001 1.638 <001 1.691 <001 ATP5E E309 0.042 1.369 0.012 BAG5 1.291 0.044 BAX 1.298 0.025 1.420 0.004 BGN E746 <001 1.755 <001 BIRC5 E480 <001 1.470 <001 1.419 <001 1.503 <001 BMP6 E536 <001 1.815 <001 1.294 0.033 1.429 0.001 BRCA2 1.184 0.037 BUB1 E288 0.001 1.391 <001 1.254 <001 1.189 0.018 CACNA1D 1.313 0.029 CADPS 1.358 0.007 1.267 0.022 CASP3 1.251 0.037 CCNB1 1.261 0.033 1.318 0.005 CCNE2 E345 0.005 1.438 <001 1.606 <001 1.426 <001 CD276 E482 0.002 1.668 <001 1.451 <001 1.302 0.011 CDC20 E417 <001 1.547 <001 1.355 <001 1.446 <001 CDC6 E340 0.011 1.265 0.046 1.367 0.002 1.272 0.025 CDH7 E402 0.003 1.409 0.002 CDKN2B E553 <001 1.746 <001 1.340 0.014 1.369 0.006 CDKN2C E411 <001 1.604 <001 1.220 0.033 CDKN3 E296 0.004 1.226 0.015 CENPF E434 0.002 1.570 <001 1.633 <001 1.610 <001 CKS2 E419 0.008 1.374 0.022 1.380 0.004 COL1A1 E677 <001 1.809 <001 1.401 <001 1.352 0.003 COL1A2 1.373 0.010 COL3A1 E669 <001 1.781 <001 1.249 0.024 1.234 0.047 COL4A1 E475 0.002 1.513 0.002 COL8A1 E506 0.001 1.691 <001 CRISP3 E406 0.004 1.471 <001 CTHRC1 E426 0.009 1.793 <001 1.311 0.019 CTNND2 1.462 <001 DDIT4 E478 0.003 1.783 <001 1.236 0.039 DYNLL1 E431 0.002 1.193 0.004 EIF3H 1.372 0.027 ENY2 1.325 0.023 1.270 0.017 ERG E303 0.041 EZH2 1.254 0.049 F2R E540 0.002 1.448 0.006 1.286 0.023 FADD E235 0.041 1.404 <001 55 2015227398 15 Sep 20

Table 5A cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value FAP E386 0.015 1.440 0.008 1.253 0.048 FASN E303 0.028 FCGR3A 1.439 0.011 1.262 0.045 FGF5 E289 0.006 GNPTAB E290 0.033 1.369 0.022 1.285 0.018 1.355 0.008 GPR68 1.396 0.005 GREM1 E341 0.022 1.502 0.003 1.366 0.006 HDAC1 1.329 0.016 HDAC9 1.378 0.012 HRAS E465 0.006 HSD17B4 1.442 <001 1.245 0.028 IGFBP3 1.366 0.019 1.302 0.011 INHBA 2.000 <001 2.336 <001 1.486 0.002 JAG1 E251 0.039 KCNN2 E347 0.020 1.524 <001 1.312 0.023 1.346 0.011 KHDRBS3 1.500 0.001 1.426 0.001 1.267 0.032 KIAA0196 1.272 0.028 KIF4A E199 0.022 1.262 0.004 KPNA2 1.252 0.016 LAMA3 1.332 0.004 1.356 0.010 LAMB1 1.317 0.028 LAMC1 E516 0.003 1.302 0.040 1.397 0.007 LIMS1 1.261 0.027 LOX 1.265 0.016 1.372 0.001 LTBP2 1.477 0.002 LUM 1.321 0.020 MANF 1.647 <001 1.284 0.027 MCM2 1.372 0.003 1.302 0.032 MCM3 1.269 0.047 MCM6 1.276 0.033 1.245 0.037 MELK 1.294 0.005 1.394 <001 MKI67 E253 0.028 1.246 0.029 MMP11 E557 <001 1.290 0.035 1.357 0.005 MRPL13 1.275 0.003 MSH2 1.355 0.009 MYBL2 E497 <001 1.509 <001 1.304 0.003 1.292 0.007 MY06 1.367 0.010 NDRG1 E270 0.042 1.314 0.025 NEK2 1.338 0.020 1.269 0.026 NET02 E434 0.004 1.303 0.033 1.283 0.012 NOX4 E413 0.006 1.308 0.037 1.444 <001 NRIP3 1.171 0.026 56 2015227398 15 Sep 20

Table 5A cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value NRP1 1.372 0.020 ODC1 1.450 <001 OR51E1 1.559 <001 1.413 0.008 PAK6 1.233 0.047 PATE1 E262 <001 1.375 <001 1.143 0.034 1.191 0.036 PCNA 1.227 0.033 1.318 0.003 PEX10 E517 <001 1.500 0.001 PGD E363 0.028 1.316 0.039 1.652 <001 PGK1 1.224 0.034 1.206 0.024 PIM1 1.205 0.042 PLA2G7 1.298 0.018 1.358 0.005 PLAU 1.242 0.032 PLK1 1.464 0.001 1.299 0.018 1.275 0.031 PLOD2 1.206 0.039 1.261 0.025 POSTN E558 0.001 1.356 0.022 1.363 0.009 PPP3CA 1.445 0.002 PSMD13 1.301 0.017 1.411 0.003 PTK2 1.318 0.031 PTK6 E582 <001 1.894 <001 1.290 0.011 1.354 0.003 PTTG1 E319 0.004 1.430 <001 1.271 0.006 1.492 <001 RAD21 1.278 0.028 1.435 0.004 1.326 0.008 RAF1 1.504 <001 RALA E374 0.028 1.459 0.001 RGS7 1.203 0.031 RRM1 E535 0.001 1.525 <001 RRM2 E302 0.003 1.197 0.047 1.342 <001 SAT1 E374 0.043 SDC1 1.344 0.011 1.473 0.008 SEC14L1 1.297 0.006 SESN3 E337 0.002 1.495 <001 1.223 0.038 SFRP4 E610 <001 1.542 0.002 1.370 0.009 SHMT2 E567 0.001 1.522 <001 1.485 0.001 1.370 <001 SKIL 1.303 0.008 SLC25A21 1.287 0.020 1.306 0.017 SLC44A1 1.308 0.045 SNRPB2 E304 0.018 SOX4 1.252 0.031 SPARC E445 0.004 1.706 <001 1.269 0.026 SPP1 1.376 0.016 SQLE 1.417 0.007 1.262 0.035 STAT1 1.209 0.029 STMN1 E315 0.029 57 2015227398 15 Sep 2015

Table 5A cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value SULF1 1.504 0.001 TAF2 1.252 0.048 1.301 0.019 TFDP1 1.395 0.010 1.424 0.002 THBS2 1.716 <001 1.719 <001 THY1 1.343 0.035 1.575 0.001 TK1 1.320 <001 1.304 <001 TOP2A 1.464 0.001 1.688 <001 1.715 <001 1.761 <001 TPD52 1.286 0.006 1.258 0.023 TPX2 1.644 <001 1.964 <001 1.699 <001 1.754 <001 TYMS 1.315 0.014 UBE2C 1.270 0.019 1.558 <001 1.205 0.027 1.333 <001 UBE2G1 1.302 0.041 UBE2T 1.451 <001 1.309 0.003 UGT2B15 1.222 0.025 UHRF1 1.370 0.003 1.520 <001 1.247 0.020 VCPIP1 1.332 0.015 VTI1B 1.237 0.036 XIAP 1.486 0.008 ZMYND8 1.408 0.007 ZNF3 1.284 0.018 ZWINT 1.289 0.028 Table 5B. Genes significantly (p<0.05) associated with cRFI or bRFI after adjustment for AUA risk group in the primary Gleason pattern or highest Gleason pattern with hazard ratio (HR) < 1.0 (increased expression is positively associated with good prognosis) Table 5B cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value AAMP 0.535 <001 0.581 <001 0.700 0.002 0.759 0.006 ABCA5 0.798 0.007 0.745 0.002 0.841 0.037 ABCC1 0.800 0.044 ABCC4 0.787 0.022 ABHD2 0.768 0.023 ACOX2 0.678 0.002 0.749 0.027 0.759 0.004 ADH5 0.645 <001 0.672 0.001 AGTR1 0.780 0.030 AKAP1 0.815 0.045 0.758 <001 AKT1 0.732 0.010 ALDH1A2 0.646 <001 0.548 <001 0.671 <001 0.713 0.001 ANPEP 0.641 <001 0.535 <001 58 2015227398 15 Sep 20

Table 5B cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value ANXA2 0.772 0.035 0.804 0.046 ATXN1 0.654 <001 0.754 0.020 0.797 0.017 AURKA 0.788 0.030 AXIN2 0.744 0.005 0.655 <001 AZGP1 0.656 <001 0.676 <001 0.754 0.001 0.791 0.004 BAD 0.700 0.004 BIN1 0.650 <001 0.764 0.013 0.803 0.015 BTG3 0.836 0.025 BTRC 0.730 0.005 C7 0.617 <001 0.680 <001 0.667 <001 0.755 0.005 CADM1 0.559 <001 0.566 <001 0.772 0.020 0.802 0.046 CAS PI 0.781 0.030 0.779 0.021 0.818 0.027 0.828 0.036 CAV1 0.775 0.034 CAV2 0.677 0.019 CCL2 0.752 0.023 CCNH 0.679 <001 0.682 <001 CD 164 0.721 0.002 0.724 0.005 CD1A 0.710 0.014 CD44 0.591 <001 0.642 <001 CD82 0.779 0.021 0.771 0.024 CDC25B 0.778 0.035 0.818 0.023 CDK14 0.788 0.011 CDK3 0.752 0.012 0.779 0.005 0.841 0.020 CDKN1A 0.770 0.049 0.712 0.014 CDKN1C 0.684 <001 0.697 <001 CHN1 0.772 0.031 COL6A1 0.648 <001 0.807 0.046 0.768 0.004 CSF1 0.621 <001 0.671 0.001 CTNNB1 0.905 0.008 CTSB 0.754 0.030 0.716 0.011 0.756 0.014 CXCL12 0.641 <001 0.796 0.038 0.708 <001 CYP3A5 0.503 <001 0.528 <001 0.791 0.028 CYR61 0.639 0.001 0.659 0.001 0.797 0.048 DARC 0.707 0.004 DDR2 0.750 0.011 DES 0.657 <001 0.758 0.022 0.699 <001 DHRS9 0.625 0.002 DHX9 0.846 <001 DIAPH1 0.682 0.007 0.723 0.008 0.780 0.026 DLC1 0.703 0.005 0.702 0.008 DLGAP1 0.703 0.008 0.636 <001 DNM3 0.701 0.001 0.817 0.042 59 2015227398 15 Sep 20

Table 5B cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value DPP4 0.686 <001 0.716 0.001 DPT 0.636 <001 0.633 <001 0.709 0.006 0.773 0.024 DUSP1 0.683 0.006 0.679 0.003 DUSP6 0.694 0.003 0.605 <001 EDN1 0.773 0.031 EDNRA 0.716 0.007 EGR1 0.575 <001 0.575 <001 0.771 0.014 EGR3 0.633 0.002 0.643 <001 0.792 0.025 EIF4E 0.722 0.002 ELK4 0.710 0.009 0.759 0.027 ENPP2 0.786 0.039 EPHA2 0.593 0.001 EPHA3 0.739 0.006 0.802 0.020 ERBB2 0.753 0.007 ERBB3 0.753 0.009 0.753 0.015 ERCC1 0.727 0.001 EREG 0.722 0.012 0.769 0.040 ESR1 0.742 0.015 FABP5 0.756 0.032 FAM107A 0.524 <001 0.579 <001 0.688 <001 0.699 0.001 FAM13C 0.639 <001 0.601 <001 0.810 0.019 0.709 <001 FAS 0.770 0.033 FAS LG 0.716 0.028 0.683 0.017 FGF10 0.798 0.045 FGF17 0.718 0.018 0.793 0.024 0.790 0.024 FGFR2 0.739 0.007 0.783 0.038 0.740 0.004 FGFR4 0.746 0.050 FKBP5 0.689 0.003 FLNA 0.701 0.006 0.766 0.029 0.768 0.037 FLNC 0.755 <001 0.820 0.022 FLT1 0.729 0.008 FOS 0.572 <001 0.536 <001 0.750 0.005 FOXQ1 0.778 0.033 0.820 0.018 FYN 0.708 0.006 GADD45B 0.577 <001 0.589 <001 GDF15 0.757 0.013 0.743 0.006 GHR 0.712 0.004 0.679 0.001 GNRH1 0.791 0.048 GPM6B 0.675 <001 0.660 <001 0.735 <001 0.823 0.049 GSK3B 0.783 0.042 GSN 0.587 <001 0.705 0.002 0.745 0.004 0.796 0.021 GSTM1 0.686 0.001 0.631 <001 0.807 0.018 60 2015227398 15 Sep 20

Table 5B cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value GSTM2 0.607 <001 0.683 <001 0.679 <001 0.800 0.027 HIRIP3 0.692 <001 0.782 0.007 HK1 0.724 0.002 0.718 0.002 HLF 0.580 <001 0.571 <001 0.759 0.008 0.750 0.004 HNF1B 0.669 <001 HPS1 0.764 0.008 HSD17B10 0.802 0.045 HSD17B2 0.723 0.048 HSD3B2 0.709 0.010 HSP90AB1 0.780 0.034 0.809 0.041 HSPA5 0.738 0.017 HSPB1 0.770 0.006 0.801 0.032 HSPB2 0.788 0.035 ICAM1 0.728 0.015 0.716 0.010 IER3 0.735 0.016 0.637 <001 0.802 0.035 IFIT1 0.647 <001 0.755 0.029 IGF1 0.675 <001 0.603 <001 0.762 0.006 0.770 0.030 IGF2 0.761 0.011 IGFBP2 0.601 <001 0.605 <001 IGFBP5 0.702 <001 IGFBP6 0.628 <001 0.726 0.003 IL1B 0.676 0.002 0.716 0.004 IL6 0.688 0.005 0.766 0.044 IL6R 0.786 0.036 IL6ST 0.618 <001 0.639 <001 0.785 0.027 0.813 0.042 IL8 0.635 <001 0.628 <001 ILK 0.734 0.018 0.753 0.026 ING5 0.684 <001 0.681 <001 0.756 0.006 ITGA4 0.778 0.040 ITGA5 0.762 0.026 ITGA6 0.811 0.038 ITGA7 0.592 <001 0.715 0.006 0.710 0.002 ITGAD 0.576 0.006 ITGB4 0.693 0.003 ITPR1 0.789 0.029 JUN 0.572 <001 0.581 <001 0.777 0.019 JUNB 0.732 0.030 0.707 0.016 KCTD12 0.758 0.036 KIT 0.691 0.009 0.738 0.028 KLC1 0.741 0.024 0.781 0.024 KLF6 0.733 0.018 0.727 0.014 KLK1 0.744 0.028 61 2015227398 15 Sep 20

Table 5B cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value KLK2 0.697 0.002 0.679 <001 KLK3 0.725 <001 0.715 <001 0.841 0.023 KRT15 0.660 <001 0.577 <001 0.750 0.002 KRT18 0.623 <001 0.642 <001 0.702 <001 0.760 0.006 KRT2 0.740 0.044 KRT5 0.674 <001 0.588 <001 0.769 0.005 KRT8 0.768 0.034 LI CAM 0.737 0.036 LAG3 0.711 0.013 0.748 0.029 LAMA4 0.649 0.009 LAMB 3 0.709 0.002 0.684 0.006 0.768 0.006 LGALS3 0.652 <001 0.752 0.015 0.805 0.028 LIG3 0.728 0.016 0.667 <001 LRP1 0.811 0.043 MDM2 0.788 0.033 MGMT 0.645 <001 0.766 0.015 MICA 0.796 0.043 0.676 <001 MPPED2 0.675 <001 0.616 <001 0.750 0.006 MRC1 0.788 0.028 MTSS1 0.654 <001 0.793 0.036 MYBPC1 0.706 <001 0.534 <001 0.773 0.004 0.692 <001 NCAPD3 0.658 <001 0.566 <001 0.753 0.011 0.733 0.009 NCOR1 0.838 0.045 NEXN 0.748 0.025 0.785 0.020 NFAT5 0.531 <001 0.626 <001 NFATC2 0.759 0.024 OAZ1 0.766 0.024 OLFML3 0.648 <001 0.748 0.005 0.639 <001 0.675 <001 OR51E2 0.823 0.034 PAGE4 0.599 <001 0.698 0.002 0.606 <001 0.726 <001 PCA3 0.705 <001 0.647 <001 PCDHGB7 0.712 <001 PGF 0.790 0.039 PLG 0.764 0.048 PLP2 0.766 0.037 PPAP2B 0.589 <001 0.647 <001 0.691 <001 0.765 0.013 PPP1R12A 0.673 0.001 0.677 0.001 0.807 0.045 PRIM A1 0.622 <001 0.712 0.008 0.740 0.013 PRKCA 0.637 <001 0.694 <001 PRKCB 0.741 0.020 0.664 <001 PROM1 0.599 0.017 0.527 0.042 0.610 0.006 0.420 0.002 PTCH1 0.752 0.027 0.762 0.011 62 2015227398 15 Sep 20

Table 5B cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value PTEN 0.779 0.011 0.802 0.030 0.788 0.009 PTGS2 0.639 <001 0.606 <001 PTHLH 0.632 0.007 0.739 0.043 0.654 0.002 0.740 0.015 PTK2B 0.775 0.019 0.831 0.028 0.810 0.017 PTPN1 0.721 0.012 0.737 0.024 PYCARD 0.702 0.005 RAB27A 0.736 0.008 RAB30 0.761 0.011 RARB 0.746 0.010 RASSF1 0.805 0.043 RHOB 0.755 0.029 0.672 0.001 RLN1 0.742 0.036 0.740 0.036 RND3 0.607 <001 0.633 <001 RNF114 0.782 0.041 0.747 0.013 SDC2 0.714 0.002 SDHC 0.698 <001 0.762 0.029 SERPINA3 0.752 0.030 SERPINB5 0.669 0.014 SH3RF2 0.705 0.012 0.568 <001 0.755 0.016 SFC22A3 0.650 <001 0.582 <001 SMAD4 0.636 <001 0.684 0.002 0.741 0.007 0.738 0.007 SMARCD1 0.757 0.001 SMO 0.790 0.049 0.766 0.013 SOD1 0.741 0.037 0.713 0.007 SORBS 1 0.684 0.003 0.732 0.008 0.788 0.049 SPDEF 0.840 0.012 SPINT1 0.837 0.048 SRC 0.674 <001 0.671 <001 SRD5A2 0.553 <001 0.588 <001 0.618 <001 0.701 <001 ST5 0.747 0.012 0.761 0.010 0.780 0.016 0.832 0.041 STAT3 0.735 0.020 STAT5A 0.731 0.005 0.743 0.009 0.817 0.027 STAT5B 0.708 <001 0.696 0.001 SUMOl 0.815 0.037 SVIE 0.689 0.003 0.739 0.008 0.761 0.011 TBP 0.792 0.037 TFF3 0.719 0.007 0.664 0.001 TGFB1I1 0.676 0.003 0.707 0.007 0.709 0.005 0.777 0.035 TGFB2 0.741 0.010 0.785 0.017 TGFBR2 0.759 0.022 TIMP3 0.785 0.037 TMPRSS2 0.780 0.012 0.742 <001 63 2015227398 15 Sep 2015

Table 5B cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value TNF 0.654 0.007 0.682 0.006 TNFRSF10 B 0.623 <001 0.681 <001 0.801 0.018 0.815 0.019 TNFSF10 0.721 0.004 TP53 0.759 0.011 TP63 0.737 0.020 0.754 0.007 TPM2 0.609 <001 0.671 <001 0.673 <001 0.789 0.031 TRAF3IP2 0.795 0.041 0.727 0.005 TRO 0.793 0.033 0.768 0.027 0.814 0.023 TUBB2A 0.626 <001 0.590 <001 VCL 0.613 <001 0.701 0.011 VIM 0.716 0.005 0.792 0.025 WFDC1 0.824 0.029 YY1 0.668 <001 0.787 0.014 0.716 0.001 0.819 0.011 ZFHX3 0.732 <001 0.709 <001 ZFP36 0.656 0.001 0.609 <001 0.818 0.045 ZNF827 0.750 0.022 [00139] Tables 6A and 6B provide genes that were significantly associated (p<0.05), positively or negatively, with recurrence (cRFI, bRFI) after adjusting for Gleason pattern in the primary and/or highest Gleason pattern. Increased expression of genes in Table 6A is negatively associated with good prognosis, while increased expression of gene in Table 6B is positively associated with good prognosis.

Table 6A. Genes significantly (p<0.05) associated with cRFI or bRFI after adjustment for Gleason pattern in the primary Gleason pattern or highest Gleason pattern with a hazard ratio (HR) > 1.0 (increased expression is negatively associated with good prognosis)

Table 6A

cRFI

Primary Pattern

cRFI

bRFI

bRFI

Official Symbol AKR1C3 ANLN AQP2 ASAP2 ASPN BAG5 BAX BGN BIRC5 HR 1.258 1.292 1.178 1.809 1.465 1.338 p-value 0.039 0.023 0.008 <001 0.009 0.008

Highest Pattern

HR 1.287 1.396 1.508 1.367 1.342 p-value <001 0.015 0.009 0.012 0.046 64

Primary Pattern

HR 1.449 1.506 1.364 p-value <001 0.002 0.004

Highest Pattern

HR 1.420 1.438 1.234 1.279 p-value 0.001 0.002 0.044 0.006 2015227398 15 Sep 20

Table 6A cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value BMP6 1.369 0.015 1.518 0.002 BUB1 1.239 0.024 1.227 0.001 1.236 0.004 CACNA1D 1.337 0.025 CADPS 1.280 0.029 CCNE2 1.256 0.043 1.577 <001 1.324 0.001 CD276 1.320 0.029 1.396 0.007 1.279 0.033 CDC20 1.298 0.016 1.334 0.002 1.257 0.032 1.279 0.003 CDH7 1.258 0.047 1.338 0.013 CDKN2B 1.342 0.032 1.488 0.009 CDKN2C 1.344 0.010 1.450 <001 CDKN3 1.284 0.012 CENPF 1.289 0.048 1.498 0.001 1.344 0.010 COL1A1 1.481 0.003 1.506 0.002 COL3A1 1.459 0.004 1.430 0.013 COL4A1 1.396 0.015 COL8A1 1.413 0.008 CRISP3 1.346 0.012 1.310 0.025 CTHRC1 1.588 0.002 DDIT4 1.363 0.020 1.379 0.028 DICER 1 1.294 0.008 ENY2 1.269 0.024 FADD 1.307 0.010 FAS 1.243 0.025 FGF5 1.328 0.002 GNPTAB 1.246 0.037 GREM1 1.332 0.024 1.377 0.013 1.373 0.011 HDAC1 1.301 0.018 1.237 0.021 HSD17B4 1.277 0.011 IFN-γ 1.219 0.048 IMMT 1.230 0.049 INHBA 1.866 <001 1.944 <001 JAG1 1.298 0.030 KCNN2 1.378 0.020 1.282 0.017 KHDRBS3 1.353 0.029 1.305 0.014 LAMA3 1.344 <001 1.232 0.048 LAMC1 1.396 0.015 LIMS1 1.337 0.004 LOX 1.355 0.001 1.341 0.002 LTBP2 1.304 0.045 MAGEA4 1.215 0.024 MANF 1.460 <001 MCM6 1.287 0.042 1.214 0.046 65 2015227398 15 Sep 2015

Table 6A cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value MELK 1.329 0.002 MMP11 1.281 0.050 MRPL13 1.266 0.021 MYBL2 1.453 <001 1.274 0.019 MYC 1.265 0.037 MY06 1.278 0.047 NET02 1.322 0.022 NFKB1 1.255 0.032 NOX4 1.266 0.041 OR51E1 1.566 <001 1.428 0.003 PATE1 1.242 <001 1.347 <001 1.177 0.011 PCNA 1.251 0.025 PEX10 1.302 0.028 PGD 1.335 0.045 1.379 0.014 1.274 0.025 PIM1 1.254 0.019 PLA2G7 1.289 0.025 1.250 0.031 PLAU 1.267 0.031 PSMD13 1.333 0.005 PTK6 1.432 <001 1.577 <001 1.223 0.040 PTTG1 1.279 0.013 1.308 0.006 RAGE 1.329 0.011 RALA 1.363 0.044 1.471 0.003 RGS7 1.120 0.040 1.173 0.031 RRM1 1.490 0.004 1.527 <001 SESN3 1.353 0.017 SFRP4 1.370 0.025 SHMT2 1.460 0.008 1.410 0.006 1.407 0.008 1.345 <001 SKIL 1.307 0.025 SLC25A21 1.414 0.002 1.330 0.004 SMARCC2 1.219 0.049 SPARC 1.431 0.005 TFDP1 1.283 0.046 1.345 0.003 THBS2 1.456 0.005 1.431 0.012 TK1 1.214 0.015 1.222 0.006 TOP2A 1.367 0.018 1.518 0.001 1.480 <001 TPX2 1.513 0.001 1.607 <001 1.588 <001 1.481 <001 UBE2T 1.409 0.002 1.285 0.018 UGT2B15 1.216 0.009 1.182 0.021 XIAP 1.336 0.037 1.194 0.043 66 2015227398 15 Sep 2015

Table 6B. Genes significantly (p<0.05) associated with cRFI or bRFI after adjustment for Gleason pattern in the primary Gleason pattern or highest Gleason pattern with hazard ration (HR) < 1.0 (increased expression is positively associated with good prognosis) Table 6B cRFI cRFI bRFI T bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value AAMP 0.660 0.001 0.675 <001 0.836 0.045 ABCA5 0.807 0.014 0.737 <001 0.845 0.030 ABCC1 0.780 0.038 0.794 0.015 ABCG2 0.807 0.035 ABHD2 0.720 0.002 ADH5 0.750 0.034 AKAP1 0.721 <001 ALDH1A2 0.735 0.009 0.592 <001 0.756 0.007 0.781 0.021 ANGPT2 0.741 0.036 ANPEP 0.637 <001 0.536 <001 ANXA2 0.762 0.044 APOE 0.707 0.013 APRT 0.727 0.004 0.771 0.006 ATXN1 0.725 0.013 AURKA 0.784 0.037 0.735 0.003 AXIN2 0.744 0.004 0.630 <001 AZGP1 0.672 <001 0.720 <001 0.764 0.001 BAD 0.687 <001 BAK1 0.783 0.014 BCL2 0.777 0.033 0.772 0.036 BIK 0.768 0.040 BIN1 0.691 <001 BTRC 0.776 0.029 C7 0.707 0.004 0.791 0.024 CADM1 0.587 <001 0.593 <001 CASP1 0.773 0.023 0.820 0.025 CAV1 0.753 0.014 CAV2 0.627 0.009 0.682 0.003 CCL2 0.740 0.019 CCNH 0.736 0.003 CCR1 0.755 0.022 CD1A 0.740 0.025 CD44 0.590 <001 0.637 <001 CD68 0.757 0.026 CD82 0.778 0.012 0.759 0.016 CDC25B 0.760 0.021 CDK3 0.762 0.024 0.774 0.007 CDKN1A 0.714 0.015 67 2015227398 15 Sep 2015

Table 6B cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value CDKN1C 0.738 0.014 0.768 0.021 COL6A1 0.690 <001 0.805 0.048 CSF1 0.675 0.002 0.779 0.036 CSK 0.825 0.004 CTNNB1 0.884 0.045 0.888 0.027 CTSB 0.740 0.017 0.676 0.003 0.755 0.010 CTSD 0.673 0.031 0.722 0.009 CTSK 0.804 0.034 CTSL2 0.748 0.019 CXCL12 0.731 0.017 CYP3A5 0.523 <001 0.518 <001 CYR61 0.744 0.041 DAP 0.755 0.011 DARC 0.763 0.029 DDR2 0.813 0.041 DES 0.743 0.020 DHRS9 0.606 0.001 DHX9 0.916 0.021 DIAPH1 0.749 0.036 0.688 0.003 DLGAP1 0.758 0.042 0.676 0.002 DLL4 0.779 0.010 DNM3 0.732 0.007 DPP4 0.732 0.004 0.750 0.014 DPT 0.704 0.014 DUSP6 0.662 <001 0.665 0.001 EBNA1BP2 0.828 0.019 EDNRA 0.782 0.048 EGF 0.712 0.023 EGR1 0.678 0.004 0.725 0.028 EGR3 0.680 0.006 0.738 0.027 EIF2C2 0.789 0.032 EIF2S3 0.759 0.012 ELK4 0.745 0.024 EPHA2 0.661 0.007 EPHA3 0.781 0.026 0.828 0.037 ERBB2 0.791 0.022 0.760 0.014 0.789 0.006 ERBB3 0.757 0.009 ERCC1 0.760 0.008 ESR1 0.742 0.014 ESR2 0.711 0.038 ETV4 0.714 0.035 FAM107A 0.619 <001 0.710 0.011 0.781 0.019 68 2015227398 15 Sep 2015

Table 6B cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value FAM13C 0.664 <001 0.686 <001 0.813 0.014 FAM49B 0.670 <001 0.793 0.014 0.815 0.044 0.843 0.047 FASLG 0.616 0.004 0.813 0.038 FGF10 0.751 0.028 0.766 0.019 FGF17 0.718 0.031 0.765 0.019 FGFR2 0.740 0.009 0.738 0.002 FKBP5 0.749 0.031 FLNC 0.826 0.029 FLT1 0.779 0.045 0.729 0.006 FLT4 0.815 0.024 FOS 0.657 0.003 0.656 0.004 FSD1 0.763 0.017 FYN 0.716 0.004 0.792 0.024 GADD45B 0.692 0.009 0.697 0.010 GDF15 0.767 0.016 GHR 0.701 0.002 0.704 0.002 0.640 <001 GNRH1 0.778 0.039 GPM6B 0.749 0.010 0.750 0.010 0.827 0.037 GRB7 0.696 0.005 GSK3B 0.726 0.005 GSN 0.660 <001 0.752 0.019 GSTM1 0.710 0.004 0.676 <001 GSTM2 0.643 <001 0.767 0.015 HK1 0.798 0.035 HLA-G 0.660 0.013 HLF 0.644 <001 0.727 0.011 HNF1B 0.755 0.013 HPS1 0.756 0.006 0.791 0.043 HSD17B10 0.737 0.006 HSD3B2 0.674 0.003 HSP90AB1 0.763 0.015 HSPB1 0.787 0.020 0.778 0.015 HSPE1 0.794 0.039 ICAM1 0.664 0.003 IER3 0.699 0.003 0.693 0.010 IFIT1 0.621 <001 0.733 0.027 IGF1 0.751 0.017 0.655 <001 IGFBP2 0.599 <001 0.605 <001 IGFBP5 0.745 0.007 0.775 0.035 IGFBP6 0.671 0.005 IL1B 0.732 0.016 0.717 0.005 IL6 0.763 0.040 69 2015227398 15 Sep 2015

Table 6B cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value IL6R 0.764 0.022 IL6ST 0.647 <001 0.739 0.012 IL8 0.711 0.015 0.694 0.006 ING5 0.729 0.007 0.727 0.003 ITGA4 0.755 0.009 ITGA5 0.743 0.018 0.770 0.034 ITGA6 0.816 0.044 0.772 0.006 ITGA7 0.680 0.004 ITGAD 0.590 0.009 ITGB4 0.663 <001 0.658 <001 0.759 0.004 JUN 0.656 0.004 0.639 0.003 KIAA0196 0.737 0.011 KIT 0.730 0.021 0.724 0.008 KLC1 0.755 0.035 KLK1 0.706 0.008 KLK2 0.740 0.016 0.723 0.001 KLK3 0.765 0.006 0.740 0.002 KRT1 0.774 0.042 KRT15 0.658 <001 0.632 <001 0.764 0.008 KRT18 0.703 0.004 0.672 <001 0.779 0.015 0.811 0.032 KRT5 0.686 <001 0.629 <001 0.802 0.023 KRT8 0.763 0.034 0.771 0.022 L1CAM 0.748 0.041 LAG3 0.693 0.008 0.724 0.020 LAMA4 0.689 0.039 LAMB 3 0.667 <001 0.645 <001 0.773 0.006 LGALS3 0.666 <001 0.822 0.047 LIG3 0.723 0.008 LRP1 0.777 0.041 0.769 0.007 MDM2 0.688 <001 MET 0.709 0.010 0.736 0.028 0.715 0.003 MGMT 0.751 0.031 MICA 0.705 0.002 MPPED2 0.690 0.001 0.657 <001 0.708 <001 MRC1 0.812 0.049 MSH6 0.860 0.049 MTSS1 0.686 0.001 MVP 0.798 0.034 0.761 0.033 MYBPC1 0.754 0.009 0.615 <001 NCAPD3 0.739 0.021 0.664 0.005 NEXN 0.798 0.037 NFAT5 0.596 <001 0.732 0.005 70 2015227398 15 Sep 20

Table 6B cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value NFATC2 0.743 0.016 0.792 0.047 NOS3 0.730 0.012 0.757 0.032 OAZ1 0.732 0.020 0.705 0.002 OCLN 0.746 0.043 0.784 0.025 OLFML3 0.711 0.002 0.709 <001 0.720 0.001 OMD 0.729 0.011 0.762 0.033 OSM 0.813 0.028 PAGE4 0.668 0.003 0.725 0.004 0.688 <001 0.766 0.005 PC A3 0.736 0.001 0.691 <001 PCDHGB7 0.769 0.019 0.789 0.022 PIK3CA 0.768 0.010 PIK3CG 0.792 0.019 0.758 0.009 PLG 0.682 0.009 PPAP2B 0.688 0.005 0.815 0.046 PPP1R12A 0.731 0.026 0.775 0.042 PRIM A1 0.697 0.004 0.757 0.032 PRKCA 0.743 0.019 PRKCB 0.756 0.036 0.767 0.029 PROM1 0.640 0.027 0.699 0.034 0.503 0.013 PTCH1 0.730 0.018 PTEN 0.779 0.015 0.789 0.007 PTGS2 0.644 <001 0.703 0.007 PTHLH 0.655 0.012 0.706 0.038 0.634 0.001 0.665 0.003 PTK2B 0.779 0.023 0.702 0.002 0.806 0.015 0.806 0.024 PYCARD 0.659 0.001 RAB30 0.779 0.033 0.754 0.014 RARB 0.787 0.043 0.742 0.009 RASSF1 0.754 0.005 RHOA 0.796 0.041 0.819 0.048 RND3 0.721 0.011 0.743 0.028 SDC1 0.707 0.011 SDC2 0.745 0.002 SDHC 0.750 0.013 SERPINA3 0.730 0.016 SERPINB5 0.715 0.041 SH3RF2 0.698 0.025 SIPA1L1 0.796 0.014 0.820 0.004 SLC22A3 0.724 0.014 0.700 0.008 SMAD4 0.668 0.002 0.771 0.016 SMARCD1 0.726 <001 0.700 0.001 0.812 0.028 SMO 0.785 0.027 SOD1 0.735 0.012 71 2015227398 15 Sep 2015

Table 6B cRFI cRFI bRFI bRFI Primary Pattern Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HR p-value SORBS 1 0.785 0.039 SPDEF 0.818 0.002 SPINT1 0.761 0.024 0.773 0.006 SRC 0.709 <001 0.690 <001 SRD5A1 0.746 0.010 0.767 0.024 0.745 0.003 SRD5A2 0.575 <001 0.669 0.001 0.674 <001 0.781 0.018 ST5 0.774 0.027 STAT1 0.694 0.004 STAT5A 0.719 0.004 0.765 0.006 0.834 0.049 STAT5B 0.704 0.001 0.744 0.012 SUMOl 0.777 0.014 SVIL 0.771 0.026 TBP 0.774 0.031 TFF3 0.742 0.015 0.719 0.024 TGFB1I1 0.763 0.048 TGFB2 0.729 0.011 0.758 0.002 TMPRSS2 0.810 0.034 0.692 <001 TNF 0.727 0.022 TNFRSF10A 0.805 0.025 TNFRSF10B 0.581 <001 0.738 0.014 0.809 0.034 TNFSF10 0.751 0.015 0.700 <001 TP63 0.723 0.018 0.736 0.003 TPM2 0.708 0.010 0.734 0.014 TRAF3IP2 0.718 0.004 TRO 0.742 0.012 TSTA3 0.774 0.028 TUBB2A 0.659 <001 0.650 <001 TYMP 0.695 0.002 VCL 0.683 0.008 VIM 0.778 0.040 WDR19 0.775 0.014 XRCC5 0.793 0.042 YY1 0.751 0.025 0.810 0.008 ZFHX3 0.760 0.005 0.726 0.001 ZFP36 0.707 0.008 0.672 0.003 ZNF827 0.667 0.002 0.792 0.039 [00140] Tables 7 A and 7B provide genes significantly associated (p<0.05), positively or negatively, with clinical recurrence (cRFI) in negative TMPRSS fusion specimens in the primary or highest Gleason pattern specimen. Increased expression of genes in Table 7A is negatively 72 2015227398 15 Sep 2015 associated with good prognosis, while increased expression of genes in Table 7B is positively associated with good prognosis.

Table 7A.

Genes significantly (p<0.05) associated with cRFI for TMPRSS2-ERG fusion negative in the primary Gleason pattern or highest Gleason pattern with hazard ratio (HR) >1.0 (increased expression is negatively associated with good prognosis)

Table 7A Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value ANLN 1.42 0.012 1.36 0.004 AQP2 1.25 0.033 ASPN 2.48 <.001 1.65 <001 BGN 2.04 <.001 1.45 0.007 BIRC5 1.59 <.001 1.37 0.005 BMP6 1.95 <.001 1.43 0.012 BMPR1B 1.93 0.002 BUB1 1.51 <.001 1.35 <001 CCNE2 1.48 0.007 CD276 1.93 <.001 1.79 <001 CDC20 1.49 0.004 1.47 <001 CDC6 1.52 0.009 1.34 0.022 CDKN2B 1.54 0.008 1.55 0.003 CDKN2C 1.55 0.003 1.57 <001 CDKN3 1.34 0.026 CENPF 1.63 0.002 1.33 0.018 CKS2 1.50 0.026 1.43 0.009 CLTC 1.46 0.014 COL1A1 1.98 <001 1.50 0.002 COL3A1 2.03 <001 1.42 0.007 COL4A1 1.81 0.002 COL8A1 1.63 0.004 1.60 0.001 CRISP3 1.31 0.016 CTHRC1 1.67 0.006 1.48 0.005 DDIT4 1.49 0.037 ENY2 1.29 0.039 EZH2 1.35 0.016 F2R 1.46 0.034 1.46 0.007 FAP 1.66 0.006 1.38 0.012 FGF5 1.46 0.001 GNPTAB 1.49 0.013 HSD17B4 1.34 0.039 1.44 0.002 INHBA 2.92 <001 2.19 <001 JAG1 1.38 0.042 KCNN2 1.71 0.002 1.73 <001 KHDRBS3 1.46 0.015 KLK14 1.28 0.034 73 2015227398 15 Sep 2015

Table 7A Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value KPNA2 1.63 0.016 LAMC1 1.41 0.044 LOX 1.29 0.036 LTBP2 1.57 0.017 MELK 1.38 0.029 MMP11 1.69 0.002 1.42 0.004 MYBL2 1.78 <001 1.49 <001 NET02 2.01 <001 1.43 0.007 NME1 1.38 0.017 PATE1 1.43 <001 1.24 0.005 PEX10 1.46 0.030 PGD 1.77 0.002 POSTN 1.49 0.037 1.34 0.026 PPFIA3 1.51 0.012 PPP3CA 1.46 0.033 1.34 0.020 PTK6 1.69 <001 1.56 <001 PTTG1 1.35 0.028 RAD51 1.32 0.048 RALBP1 1.29 0.042 RGS7 1.18 0.012 1.32 0.009 RRM1 1.57 0.016 1.32 0.041 RRM2 1.30 0.039 SAT1 1.61 0.007 SESN3 1.76 <001 1.36 0.020 SFRP4 1.55 0.016 1.48 0.002 SHMT2 2.23 <001 1.59 <001 SPARC 1.54 0.014 SQLE 1.86 0.003 STMN1 2.14 <001 THBS2 1.79 <001 1.43 0.009 TK1 1.30 0.026 TOP2A 2.03 <001 1.47 0.003 TPD52 1.63 0.003 TPX2 2.11 <001 1.63 <001 TRAP1 1.46 0.023 UBE2C 1.57 <001 1.58 <001 UBE2G1 1.56 0.008 UBE2T 1.75 <001 UGT2B15 1.31 0.036 1.33 0.004 UHRF1 1.46 0.007 UTP23 1.52 0.017 74 2015227398 15 Sep 2015

Table 7B. Genes significantly (p<0.05) associated with cRFI for TMPRSS2-ERG fusion negative in the primary Gleason pattern or highest Gleason pattern with hazard ratio (HR) < 1.0 (increased expression is positively associated with good prognosis) Table 7B Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value AAMP 0.56 <001 0.65 0.001 ABCA5 0.64 <001 0.71 <001 ABCB1 0.62 0.004 ABCC3 0.74 0.031 ABCG2 0.78 0.050 ABHD2 0.71 0.035 ACOX2 0.54 <001 0.71 0.007 ADH5 0.49 <001 0.61 <001 AKAP1 0.77 0.031 0.76 0.013 AKR1C1 0.65 0.006 0.78 0.044 AKT1 0.72 0.020 AKT3 0.75 <001 ALDH1A2 0.53 <001 0.60 <001 AMPD3 0.62 <001 0.78 0.028 ANPEP 0.54 <001 0.61 <001 ANXA2 0.63 0.008 0.74 0.016 ARHGAP29 0.67 0.005 0.77 0.016 ARHGDIB 0.64 0.013 ATP5J 0.57 0.050 ATXN1 0.61 0.004 0.77 0.043 AXIN2 0.51 <001 0.62 <001 AZGP1 0.61 <001 0.64 <001 BCL2 0.64 0.004 0.75 0.029 BIN1 0.52 <001 0.74 0.010 BTG3 0.75 0.032 0.75 0.010 BTRC 0.69 0.011 C7 0.51 <001 0.67 <001 CADM1 0.49 <001 0.76 0.034 CASP1 0.71 0.010 0.74 0.007 CAV1 0.73 0.015 CCL5 0.67 0.018 0.67 0.003 CCNH 0.63 <001 0.75 0.004 CCR1 0.77 0.032 CD 164 0.52 <001 0.63 <001 CD44 0.53 <001 0.74 0.014 CDH10 0.69 0.040 CDH18 0.40 0.011 CDK14 0.75 0.013 CDK2 0.81 0.031 CDK3 0.73 0.022 75 2015227398 15 Sep 20

Table 7B Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value CDKN1A 0.68 0.038 CDKN1C 0.62 0.003 0.72 0.005 COL6A1 0.54 <001 0.70 0.004 COL6A3 0.64 0.004 CSF1 0.56 <001 0.78 0.047 CSRP1 0.40 <001 0.66 0.002 CTGF 0.66 0.015 0.74 0.027 CTNNB1 0.69 0.043 CTSB 0.60 0.002 0.71 0.011 CTSS 0.67 0.013 CXCL12 0.56 <001 0.77 0.026 CYP3A5 0.43 <001 0.63 <001 CYR61 0.43 <001 0.58 <001 DAG1 0.72 0.012 DARC 0.66 0.016 DDR2 0.65 0.007 DES 0.52 <001 0.74 0.018 DHRS9 0.54 0.007 DICER 1 0.70 0.044 DLC1 0.75 0.021 DLGAP1 0.55 <001 0.72 0.005 DNM3 0.61 0.001 DPP4 0.55 <001 0.77 0.024 DPT 0.48 <001 0.61 <001 DUSP1 0.47 <001 0.59 <001 DUSP6 0.65 0.009 0.65 0.002 DYNLL1 0.74 0.045 EDNRA 0.61 0.002 0.75 0.038 EFNB2 0.71 0.043 EGR1 0.43 <001 0.58 <001 EGR3 0.47 <001 0.66 <001 EIF5 0.77 0.028 ELK4 0.49 <001 0.72 0.012 EPHA2 0.70 0.007 EPHA3 0.62 <001 0.72 0.009 EPHB2 0.68 0.009 ERBB2 0.64 <001 0.63 <001 ERBB3 0.69 0.018 ERCC1 0.69 0.019 0.77 0.021 ESR2 0.61 0.020 FAAH 0.57 <001 0.77 0.035 FABP5 0.67 0.035 FAM107A 0.42 <001 0.59 <001 FAM13C 0.53 <001 0.59 <001 76 2015227398 15 Sep 20

Table 7B Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value FAS 0.71 0.035 FASLG 0.56 0.017 0.67 0.014 FGF10 0.57 0.002 FGF17 0.70 0.039 0.70 0.010 FGF7 0.63 0.005 0.70 0.004 FGFR2 0.63 0.003 0.71 0.003 FKBP5 0.72 0.020 FLNA 0.48 <001 0.74 0.022 FOS 0.45 <001 0.56 <001 FOXOl 0.59 <001 FOXQ1 0.57 <001 0.69 0.008 FYN 0.62 0.001 0.74 0.013 G6PD 0.77 0.014 GADD45A 0.73 0.045 GADD45B 0.45 <001 0.64 0.001 GDF15 0.58 <001 GHR 0.62 0.008 0.68 0.002 GPM6B 0.60 <001 0.70 0.003 GSK3B 0.71 0.016 0.71 0.006 GSN 0.46 <001 0.66 <001 GSTM1 0.56 <001 0.62 <001 GSTM2 0.47 <001 0.67 <001 HGD 0.72 0.002 HIRIP3 0.69 0.021 0.69 0.002 HK1 0.68 0.005 0.73 0.005 HLA-G 0.54 0.024 0.65 0.013 HLF 0.41 <001 0.68 0.001 HNF1B 0.55 <001 0.59 <001 HPS1 0.74 0.015 0.76 0.025 HSD17B3 0.65 0.031 HSPB2 0.62 0.004 0.76 0.027 ICAM1 0.61 0.010 IER3 0.55 <001 0.67 0.003 IFIT1 0.57 <001 0.70 0.008 IFNG 0.69 0.040 IGF1 0.63 <001 0.59 <001 IGF2 0.67 0.019 0.70 0.005 IGFBP2 0.53 <001 0.63 <001 IGFBP5 0.57 <001 0.71 0.006 IGFBP6 0.41 <001 0.71 0.012 IL10 0.59 0.020 IL1B 0.53 <001 0.70 0.005 IL6 0.55 0.001 IL6ST 0.45 <001 0.68 <001 77 2015227398 15 Sep 2015

Table 7B Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value IL8 0.60 0.005 0.70 0.008 ILK 0.68 0.029 0.76 0.036 ING5 0.54 <001 0.82 0.033 ITGA1 0.66 0.017 ITGA3 0.70 0.020 ITGA5 0.64 0.011 ITGA6 0.66 0.003 0.74 0.006 ITGA7 0.50 <001 0.71 0.010 ITGB4 0.63 0.014 0.73 0.010 ITPR1 0.55 <001 ITPR3 0.76 0.007 JUN 0.37 <001 0.54 <001 JUNB 0.58 0.002 0.71 0.016 KCTD12 0.68 0.017 KIT 0.49 0.002 0.76 0.043 KLC1 0.61 0.005 0.77 0.045 KLF6 0.65 0.009 KLK1 0.68 0.036 KLK10 0.76 0.037 KLK2 0.64 <001 0.73 0.006 KLK3 0.65 <001 0.76 0.021 KLRK1 0.63 0.005 KRT15 0.52 <001 0.58 <001 KRT18 0.46 <001 KRT5 0.51 <001 0.58 <001 KRT8 0.53 <001 L1CAM 0.65 0.031 LAG3 0.58 0.002 0.76 0.033 LAMA4 0.52 0.018 LAMB 3 0.60 0.002 0.65 0.003 LGALS3 0.52 <001 0.71 0.002 LIG3 0.65 0.011 LRP1 0.61 0.001 0.75 0.040 MGMT 0.66 0.003 MICA 0.59 0.001 0.68 0.001 MLXIP 0.70 0.020 MMP2 0.68 0.022 MMP9 0.67 0.036 MPPED2 0.57 <001 0.66 <001 MRC1 0.69 0.028 MTSS1 0.63 0.005 0.79 0.037 MVP 0.62 <001 MYBPC1 0.53 <001 0.70 0.011 NCAM1 0.70 0.039 0.77 0.042 78 2015227398 15 Sep 20

Table 7B Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value NCAPD3 0.52 <001 0.59 <001 NDRG1 0.69 0.008 NEXN 0.62 0.002 NFAT5 0.45 <001 0.59 <001 NFATC2 0.68 0.035 0.75 0.036 NFKBIA 0.70 0.030 NRG1 0.59 0.022 0.71 0.018 OAZ1 0.69 0.018 0.62 <001 OFFMF3 0.59 <001 0.72 0.003 OR51E2 0.73 0.013 PAGE4 0.42 <001 0.62 <001 PC A3 0.53 <001 PCDHGB7 0.70 0.032 PGF 0.68 0.027 0.71 0.013 PGR 0.76 0.041 PIK3C2A 0.80 <001 PIK3CA 0.61 <001 0.80 0.036 PIK3CG 0.67 0.001 0.76 0.018 PFP2 0.65 0.015 0.72 0.010 PPAP2B 0.45 <001 0.69 0.003 PPP1R12A 0.61 0.007 0.73 0.017 PRIM A1 0.51 <001 0.68 0.004 PRKCA 0.55 <001 0.74 0.009 PRKCB 0.55 <001 PROM1 0.67 0.042 PROS1 0.73 0.036 PTCH1 0.69 0.024 0.72 0.010 PTEN 0.54 <001 0.64 <001 PTGS2 0.48 <001 0.55 <001 PTH1R 0.57 0.003 0.77 0.050 PTHLH 0.55 0.010 PTK2B 0.56 <001 0.70 0.001 PYCARD 0.73 0.009 RAB27A 0.65 0.009 0.71 0.014 RAB30 0.59 0.003 0.72 0.010 RAGE 0.76 0.011 RARB 0.59 <001 0.63 <001 RASSF1 0.67 0.003 RBI 0.67 0.006 RFX1 0.71 0.040 0.70 0.003 RHOA 0.71 0.038 0.65 <001 RHOB 0.58 0.001 0.71 0.006 RND3 0.54 <001 0.69 0.003 RNF114 0.59 0.004 0.68 0.003 79 2015227398 15 Sep 20

Table 7B Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value SCUBE2 0.77 0.046 SDHC 0.72 0.028 0.76 0.025 SEC23A 0.75 0.029 SEMA3A 0.61 0.004 0.72 0.011 SEPT9 0.66 0.013 0.76 0.036 SERPINB5 0.75 0.039 SH3RF2 0.44 <001 0.48 <001 SHH 0.74 0.049 SLC22A3 0.42 <001 0.61 <001 SMAD4 0.45 <001 0.66 <001 SMARCD1 0.69 0.016 SOD1 0.68 0.042 SORBS 1 0.51 <001 0.73 0.012 SPARCL1 0.58 <001 0.77 0.040 SPDEF 0.77 <001 SPINT1 0.65 0.004 0.79 0.038 SRC 0.61 <001 0.69 0.001 SRD5A2 0.39 <001 0.55 <001 ST5 0.61 <001 0.73 0.012 STAT1 0.64 0.006 STAT3 0.63 0.010 STAT5A 0.62 0.001 0.70 0.003 STAT5B 0.58 <001 0.73 0.009 SUMOl 0.66 <001 SVIL 0.57 0.001 0.74 0.022 TBP 0.65 0.002 TFF1 0.65 0.021 TFF3 0.58 <001 TGFB1I1 0.51 <001 0.75 0.026 TGFB2 0.48 <001 0.62 <001 TGFBR2 0.61 0.003 TIAM1 0.68 0.019 TIMP2 0.69 0.020 TIMP3 0.58 0.002 TNFRSF10A 0.73 0.047 TNFRSF10B 0.47 <001 0.70 0.003 TNFSF10 0.56 0.001 TP63 0.67 0.001 TPM1 0.58 0.004 0.73 0.017 TPM2 0.46 <001 0.70 0.005 TRA2A 0.68 0.013 TRAF3IP2 0.73 0.041 0.71 0.004 TRO 0.72 0.016 0.71 0.004 TUBB2A 0.53 <001 0.73 0.021 80 2015227398 15 Sep 2015

Table 7B Primary Pattern Highest Pattern Official Symbol HR p-value HR P-value TYMP 0.70 0.011 VC AMI 0.69 0.041 VCL 0.46 <001 VEGFA 0.77 0.039 VEGFB 0.71 0.035 VIM 0.60 0.001 XRCC5 0.75 0.026 YY1 0.62 0.008 0.77 0.039 ZFHX3 0.53 <001 0.58 <001 ZFP36 0.43 <001 0.54 <001 ZNF827 0.55 0.001 [00141] Tables 8A and 8B provide genes that were significantly associated (p<0.05), positively or negatively, with clinical recurrence (cRFI) in positive TMPRSS fusion specimens in the primary or highest Gleason pattern specimen. Increased expression of genes in Table 8A is negatively associated with good prognosis, while increased expression of genes in Table 8B is positively associated with good prognosis.

Table 8A. Genes significantly (p<0.05) associated with cRFI for TMPRSS2-ERG fusion positive in the primary Gleason pattern or highest Gleason pattern with hazard ratio (HR) >1.0 (increased expression is negatively associated with good prognosis) Table 8A Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value ACTR2 1.78 0.017 AKR1C3 1.44 0.013 ALCAM 1.44 0.022 ANLN 1.37 0.046 1.81 <001 APOE 1.49 0.023 1.66 0.005 AQP2 1.30 0.013 ARHGDIB 1.55 0.021 ASPN 2.13 <001 2.43 <001 ATP5E 1.69 0.013 1.58 0.014 BGN 1.92 <001 2.55 <001 BIRC5 1.48 0.006 1.89 <001 BMP6 1.51 0.010 1.96 <001 BRCA2 1.41 0.007 BUB1 1.36 0.007 1.52 <001 CCNE2 1.55 0.004 1.59 <001 CD276 1.65 <001 CDC20 1.68 <001 1.74 <001 CDH11 1.50 0.017 CDH18 1.36 <001 81 2015227398 15 Sep 20

Table 8A Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value CDH7 1.54 0.009 1.46 0.026 CDKN2B 1.68 0.008 1.93 0.001 CDKN2C 2.01 <001 1.77 <001 CDKN3 1.51 0.002 1.33 0.049 CENPF 1.51 0.007 2.04 <001 CKS2 1.43 0.034 1.56 0.007 COL1A1 2.23 <001 3.04 <001 COL1A2 1.79 0.001 2.22 <001 COL3A1 1.96 <001 2.81 <001 COL4A1 1.52 0.020 COL5A1 1.50 0.020 COL5A2 1.64 0.017 1.55 0.010 COL8A1 1.96 <001 2.38 <001 CRISP3 1.68 0.002 1.67 0.002 CTHRC1 2.06 <001 CTNND2 1.42 0.046 1.50 0.025 CTSK 1.43 0.049 CXCR4 1.82 0.001 1.64 0.007 DDIT4 1.54 0.016 1.58 0.009 DLL4 1.51 0.007 DYNLL1 1.50 0.021 1.22 0.002 F2R 2.27 <001 2.02 <001 FAP 2.12 <001 FCGR3A 1.94 0.002 FGF5 1.23 0.047 FOXP3 1.52 0.006 1.48 0.018 GNPTAB 1.44 0.042 GPR68 1.51 0.011 GREM1 1.91 <001 2.38 <001 HDAC1 1.43 0.048 HDAC9 1.65 <001 1.67 0.004 HRAS 1.65 0.005 1.58 0.021 IGFBP3 1.94 <001 1.85 <001 INHBA 2.03 <001 2.64 <001 JAG1 1.41 0.027 1.50 0.008 KCTD12 1.51 0.017 KHDRBS3 1.48 0.029 1.54 0.014 KPNA2 1.46 0.050 FAMA3 1.35 0.040 FAMC1 1.77 0.012 FTBP2 1.82 <001 FUM 1.51 0.021 1.53 0.009 MEEK 1.38 0.020 1.49 0.001 MKI67 1.37 0.014 82 2015227398 15 Sep 2015

Table 8A Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value MMP11 1.73 <001 1.69 <001 MRPL13 1.30 0.046 MYBL2 1.56 <001 1.72 <001 MYLK3 1.17 0.007 NOX4 1.58 0.005 1.96 <001 NRIP3 1.30 0.040 NRP1 1.53 0.021 OLFML2B 1.54 0.024 OSM 1.43 0.018 PATE1 1.20 <001 1.33 <001 PCNA 1.64 0.003 PEX10 1.41 0.041 1.64 0.003 PIK3CA 1.38 0.037 PLK1 1.52 0.009 1.67 0.002 PLOD2 1.65 0.002 POSTN 1.79 <001 2.06 <001 PTK6 1.67 0.002 2.38 <001 PTTG1 1.56 0.002 1.54 0.003 RAD21 1.61 0.036 1.53 0.005 RAD51 1.33 0.009 RALA 1.95 0.004 1.60 0.007 REG4 1.43 0.042 R0B02 1.46 0.024 RRM1 1.44 0.033 RRM2 1.50 0.003 1.48 <001 SAT1 1.42 0.009 1.43 0.012 SEC14L1 1.64 0.002 SFRP4 2.07 <001 2.40 <001 SHMT2 1.52 0.030 1.60 0.001 SLC44A1 1.42 0.039 SPARC 1.93 <001 2.21 <001 SULF1 1.63 0.006 2.04 <001 THBS2 1.95 <001 2.26 <001 THY1 1.69 0.016 1.95 0.002 TK1 1.43 0.003 TOP2A 1.57 0.002 2.11 <001 TPX2 1.84 <001 2.27 <001 UBE2C 1.41 0.011 1.44 0.006 UBE2T 1.63 0.001 UHRF1 1.51 0.007 1.69 <001 WISP1 1.47 0.045 WNT5A 1.35 0.027 1.63 0.001 ZWINT 1.36 0.045 83 2015227398 15 Sep 2015

Table 8B. Genes significantly (p<0.05) associated with cRFI for TMPRSS2-ERG fusion positive in the primary Gleason pattern or highest Gleason pattern with hazard ratio (HR) < 1.0 (increased expression is positively associated with good prognosis) Table 8B Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value AAMP 0.57 0.007 0.58 <001 ABCA5 0.80 0.044 ACE 0.65 0.023 0.55 <001 ACOX2 0.55 <001 ADH5 0.68 0.022 AKAP1 0.81 0.043 ALDH1A2 0.72 0.036 0.43 <001 ANPEP 0.66 0.022 0.46 <001 APRT 0.73 0.040 AXIN2 0.60 <001 AZGP1 0.57 <001 0.65 <001 BCL2 0.69 0.035 BIK 0.71 0.045 BIN1 0.71 0.004 0.71 0.009 BTRC 0.66 0.003 0.58 <001 C7 0.64 0.006 CADM1 0.61 <001 0.47 <001 CCL2 0.73 0.042 CCNH 0.69 0.022 CD44 0.56 <001 0.58 <001 CD82 0.72 0.033 CDC25B 0.74 0.028 CDH1 0.75 0.030 0.72 0.010 CDH19 0.56 0.015 CDK3 0.78 0.045 CDKN1C 0.74 0.045 0.70 0.014 CSF1 0.72 0.037 CTSB 0.69 0.048 CTSL2 0.58 0.005 CYP3A5 0.51 <001 0.30 <001 DHX9 0.89 0.006 0.87 0.012 DLC1 0.64 0.023 DLGAP1 0.69 0.010 0.49 <001 DPP4 0.64 <001 0.56 <001 DPT 0.63 0.003 EGR1 0.69 0.035 EGR3 0.68 0.025 EIF2S3 0.70 0.021 EIF5 0.71 0.030 ELK4 0.71 0.041 0.60 0.003 84 2015227398 15 Sep 2015

Table 8B Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value EPHA2 0.72 0.036 0.66 0.011 EPHB4 0.65 0.007 ERCC1 0.68 0.023 ESR2 0.64 0.027 FAM107A 0.64 0.003 0.61 0.003 FAM13C 0.68 0.006 0.55 <001 FGFR2 0.73 0.033 0.59 <001 FKBP5 0.60 0.006 FLNC 0.68 0.024 0.65 0.012 FLT1 0.71 0.027 FOS 0.62 0.006 FOXOl 0.75 0.010 GADD45B 0.68 0.020 GHR 0.62 0.006 GPM6B 0.57 <001 GSTM1 0.68 0.015 0.58 <001 GSTM2 0.65 0.005 0.47 <001 HGD 0.63 0.001 0.71 0.020 HK1 0.67 0.003 0.62 0.002 HFF 0.59 <001 HNF1B 0.66 0.004 0.61 0.001 IER3 0.70 0.026 IGF1 0.63 0.005 0.55 <001 IGF1R 0.76 0.049 IGFBP2 0.59 0.007 0.64 0.003 IF6ST 0.65 0.005 IF8 0.61 0.005 0.66 0.019 IFK 0.64 0.015 ING5 0.73 0.033 0.70 0.009 ITGA7 0.72 0.045 0.69 0.019 ITGB4 0.63 0.002 KFC1 0.74 0.045 KFK1 0.56 0.002 0.49 <001 KFK10 0.68 0.013 KFK11 0.66 0.003 KFK2 0.66 0.045 0.65 0.011 KFK3 0.75 0.048 0.77 0.014 KRT15 0.71 0.017 0.50 <001 KRT5 0.73 0.031 0.54 <001 FAMA5 0.70 0.044 FAMB3 0.70 0.005 0.58 <001 EG AES 3 0.69 0.025 LIG3 0.68 0.022 MDK 0.69 0.035 85 2015227398 15 Sep 20

Table 8B Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value MGMT 0.59 0.017 0.60 <001 MGST1 0.73 0.042 MICA 0.70 0.009 MPPED2 0.72 0.031 0.54 <001 MTSS1 0.62 0.003 MYBPC1 0.50 <001 NCAPD3 0.62 0.007 0.38 <001 NCOR1 0.82 0.048 NFAT5 0.60 0.001 0.62 <001 NRG1 0.66 0.040 0.61 0.029 NUP62 0.75 0.037 OMD 0.54 <001 PAGE4 0.64 0.005 PCA3 0.66 0.012 PCDHGB7 0.68 0.018 PGR 0.60 0.012 PPAP2B 0.62 0.010 PPP1R12A 0.73 0.031 0.58 0.003 PRIM A1 0.65 0.013 PROM1 0.41 0.013 PTCH1 0.64 0.006 PTEN 0.75 0.047 PTGS2 0.67 0.011 PTK2B 0.66 0.005 PTPN1 0.71 0.026 RAGE 0.70 0.012 RARB 0.68 0.016 RGS10 0.84 0.034 RHOB 0.66 0.016 RND3 0.63 0.004 SDHC 0.73 0.044 0.69 0.016 SERPINA3 0.67 0.011 0.51 <001 SERPINB5 0.42 <001 SH3RF2 0.66 0.012 0.51 <001 SLC22A3 0.59 0.003 0.48 <001 SMAD4 0.64 0.004 0.49 <001 SMARCC2 0.73 0.042 SMARCD1 0.73 <001 0.76 0.035 SMO 0.64 0.006 SNAI1 0.53 0.008 SOD1 0.60 0.003 SRC 0.64 <001 0.61 <001 SRD5A2 0.63 0.004 0.59 <001 STAT3 0.64 0.014 86 2015227398 15 Sep 2015

Table 8B Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value STAT5A 0.70 0.032 STAT5B 0.74 0.034 0.63 0.003 SVIL 0.71 0.028 TGFB1I1 0.68 0.036 TMPRSS 2 0.72 0.015 0.67 <.001 TNFRSF10A 0.69 0.010 TNFRSF10B 0.67 0.007 0.64 0.001 TNFRSF18 0.38 0.003 TNFSF10 0.71 0.025 TP53 0.68 0.004 0.57 <001 TP63 0.75 0.049 0.52 <001 TPM2 0.62 0.007 TRAF3IP2 0.71 0.017 0.68 0.005 TRO 0.72 0.033 TUBB2A 0.69 0.038 VCL 0.62 <001 VEGFA 0.71 0.037 WWOX 0.65 0.004 ZFHX3 0.77 0.011 0.73 0.012 ZFP36 0.69 0.018 ZNF827 0.68 0.013 0.49 <001 )0142] Tables 9A and 9B provide genes significantly associated (p<0.( )5), positively or negatively, with TMPRSS fusion status in the primary Gleason pattern. Increased expression of genes in Table 9A are positively associated with TMPRSS fusion positivity, while increased expression of genes in Table 10A are negatively associated with TMPRSS fusion positivity.

Table 9A.

Genes significantly (p<0.05) associated with TMPRSS fusion status in the primary Gleason pattern with odds ratio (OR) > 1.0 (increased expression is positively associated with TMPRSS fusion positivity

Table 9A Official Symbol p-value Odds Ratio Official Symbol p-value Odds Ratio ABCC8 <001 1.86 MAP3K5 <001 2.06 ALDH18A1 0.005 1.49 MAP7 <001 2.74 ALKBH3 0.043 1.30 MSH2 0.005 1.59 ALOX5 <001 1.66 MSH3 0.006 1.45 AMPD3 <001 3.92 MUC1 0.012 1.42 APEX1 <001 2.00 MY06 <001 3.79 ARHGDIB <001 1.87 NCOR2 0.001 1.62 ASAP2 0.019 1.48 NDRG1 <001 6.77 ATXN1 0.013 1.41 NET02 <001 2.63 BMPR1B <001 2.37 ODC1 <001 1.98 CACNA1D <001 9.01 OR51E1 <001 2.24 87 2015227398 15 Sep 2015

Table 9A Official Symbol p-value Odds Ratio Official Symbol p-value Odds Ratio CADPS 0.015 1.39 PDE9A <001 2.21 CD276 0.003 2.25 PEX10 <001 3.41 CDH1 0.016 1.37 PGK1 0.022 1.33 CDH7 <001 2.22 PLA2G7 <001 5.51 CDK7 0.025 1.43 PPP3CA 0.047 1.38 COL9A2 <001 2.58 PSCA 0.013 1.43 CRISP3 <001 2.60 PSMD13 0.004 1.51 CTNND1 0.033 1.48 PTCH1 0.022 1.38 ECE1 <001 2.22 PTK2 0.014 1.38 EIF5 0.023 1.34 PTK6 <001 2.29 EPHB4 0.005 1.51 PTK7 <001 2.45 ERG <001 14.5 PTPRK <001 1.80 FAM171B 0.047 1.32 RAB30 0.001 1.60 FAM73A 0.008 1.45 REG4 0.018 1.58 FASN 0.004 1.50 RELA 0.001 1.62 GNPTAB <001 1.60 RFX1 0.020 1.43 GPS1 0.006 1.45 RGS10 <001 1.71 GRB7 0.023 1.38 SCUBE2 0.009 1.48 HDAC1 <001 4.95 SEPT9 <001 3.91 HGD <001 1.64 SH3RF2 0.004 1.48 HIP1 <001 1.90 SH3YL1 <001 1.87 HNF1B <001 3.55 SHH <001 2.45 HSPA8 0.041 1.32 SIM2 <001 1.74 IGF1R 0.001 1.73 SIPA1L1 0.021 1.35 ILF3 <001 1.91 SLC22A3 <001 1.63 IMMT 0.025 1.36 SLC44A1 <001 1.65 ITPR1 <001 2.72 SPINT1 0.017 1.39 ITPR3 <001 5.91 TFDP1 0.005 1.75 JAG1 0.007 1.42 TMPRS S 2ERG A 0.002 14E5 KCNN2 <001 2.80 TMPRS S 2ERGB <001 1.97 KHDRBS3 <001 2.63 TRIM 14 <001 1.65 KIAA0247 0.019 1.38 TSTA3 0.018 1.38 KLK11 <001 1.98 UAP1 0.046 1.39 LAMC1 0.008 1.56 UBE2G1 0.001 1.66 LAMC2 <001 3.30 UGDH <001 2.22 LOX 0.009 1.41 XRCC5 <001 1.66 LRP1 0.044 1.30 ZMYND8 <001 2.19 88 2015227398 15 Sep 2015

Table 9B. Genes significantly (p<0.05) associated with TMPRSS fusion status in the primary Gleason pattern with odds ratio (OR) < 1.0 (increased expression is negatively associated with TMPRSS fusion positivity) Table 9B Official Symbol p-value Odds Ratio ABCC4 0.045 0.77 ABHD2 <001 0.38 ACTR2 0.027 0.73 AD AMTS 1 0.024 0.58 ADH5 <001 0.58 AGTR2 0.016 0.64 AKAP1 0.013 0.70 AKT2 0.015 0.71 ALCAM <001 0.45 ALDH1A2 0.004 0.70 ANPEP <001 0.43 ANXA2 0.010 0.71 APC 0.036 0.73 APOC1 0.002 0.56 APOE <001 0.44 ARF1 0.041 0.77 ATM 0.036 0.74 AURKB <001 0.62 AZGP1 <001 0.54 BBC3 0.030 0.74 BCL2 0.012 0.70 BIN 1 0.021 0.74 BTG1 0.004 0.67 BTG3 0.003 0.63 C7 0.023 0.74 CADM1 0.007 0.69 CAS PI 0.011 0.70 CAV1 0.011 0.71 CCND1 0.019 0.72 CCR1 0.022 0.73 CD44 <001 0.57 CD68 <001 0.54 CD82 0.002 0.66 CDH5 0.007 0.66 CDKN1A <001 0.60 CDKN2B <001 0.54 CDKN2C 0.012 0.72 CDKN3 0.037 0.77 CHN1 0.038 0.75 CKS2 <001 0.48 89 2015227398 15 Sep 20

Table 9B Official Symbol p-value Odds Ratio COL11A1 0.017 0.72 COL1A1 <001 0.59 COL1A2 0.001 0.62 COL3A1 0.027 0.73 COL4A1 0.043 0.76 COL5A1 0.039 0.74 COL5A2 0.026 0.73 COL6A1 0.008 0.66 COL6A3 <001 0.59 COL8A1 0.022 0.74 CSF1 0.011 0.70 CTNNB1 0.021 0.69 CTSB <001 0.62 CTSD 0.036 0.68 CTSK 0.007 0.70 CTSS 0.002 0.64 CXCL12 <001 0.48 CXCR4 0.005 0.68 CXCR7 0.046 0.76 CYR61 0.004 0.65 DAP 0.002 0.64 DARC 0.021 0.73 DDR2 0.021 0.73 DHRS9 <001 0.52 DIAPH1 <001 0.56 DICER 1 0.029 0.75 DLC1 0.013 0.72 DLGAP1 <001 0.60 DLL4 <001 0.57 DPT 0.006 0.68 DUSP1 0.012 0.68 DUSP6 0.001 0.62 DVL1 0.037 0.75 EFNB2 <001 0.32 EGR1 0.003 0.65 ELK4 <001 0.60 ERBB2 <001 0.61 ERBB3 0.045 0.76 ESR2 0.010 0.70 ETV1 0.042 0.74 FABP5 <001 0.21 FAM13C 0.006 0.67 FCGR3A 0.018 0.72 FGF17 0.009 0.71 90 2015227398 15 Sep 20

Table 9B Official Symbol p-value Odds Ratio FGF6 0.011 0.70 FGF7 0.003 0.63 FN1 0.006 0.69 FOS 0.035 0.74 FOXP3 0.010 0.71 GABRG2 0.029 0.74 GADD45B 0.003 0.63 GDF15 <001 0.54 GPM6B 0.004 0.67 GPNMB 0.001 0.62 GSN 0.009 0.69 HLA-G 0.050 0.74 HLF 0.018 0.74 HPS1 <001 0.48 HSD17B3 0.003 0.60 HSD17B4 <001 0.56 HSPB1 <001 0.38 HSPB2 0.002 0.62 IFI30 0.049 0.75 IFNG 0.006 0.64 IGF1 0.016 0.73 IGF2 0.001 0.57 IGFBP2 <001 0.51 IGFBP3 <001 0.59 IGFBP6 <001 0.57 IL10 <001 0.62 IL17A 0.012 0.63 ILIA 0.011 0.59 IL2 0.001 0.63 IL6ST <001 0.52 INSL4 0.014 0.71 ITGA1 0.009 0.69 ITGA4 0.007 0.68 JUN <001 0.59 KIT <001 0.64 KRT76 0.016 0.70 LAG3 0.002 0.63 LAPTM5 <001 0.58 LGALS3 <001 0.53 LTBP2 0.011 0.71 LUM 0.012 0.70 MAOA 0.020 0.73 MAP4K4 0.007 0.68 MGST1 <001 0.54 91 2015227398 15 Sep 20

Table 9B Official Symbol p-value Odds Ratio MMP2 <001 0.61 MPPED2 <001 0.45 MRC1 0.005 0.67 MTPN 0.002 0.56 MTSS1 <001 0.53 MVP 0.009 0.72 MYBPC1 <001 0.51 MYLK3 0.001 0.58 NCAM1 <001 0.59 NCAPD3 <001 0.40 NCOR1 0.004 0.69 NFKBIA <001 0.63 NNMT 0.006 0.66 NPBWR1 0.027 0.67 OAZ1 0.049 0.64 OLFML3 <001 0.56 OSM <001 0.64 PAGE1 0.012 0.52 PDGFRB 0.016 0.73 PEC AMI <001 0.55 PGR 0.048 0.77 PIK3CA <001 0.55 PIK3CG 0.008 0.71 PLAU 0.044 0.76 PLK1 0.006 0.68 PLOD2 0.013 0.71 PLP2 0.024 0.73 PNLIPRP2 0.009 0.70 PPAP2B <001 0.62 PRKAR2B <001 0.61 PRKCB 0.044 0.76 PROS1 0.005 0.67 PTEN <001 0.47 PTGER3 0.007 0.69 PTH1R 0.011 0.70 PTK2B <001 0.61 PTPN1 0.028 0.73 RAB27A <001 0.21 RAD51 <001 0.51 RAD9A 0.030 0.75 RARB <001 0.62 RASSF1 0.038 0.76 RECK 0.009 0.62 RHOB 0.004 0.64 92 2015227398 15 Sep 2015

Table 9B Official Symbol p-value Odds Ratio RHOC <001 0.56 RLN1 <001 0.30 RND3 0.014 0.72 S100P 0.002 0.66 SDC2 <001 0.61 SEMA3A 0.001 0.64 SMAD4 <001 0.64 SPARC <001 0.59 SPARCL1 <001 0.56 SPINK1 <001 0.26 SRD5A1 0.039 0.76 STAT1 0.026 0.74 STS 0.006 0.64 SULF1 <001 0.53 TFF3 <001 0.19 TGFA 0.002 0.65 TGFB1I1 0.040 0.77 TGFB2 0.003 0.66 TGFB3 <001 0.54 TGFBR2 <001 0.61 THY1 <001 0.63 TIMP2 0.004 0.66 TIMP3 <001 0.60 TMPRSS2 <001 0.40 TNFSF11 0.026 0.63 TPD52 0.002 0.64 TRAM1 <001 0.45 TRPC6 0.002 0.64 TUBB2A <001 0.49 VCL <001 0.57 VEGFB 0.033 0.73 VEGFC <001 0.61 VIM 0.012 0.69 WISP1 0.030 0.75 WNT5A <001 0.50 [00143] A molecular field effect was investigated, and determined that the expression levels of histologically normal-appearing cells adjacent to the tumor exhibited a molecular signature of prostate cancer. Tables 10A and 10B provide genes significantly associated (p<0.05), positively or negatively, with cRFI or bRFI in non-tumor samples. Table 10A is negatively associated with good prognosis, while increased expression of genes in Table 10B is positively associated with good prognosis. 93 2015227398 15 Sep 2015

Table 10A Genes significantly (p<0.05) associated with cRFI or bRFI in Non-Tumor Samples with hazard ratio (HR) >1.0 (increased expression is negatively associated with good prognosis) Table 10A cRFI bRFI Official Symbol HR p-value HR p-value ALCAM 1.278 0.036 ASPN 1.309 0.032 BAG5 1.458 0.004 BRCA2 1.385 <001 CACNA1D 1.329 0.035 CD 164 1.339 0.020 CDKN2B 1.398 0.014 COL3A1 1.300 0.035 COL4A1 1.358 0.019 CTNND2 1.370 0.001 DARC 1.451 0.003 DICER 1 1.345 <001 DPP4 1.358 0.008 EFNB2 1.323 0.007 FASN 1.327 0.035 GHR 1.332 0.048 HSPA5 1.260 0.048 INHBA 1.558 <001 KCNN2 1.264 0.045 KRT76 1.115 <001 LAMC1 1.390 0.014 LAMC2 1.216 0.042 LIG3 1.313 0.030 MAOA 1.405 0.013 MCM6 1.307 0.036 MKI67 1.271 0.008 NEK2 1.312 0.016 NPBWR1 1.278 0.035 ODC1 1.320 0.010 PEX10 1.361 0.014 PGK1 1.488 0.004 PLA2G7 1.337 0.025 POSTN 1.306 0.043 PTK6 1.344 0.005 REG4 1.348 0.009 RGS7 1.144 0.047 SFRP4 1.394 0.009 TARP 1.412 0.011 TFF1 1.346 0.010 TGFBR2 1.310 0.035 94 2015227398 15 Sep 2015

Table 10A cRFI bRFI Official Symbol HR p-value HR p-value THY1 1.300 0.038 TMPRS S 2ERG A 1.333 <001 TPD52 1.374 0.015 TRPC6 1.272 0.046 UBE2C 1.323 0.007 UHRF1 1.325 0.021 Table 10B Genes significantly (p<0.05) associated with cRFI or bRFI in Non-Tumor Samples with hazard ratio (HR) < 1.0 (increased expression is positively associated with good prognosis)

Table 10B cRFI bRFI Official Symbol HR p-value HR p-value ABCA5 0.807 0.028 ABCC3 0.760 0.019 0.750 0.003 ABHD2 0.781 0.028 ADAM 15 0.718 0.005 AKAP1 0.740 0.009 AMPD3 0.793 0.013 ANGPT2 0.752 0.027 ANXA2 0.776 0.035 APC 0.755 0.014 APRT 0.762 0.025 AR 0.752 0.015 ARHGDIB 0.753 <001 BIN 1 0.738 0.016 CADM1 0.711 0.004 CCNH 0.820 0.041 CCR1 0.749 0.007 CDK14 0.772 0.014 CDK3 0.819 0.044 CDKN1C 0.808 0.038 CHAF1A 0.634 0.002 0.779 0.045 CHN1 0.803 0.034 CHRAC1 0.751 0.014 0.779 0.021 COF5A1 0.736 0.012 COF5A2 0.762 0.013 COF6A1 0.757 0.032 COF6A3 0.757 0.019 CSK 0.663 <001 0.698 <001 CTSK 0.782 0.029 CXCF12 0.771 0.037 CXCR7 0.753 0.008 95 2015227398 15 Sep 20

Table 10B cRFI bRFI Official Symbol HR p-value HR p-value CYP3A5 0.790 0.035 DDIT4 0.725 0.017 DIAPH1 0.771 0.015 DLC1 0.744 0.004 0.807 0.015 DLGAP1 0.708 0.004 DUSP1 0.740 0.034 EDN1 0.742 0.010 EGR1 0.731 0.028 EIF3H 0.761 0.024 EIF4E 0.786 0.041 ERBB2 0.664 0.001 ERBB4 0.764 0.036 ERCC1 0.804 0.041 ESR2 0.757 0.025 EZH2 0.798 0.048 FAAH 0.798 0.042 FAM13C 0.764 0.012 FAM171B 0.755 0.005 FAM49B 0.811 0.043 FAM73A 0.778 0.015 FAS LG 0.757 0.041 FGFR2 0.735 0.016 FOS 0.690 0.008 FYN 0.788 0.035 0.777 0.011 GPNMB 0.762 0.011 GSK3B 0.792 0.038 HGD 0.774 0.017 HIRIP3 0.802 0.033 HSP90AB1 0.753 0.013 HSPB1 0.764 0.021 HSPE1 0.668 0.001 IFI30 0.732 0.002 IGF2 0.747 0.006 IGFBP5 0.691 0.006 IL6ST 0.748 0.010 IL8 0.785 0.028 IMMT 0.708 <001 ITGA6 0.747 0.008 ITGAV 0.792 0.016 ITGB3 0.814 0.034 ITPR3 0.769 0.009 JUN 0.655 0.005 KHDRBS3 0.764 0.012 KLF6 0.714 <001 96 2015227398 15 Sep 20

Table 10B cRFI bRFI Official Symbol HR p-value HR p-value KLK2 0.813 0.048 LAMA4 0.702 0.009 LAMA5 0.744 0.011 LAPTM5 0.740 0.009 LGALS3 0.773 0.036 0.788 0.024 LIMS1 0.807 0.012 MAP3K5 0.815 0.034 MAP3K7 0.809 0.032 MAP4K4 0.735 0.018 0.761 0.010 MAPKAPK3 0.754 0.014 MICA 0.785 0.019 MTA1 0.808 0.043 MVP 0.691 0.001 MYLK3 0.730 0.039 MY06 0.780 0.037 NCOA1 0.787 0.040 NCOR1 0.876 0.020 NDRG1 0.761 <001 NFAT5 0.770 0.032 NFKBIA 0.799 0.018 NME2 0.753 0.005 NUP62 0.842 0.032 OAZ1 0.803 0.043 OLFML2B 0.745 0.023 OLFML3 0.743 0.009 OSM 0.726 0.018 PCA3 0.714 0.019 PEC AMI 0.774 0.023 PIK3C2A 0.768 0.001 PIM1 0.725 0.011 PLOD2 0.713 0.008 PPP3CA 0.768 0.040 PROM1 0.482 <001 PTEN 0.807 0.012 PTGS2 0.726 0.011 PTTG1 0.729 0.006 PYCARD 0.783 0.012 RAB30 0.730 0.002 RAGE 0.792 0.012 RFX1 0.789 0.016 0.792 0.010 RGS10 0.781 0.017 RUNX1 0.747 0.007 SDHC 0.827 0.036 SEC23A 0.752 0.010 97 2015227398 15 Sep 2015

Table 10B cRFI bS .FI Official Symbol HR p-value HR p-value SEPT9 0.889 0.006 SERPINA3 0.738 0.013 SLC25A21 0.788 0.045 SMARCD1 0.788 0.010 0.733 0.007 SMO 0.813 0.035 SRC 0.758 0.026 SRD5A2 0.738 0.005 ST5 0.767 0.022 STAT5A 0.784 0.039 TGFB2 0.771 0.027 TGFB3 0.752 0.036 THBS2 0.751 0.015 TNFRSF10B 0.739 0.010 TPX2 0.754 0.023 TRAF3IP2 0.774 0.015 TRAM1 0.868 <.001 0.880 <.001 TRIM 14 0.785 0.047 TUBB2A 0.705 0.010 TYMP 0.778 0.024 UAP1 0.721 0.013 UTP23 0.763 0.007 0.826 0.018 VCL 0.837 0.040 VEGFA 0.755 0.009 WDR19 0.724 0.005 YBX1 0.786 0.027 ZFP36 0.744 0.032 ZNF827 0.770 0.043 [00144] Table 11 provides genes that are significantly associated (p<0.05) with cRFI or bRFI after adjustment for Gleason pattern or highest Gleason pattern.

Table 11

Genes significantly (p<0.05) associated with cRFI or bRFI after adjustment for Gleason pattern in the primary Gleason pattern or highest Gleason pattern Some HR <= 1.0 and some HR > 1.0

Table 11 cRFI bRFI bRFI Highest Pattern Primary Pattern Highest Pattern Official Symbol HR p-value HR p-value HR p-value HSPA5 0.710 0.009 1.288 0.030 ODC1 0.741 0.026 1.343 0.004 1.261 0.046 [00145] Tables 12A and 12B provide genes that are significantly associated (p<0.05) with prostate cancer specific survival (PCSS) in the primary Gleason pattern. Increased expression of 98 2015227398 15 Sep 2015 genes in Table 12A is negatively associated with good prognosis, while increased expression of genes in Table 12B is positively associated with good prognosis. Table 12A Genes significantly (p<0.05) associated with prostate cancer specific survival (PCSS) in the Primary Gleason Pattern HR >1.0 (Increased expression is negatively associated with good prognosis)_

Table 12A Official Symbol HR p-value Official Symbol HR p-value AKR1C3 1.476 0.016 GREM1 1.942 <001 ANLN 1.517 0.006 IFI30 1.482 0.048 APOC1 1.285 0.016 IGFBP3 1.513 0.027 APOE 1.490 0.024 INHBA 3.060 <001 ASPN 3.055 <001 KIF4A 1.355 0.001 ATP5E 1.788 0.012 KLK14 1.187 0.004 AURKB 1.439 0.008 LAPTM5 1.613 0.006 BGN 2.640 <001 LTBP2 2.018 <001 BIRC5 1.611 <001 MMP11 1.869 <001 BMP6 1.490 0.021 MYBL2 1.737 0.013 BRCA1 1.418 0.036 NEK2 1.445 0.028 CCNB1 1.497 0.021 NOX4 2.049 <001 CD276 1.668 0.005 OLFML2B 1.497 0.023 CDC20 1.730 <001 PLK1 1.603 0.006 CDH11 1.565 0.017 POSTN 2.585 <001 CDH7 1.553 0.007 PPFIA3 1.502 0.012 CDKN2B 1.751 0.003 PTK6 1.527 0.009 CDKN2C 1.993 0.013 PTTG1 1.382 0.029 CDKN3 1.404 0.008 RAD51 1.304 0.031 CENPF 2.031 <001 RGS7 1.251 <001 CHAF1A 1.376 0.011 RRM2 1.515 <001 CKS2 1.499 0.031 SAT1 1.607 0.004 COL1A1 2.574 <001 SDC1 1.710 0.007 COL1A2 1.607 0.011 SESN3 1.399 0.045 COL3A1 2.382 <001 SFRP4 2.384 <001 COL4A1 1.970 <001 SHMT2 1.949 0.003 COL5A2 1.938 0.002 SPARC 2.249 <001 COL8A1 2.245 <001 STMN1 1.748 0.021 CTHRC1 2.085 <001 SULF1 1.803 0.004 CXCR4 1.783 0.007 THBS2 2.576 <001 DDIT4 1.535 0.030 THY1 1.908 0.001 DYNLL1 1.719 0.001 TK1 1.394 0.004 F2R 2.169 <001 TOP2A 2.119 <001 FAM171B 1.430 0.044 TPX2 2.074 0.042 FAP 1.993 0.002 UBE2C 1.598 <001 99 2015227398 15 Sep 2015

Table 12A Official Symbol HR p-value Official Symbol HR p-value FCGR3A 2.099 <001 UGT2B15 1.363 0.016 FN1 1.537 0.024 UHRF1 1.642 0.001 GPR68 1.520 0.018 ZWINT 1.570 0.010 Table 12B Genes significantly (p<0.05) associated with prostate cancer specific survival (PCSS) in the Primary Gleason Pattern HR < 1.0 (Increased expression is positively associated with good prognosis)

Table 12B Official Official Symbol HR p-value Symbol HR p-value AAMP 0.649 0.040 IGFBP6 0.578 0.003 ABCA5 0.777 0.015 IL2 0.528 0.010 ABCG2 0.715 0.037 IL6ST 0.574 <001 ACOX2 0.673 0.016 IL8 0.540 0.001 ADH5 0.522 <001 ING5 0.688 0.015 ALDH1A2 0.561 <001 ITGA6 0.710 0.005 AMACR 0.693 0.029 ITGA7 0.676 0.033 AMPD3 0.750 0.049 JUN 0.506 0.001 ANPEP 0.531 <001 KIT 0.628 0.047 ATXN1 0.640 0.011 KLK1 0.523 0.002 AXIN2 0.657 0.002 KLK2 0.581 <001 AZGP1 0.617 <001 KLK3 0.676 <001 BDKRB1 0.553 0.032 KRT15 0.684 0.005 BIN1 0.658 <001 KRT18 0.536 <001 BTRC 0.716 0.011 KRT5 0.673 0.004 C7 0.531 <001 KRT8 0.613 0.006 CADM1 0.646 0.015 LAMB 3 0.740 0.027 CASP7 0.538 0.029 LGALS3 0.678 0.007 CCNH 0.674 0.001 MGST1 0.640 0.002 CD 164 0.606 <001 MPPED2 0.629 <001 CD44 0.687 0.016 MTS SI 0.705 0.041 CDK3 0.733 0.039 MYBPC1 0.534 <001 CHN1 0.653 0.014 NCAPD3 0.519 <001 COL6A1 0.681 0.015 NFAT5 0.536 <001 CSF1 0.675 0.019 NRG1 0.467 0.007 CSRP1 0.711 0.007 OLFML3 0.646 0.001 CXCL12 0.650 0.015 OMD 0.630 0.006 CYP3A5 0.507 <001 OR51E2 0.762 0.017 CYR61 0.569 0.007 PAGE4 0.518 <001 DLGAP1 0.654 0.004 PCA3 0.581 <001 DNM3 0.692 0.010 PGF 0.705 0.038 DPP4 0.544 <001 PPAP2B 0.568 <001 DPT 0.543 <001 PPP1R12A 0.694 0.017 DUSP1 0.660 0.050 PRIM A1 0.678 0.014 100 2015227398 15 Sep 2015

Table 12B Official Symbol HR p-value Official Symbol HR p-value DUSP6 0.699 0.033 PRKCA 0.632 0.001 EGR1 0.490 <001 PRKCB 0.692 0.028 EGR3 0.561 <001 PROM1 0.393 0.017 EIF5 0.720 0.035 PTEN 0.689 0.002 ERBB3 0.739 0.042 PTGS2 0.611 0.004 FAAH 0.636 0.010 PTH1R 0.629 0.031 FAM107A 0.541 <001 RAB27A 0.721 0.046 FAM13C 0.526 <001 RND3 0.678 0.029 FAS 0.689 0.030 RNF114 0.714 0.035 FGF10 0.657 0.024 SDHC 0.590 <001 FKBP5 0.699 0.040 SERPINA3 0.710 0.050 FLNC 0.742 0.036 SH3RF2 0.570 0.005 FOS 0.556 0.005 SLC22A3 0.517 <001 FOXQ1 0.666 0.007 SMAD4 0.528 <001 GADD45B 0.554 0.002 SMO 0.751 0.026 GDF15 0.659 0.009 SRC 0.667 0.004 GHR 0.683 0.027 SRD5A2 0.488 <001 GPM6B 0.666 0.005 STAT5B 0.700 0.040 GSN 0.646 0.006 SVIL 0.694 0.024 GSTM1 0.672 0.006 TFF3 0.701 0.045 GSTM2 0.514 <001 TGFB1I1 0.670 0.029 HGD 0.771 0.039 TGFB2 0.646 0.010 HIRIP3 0.730 0.013 TNFRSF10B 0.685 0.014 HK1 0.778 0.048 TNFSF10 0.532 <001 HLF 0.581 <001 TPM2 0.623 0.005 HNF1B 0.643 0.013 TRO 0.767 0.049 HSD17B10 0.742 0.029 TUBB2A 0.613 0.003 IER3 0.717 0.049 VEGFB 0.780 0.034 IGF1 0.612 <001 ZFP36 0.576 0.001 ZNF827 0.644 0.014 [00146] Analysis of gene expression and upgrading/upstaging was based on univariate ordinal logistic regression models using weighted maximum likelihood estimators for each gene in the gene list (727 test genes and 5 reference genes). P-values were generated using a Wald test of the null hypothesis that the odds ratio (OR) is one. Both unadjusted p-values and the q-value (smallest FDR at which the hypothesis test in question is rejected) were reported. Unadjusted p-values <0.05 were considered statistically significant. Since two tumor specimens were selected for each patient, this analysis was performed using the 2 specimens from each patient as follows: (1) analysis using the primary Gleason pattern specimen from each patient 101 2015227398 15 Sep 2015 (Specimens A1 and B2 as described in Table 2); and (2) analysis using the highest Gleason pattern specimen from each patient (Specimens A1 and B1 as described in Table 2). 200 genes were found to be significantly associated (p<0.05) with upgrading/upstaging in the primary Gleason pattern sample (PGP) and 203 genes were found to be significantly associated (p<0.05) with upgrading/upstaging in the highest Gleason pattern sample (HGP).

[00147] Tables 13A and 13B provide genes significantly associated (p<0.05), positively or negatively, with upgrading/upstaging in the primary and/or highest Gleason pattern. Increased expression of genes in Table 13A is positively associated with higher risk of upgrading/upstaging (poor prognosis), while increased expression of genes in Table 13B is negatively associated with risk of upgrading/upstaging (good prognosis).

TABLE13A

Genes significantly (p<0.05) associated with upgrading/upstaging in the Primary Gleason Pattern (PGP) and Highest Gleason Pattern (HGP) OR >1.0 (Increased expression is positively associated with higher risk of upgrading/upstaging (poor prognosis))

Table 13A PGP HGP Gene OR p-value OR p-value ALCAM 1.52 0.0179 1.50 0.0184 ANLN 1.36 0.0451 , APOE 1.42 0.0278 1.50 0.0140 ASPN 1.60 0.0027 2.06 0.0001 AURKA 1.47 0.0108 , AURKB , 1.52 0.0070 BAX 1.48 0.0095 BGN 1.58 0.0095 1.73 0.0034 BIRC5 1.38 0.0415 BMP6 1.51 0.0091 1.59 0.0071 BUB1 1.38 0.0471 1.59 0.0068 CACNA1D 1.36 0.0474 1.52 0.0078 CASP7 1.32 0.0450 CCNE2 1.54 0.0042 CD276 , 1.44 0.0265 CDC20 1.35 0.0445 1.39 0.0225 CDKN2B , 1.36 0.0415 CENPF 1.43 0.0172 1.48 0.0102 CLTC 1.59 0.0031 1.57 0.0038 102 2015227398 15 Sep 2015

Table 13A PGP HGP Gene OR p-value OR p-value COL1A1 1.58 0.0045 1.75 0.0008 COL3A1 E45 0.0143 1.47 0.0131 COL8A1 1.40 0.0292 1.43 0.0258 CRISP3 1.40 0.0256 CTHRC1 E56 0.0092 DBN1 1.43 0.0323 1.45 0.0163 DIAPH1 E51 0.0088 1.58 0.0025 DICER 1 1.40 0.0293 DI02 1.49 0.0097 DVL1 E53 0.0160 F2R 1.46 0.0346 1.63 0.0024 FAP 1.47 0.0136 1.74 0.0005 FCGR3A 1.42 0.0221 HPN E36 0.0468 HSD17B4 1.47 0.0151 HSPA8 E65 0.0060 1.58 0.0074 IF11 E50 0.0100 1.48 0.0113 IF1B 1.41 0.0359 INHBA E56 0.0064 1.71 0.0042 KHDRBS3 1.43 0.0219 1.59 0.0045 KIF4A E50 0.0209 KPNA2 1.40 0.0366 KRT2 1.37 0.0456 KRT75 1.44 0.0389 MANF E39 0.0429 MEEK 1.74 0.0016 MKI67 E35 0.0408 MMP11 E56 0.0057 NOX4 1.49 0.0105 1.49 0.0138 PLAUR 1.44 0.0185 PLK1 1.41 0.0246 PTK6 E36 0.0391 RAD51 E39 0.0300 RAF1 E58 0.0036 RRM2 E57 0.0080 SESN3 E33 0.0465 SFRP4 2.33 <0.0001 2.51 0.0015 SKIL 1.44 0.0288 1.40 0.0368 SOX4 E50 0.0087 1.59 0.0022 103 2015227398 15 Sep 2015

Table 13A PGP HGP Gene OR p-value OR p-value SPINK1 1.52 0.0058 SPP1 1.42 0.0224 THBS2 1.36 0.0461 TK1 1.38 0.0283 TOP2A 1.85 0.0001 1.66 0.0011 TPD52 1.78 0.0003 1.64 0.0041 TPX2 1.70 0.0010 UBE2G1 1.38 0.0491 UBE2T 1.37 0.0425 1.46 0.0162 UHRF1 1.43 0.0164 VCPIP1 1.37 0.0458 TABLE 13B

Genes significantly (p<0.05) associated with upgrading/upstaging in the Primary Gleason Pattern (PGP) and Highest Gleason Pattern (HGP) OR < 1.0 (Increased expression is negatively associated with higher risk of upgrading/upstaging (good prognosis)) Table 13B PGP HGP Gene OR p-value OR p-value ABCC3 0.70 0.0216 ABCC8 0.66 0.0121 ABCG2 0.67 0.0208 0.61 0.0071 ACE 0.73 0.0442 ACOX2 0.46 0.0000 0.49 0.0001 ADH5 0.69 0.0284 0.59 0.0047 AIG1 0.60 0.0045 AKR1C1 0.66 0.0095 ALDH1A2 0.36 <0.0001 0.36 <0.0001 ALKBH3 0.70 0.0281 0.61 0.0056 ANPEP 0.68 0.0109 ANXA2 0.73 0.0411 0.66 0.0080 APC 0.68 0.0223 ATXN1 0.70 0.0188 AXIN2 0.60 0.0072 0.68 0.0204 AZGP1 0.66 0.0089 0.57 0.0028 BCL2 0.71 0.0182 BIN 1 0.55 0.0005 BTRC 0.69 0.0397 0.70 0.0251 C7 0.53 0.0002 0.51 <0.0001 104 2015227398 15 Sep 2015

Table 13B PGP HGP Gene OR p-value OR p-value CADM1 0.57 0.0012 0.60 0.0032 CAS PI 0.64 0.0035 0.72 0.0210 CAV1 0.64 0.0097 0.59 0.0032 CAV2 0.58 0.0107 CD 164 0.69 0.0260 CD82 0.67 0.0157 0.69 0.0167 CDH1 0.61 0.0012 0.70 0.0210 CDK14 0.70 0.0354 CDK3 0.72 0.0267 CDKN1C 0.61 0.0036 0.56 0.0003 CHN1 0.71 0.0214 COL6A1 0.62 0.0125 0.60 0.0050 COL6A3 0.65 0.0080 0.68 0.0181 CSRP1 0.43 0.0001 0.40 0.0002 CTSB 0.66 0.0042 0.67 0.0051 CTSD 0.64 0.0355 CTSK 0.69 0.0171 CTSL1 0.72 0.0402 CUL1 0.61 0.0024 0.70 0.0120 CXCL12 0.69 0.0287 0.63 0.0053 CYP3A5 0.68 0.0099 0.62 0.0026 DDR2 0.68 0.0324 0.62 0.0050 DES 0.54 0.0013 0.46 0.0002 DHX9 0.67 0.0164 DLGAP1 0.66 0.0086 DPP4 0.69 0.0438 0.69 0.0132 DPT 0.59 0.0034 0.51 0.0005 DUSP1 0.67 0.0214 EDN1 0.66 0.0073 EDNRA 0.66 0.0148 0.54 0.0005 EIF2C2 0.65 0.0087 ELK4 0.55 0.0003 0.58 0.0013 ENPP2 0.65 0.0128 0.59 0.0007 EPHA3 0.71 0.0397 0.73 0.0455 EPHB2 0.60 0.0014 EPHB4 0.73 0.0418 EPHX3 0.71 0.0419 ERCC1 0.71 0.0325 FAM107A 0.56 0.0008 0.55 0.0011 105 2015227398 15 Sep 2015

Table 13B PGP HGP Gene OR p-value OR p-value FAM13C 0.68 0.0276 0.55 0.0001 FAS 0.72 0.0404 FBN1 0.72 0.0395 FBXW7 0.69 0.0417 FGF10 0.59 0.0024 0.51 0.0001 FGF7 0.51 0.0002 0.56 0.0007 FGFR2 0.54 0.0004 0.47 <0.0001 FLNA 0.58 0.0036 0.50 0.0002 FLNC 0.45 0.0001 0.40 <0.0001 FLT4 0.61 0.0045 F0X01 0.55 0.0005 0.53 0.0005 FOXP3 0.71 0.0275 0.72 0.0354 GHR 0.59 0.0074 0.53 0.0001 GNRH1 0.72 0.0386 GPM6B 0.59 0.0024 0.52 0.0002 GSN 0.65 0.0107 0.65 0.0098 GSTM1 0.44 <0.0001 0.43 <0.0001 GSTM2 0.42 <0.0001 0.39 <0.0001 HLF 0.46 <0.0001 0.47 0.0001 HPS1 0.64 0.0069 0.69 0.0134 HSPA5 0.68 0.0113 HSPB2 0.61 0.0061 0.55 0.0004 HSPG2 0.70 0.0359 ID3 0.70 0.0245 IGF1 0.45 <0.0001 0.50 0.0005 IGF2 0.67 0.0200 0.68 0.0152 IGFBP2 0.59 0.0017 0.69 0.0250 IGFBP6 0.49 <0.0001 0.64 0.0092 IL6ST 0.56 0.0009 0.60 0.0012 ILK 0.51 0.0010 0.49 0.0004 ITGA1 0.58 0.0020 0.58 0.0016 ITGA3 0.71 0.0286 0.70 0.0221 ITGA5 0.69 0.0183 ITGA7 0.56 0.0035 0.42 <0.0001 ITGB1 0.63 0.0095 0.68 0.0267 ITGB3 0.62 0.0043 0.62 0.0040 ITPR1 0.62 0.0032 JUN 0.73 0.0490 0.68 0.0152 KIT 0.55 0.0003 0.57 0.0005 106 2015227398 15 Sep 2015

Table 13B PGP HGP Gene OR p-value OR p-value KLC1 0.70 0.0248 KLK1 0.60 0.0059 KRT15 0.58 0.0009 0.45 <0.0001 KRT5 0.70 0.0262 0.59 0.0008 LAMA4 0.56 0.0359 0.68 0.0498 LAMB 3 0.60 0.0017 LGALS3 0.58 0.0007 0.56 0.0012 LRP1 0.69 0.0176 MAP3K7 0.70 0.0233 0.73 0.0392 MCM3 0.72 0.0320 MMP2 0.66 0.0045 0.60 0.0009 MMP7 0.61 0.0015 0.65 0.0032 MMP9 0.64 0.0057 0.72 0.0399 MPPED2 0.72 0.0392 0.63 0.0042 MTA1 0.68 0.0095 MTSS1 0.58 0.0007 0.71 0.0442 MVP 0.57 0.0003 0.70 0.0152 MYBPC1 0.70 0.0359 NCAM1 0.63 0.0104 0.64 0.0080 NCAPD3 0.67 0.0145 0.64 0.0128 NEXN 0.54 0.0004 0.55 0.0003 NFAT5 0.72 0.0320 0.70 0.0177 NUDT6 0.66 0.0102 OLFML3 0.56 0.0035 0.51 0.0011 OMD 0.61 0.0011 0.73 0.0357 PAGE4 0.42 <0.0001 0.36 <0.0001 PAK6 0.72 0.0335 PCDHGB7 0.70 0.0262 0.55 0.0004 PGF 0.72 0.0358 0.71 0.0270 PLP2 0.66 0.0088 0.63 0.0041 PPAP2B 0.44 <0.0001 0.50 0.0001 PPP1R12A 0.45 0.0001 0.40 <0.0001 PRIM A1 0.63 0.0102 PRKAR2B 0.71 0.0226 PRKCA 0.34 <0.0001 0.42 <0.0001 PRKCB 0.66 0.0120 0.49 <0.0001 PROM1 0.61 0.0030 PTEN 0.59 0.0008 0.55 0.0001 PTGER3 0.67 0.0293 107 2015227398 15 Sep 2015

Table 13B PGP HGP Gene OR p-value OR p-value PTH1R 0.69 0.0259 0.71 0.0327 PTK2 0.75 0.0461 PTK2B 0.70 0.0244 0.74 0.0388 PYCARD 0.73 0.0339 0.67 0.0100 RAD9A 0.64 0.0124 RARB 0.67 0.0088 0.65 0.0116 RGS10 0.70 0.0219 RHOB 0.72 0.0475 RND3 0.67 0.0231 SDHC 0.72 0.0443 SEC23A 0.66 0.0101 0.53 0.0003 SEMA3A 0.51 0.0001 0.69 0.0222 SH3RF2 0.55 0.0002 0.54 0.0002 SLC22A3 0.48 0.0001 0.50 0.0058 SMAD4 0.49 0.0001 0.50 0.0003 SMARCC2 0.59 0.0028 0.65 0.0052 SMO 0.60 0.0048 0.52 <0.0001 SORBS 1 0.56 0.0024 0.48 0.0002 SPARCL1 0.43 0.0001 0.50 0.0001 SRD5A2 0.26 <0.0001 0.31 <0.0001 ST5 0.63 0.0103 0.52 0.0006 STAT5A 0.60 0.0015 0.61 0.0037 STAT5B 0.54 0.0005 0.57 0.0008 SUMOl 0.65 0.0066 0.66 0.0320 SVIL 0.52 0.0067 0.46 0.0003 TGFB1I1 0.44 0.0001 0.43 0.0000 TGFB2 0.55 0.0007 0.58 0.0016 TGFB3 0.57 0.0010 0.53 0.0005 TEMPI 0.72 0.0224 TIMP2 0.68 0.0198 0.69 0.0206 TIMP3 0.67 0.0105 0.64 0.0065 TMPRSS2 0.72 0.0366 TNFRSF10A 0.71 0.0181 TNFSF10 0.71 0.0284 TOP2B 0.73 0.0432 TP63 0.62 0.0014 0.50 <0.0001 TPM1 0.54 0.0007 0.52 0.0002 TPM2 0.41 <0.0001 0.40 <0.0001 TPP2 0.65 0.0122 108

Table 13B PGP HGP Gene OR p-value OR p-value TRA2A 0.72 0.0318 TRAF3IP2 0.62 0.0064 0.59 0.0053 TRO 0.57 0.0003 0.51 0.0001 VCL 0.52 0.0005 0.52 0.0004 VIM 0.65 0.0072 0.65 0.0045 WDR19 0.66 0.0097 WFDC1 0.58 0.0023 0.60 0.0026 ZFHX3 0.69 0.0144 0.62 0.0046 ZNF827 0.62 0.0030 0.53 0.0001 2015227398 15 Sep 2015

Example 3: Identification of MicroRNAs Associated with Clinical Recurrence and Death Due to Prostate Cancer [00148] MicroRNAs function by binding to portions of messenger RNA (mRNA) and changing how frequently the mRNA is translated into protein. They can also influence the turnover of mRNA and thus how long the mRNA remains intact in the cell. Since microRNAs function primarily as an adjunct to mRNA, this study evaluated the joint prognostic value of microRNA expression and gene (mRNA) expression. Since the expression of certain microRNAs may be a surrogate for expression of genes that are not in the assessed panel, we also evaluated the prognostic value of microRNA expression by itself.

Patients and Samples [00149] Samples from the 127 patients with clinical recurrence and 374 patients without clinical recurrence after radical prostatectomy described in Example 2 were used in this study. The final analysis set comprised 416 samples from patients in which both gene expression and microRNA expression were successfully assayed. Of these, 106 patients exhibited clinical recurrence and 310 did not have clinical recurrence. Tissue samples were taken from each prostate sample representing (1) the primary Gleason pattern in the sample, and (2) the highest Gleason pattern in the sample. In addition, a sample of histologically normal-appearing tissue adjacent to the tumor (NAT) was taken. The number of patients in the analysis set for each tissue type and the number of them who experienced clinical recurrence or death due to prostate cancer are shown in Table 14. 109 2015227398 15 Sep 2015

Table 14. Number of Patients and Events in Analysis Set Patients Clinical Recurrences Deaths Due to Prostate Cancer Primary Gleason Pattern Tumor Tissue 416 106 36 Highest Gleason Pattern Tumor Tissue 405 102 36 Normal Adjacent Tissue 364 81 29

Assay Method [00150] Expression of 76 test microRNAs and 5 reference microRNAs were determined from RNA extracted from fixed paraffin-embedded (FPE) tissue. MicroRNA expression in all three tissue type was quantified by reverse transcriptase polymerase chain reaction (RT-PCR) using the crossing point (Cp) obtained from the Taqman® MicroRNA Assay kit (Applied Biosystems, Inc., Carlsbad, CA).

Statistical Analysis [00151] Using univariate proportional hazards regression (Cox DR, Journal of the Royal Statistical Society, Series B 34:187-220, 1972), applying the sampling weights from the cohort sampling design, and using variance estimation based on the Lin and Wei method (Lin and Wei, Journal of the American Statistical Association 84:1074-1078, 1989), microRNA expression, normalized by the average expression for the 5 reference microRNAs hsa-miR-106a, hsa-miR-146b-5p, hsa-miR-191, hsa-miR-19b, and hsa-miR-92a, and reference-normalized gene expression of the 733 genes (including the reference genes) discussed above, were assessed for association with clinical recurrence and death due to prostate cancer. Standardized hazard ratios (the proportional change in the hazard associated with a change of one standard deviation in the covariate value) were calculated.

[00152] This analysis included the following classes of predictors: [00153] l. MicroRNAs alone [00154] 2. MicroRNA-gene pairs Tier 1 110 [00155] 3. MicroRNA-gene pairs Tier 2 [00156] 4. MicroRNA-gene pairs Tier 3 [00157] 5. All other microRNA-gene pairs Tier 4 [00158] The four tiers were pre-determined based on the likelihood (Tier 1 representing 2015227398 15 Sep 2015 the highest likelihood) that the gene-microRNA pair functionally interacted or that the microRNA was related to prostate cancer based on a review of the literature and existing microarray data sets.

[00159] False discovery rates (FDR) (Benjamini and Hochberg, Journal of the Royal Statistical Society, Series B 57:289-300, 1995) were assessed using Efron’s separate class methodology (Efron, Annals of Applied Statistics 2:197-223., 2008). The false discovery rate is the expected proportion of the rejected null hypotheses that are rejected incorrectly (and thus are false discoveries). Efron’s methodology allows separate FDR assessment (^-values) (Storey, Journal of the Royal Statistical Society, Series B 64:479-498, 2002) within each class while utilizing the data from all the classes to improve the accuracy of the calculation. In this analysis, the q-value for a microRNA or microRNA-gene pair can be interpreted as the empirical Bayes probability that the microRNA or microRNA-gene pair identified as being associated with clinical outcome is in fact a false discovery given the data. The separate class approach was applied to a true discovery rate degree of association (TDRDA) analysis (Crager, Statistics in Medicine 29:33-45, 2010) to determine sets of microRNAs or microRNA-gene pairs that have standardized hazard ratio for clinical recurrence or prostate cancer-specific death of at least a specified amount while controlling the FDR at 10%. For each microRNA or microRNA-gene pair, a maximum lower bound (MLB) standardized hazard ratio was computed, showing the highest lower bound for which the microRNA or microRNA-gene pair was included in a TDRDA set with 10% FDR. Also calculated was an estimate of the true standardized hazard ratio corrected for regression to the mean (RM) that occurs in subsequent studies when the best predictors are selected from a long list (Crager, 2010 above). The RM-corrected estimate of the standardized hazard ratio is a reasonable estimate of what could be expected if the selected microRNA or microRNA-gene pair were studied in a separate, subsequent study.

[00160] These analyses were repeated adjusting for clinical and pathology covariates available at the time of patient biopsy: biopsy Gleason score, baseline PSA level, and clinical T-111 2015227398 15 Sep 2015 stage (T1-T2A vs. T2B or T2C) to assess whether the microRNAs or microRNA-gene pairs have predictive value independent of these clinical and pathology covariates.

Results [00161] The analysis identified 21 microRNAs assayed from primary Gleason pattern tumor tissue that were associated with clinical recurrence of prostate cancer after radical prostatectomy, allowing a false discovery rate of 10% (Table 15). Results were similar for microRNAs assessed from highest Gleason pattern tumor tissue (Table 16), suggesting that the association of microRNA expression with clinical recurrence does not change markedly depending on the location within a tumor tissue sample. No microRNA assayed from normal adjacent tissue was associated with the risk of clinical recurrence at a false discovery rate of 10%. The sequences of the microRNAs listed in Tables 15-21 are shown in Table B. 112 2015227398 15 Sep 2015

Table 15. MicroRNAs Associated with Clinical Recurrence of Prostate Cancer Primary Gleason Pattern Tumor Tissue Absolute Standardized Hazard Ratio MicroRNA p-value #-valuea (FDR) Direction of Association’’ Uncor rected Estimate 95% Confidence Interval Max. Lower Bound @10% FDR RM- Corrected Estimate0 hsa-miR-93 <0.0001 0.0% (+) 1.79 (1.38, 2.32) 1.19 1.51 hsa-miR-106b <0.0001 0.1% (+) 1.80 (1.38, 2.34) 1.19 1.51 hsa-miR-30e-5p <0.0001 0.1% (-) 1.63 (1.30, 2.04) 1.18 1.46 hsa-miR-21 <0.0001 0.1% (+) 1.66 (1.31, 2.09) 1.18 1.46 hsa-miR-133a <0.0001 0.1% (-) 1.72 (1.33,2.21) 1.18 1.48 hsa-miR-449a <0.0001 0.1% (+) 1.56 (1.26, 1.92) 1.17 1.42 hsa-miR-30a 0.0001 0.1% (-) 1.56 (1.25, 1.94) 1.16 1.41 hsa-miR-182 0.0001 0.2% (+) 1.74 (1.31,2.31) 1.17 1.45 hsa-miR-27a 0.0002 0.2% (+) 1.65 (1.27, 2.14) 1.16 1.43 hsa-miR-222 0.0006 0.5% (-) 1.47 (1.18, 1.84) 1.12 1.35 hsa-miR-103 0.0036 2.1% (+) 1.77 (1.21,2.61) 1.12 1.36 hsa-miR-1 0.0037 2.2% (-) 1.32 (1.10, 1.60) 1.07 1.26 hsa-miR-145 0.0053 2.9% (-) 1.34 (1.09, 1.65) 1.07 1.27 hsa-miR-141 0.0060 3.2% (+) 1.43 (1.11, 1.84) 1.07 1.29 hsa-miR-92a 0.0104 4.8% (+) 1.32 (1.07, 1.64) 1.05 1.25 hsa-miR-22 0.0204 7.7% (+) 1.31 (1.03, 1.64) 1.03 1.23 hsa-miR-29b 0.0212 7.9% (+) 1.36 (1.03, 1.76) 1.03 1.24 hsa-miR-210 0.0223 8.2% (+) 1.33 (1.03, 1.70) 1.00 1.23 hsa-miR-486-5p 0.0267 9.4% (-) 1.25 (1.00, 1.53) 1.00 1.20 hsa-miR-19b 0.0280 9.7% (-) 1.24 (1.00, 1.50) 1.00 1.19 hsa-miR-205 0.0289 10.0% (-) 1.25 (1.00, 1.53) 1.00 1.20 aThe #-value is the empirical Bayes probability that the microRNA’s association with clinical recurrence is a false discovery, given the data. bDirection of association indicates where higher microRNA expression is associated with higher (+) or lower (-) risk of clinical recurrence. CRM: regression to the mean. 113 2015227398 15 Sep 2015

Table 16. MicroRNAs Associated with Clinical Recurrence of Prostate Cancer Highest Gleason Pattern Tumor Tissue Absolute Standardized Hazard Ratio MicroRNA p-value r/-value3 (FDR) Direction of Association’’ Uncor rected Estimate 95% Confidence Interval Max. Lower Bound @10% FDR RM- Corrected Estimatec hsa-miR-93 <0.0001 0.0% (+) 1.91 (1.48, 2.47) 1.24 1.59 hsa-miR-449a <0.0001 0.0% (+) 1.75 (1.40, 2.18) 1.23 1.54 hsa-miR-205 <0.0001 0.0% (-) 1.53 (1.29, 1.81) 1.20 1.43 hsa-miR-19b <0.0001 0.0% (-) 1.37 (1.19, 1.57) 1.15 1.32 hsa-miR-106b <0.0001 0.0% (+) 1.84 (1.39, 2.42) 1.22 1.51 hsa-miR-21 <0.0001 0.0% (+) 1.68 (1.32, 2.15) 1.19 1.46 hsa-miR-30a 0.0005 0.4% (-) 1.44 (1.17, 1.76) 1.13 1.33 hsa-miR-30e-5p 0.0010 0.6% (-) 1.37 (1.14, 1.66) 1.11 1.30 hsa-miR-133a 0.0015 0.8% (-) 1.57 (1.19, 2.07) 1.13 1.36 hsa-miR-1 0.0016 0.8% (-) 1.42 (1.14, 1.77) 1.11 1.31 hsa-miR-103 0.0021 1.1% (+) 1.69 (1.21, 2.37) 1.13 1.37 hsa-miR-210 0.0024 1.2% (+) 1.43 (1.13, 1.79) 1.11 1.31 hsa-miR-182 0.0040 1.7% (+) 1.48 (1.13, 1.93) 1.11 1.31 hsa-miR-27a 0.0055 2.1% (+) 1.46 (1.12, 1.91) 1.09 1.30 hsa-miR-222 0.0093 3.2% (-) 1.38 (1.08, 1.77) 1.08 1.27 hsa-miR-331 0.0126 3.9% (+) 1.38 (1.07, 1.77) 1.07 1.26 hsa-miR-191* 0.0143 4.3% (+) 1.38 (1.06, 1.78) 1.07 1.26 hsa-miR-425 0.0151 4.5% (+) 1.40 (1.06, 1.83) 1.07 1.26 hsa-miR-31 0.0176 5.1% (-) 1.29 (1.04, 1.60) 1.05 1.22 hsa-miR-92a 0.0202 5.6% (+) 1.31 (1.03, 1.65) 1.05 1.23 hsa-miR-155 0.0302 7.6% (-) 1.32 (1.00, 1.69) 1.03 1.22 hsa-miR-22 0.0437 9.9% (+) 1.30 (1.00, 1.67) 1.00 1.21 aThe ('/-value is the empirical Bayes probability that the microRNA’s association with death due to prostate cancer is a false discovery, given the data. bDirection of association indicates where higher microRNA expression is associated with higher (+) or lower (-) risk of clinical recurrence. CRM: regression to the mean. 114 2015227398 15 Sep 2015 [00162] Table 17 shows microRNAs assayed from primary Gleason pattern tissue that were identified as being associated with the risk of prostate-cancer-specific death, with a false discovery rate of 10%. Table 18 shows the corresponding analysis for microRNAs assayed from highest Gleason pattern tissue. No microRNA assayed from normal adjacent tissue was associated with the risk of prostate-cancer-specific death at a false discovery rate of 10%.

Table 17. MicroRNAs Associated with Death Due to Prostate Cancer Primary Gleason Pattern Tumor Tissue

Absolute Standardized Hazard Ratio

Max.

Lower

MicroRNA p-value #-valuea (FDR) Direction of Association’’ Uncor rected Estimate 95% Confidence Interval Bound @10% FDR RM- Corrected Estimatec hsa-miR-30e-5p 0.0001 0.6% (-) 1.88 (1.37, 2.58) 1.15 1.46 hsa-miR-30a 0.0001 0.7% (-) 1.78 (1.33, 2.40) 1.14 1.44 hsa-miR-133a 0.0005 1.2% (-) 1.85 (1.31,2.62) 1.13 1.41 hsa-miR-222 0.0006 1.4% (-) 1.65 (1.24, 2.20) 1.12 1.38 hsa-miR-106b 0.0024 2.7% (+) 1.85 (1.24, 2.75) 1.11 1.35 hsa-miR-1 0.0028 3.0% (-) 1.43 (1.13, 1.81) 1.08 1.30 hsa-miR-21 0.0034 3.3% (+) 1.63 (1.17, 2.25) 1.09 1.33 hsa-miR-93 0.0044 3.9% (+) 1.87 (1.21, 2.87) 1.09 1.32 hsa-miR-26a 0.0072 5.3% (-) 1.47 (1.11, 1.94) 1.07 1.29 hsa-miR-152 0.0090 6.0% (-) 1.46 (1.10, 1.95) 1.06 1.28 hsa-miR-331 0.0105 6.5% (+) 1.46 (1.09, 1.96) 1.05 1.27 hsa-miR-150 0.0159 8.3% (+) 1.51 (1.07, 2.10) 1.03 1.27 hsa-miR-27b 0.0160 8.3% (+) 1.97 (1.12, 3.42) 1.05 1.25 aThe q-value is the empirical Bayes probability that the microRNA’s association with death due to prostate cancer endpoint is a false discovery, given the data. bDirection of association indicates where higher microRNA expression is associated with higher (+) or lower (-) risk of death due to prostate cancer. °RM: regression to the mean. 115 2015227398 15 Sep 2015

Table 18. MicroRNAs Associated with Death Due to Prostate Cancer Highest Gleason Pattern Tumor Tissue MicroRNA p-value #-valuea (FDR) Direction of Association’’ Absolute Standardized Hazard Ratio Max. Lower Uncor- Bound rected 95% Confidence @10% Estimate Interval FDR RM-Corrected Estimatec hsa-miR-27b 0.0016 6.1% (+) 2.66 (1.45, 4.88) 1.07 1.32 hsa-miR-21 0.0020 6.4% (+) 1.66 (1.21, 2.30) 1.05 1.34 hsa-miR-lOa 0.0024 6.7% (+) 1.78 (1.23, 2.59) 1.05 1.34 hsa-miR-93 0.0024 6.7% (+) 1.83 (1.24, 2.71) 1.05 1.34 hsa-miR-106b 0.0028 6.8% (+) 1.79 (1.22, 2.63) 1.05 1.33 hsa-miR-150 0.0035 7.1% (+) 1.61 (1.17, 2.22) 1.05 1.32 hsa-miR-1 0.0104 9.0% (-) 1.52 (1.10, 2.09) 1.00 1.28 aThe g-value is the empirical Bayes probability that the microRNA’s association with clinical endpoint is a false discovery, given the data. bDirection of association indicates where higher microRNA expression is associated with higher (+) or lower (-) risk of death due to prostate cancer. CRM: regression to the mean.

[00163] Table 19 and Table 20 shows the microRNAs that can be identified as being associated with the risk of clinical recurrence while adjusting for the clinical and pathology covariates of biopsy Gleason score, baseline PSA level, and clinical T-stage. The distributions of these covariates are shown in Figure 1. Fifteen (15) of the microRNAs identified in Table 15 are also present in Table 19, indicating that these microRNAs have predictive value for clinical recurrence that is independent of the Gleason score, baseline PSA, and clinical T-stage.

[00164] Two microRNAs assayed from primary Gleason pattern tumor tissue were found that had predictive value for death due to prostate cancer independent of Gleason score, baseline PSA, and clinical T-stage (Table 21). 116 2015227398 15 Sep 2015

Absolute Standardized Hazard Ratio

Table 19. MicroRNAs Associated with Clinical Recurrence of Prostate Cancer Adjusting for Biopsy Gleason Score, Baseline PSA Level, and Clinical T-Stage Primary Gleason Pattern Tumor Tissue

Max.

Lower

MicroRNA p-value cy-value3 (FDR) Direction of Asso-ciationb Uncor rected Estimate 95% Confidence Interval Bound @10% FDR RM- Corrected Estimate3 hsa-miR-30e-5p <0.0001 0.0% (-) 1.80 (1.42, 2.27) 1.23 1.53 hsa-miR-30a <0.0001 0.0% (-) 1.75 (1.40, 2.19) 1.22 1.51 hsa-miR-93 <0.0001 0.1% (+) 1.70 (1.32, 2.20) 1.19 1.44 hsa-miR-449a 0.0001 0.1% (+) 1.54 (1.25, 1.91) 1.17 1.39 hsa-miR-133a 0.0001 0.1% (-) 1.58 (1.25,2.00) 1.17 1.39 hsa-miR-27a 0.0002 0.1% (+) 1.66 (1.28, 2.16) 1.17 1.41 hsa-miR-21 0.0003 0.2% (+) 1.58 (1.23,2.02) 1.16 1.38 hsa-miR-182 0.0005 0.3% (+) 1.56 (1.22, 1.99) 1.15 1.37 hsa-miR-106b 0.0008 0.5% (+) 1.57 (1.21,2.05) 1.15 1.36 hsa-miR-222 0.0028 1.1% (-) 1.39 (1.12, 1.73) 1.11 1.28 hsa-miR-103 0.0048 1.7% (+) 1.69 (1.17, 2.43) 1.13 1.32 hsa-miR-486-5p 0.0059 2.0% (-) 1.34 (1.09, 1.65) 1.09 1.25 hsa-miR-1 0.0083 2.7% (-) 1.29 (1.07, 1.57) 1.07 1.23 hsa-miR-141 0.0088 2.8% (+) 1.43 (1.09, 1.87) 1.09 1.27 hsa-miR-200c 0.0116 3.4% (+) 1.39 (1.07, 1.79) 1.07 1.25 hsa-miR-145 0.0201 5.1% (-) 1.27 (1.03, 1.55) 1.05 1.20 hsa-miR-206 0.0329 7.2% (-) 1.40 (1.00, 1.91) 1.05 1.23 hsa-miR-29b 0.0476 9.4% (+) 1.30 (1.00, 1.69) 1.00 1.20 aThe g-value is the empirical Bayes probability that the microRNA’s association with clinical recurrence is a false discovery, given the data. bDirection of association indicates where higher microRNA expression is associated with higher (+) or lower (-) risk of clinical recurrence. CRM: regression to the mean. 117 2015227398 15 Sep 2015

Table 20. MicroRNAs Associated with Clinical Recurrence of Prostate Cancer Adjusting for Biopsy Gleason Score, Baseline PSA Level, and Clinical T-Stage Highest Gleason Pattern Tumor Tissue

Absolute Standardized Hazard Ratio

Max.

Lower

MicroRNA p-value ¢-value3 (FDR) Direction of Association’’ Uncor rected Estimate 95% Confidence Interval Bound @10% FDR RM- Corrected Estimate0 hsa-miR-30a <0.0001 0.0% (-) 1.62 (1.32,1.99) 1.20 1.43 hsa-miR-30e-5p <0.0001 0.0% (-) 1.53 (1.27,1.85) 1.19 1.39 hsa-miR-93 <0.0001 0.0% (+) 1.76 (1.37,2.26) 1.20 1.45 hsa-miR-205 <0.0001 0.0% (-) 1.47 (1.23,1.74) 1.18 1.36 hsa-miR-449a 0.0001 0.1% (+) 1.62 (1.27, 2.07) 1.18 1.38 hsa-miR-106b 0.0003 0.2% (+) 1.65 (1.26, 2.16) 1.17 1.36 hsa-miR-133a 0.0005 0.2% (-) 1.51 (1.20,1.90) 1.16 1.33 hsa-miR-1 0.0007 0.3% (-) 1.38 (1.15, 1.67) 1.13 1.28 hsa-miR-210 0.0045 1.2% (+) 1.35 (1.10, 1.67) 1.11 1.25 hsa-miR-182 0.0052 1.3% (+) 1.40 (1.10,1.77) 1.11 1.26 hsa-miR-425 0.0066 1.6% (+) 1.48 (1.12,1.96) 1.12 1.26 hsa-miR-155 0.0073 1.8% (-) 1.36 (1.09,1.70) 1.10 1.24 hsa-miR-21 0.0091 2.1% (+) 1.42 (1.09, 1.84) 1.10 1.25 hsa-miR-222 0.0125 2.7% (-) 1.34 (1.06, 1.69) 1.09 1.23 hsa-miR-27a 0.0132 2.8% (+) 1.40 (1.07, 1.84) 1.09 1.23 hsa-miR-191* 0.0150 3.0% (+) 1.37 (1.06, 1.76) 1.09 1.23 hsa-miR-103 0.0180 3.4% (+) 1.45 (1.06, 1.98) 1.09 1.23 hsa-miR-31 0.0252 4.3% (-) 1.27 (1.00, 1.57) 1.07 1.19 hsa-miR-19b 0.0266 4.5% (-) 1.29 (1.00, 1.63) 1.07 1.20 hsa-miR-99a 0.0310 5.0% (-) 1.26 (1.00, 1.56) 1.06 1.18 hsa-miR-92a 0.0348 5.4% (+) 1.31 (1.00, 1.69) 1.06 1.19 hsa-miR-146b-5p 0.0386 5.8% (-) 1.29 (1.00, 1.65) 1.06 1.19 hsa-miR-145 0.0787 9.7% (-) 1.23 (1.00, 1.55) 1.00 1.15 aThe g-value is the empirical Bayes probability that the microRNA’s association with clinical clinical recurrence is a false discovery, given the data. bDirection of association indicates where higher microRNA expression is associated with higher (+) or lower (-) risk of clinical recurrence. CRM: regression to the mean. 118 2015227398 15 Sep 2015

Table 21. MicroRNAs Associated with Death Due to Prostate Cancer Adjusting for Biopsy Gleason Score, Baseline PSA Level, and Clinical T-Stage Primary Gleason Pattern Tumor Tissue Absolute Standardized Hazard Ratio MicroRNA p-value r/-value3 (FDR) Direction of Association’’ Uncor rected Estimate 95% Confidence Interval Max. Lower Bound @10% FDR RM- Corrected Estimatec hsa-miR-30e-5p 0.0001 2.9% (-) 1.97 (1.40, 2.78) 1.09 1.39 hsa-miR-30a 0.0002 3.3% (-) 1.90 (1.36, 2.65) 1.08 1.38 aThe ('/-value is the empirical Bayes probability that the microRNA’ s association with clinical recurrence is a false discovery, given the data. bDirection of association indicates where higher microRNA expression is associated with higher (+) or lower (-) risk of clinical recurrence. CRM: regression to the mean.

[00165] Accordingly, the normalized expression levels of hsa-miR-93; hsa-miR-106b; hsa-miR-21; hsa-miR-449a; hsa-miR-182; hsa-miR-27a; hsa-miR-103; hsa-miR-141; hsa-miR-92a; hsa-miR-22; hsa-miR-29b; hsa-miR-210; hsa-miR-331; hsa-miR-191; hsa-miR-425; and hsa-miR-200c are positively associated with an increased risk of recurrence; and hsa-miR-30e-5p; hsa-miR-133a; hsa-miR-30a; hsa-miR-222; hsa-miR-1; hsa-miR-145; hsa-miR-486-5p; hsa-miR-19b; hsa-miR-205; hsa-miR-31; hsa-miR-155; hsa-miR-206; hsa-miR-99a; andhsa-miR-146b-5p are negatively associated with an increased risk of recurrence.

[00166] Furthermore, the normalized expression levels of hsa-miR-106b; hsa-miR-21; hsa-miR-93; hsa-miR-331; hsa-miR-150; hsa-miR-27b; and hsa-miR-lOa are positively associated with an increased risk of prostate cancer specific death; and the normalized expression levels of hsa-miR-30e-5p; hsa-miR-30a; hsa-miR-133a; hsa-miR-222; hsa-miR-1; hsa-miR-26a; and hsa-miR-152 are negatively associated with an increased risk of prostate cancer specific death.

[00167] Table 22 shows the number of microRNA-gene pairs that were grouped in each tier (Tiers 1-4) and the number and percentage of those that were predictive of clinical recurrence at a false discovery rate of 10%. 119 2015227398 15 Sep 2015

Table 22. Tier Total Number of MicroRNA-Gene Pairs Number of Pairs Predictive of Clinical Recurrence at False Discovery Rate 10% (%) Tier 1 80 46 (57.5%) Tier 2 719 591 (82.2%) Tier 3 3,850 2,792 (72.5%) Tier 4 54,724 38,264 (69.9%) 120 121 2015227398 15 Sep 2015

Offlclul wm hoi. VctWsloft Numbers smm NO forward Printer JimHefler: V) Reverse Primer Wittienev. M :111 Pr»be Sequence: 111 Am plfatn Aet|«eftee; AAMP NM_001087 1 GTGTGGCAGGTGG ACACTAA 2 CTCCATCCACTCCAG GTCTC 3 CGCTTCAAAGGACCAGACCT CCTC 4 GTGTGGCAGGTGGACACTAAGGAGGAGGTCTGGTCC TTTGAAGCGGGAGACCTGGAGTGGATGGAG ABCA5 NM_172232 5 GGTATGGATCCCA AAGCCA 6 CAGCCCGCTTTCTGTT TTTA 7 CACATGTGGCGAGCAATTCG AACT 8 GGTATGGATCCCAAAGCCAAACAGCACATGTGGCGA GCAATTCGAACTGCATTTAAAAACAGAAAGCGGGCT ABCB1 NM_000927 9 AAACACCACTGGA GCATTGA 10 CAAGCCTGGAACCTA TAGCC 11 CAAGCCTGGAACCTATAGCC 12 AAACACCACTGGAGCATTGACTACCAGGCTCGCCAA TGATGCTGCTCAAGTTAAAGGGGCTATAGGTTCCAG ABCC1 NM_004996 13 TCATGGTGCCCGT CAATG 14 CGATTGTCTTTGCTCT TCATGTG 15 ACCTGATACGTCTTGGTCTT CATCGCCAT 16 TCATGGTGCCCGTCAATGCTGTGATGGCGATGAAGA CCAAGACGTATCAGGTGGCCCACATGAAGAGCAAAG ABCC3 NM_003786 17 TCATCCTGGCGAT CTACTTCCT 18 CCGTTGAGTGGAATC AGCAA 19 TCTGTCCTGGCTGGAGTCGC TTTCAT 20 TCATCCTGGCGATCTACTTCCTCTGGCAGAACCTAGG TCCCTCTGTCCTGGCTGGAGTCGCTTTCATGGTCTTG CTGATTCCACTCAACGG ABCC4 NM_005845 21 AGCGCCTGGAATC TACAACT 22 AGAGCCCCTGGAGAG AAGAT 23 CGGAGTCCAGTGTTTTCCCA CTTA 24 AGCGCCTGGAATCTACAACTCGGAGTCCAGTGTTTTC CCACTTATCATCTTCTCTCCAGGGGCTCT ABCC8 NM_000352 25 CGTCTGTCACTGT GGAGTGG 26 TGATCCGGTTT AGC A GGC 27 AGTCTCTTGGCCACCTTCAG CCCT 28 CGTCTGTCACTGTGGAGTGGACAGGGCTGAAGGTGG CCAAGAGACTGCACCGCAGCCTGCTAAACCGGATCA ABCG2 NM_004827 29 GGTCTCAACGCCA TCCTG 30 CTTGGATCTTTCCTTG CAGC 31 ACGAAGATTTGCCTCCACCT GTGG 32 GGTCTCAACGCCATCCTGGGACCCACAGGTGGAGGC AAATCTTCGTTATTAGATGTCTTAGCTGCAAGGAAAG ABHD2 NM_007011 33 GTAGTGGGTCTGC ATGGATGT 34 TGAGGGTTGGCACTC AGG 35 CAGGTGGCTCCTTTGATCCC TGA 36 GTAGTGGGTCTGCATGGATGTTTCAGGGATCAAAGG AGCCACCTGGGCGCCTGAGTGCCAACCCTCA ACE NM_000789 37 CCGCTGTACGAGG ATTTCA 38 CCGTGTCTGTGAAGC CGT 39 TGCCCTCAGCAATGAAGCCT ACAA 40 CCGCTGTACGAGGATTTCACTGCCCTCAGCAATGAA GCCTACAAGCAGGACGGCTTCACAGACACGG ACOX2 NM_003500 41 ATGGAGGTGCCCA GAACAC 42 ACTCCGGGTAACTGT GGATG 43 TGCTCTCAACTTTCCTGCGG AGTG 44 ATGGAGGTGCCCAGAACACTGCACTCCGCAGGAAAG TTGAGAGCATCATCCACAGTTACCCGGAGT ACTR2 NM 005722 45 ATCCGCATTGAAG ACCCA 46 ATCCGCTAGAACTGC ACCAC 47 CCCGCAGAAAGCACATGGT A TTCC 48 ATCCGCATTGAAGACCCACCCCGCAGAAAGCACATG GTATTCCTGGGTGGTGCAGTTCTAGCGGAT ADAM 15 NM 003815 49 GGCGGGATGTGGT 50 ATTTCTGGGCCTCCG 51 TCAGCCACAATCACCAACTC 52 GGCGGGATGTGGTAACAGAGACCAAGACTGTGGAGT ADAMT SI NM 006988 53 GGACAGGTGCAAG CTCATCTG 54 ATCTACAACCTTGGG CTGCAA 55 CAAGCCAAAGGCATTGGCTA CTTCTTCG 56 GGACAGGTGCAAGCTCATCTGCCAAGCCAAAGGCAT TGGCTACTTCTTCGTTTTGCAGCCCAAGGTTGTAGAT ADH5 NM 000671 57 ATGCTGTCATCATT 58 CTGCTTCCTTTCCCTT 59 TGTCTGCCCATTATCTTCAT 60 ATGCTGTCATCATTGTCACGGTTTGTCTGCCCATTAT AFAP1 NM 198595 61 GATGTCCATCCTT 62 CAACCCTGATGCCTG 63 CCTCCAGTGCTGTGTTCCCA 64 GATGTCCATCCTTGAAACAGCCTCTTCTGGGAACACA AGTR1 NM 000685 65 AGCATTGATCGAT 66 CTACAAGCATTGTGC 67 ATTGTTCACCCAATGAAGTC 68 AGCATTGATCGATACCTGGCTATTGTTCACCCAATGA AGTR2 NM 000686 69 ACTGGCATAGGAA 70 ATTGACTGGGTCTCTT 71 CCACCCAGACCCCATGTAGC 72 ACTGGCATAGGAAATGGTATCCAGAATGGAATTTTG AIG1 NM 016108 73 CGACGGTTCTGCC 74 TGCTCCTGCTGGGAT 75 AATCGAGATGAGGACATCGC 76 CGACGGTTCTGCCCTTTATATTAATCGAGATGAGGAC AKAP1 NM 003488 77 TGTGGTTGGAGAT 78 GTCTACCCACTGGGC 79 CTCCACCAGGGACCGGTTTA 80 TGTGGTTGGAGATGAAGTGGTGTTGATAAACCGGTC AKR1C1 BC040210 81 GTGTGTGAAGCTG 82 CTCTGCAGGCGCATA 83 CCAAATCCCAGGACAGGCAT 84 GTGTGTGAAGCTGAATGATGGTCACTTCATGCCTGTC AKR1C3 NM 003739 85 GCTTTGCCTGATG TCTACCAGAA 86 GTCCAGTCACCGGCA TAGAGA 87 TGCGTCACCATCCACACACA GGG 88 GCTTTGCCTGATGTCTACCAGAAGCCCTGTGTGTGGA TGGTGACGCAGAGGACGTCTCTATGCCGGTGACTGG AKT1 NM 005163 89 CGCTTCTATGGCG 90 TCCCGGTACACCACG 91 CAGCCCTGGACTACCTGCAC 92 CGCTTCTATGGCGCTGAGATTGTGTCAGCCCTGGACT AKT2 NM 001626 93 TCCTGCCACCCTTC 94 GGCGGTAAATTCATC 95 CAGGTCACGTCCGAGGTCGA 96 TCCTGCCACCCTTCAAACCTCAGGTCACGTCCGAGGT AKT3 NM 005465 97 TTGTCTCTGCCTTG GACTATCTACA 98 CCAGCATTAGATTCTC CAACTTGA 99 TCACGGTACACAATCTTTCC GGA 100 TTGTCTCTGCCTTGGACTATCTACATTCCGGAAAGAT TGTGTACCGTGATCTCAAGTTGGAGAATCTAATGCTG ALCAM NM 001627 101 GAGGAATATGGAA 102 GTGGCGGAGATCAAG 103 CCAGTTCCTGCCGTCTGCTC 104 GAGGAATATGGAATCCAAGGGGGCCAGTTCCTGCCG ALDH18 A1 NM 002860 105 GATGCAGCTGGAA CCCAA 106 CTCCAGCTCAGTGGG GAA 107 CCTGAAACTTGCATCTCCTG CTGC 108 GATGCAGCTGGAACCCAAGCTGCAGCAGGAGATGCA AGTTTCAGGATGTTCCCCACTGAGCTGGAG ALDH1A NM 170696 109 CACGTCTGTCCCT 110 GACCGTGGCTCAACT 111 TCTCTGTAGGGCCCAGCTCT 112 CACGTCTGTCCCTCTCTGCTTTCTCTGTAGGGCCCAG ALKBH3 NM 139178 113 TCGCTTAGTCTGC 114 TCTGAGCCCCAGTTTT 115 TAAACAGGGCAGTCACTTTC 116 TCGCTTAGTCTGCACCTCAACCGTGCGGAAAGTGACT ALOX12 NM 000697 117 AGTTCCTCAATGG 118 AGCACTAGCCTGGAG 119 CATGCTGTTGAGACGCTCGA 120 AGTTCCTCAATGGTGCCAACCCCATGCTGTTGAGACG ALOX5 NM 000698 121 GAGCTGCAGGACT 122 GAAGCCTGAGGACTT 123 CCGCATGCCGTACACGTAGA 124 GAGCTGCAGGACTTCGTGAACGATGTCTACGTGTAC AMACR NM 203382 125 GTCTCTGGGCTGT CAGCTTT 126 TGGGTATAAGATCCA GAACTTGC 127 TCCATGTGTTTGATTTCTCCT CAGGC 128 GTCTCTGGGCTGTCAGCTTTCCTTTCTCCATGTGTTT GATTTCTCCTCAGGCTGGTAGCAAGTTCTGGATCTTA 122 2015227398 15 Sep 2015

Official wm hoi. WcVslod Numbers K nisi! forward Primer Seriuencc: Hi NO Sever*’ Primer Secunwr; ise Prnhr Sequence: IB WO Ampllcon .Sequence: AMPD3 NM_000480 129 TGGTTCATCCAGC ACAAGG 130 CATAAATCCGGGGCA CCT 131 TACTCTCCCAACATGCGCTG GATC 132 TGGTTCATCCAGCACAAGGTCTACTCTCCCAACATGC GCTGGATCATCCAGGTGCCCCGGATTTATG ANGPT2 NM 001147 133 CCGTGAAAGCTGC 134 TTGCAGTGGGAAGAA 135 AAGCTGACACAGCCCTCCCA 136 CCGTGAAAGCTGCTCTGTAAAAGCTGACACAGCCCT ANLN NM 018685 137 TGAAAGTCCAAAA 138 CAGAACCAAGGCTAT 139 CCAAAGAACTCGTGTCCCTC 140 TGAAAGTCCAAAACCAGGAAAATTCCAAAGAACTCG ANPEP NM001150 141 CCACCTTGGACCA AAGTAAAGC 142 TCTCAGCGTCACCTG GTAGGA 143 CTCCCCAACACGCTGAAACC CG 144 CCACCTTGGACCAAAGTAAAGCGTGGAATCGTTACC GCCTCCCCAACACGCTGAAACCCGATTCCTACCGGG ANXA2 NM 004039 145 CAAGACACTAAGG GCGACTACCA 146 CGTGTCGGGCTTCAG TCAT 147 CCACCACACAGGTACAGCAG CGCT 148 CAAGACACTAAGGGCGACTACCAGAAAGCGCTGCTG TACCTGTGTGGTGGAGATGACTGAAGCCCGACACG APC NM 000038 149 GGACAGCAGGAAT 150 ACCCACTCGATTTGTT 151 CATTGGCTCCCCGTGACCTG 152 GGACAGCAGGAATGTGTTTCTCCATACAGGTCACGG APEX1 NM 001641 153 GATGAAGCCTTTC 154 AGGTCTCCACACAGC 155 CTTTCGGGAAGCCAGGCCCT 156 GATGAAGCCTTTCGCAAGTTCCTGAAGGGCCTGGCTT APOC1 NM 001645 157 CCAGCCTGATAAA 158 CACTCTGAATCCTTGC 159 AGGACAGGACCTCCCAACCA 160 CCAGCCTGATAAAGGTCCTGCGGGCAGGACAGGACC APOE NMJ100041 161 GCCTCAAGAGCTG GTTCG 162 CCTGCACCTTCTCCAC CA 163 ACTGGCGCTGCATGTCTTCC AC 164 GCCTCAAGAGCTGGTTCGAGCCCCTGGTGGAAGACA TGCAGCGCCAGTGGGCCGGGCTGGTGGAGAAGGTGC APRT NM 000485 165 GAGGTCCTGGAGT 166 AGGTGCCAGCTTCTC 167 CCTTAAGCGAGGTCAGCTCC 168 GAGGTCCTGGAGTGCGTGAGCCTGGTGGAGCTGACC AOP2 NM 000486 169 GTGTGGGTGCCAG 170 CCCTTCAGCCCTCTCA 171 CTCCTTCCCTTCCCCTTCTCC 172 GTGTGGGTGCCAGTCCTCCTCAGGAGAAGGGGAAGG AR NM 000044 173 CGACTTCACCGCA 174 TGACACAAGTGGGAC 175 ACCATGCCGCCAGGGTACCA 176 CGACTTCACCGCACCTGATGTGTGGTACCCTGGCGG ARF1 NM 001658 177 CAGTAGAGATCCC 178 ACAAGCACATGGCTA 179 CTTGTCCTTGGGTCACCCTG 180 CAGTAGAGATCCCCGCAACTCGCTTGTCCTTGGGTCA ARHGAP 29 NM_004815 181 CACGGTCTCGTGG TGAAGT 182 CAGTTGCTTGCCCAG GAC 183 ATGCCAGACCCAGACAAAG CATCA 184 CACGGTCTCGTGGTGAAGTCAATGCCAGACCCAGAC AAAGCATCAGCTTGTCCTGGGCAAGCAACTG ARHGDI NM_001175 185 TGGTCCCTAGAAC 186 TGATGGAGGATCAGA 187 TAAAACCGGGCTTTCACCCA 188 TGGTCCCTAGAACAAGAGGCTTAAAACCGGGCTTTC ASAP2 NM 003887 189 CGGCCCATCAGCT 190 CTCTGGCCAAAGATA 191 CTGGGCTCCAACCAGCTTCA 192 CGGCCCATCAGCTTCTACCAGCTGGGCTCCAACCAG ASPN NM 017680 193 TGGACTAATCTGT 194 AAACACCCTTCAACA 195 AGTATCACCCAGGGTGCAGC 196 TGGACTAATCTGTGGGAGCAGTTTATTCCAGTATCAC ATM NM 000051 197 TGCTTTCTACACAT 198 GTTGTGGATCGGCTC 199 CCAGCTGTCTTCGACACTTC 200 TGCTTTCTACACATGTTCAGGGATTTTTCACCAGCTG ATP5E NM 006886 201 CCGCTTTCGCTAC 202 TGGGAGTATCGGATG 203 TCCAGCCTGTCTCCAGTAGG 204 CCGCTTTCGCTACAGCATGGTGGCCTACTGGAGACA ATP5J NM_0010037 03 205 GTCGACCGACTGA AACGG 206 CTCTACTTCCGGCCCT GG 207 CTACCCGCCATCGCAATGCA TTAT 208 GTCGACCGACTGAAACGGCGGCCCATAATGCATTGC GATGGCGGGTAGGCGTGTGGGGGCGGAGCCAGGGCC ATXN1 NM 000332 209 GATCGACTCCAGC 210 GAACTGTATCACGGC 211 CGGGCTATGGCTGTCTTCAA 212 GATCGACTCCAGCACCGTAGAGAGGATTGAAGACAG AURKA NM 003600 213 CATCTTCCAGGAG 214 TCCGACCTTCAATCAT 215 CTCTGTGGCACCCTGGACTA 216 CATCTTCCAGGAGGACCACTCTCTGTGGCACCCTGGA AURKB NM 004217 217 AGCTGCAGAAGAG 218 GCATCTGCCAACTCC 219 TGACGAGCAGCGAACAGCC 220 AGCTGCAGAAGAGCTGCACATTTGACGAGCAGCGAA AXIN2 NM 004655 221 GGCTATGTCTTTG 222 ATCCGTCAGCGCATC 223 ACCAGCGCCAACGACAGTG 224 GGCTATGTCTTTGCACCAGCCACCAGCGCCAACGAC AZGP1 NM 001185 225 GAGGCCAGCTAGG 226 CAGGAAGGGCAGCTA 227 TCTGAGATCCCACATTGCCT 228 GAGGCCAGCTAGGAAGCAAGGGTTGGAGGCAATGTG BAD NM 032989 229 GGGTCAGGGGCCT 230 CTGCTCACTCGGCTC 231 TGGGCCCAGAGCATGTTCCA 232 GGGTCAGGGGCCTCGAGATCGGGCTTGGGCCCAGAG BAG5 NM0010150 49 233 ACTCCTGCAATGA ACCCTGT 234 ACAAACAGCTCCCCA CGA 235 ACACCGGATTTAGCTCTTGT CGGC 236 ACTCCTGCAATGAACCCTGTTGACACCGGATTTAGCT CTTGTCGGCCTTCGTGGGGAGCTGTTTGT BAK1 NM001188 237 CCATTCCCACCATT 238 GGGAACATAGACCCA 239 ACACCCCAGACGTCCTGGCC 240 CCATTCCCACCATTCTACCTGAGGCCAGGACGTCTGG BAX NM 004324 241 CCGCCGTGGACAC 242 TTGCCGTCAGAAAAC 243 TGCCACTCGGAAAAAGACCT 244 CCGCCGTGGACACAGACTCCCCCCGAGAGGTCTTTTT BBC3 NM_014417 245 CCTGGAGGGTCCT GTACAAT 246 CTAATTGGGCTCCAT CTCG 247 CATCATGGGACTCCTGCCCT TACC 248 CCTGGAGGGTCCTGTACAATCTCATCATGGGACTCCT GCCCTTACCCAGGGGCCACAGAGCCCCCGAGATGGA BCL2 NM_000633 249 CAGATGGACCTAG TACCCACTGAGA 250 CCTATGATTTAAGGG CATTTTTCC 251 TTCCACGCCGAAGGACAGCG AT 252 CAGATGGACCTAGTACCCACTGAGATTTCCACGCCG AAGGACAGCGATGGGAAAAATGCCCTTAAATCATAG BDKRB1 NM_000710 253 GTGGCAGAAATCT 254 GAAGGGCAAGCCCAA 255 ACCTGGCAGCCTCTGATCTG 256 GTGGCAGAAATCTACCTGGCCAACCTGGCAGCCTCT BGN NM_001711 257 GAGCTCCGCAAGG 258 CTTGTTGTTCACCAGG 259 CAAGGGTCTCCAGCACCTCT 260 GAGCTCCGCAAGGATGACTTCAAGGGTCTCCAGCAC BIK NM_001197 261 ATTCCTATGGCTCT GCAATTGTC 262 GGCAGGAGTGAATGG CTCTTC 263 CCGGTTAACTGTGGCCTGTG CCC 264 ATTCCTATGGCTCTGCAATTGTCACCGGTTAACTGTG GCCTGTGCCCAGGAAGAGCCATTCACTCCTGCC BIN1 NM 004305 265 CCTGCAAAAGGGA ACAAGAG 266 CGTGGTTGACTCTGA TCTCG 267 CTTCGCCTCCAGATGGCTCC C 268 CCTGCAAAAGGGAACAAGAGCCCTTCGCCTCCAGAT GGCTCCCCTGCCGCCACCCCCGAGATCAGAGTCAAC BIRC5 NM0010122 71 269 TTCAGGTGGATGA GGAGACA 270 CACACAGCAGTGGCA AAAG 271 TCTGCCAGACGCTTCCTATC ACTCTATTC 272 TTCAGGTGGATGAGGAGACAGAATAGAGTGATAGGA AGCGTCTGGCAGATACTCCTTTTGCCACTGCTGTGTG BMP6 NM001718 273 GTGCAGACCTTGG 274 CTTAGTTGGCGCACA 275 TGAACCCCGAGTATGTCCCC 276 GTGCAGACCTTGGTTCACCTTATGAACCCCGAGTATG BMPR1B NM 001203 277 ACCACTTTGGCCA 278 GCGGTGTTTGTACCC 279 ATTCACATTACCATAGCGGC 280 ACCACTTTGGCCATCCCTGCATTTGGGGCCGCTATGG 123 2015227398 15 Sep 2015

offtriul Nv in hill Wcv>j<>n m&amp;m forward Primer .Srrincnec: Μϋϋ Sevrrw Primer Sectunirr; ιιβ Probe Sequence: SIX? II) M> AmpScen .Sequence: BRCA1 NM 007294 281 TCAGGGGGCT AGA 282 CCATTCCAGTTGATCT 283 CTATGGGCCCTTCACCAACA 284 TCAGGGGGCTAGAAATCTGTTGCTATGGGCCCTTCAC BRCA2 NM 000059 285 AGTTCGTGCTTTG 286 AAGGTAAGCTGGGTC 287 CATTCTTCACTGCTTCATAA 288 AGTTCGTGCTTTGCAAGATGGTGCAGAGCTTTATGAA BTG1 NM 001731 289 GAGGTCCGAGCGA 290 AGTTATTTTCGAGAC 291 CGCTCGTCTCTTCCTCTCTC 292 GAGGTCCGAGCGATGTGACCAGGCCGCCATCGCTCG BTG3 NM 006806 293 CCATATCGCCCAA 294 CCAGTGATTCCGGTC 295 CATGGGTACCTCCTCCTGGA 296 CCATATCGCCCAATTCCAGTGACATGGGTACCTCCTC BTRC NM 033637 297 GTTGGGACACAGT 298 TGAAGCAGTCAGTTG 299 CAGTCGGCCCAGGACGGTCT 300 GTTGGGACACAGTTGGTCTGCAGTCGGCCCAGGACG BUB1 NM 004336 301 CCGAGGTTAATCC 302 AAGACATGGCGCTCT 303 TGCTGGGAGCCTACACTTGG 304 CCGAGGTTAATCCAGCACGTATGGGGCCAAGTGTAG Cl NM 000587 305 ATGTCTGAGTGTG 306 AGGCCTTATGCTGGT 307 ATGCTCTGCCCTCTGCATCT 308 ATGTCTGAGTGTGAGGCGGGCGCTCTGAGATGCAGA CACNA1 D NM_000720 309 AGGACCCAGCTCC ATGTG 310 CCTACATTCCGTGCC ATTG 311 CAGTACACTGGCGTCCATTC CCTG 312 AGGACCCAGCTCCATGTGCGTTCTCAGGGAATGGAC GCCAGTGTACTGCCAATGGCACGGAATGTAGG CADM1 NM 014333 313 CCACCACCATCCT 314 GATCCACTGCCCTGA 315 TCTTCACCTGCTCGGGAATC 316 CCACCACCATCCTTACCATCATCACAGATTCCCGAGC CADPS NM 003716 317 CAGCAAGGAGACT 318 GGTCCTCTTCTCCACG 319 CTCCTGGATGGCCAAATTTG 320 CAGCAAGGAGACTGTGCTGAGCTCCTGGATGGCCAA CASP1 NM 001223 321 AACTGGAGCTGAG 322 CATCTACGCTGTACC 323 TCACAGGCATGACAATGCTG 324 AACTGGAGCTGAGGTTGACATCACAGGCATGACAAT CASP3 NM 032991 325 TGAGCCTGAGCAG 326 CCTTCCTGCGTGGTCC 327 TCAGCCTGTTCCATGAAGGC 328 TGAGCCTGAGCAGAGACATGACTCAGCCTGTTCCAT CASP7 NM 033338 329 GCAGCGCCGAGAC 330 AGTCTCTCTCCGTCGC 331 CTTTCGCTAAAGGGGCCCCA 332 GCAGCGCCGAGACTTTTAGTTTCGCTTTCGCTAAAGG CAV1 NM 001753 333 GTGGCTCAACATT 334 CAATGGCCTCCATTTT 335 ATTTCAGCTGATCAGTGGGC 336 GTGGCTCAACATTGTGTTCCCATTTCAGCTGATCAGT CAV2 NM 198212 337 CTTCCCTGGGACG 338 CTCCTGGTCACCCTTC 339 CCCGTACTGTCATGCCTCAG 340 CTTCCCTGGGACGACTTGCCAGCTCTGAGGCATGAC CCL2 NM 002982 341 CGCTCAGCCAGAT GCAATC 342 GCACTGAGATCTTCC TATTGGTGAA 343 TGCCCCAGTCACCTGCTGTT A 344 CGCTCAGCCAGATGCAATCAATGCCCCAGTCACCTG CTGTTATAACTTCACCAATAGGAAGATCTCAGTGC CCL5 NM 002985 345 AGGTTCTGAGCTC 346 ATGCTGACTTCCTTCC 347 ACAGAGCCCTGGCAAAGCC 348 AGGTTCTGAGCTCTGGCTTTGCCTTGGCTTTGCCAGG CCNB1 NM 031966 349 TTCAGGTTGTTGC AGGAGAC 350 CATCTTCTTGGGCAC ACAAT 351 TGTCTCCATTATTGATCGGT TCATGCA 352 TTCAGGTTGTTGCAGGAGACCATGTACATGACTGTCT CCATTATTGATCGGTTCATGCAGAATAATTGTGTGCC CCND1 NM 001758 353 GCATGTTCGTGGC 354 CGGTGTAGATGCACA 355 AAGGAGACCATCCCCCTGAC 356 GCATGTTCGTGGCCTCTAAGATGAAGGAGACCATCC CCNE2 NM 057749 357 ATGCTGTGGCTCC TTCCTAACT 358 ACCCAAATTGTGATA T ACAAAAAGGTT 359 TACCAAGCAACCTACATGTC AAGAAAGCCC 360 ATGCTGTGGCTCCTTCCTAACTGGGGCTTTCTTGACA TGTAGGTTGCTTGGTAATAACCTTTTTGTATATCACA CCNH NM 001239 361 GAGATCTTCGGTG 362 CTGCAGACGAGAACC 363 CATCAGCGTCCTGGCGTAAA 364 GAGATCTTCGGTGGGGGTACGGGTGTTTTACGCCAG CCR1 NM 001295 365 TCCAAGACCCAAT 366 TCGTAGGCTTTCGTG 367 ACTCACCACACCTGCAGCCT 368 TCCAAGACCCAATGGGAATTCACTCACCACACCTGC CD164 NM 006016 369 CAACCTGTGCGAA 370 ACACCCAAGACCAGG 371 CCTCCAATGAAACTGGCTGC 372 CAACCTGTGCGAAAGTCTACCTTTGATGCAGCCAGTT CD1A NM_001763 373 GGAGTGGAAGGAA CTGGAAA 374 TCATGGGCGTATCTA CGAAT 375 CGCACCATTCGGTCATTTGA GG 376 GGAGTGGAAGGAACTGGAAACATTATTCCGTATACG CACCATTCGGTCATTTGAGGGAATTCGTAGATACGCC CD276 ΝΜ 0010247 36 377 CCAAAGGATGCGA TACACAG 378 GGATGACTTGGGAAT CATGTC 379 CCACTGTGCAGCCTTATTTC TCCAATG 380 CCAAAGGATGCGATACACAGACCACTGTGCAGCCTT ATTTCTCCAATGGACATGATTCCCAAGTCATCC CD44 NM 000610 381 GGCACCACTGCTT 382 GATGCTCATGGTGAA 383 ACTGGAACCCAGAAGCACA 384 GGCACCACTGCTTATGAAGGAAACTGGAACCCAGAA CD68 NM 001251 385 TGGTTCCCAGCCC 386 CTCCTCCACCCTGGGT 387 CTCCAAGCCCAGATTCAGAT 388 TGGTTCCCAGCCCTGTGTCCACCTCCAAGCCCAGATT CD82 NM 002231 389 GTGCAGGCTCAGG TGAAGTG 390 GACCTCAGGGCGATT CATGA 391 TCAGCTTCTACAACTGGACA GACAACGCTG 392 GTGCAGGCTCAGGTGAAGTGCTGCGGCTGGGTCAGC TTCTACAACTGGACAGACAACGCTGAGCTCATGAAT CDC20 NM 001255 393 TGGATTGGAGTTC 394 GCTTGCACTCCACAG 395 ACTGGCCGTGGCACTGGACA 396 TGGATTGGAGTTCTGGGAATGTACTGGCCGTGGCAC CDC25B NM 021873 397 GCTGCAGGACCAG 398 TAGGGCAGCTGGCTT 399 CTGCTACCTCCCTTGCCTTT 400 GCTGCAGGACCAGTGAGGGGCCTGCGCCAGTCCTGC CDC6 NM 001254 401 GCAACACTCCCCA 402 TGAGGGGGACCATTC 403 TTGTTCTCCACCAAAGCAAG 404 GCAACACTCCCCATTTACCTCCTTGTTCTCCACCAAA CDH1 NM 004360 405 TGAGTGTCCCCCG GTATCTTC 406 CAGCCGCTTTCAGAT TTTCAT 407 TGCCAATCCCGATGAAATTG GAAATTT 408 TGAGTGTCCCCCGGTATCTTCCCCGCCCTGCCAATCC CGATGAAATTGGAAATTTTATTGATGAAAATCTGAAA CDH10 NM 006727 409 TGTGGTGCAAGTC 410 TGTAAATGACTCTGG 411 ATGCCGATGACCCTTCATAT 412 TGTGGTGCAAGTCACAGCTACAGATGCCGATGACCC CDH11 NM 001797 413 GTCGGCAGAAGCA 414 CTACTCATGGGCGGG 415 CCTTCTGCCCATAGTGATCA 416 GTCGGCAGAAGCAGGACTTGTACCTTCTGCCCATAG CDH19 NM 021153 417 AGTACCATAATGC 418 AGACTGCCTGTATAG 419 ACTCGGAAAACCACAAGCG 420 AGTACCATAATGCGGGAACGCAAGACTCGGAAAACC CDH5 NM 001795 421 ACAGGAGACGTGT 422 CAGCAGTGAGGTGGT 423 T ATTCTCCCGGTCCAGCCTC 424 ACAGGAGACGTGTTCGCCATTGAGAGGCTGGACCGG CDH7 NM 033646 425 GTTTGACATGGCT 426 AGTCACATCCCTCCG 427 ACCTCAACGTCATCCGAGAC 428 GTTTGACATGGCTGCACTGAGAAACCTCAACGTCATC CDK14 NM 012395 429 GCAAGGTAAATGG 430 GATAGCTGTGAAAGG 431 CTTCCTGCAGCCTGATCACC 432 GCAAGGTAAATGGGAAGTTGGTAGCTCTGAAGGTGA CDK2 NM_001798 433 AATGCTGCACTAC GACCCTA 434 TTGGTCACATCCTGG AAGAA 435 CCTTGGCCGAAATCCGCTTG T 436 AATGCTGCACTACGACCCTAACAAGCGGATTTCGGC CAAGGCAGCCCTGGCTCACCCTTTCTTCCAGGATGTG CDK3 NM 001258 437 CCAGGAAGGGACT 438 GTTGCATGAGCAGGT 439 CTCTGGCTCCAGATTGGGCA 440 CCAGGAAGGGACTGGAAGAGATTGTGCCCAATCTGG CDK7 NM_001799 441 GTCTCGGGCAAAG 442 CTCTGGCCTTGTAAA 443 CCTCCCCAAGGAAGTCCAGC 444 GTCTCGGGCAAAGCGTTATGAGAAGCTGGACTTCCT 2015227398 15 Sep 2015 124

offtrial Nt in hul Xm'Mon \t> forward Printer .Smlieflee: p V» Srtrtv Primer Swtunirr; ΙΙϋΙ iiiiii 1’r< 4>e ?><.< (lu-iice: il Amplhwt Aei|«emv; CDKN1A NM 000389 445 TGGAGACTCTCAG 446 GGCGTTTGGAGTGGT 447 CGGCGGCAGACCAGCATGA 448 TGGAGACTCTCAGGGTCGAAAACGGCGGCAGACCAG CDKN1C NM 000076 449 CGGCGATCAAGAA 450 CAGGCGCTGATCTCT 451 CGGGCCTCTGATCTCCGATT 452 CGGCGATCAAGAAGCTGTCCGGGCCTCTGATCTCCG CDKN2B NM 004936 453 GACGCTGCAGAGC 454 GCGGGAATCTCTCCT 455 CACAGGATGCTGGCCTTTGC 456 GACGCTGCAGAGCACCTTTGCACAGGATGCTGGCCT CDKN2C NM 001262 457 GAGCACTGGGCAA 458 CAAAGGCGAACGGGA 459 CCTGTAACTTGAGGGCCACC 460 GAGCACTGGGCAATCGTTACGACCTGTAACTTGAGG CDKN3 NM 005192 461 TGGATCTCTACCA 462 ATGTCAGGAGTCCCT 463 ATCACCCATCATCATCCAAT 464 TGGATCTCTACCAGCAATGTGGAATTATCACCCATCA CDS2 NM 003818 465 GGGCTTCTTTGCT 466 ACAGGGCAGACAAAG 467 CCCGGACATCACATAGGACA 468 GGGCTTCTTTGCTACTGTGGTGTTTGGCCTTCTGCTG CENPF NM 016343 469 CTCCCGTCAACAG 470 GGGTGAGTCTGGCCT 471 ACACTGGACCAGGAGTGCAT 472 CTCCCGTCAACAGCGTTCTTTCCAAACACTGGACCAG CHAF1A NM 005483 473 GAACTCAGTGTAT 474 GCTCTGTAGCACCTG 475 TGCACGTACCAGCACATCCT 476 GAACTCAGTGTATGAGAAGCGGCCTGACTTCAGGAT CHN1 NM 001822 477 TTACGACGCTCGT 478 TCTCCCTGATGCACAT 479 CCACCATTGGCCGCTTAGTG 480 TTACGACGCTCGTGAAAGCACATACCACTAAGCGGC CHRAC1 NM 017444 481 TCTCGCTGCCTCTA 482 CCTGGTTGATGCTGG 483 ATCCGGGTCATCATGAAGAG 484 TCTCGCTGCCTCTATCCCGCATCCGGGTCATCATGAA CKS2 NM 001827 485 GGCTGGACGTGGT 486 CGCTGCAGAAAATGA 487 CTGCGCCCGCTCTTCGCG 488 GGCTGGACGTGGTTTTGTCTGCTGCGCCCGCTCTTCG CLDN3 NM 001306 489 ACCAACTGCGTGC 490 GGCGAGAAGGAACAG 491 CAAGGCCAAGATCACCATCG 492 ACCAACTGCGTGCAGGACGACACGGCCAAGGCCAAG CLTC NM 004859 493 ACCGTATGGACAG 494 TGACTACAGGATCAG 495 TCTCACATGCTGTACCCAAA 496 ACCGTATGGACAGCCACAGCCTGGCTTTGGGTACAG COL11A NM 001854 497 GCCCAAGAGGGGA 498 GGACCTGGGTCTCCA 499 CTGCTCGACCTTTGGGTCCT 500 GCCCAAGAGGGGAAGATGGCCCTGAAGGACCCAAAG COL1A1 NM 000088 501 GTGGCCATCCAGC 502 CAGTGGTAGGTGATG 503 TCCTGCGCCTGATGTCCACC 504 GTGGCCATCCAGCTGACCTTCCTGCGCCTGATGTCCA COL1A2 NM_000089 505 CAGCCAAGAACTG GTATAGGAGCT 506 AAACTGGCTGCCAGC ATTG 507 TCTCCTAGCCAGACGTGTTT CTTGTCCTTG 508 CAGCCAAGAACTGGTATAGGAGCTCCAAGGACAAGA AACACGTCTGGCTAGGAGAAACTATCAATGCTGGCA COL3A1 NM 000090 509 GGAGGTTCTGGAC 510 ACCAGGACTGCCACG 511 CTCCTGGTCCCCAAGGTGTC 512 GGAGGTTCTGGACCTGCTGGTCCTCCTGGTCCCCAAG COL4A1 NM 001845 513 ACAAAGGCCTCCC 514 GAGTCCCAGGAAGAC 515 CTCCTTTGACACCAGGGATG 516 ACAAAGGCCTCCCAGGATTGGATGGCATCCCTGGTG COL5A1 NM 000093 517 CTCCCTGGGAAAG 518 CTGGACCAGGAAGCC 519 CCAGGGAAACCACGTAATCC 520 CTCCCTGGGAAAGATGGCCCTCCAGGATTACGTGGT COL5A2 NM 000393 521 GGTCGAGGAACCC 522 GCCTGGAGGTCCAAC 523 CCAGGAAATCCTGTAGCACC 524 GGTCGAGGAACCCAAGGTCCGCCTGGTGCTACAGGA COL6A1 NM 001848 525 GGAGACCCTGGTG 526 TCTCCAGGGACACCA 527 CTTCTCTTCCCTGATCACCC 528 GGAGACCCTGGTGAAGCTGGCCCGCAGGGTGATCAG COL6A3 NM 004369 529 GAGAGCAAGCGAG 530 AACAGGGAACTGGCC 531 CCTCTTTGACGGCTCAGCCA 532 GAGAGCAAGCGAGACATTCTGTTCCTCTTTGACGGCT COL8A1 NM 001850 533 TGGTGTTCCAGGG 534 CCCTGTAAACCCTGA 535 CCT AAGGGAGAGCCAGGAA 536 TGGTGTTCCAGGGCTTCTCGGACCTAAGGGAGAGCC COL9A2 NM 001852 537 GGGAACCATCCAG 538 ATTCCGGGTGGACAG 539 ACACAGGAAATCCGCACTGC 540 GGGAACCATCCAGGGTCTGGAAGGCAGTGCGGATTT CRISP3 NM 006061 541 TCCCTTATGAACA 542 AACCATTGGTGCATA 543 TGCCAGTTGCCCAGATAACT 544 TCCCTTATGAACAAGGAGCACCTTGTGCCAGTTGCCC CSF1 NM 000757 545 TGCAGCGGCTGAT TGACA 546 CAACTGTTCCTGGTCT ACAAACTCA 547 TCAGATGGAGACCTCGTGCC AAATTACA 548 TGCAGCGGCTGATTGACAGTCAGATGGAGACCTCGT GCCAAATTACATTTGAGTTTGTAGACCAGGAACAGTT CSK NM 004383 549 CCTGAACATGAAG 550 CATCACGTCTCCGAA 551 TCCCGATGGTCTGCAGCAGC 552 CCTGAACATGAAGGAGCTGAAGCTGCTGCAGACCAT CSRP1 NM 004078 553 ACCCAAGACCCTG 554 GCAGGGGTGGAGTGA 555 CCACCCTTCTCCAGGGACCC 556 ACCCAAGACCCTGCCTCTTCCACTCCACCCTTCTCCA CTGF NM 001901 557 GAGTTCAAGTGCC CTGACG 558 AGTTGTAATGGCAGG CACAO 559 AACATCATGTTCTTCTTCAT GACCTCGC 560 GAGTTCAAGTGCCCTGACGGCGAGGTCATGAAGAAG AACATGATGTTCATCAAGACCTGTGCCTGCCATTACA CTHRC1 NM 138455 561 TGGCTCACTTCGG 562 TCAGCTCCATTGAAT 563 CAACGCTGACAGCATGCATT 564 TGGCTCACTTCGGCTAAAATGCAGAAATGCATGCTGT CTNNA1 NM_001903 565 CGTTCCGATCCTCT ATACTGCAT 566 AGGTCCCTGTTGGCC TTATAGG 567 ATGCCTACAGCACCCTGATG TCGCA 568 CGTTCCGATCCTCTATACTGCATCCCAGGCATGCCTA CAGCACCCTGATGTCGCAGCCTATAAGGCCAACAGG CTNNB1 NM_001904 569 GGCTCTTGTGCGT ACTGTCCTT 570 TCAGATGACGAAGAG CACAGATG 571 AGGCTCAGTGATGTCTTCCC TGTCACCAG 572 GGCTCTTGTGCGTACTGTCCTTCGGGCTGGTGACAGG GAAGACATCACTGAGCCTGCCATCTGTGCTCTTCGTC CTNND1 NM 001331 573 CGGAAACTTCGGG 574 CTGAATCCTTCTGCCC 575 TTGATGCCCTCATTTTCATT 576 CGGAAACTTCGGGAATGTGATGGTTTAGTTGATGCC CTNND2 NM 001332 577 GCCCGTCCCTACA 578 CTCACACCCAGGAGT 579 CTATGAAACGAGCCACTACC 580 GCCCGTCCCTACAGTGAACTGAACTATGAAACGAGC CTSB NM 001908 581 GGCCGAGATCTAC 582 GCAGGAAGTCCGAAT 583 CCCCGTGGAGGGAGCTTTCT 584 GGCCGAGATCTACAAAAACGGCCCCGTGGAGGGAGC CTSD NM_001909 585 GTACATGATCCCC TGTGAGAAGGT 586 GGGACAGCTTGTAGC CTTTGC 587 ACCCTGCCCGCGATCACACT GA 588 GTACATGATCCCCTGTGAGAAGGTGTCCACCCTGCCC GCGATCACACTGAAGCTGGGAGGCAAAGGCTACAAG CTSK NM 000396 589 AGGCTTCTCTTGG 590 CCACCTCTTCACTGGT 591 CCCCAGGTGGTTCATAGCCA 592 AGGCTTCTCTTGGTGTCCATACATATGAACTGGCTAT CTSL2 NM 001333 593 TGTCTCACTGAGC 594 ACCATTGCAGCCCTG 595 CTTGAGGACGCGAACAGTCC 596 TGTCTCACTGAGCGAGCAGAATCTGGTGGACTGTTC CTSS NM 004079 597 TGACAACGGCTTT 598 TCCATGGCTTTGTAG 599 TGATAACAAGGGCATCGACT 600 TGACAACGGCTTTCCAGTACATCATTGATAACAAGG CUL1 NM 003592 601 ATGCCCTGGTAAT 602 GCGACCACAAGCCTT 603 CAGCCACAAAGCCAGCGTCA 604 ATGCCCTGGTAATGTCTGCATTCAACAATGACGCTGG CXCL12 NM 000609 605 GAGCTACAGATGC 606 TTTGAGATGCTTGAC 607 TTCTTCGAAAGCCATGTTGC 608 GAGCTACAGATGCCCATGCCGATTCTTCGAAAGCCA CXCR4 NM 003467 609 TGACCGCTTCTAC 610 AGGATAAGGCCAACC 611 CTGAAACTGGAACACAACCA 612 TGACCGCTTCTACCCCAATGACTTGTGGGTGGTTGTG CXCR7 NM_020311 613 CGCCTCAGAACGA TGGAT 614 GTTGCATGGCCAGCT GAT 615 CTCAGAGCCAGGGAACTTCT CGGA 616 CGCCTCAGAACGATGGATCTGCATCTCTTCGACTACT CAGAGCCAGGGAACTTCTCGGACATCAGCTGGCCAT 125 2015227398 15 Sep 2015

Ofttti .1 ΝνίιιΕκΊ Vl't >11 1 mm \o I urn.inJ Ptirm-r Λ(·(ΐιι»Ί«(·: 1 ϋφ iii ϋ: i Eii'i c’Csi· |'nTurr NifliH'inv: M Q ϋϋ i >i:q Hi NO \ηΐ)>ΙΓι'»ιι N<-(|m-Nn·: CYP3A5 NM 000777 617 TCATTGCCCAGTA 618 GACAGGCTTGCCTTT 619 T CCCGCCTCAAGTTTCTCAC 620 TCAr rTGCCCAGTATGGAGATGTATTGGTGAGAAACTT CYR61 NM 001554 621 TGCTCATTCTTGAG 622 GTGGCTGCATTAGTG 623 C AGCACCCTTGGCAGTTTCG 624 TGCr rCATTCTTGAGGAGCATTAAGGTATTTCGAAACT DAG1 NM 004393 625 GTGACTGGGCTCA 626 ATCCCACTTGTGCTCC 627 C AAGTCAGAGTTTCCCTGGT 628 GTG \CTGGGCTCATGCCTCCAAGTCAGAGTTTCCCTG DAP NM 004394 629 CCAGCCTTTCTGG 630 GACCAGGTCTGCCTC 631 C TCACCAGCTGGCAGACGTG 632 CCA 3CCTTTCTGGTGCTGTTCTCCAGTTCACGTCTGC DAPK1 NM 004938 633 CGCTGACATCATG 634 TCTCTTTCAGCAACGA 635 T CATATCCAAACTCGCCTCC 636 CGC TGACATCATGAATGTTCCTCGACCGGCTGGAGG DARC NM 002036 637 GCCCTCATTAGTC 638 CAGACAGAAGGGCTG 639 T CAGCGCCTGTGCTTCCAAG 640 GCC CTCATTAGTCCTTGGCTCTTATCTTGGAAGCACA DDIT4 NM 019058 641 CCTGGCGTCTGTC 642 CGAAGAGGAGGTGGA 643 C TAGCCTTTGGGACCGCTTC 644 CCT( 3GCGTCTGTCCTCACCATGCCTAGCCTTTGGGAC DDR2 NM_0010147 96 645 CTATTACCGGATC CAGGGC 646 CCCAGCAAGATACTC TCCCA 647 A T GTGCTCCCTATCCGCTGGA GTC 648 CTAr CGC rTACCGGATCCAGGGCCGGGCAGTGCTCCCTATC TGGATGTCTTGGGAGAGTATCTTGCTGGG DES NM_001927 649 ACTTCTCACTGGC 650 GCTCCACCTTCTCGTT 651 T GAACCAGGAGTTTCTGACC 652 ACT" rCTCACTGGCCGACGCGGTGAACCAGGAGTTTCT DHRS9 NM_005771 653 GGAGAAAGGTCTC 654 CAGTCAGTGGGAGCC 655 A TCAATAATGCTGGTGTTCC 656 GGA GAAAGGTCTCTGGGGTCTGATCAATAATGCTGG DHX9 NM_001357 657 GTTCGAACCATCT 658 TCCAGTTGGATTGTG 659 C CAAGGAACCACACCCACTT 660 GTT< 3GAACCATCTCAGCGACAAAACCAAGTGGGTGT DIAPH1 NM_005219 661 CAAGCAGTCAAGG 662 AGTTTTGCTCGCCTCA 663 T TCTTCTGTCTCCCGCCGCT 664 CAA 3CAGTCAAGGAGAACCAGAAGCGGCGGGAGAC DICER 1 NM 177438 665 TCCAATTCCAGCA 666 GGCAGTGAAGGCGAT 667 A GAAAAGCTGTTTGTCTCCC 668 TCC \ATTCCAGCATCACTGTGGAGAAAAGCTGTTTGT DI02 NM 013989 669 CTCCTTTCACGAG 670 AGGAAGTCAGCCACT 671 A CTCTTCCACCAGTTTGCGG 672 CTC< :tttcacgagccagctgccagccttccgcaaact DLC1 NM 006094 673 GATTCAGACGAGG 674 CACCTCTTGCTGTCCC 675 A AAGTCCATTTGCCACTGAT 676 GAT rCAGACGAGGATGAGCCTTGTGCCATCAGTGGC DLGAP1 NM 004746 677 CTGCTGAGCCCAG 678 AGCCTGGAAGGAGTT 679 C GCAGACCACCCATACTACA 680 CTG< 3TGAGCCCAGTGGAGCACCACCCCGCAGACCAC DLL4 NM 019074 681 CACGGAGGT AT AA 682 AGAAGGAAGGTCCAG 683 C TACCTGGACATCCCTGCTC 684 CAC 3GAGGTATAAGGCAGGAGCCTACCTGGACATCC DNM3 NM 015569 685 CTTTCCCACCCGG 686 AAGGACCTTCTGCAG 687 C ATATCGCTGACCGAATGGG 688 cm rCCCACCCGGCTTACAGACATATCGCTGACCGAA DPP4 NM 001935 689 GTCCTGGGATCGG 690 GTACTCCCACCGGGA 691 C GGCTATTCCACACTTGAAC 692 GTCI 3TGGGATCGGGAAGTGGCGTGTTCAAGTGTGGA DPT NM 001937 693 CACCTAGAAGCCT 694 CAGTAGCTCCCCAGG 695 T TCCTAGGAAGGCTGGCAGA 696 CAC CTAGAAGCCTGCCCACGATTCCTAGGAAGGCTG DUSP1 NM 004417 697 AGACATCAGCTCC 698 GACAAACACCCTTCC 699 C GAGGCCATTGACTTCATAG 700 AGA CATCAGCTCCTGGTTCAACGAGGCCATTGACTTC DUSP6 NM 001946 701 CATGCAGGGACTG 702 TGCTCCTACCCTATCA 703 T CTACCCTATGCGCCTGGAA 704 CAT< 3CAGGGACTGGGATTCGAGGACTTCCAGGCGCA DVL1 NM_004421 705 TCTGTCCCACCTG 706 TCAGACTGTTGCCGG 707 C TTGGAGCAGCCTGCACCTT 708 TCT( 3TCCCACCTGCTGCTGCCCCTTGGAGCAGCCTGC DYNLL1 NM 0010374 94 709 GCCGCCTACCTCA CAGAC 710 GCCTGACTCCAGCTC TCCT 711 A A CCCACGTCAGTGAGTGCTC CAA 712 GCC GTGi GCCTACCTCACAGACTTGTGAGCACTCACTGAC GGTAGCGCCCAGGGCCTGCGGGGCGCAGGAGAG EBNA1B P2 NM 006824 713 TGCGGCGAGATGG ACACT 714 GTGACAAGGGATTCA TCGGATT 715 C C CCGCTCTCGGATTCGGAGT G 716 TGC< GAG 3GCGAGATGGACACTCCCCCGCTCTCGGATTCG TCGGAATCCGATGAATCCCTTGTCAC ECE1 NM 001397 717 ACCTTGGGATCTG 718 GGACCAGGACCTCCA 719 T CCACTCTCGATACCCTGCA 720 ACC rTGGGATCTGCCTCCAAGCTGGTGCAGGGTATC EDN1 NM 001955 721 TGCCACCTGGACA 722 TGGACCTAGGGCTTC 723 C ACTCCCGAGCACGTTGTTC 724 TGC< 3ACCTGGACATCATTTGGGTCAACACTCCCGAGC EDNRA NM 001957 725 TTTCCTCAAATTTG 726 TTACACATCCAACCA 727 C CTTTGCCTCAGGGCATCCT 728 TTT( XTCAAATTTGCCTCAAGATGGAAACCCTTTGCC EFNB2 NM 004093 729 TGACATTATCATC CCGCTAAGGA 730 GTAGTCCCCGCTGAC CTTCTC 731 C C GGACAGCGTCTTCTGCCCT ACT 732 TGA CTO CATTATCATCCCGCTAAGGACTGCGGACAGCGT :tgccctcactacgagaaggtcagcggggacta EOF NM_001963 733 CTTTGCCTTGCTCT GTCACAGT 734 AAATACCTGACACCC TTATGACAAATT 735 A T GAGTTTAACAGCCCTGCTC GGCTGACTT 736 cm AGO rGCCTTGCTCTGTCACAGTGAAGTCAGCCAGAGC GCTGTTAAACTCTGTGAAATTTGTCATAAGGGTG EGR1 NM_001964 737 GTCCCCGCTGCAG ATCTCT 738 CTCCAGCTTAGGGTA GTTGTCCAT 739 C C GGATCCTTTCCTCACTCGC CA 740 GTCI CCT< 3CCGCTGCAGATCTCTGACCCGTTCGGATCCTTT :actcgcccaccatggacaactaccctaagctgg EGR3 NM_004430 741 CCATGTGGATGAA TGAGGTG 742 TGCCTGAGAAGAGGT GAGGT 743 A C CCCAGTCTCACCTTCTCCC ACC 744 CCA GTC TGTGGATGAATGAGGTGTCTCCTTTCCATACCCA rCACCTTCTCCCCACCCTACCTCACCTCTTCTCA EIF2C2 NM_012154 745 GCACTGTGGGCAG 746 ATGTTTGGTGACTGG 747 C GGGTCACATTGCAGACACG 748 GCA CTGTGGGCAGATGAAGAGGAAGTACCGCGTCTG EIF2S3 NM_001415 749 CTGCCTCCCTGATT 750 GGTGGCAAGTGCCTG 751 T CTCGTGCTTCAGCCTCCCA 752 CTG< 3CTCCCTGATTCAAGTGATTCTCGTGCTTCAGCC EIF3H NM_003756 753 CTCATTGCAGGCC AGATAAA 754 GCCATGAAGAGCTTG CCTA 755 C T AGAACATCAAGGAGTTCAC GCCCA 756 CTC AI'C, \TT GC AGGC'CAGAT AAACACTT ACT GC'CAGAAC \AGGAGTTCACTGCCCAAAACTTAGGCAAGCTC EIF4E NM_001968 757 GATCTAAGATGGC GACTGTCGAA 758 TTAGATTCCGTTTTCT CCTCTTCTG 759 A C CCACCCCTACTCCTAATCC CCGACT 760 GAT CTA( 3TAAGATGGCGACTGTCGAACCGGAAACCACCC 3TCCTAATCCCCCGACTACAGAAGAGGAGAAAA EIF5 NM 001969 761 GAATTGGTCTCCA 762 TCCAGGTATATGGCT 763 C CACTTGCACCCGAATCTTG 764 GAA TTGGTCTCCAGCTGCCTTTGATCAAGATTCGGGT ET.K4 NM 001973 765 GATGTGGAGAATG 766 AGTCATTGCGGCTAG 767 A TAAACCACCTCAGCCTGGT 768 GAT 3TGGAGAATGGAGGGAAAGATAAACCACCTCAG ENPP2 NM 006209 769 CTCCTGCGCACTA 770 TCCCTGGATAATTGG 771 T AACTTCCTCTGGCATGGTT 772 CTC< 3TGCGCACTAATACCTTCAGGCCAACCATGCCAG ENY2 NM 020189 773 CCTCAAAGAGTTG 774 CCTCTTTACAGTGTGC 775 C TGATCCTTCCAGCCACATT 776 CCT< 3AAAGAGTTGCTGAGAGCTAAATTAATTGAATGT EPHA2 NM 004431 777 CGCCTGTTCACCA 778 GTGGCGTGCCTCGAA 779 T GCGCCCGATGAGATCACCG 780 CGC 3T GTT CACCAAGAITGACACCATT GCGCCCGAT G 126 2015227398 15 Sep 2015

Offlclul wm hoi. VctWsloft Nuiiihcrs forward Primer KroUeflcr: i V) Reverse Primer Swtumrr; ιιβ Pr»be Sequence: si:q IB WO Ampllcon Sequence: EPHA3 NM_005233 781 CAGTAGCCTCAAG 782 TTCGTCCCATATCCAG 783 TATTCCAAATCCGAGCCCGA 784 CAGTAGCCTCAAGCCTGACACTATATACGTATTCCAA EPHB2 NM_004442 785 CAACCAGGCAGCT 786 GTAATGCTGTCCACG 787 CACCTGATGCATGATGGACA 788 CAACCAGGCAGCTCCATCGGCAGTGTCCATCATGCA EPHB4 NM_004444 789 TGAACGGGGTATC CTCCTTA 790 AGGTACCTCTCGGTC AGTGG 791 CGTCCCATTTGAGCCTGTCA ATGT 792 TGAACGGGGTATCCTCCTTAGCCACGGGGCCCGTCC CATTTGAGCCTGTCAATGTCACCACTGACCGAGAGGT ERBB2 NM_004448 793 CGGTGTGAGAAGT 794 CCTCTCGCAAGTGCT 795 CCAGACCATAGCACACTCGG 796 CGGTGTGAGAAGTGCAGCAAGCCCTGTGCCCGAGTG ERBB3 NM_001982 797 CGGTTATGTCATG CCAGATACAC 798 GAACTGAGACCCACT GAAGAAAGG 799 CCTCAAAGGTACTCCCTCCT CCCGG 800 CGGTTATGTCATGCCAGATACACACCTCAAAGGTACT CCCTCCTCCCGGGAAGGCACCCTTTCTTCAGTGGGTC ERBB4 NM_005235 801 TGGCTCTTAATCA GTTTCGTTACCT 802 CAAGGCATATCGATC CTCATAAAGT 803 TGTCCCACGAATAATGCGTA AATTCTCCAG 804 TGGCTCTTAATCAGTTTCGTTACCTGCCTCTGGAGAA TTTACGCATTATTCGTGGGACAAAACTTTATGAGGAT ERCC1 NM_001983 805 GTCCAGGTGGATG 806 CGGCCAGGATACACA 807 CAGCAGGCCCTCAAGGAGCT 808 GTCCAGGTGGATGTGAAAGATCCCCAGCAGGCCCTC EREG NM_001432 809 TGCTAGGGTAAAC 810 TGGAGACAAGTCCTG 811 TAAGCCATGGCTGACCTCTG 812 TGCTAGGGTAAACGAAGGCATAATAAGCCATGGCTG ERG NM_004449 813 CCAACACTAGGCT 814 CCTCCGCCAGGTCTTT 815 AGCCATATGCCTTCTCATCT 816 CCAACACTAGGCTCCCCACCAGCCATATGCCTTCTCA ESR1 NM_000125 817 CGTGGTGCCCCTC 818 GGCTAGTGGGCGCAT 819 CTGGAGATGCTGGACGCCC 820 CGTGGTGCCCCTCTATGACCTGCTGCTGGAGATGCTG ESR2 NM_001437 821 TGGTCCATCGCCA GTTATCA 822 TGTTCTAGCGATCTTG CTTCACA 823 ATCTGTATGCGGAACCTCAA AAGAGTCCCT 824 TGGTCCATCGCCAGTTATCACATCTGTATGCGGAACC TCAAAAGAGTCCCTGGTGTGAAGCAAGATCGCTAGA ETV1 NM 004956 825 TCAAACAAGAGCC 826 AACTGCCAGAGCTGA 827 ATCGGGAAGGACCCACATAC 828 TCAAACAAGAGCCAGGAATGTATCGGGAAGGACCCA ETV4 NM 001986 829 TCCAGTGCCTATG 830 ACTGTCCAAGGGCAC 831 CAGACAAATCGCCATCAAGT 832 TCCAGTGCCTATGACCCCCCCAGACAAATCGCCATCA EZH2 NM_004456 833 TGGAAACAGCGAA GGATACA 834 CACCGAACACTCCCT AGTCC 835 TCCTGACTTCTGTGAGCTCA TTGCG 836 TGGAAACAGCGAAGGATACAGCCTGTGCACATCCTG ACTTCTGTGAGCTCATTGCGCGGGACTAGGGAGTGTT F2R NM_001992 837 AAGGAGCAAACCA 838 GCAGGGTTTCATTGA 839 CCCGGGCTCAACATCACTAC 840 AAGGAGCAAACCATCCAGGTGCCCGGGCTCAACATC FAAH NM 001441 841 GACAGCGTAGTGG TGCATGT 842 AGCTGAACATGGACT GTGGA 843 TGCCCTTCGTGCACACCAAT G 844 GACAGCGTAGTGGTGCATGTGCTGAAGCTGCAGGGT GCCGTGCCCTTCGTGCACACCAATGTTCCACAGTCCA FABP5 NM 001444 845 GCTGATGGCAGAA AAACTCA 846 CTTTCCTTCCCATCCC ACT 847 CCTGATGCTGAACCAATGCA CCAT 848 GCTGATGGCAGAAAAACTCAGACTGTCTGCAACTTT ACAGATGGTGCATTGGTTCAGCATCAGGAGTGGGAT FADD NM_003824 849 GTTTTCGCGAGAT 850 CTCCGGTGCCTGATTC 851 AACGCGCTCTTGTCGATTTC 852 GTTTTCGCGAGATAACGGTCGAAAACGCGCTCTTGTC FAM107 NM_007177 853 AAGTCAGGGAAAA 854 GCTGGCCCTACAGCT 855 AATTGCCACACTGACCAGCG 856 AAGTCAGGGAAAACCTGCGGAGAATTGCCACACTGA FAM13C NM_198215 857 ATCTTCAAAGCGG 858 GCTGGATACCACATG 859 TCCTGACTTTCTCCGTGGCT 860 ATCTTCAAAGCGGAGAGCGGGAGGAGCCACGGAGAA FAM171 B NM_177454 861 CCAGGAAGGAAAA GCACTGT 862 GTGGTCTGCCCCTTCT TTTA 863 TGAAGATTTTGAAGCTAATA CATCCCCCAC 864 CCAGGAAGGAAAAGCACTGTTGAAGATTTTGAAGCT AATACATCCCCCACTAAAAGAAGGGGCAGACCAC FAM49B NM 016623 865 AGATGCAGAAGGC 866 GCTGGATTGCCTCTC 867 TGGCCAGCTCCTCTGTATGA 868 AGATGCAGAAGGCATCTTGGAGGACTTGCAGTCATA FAM73A NM_198549 869 TGAGAAGGTGCGC TATTCAA 870 GGCCATTAAAAGCTC AGTGC 871 AAGACCTCATGCAGTTACTC ATTCGCC 872 TGAGAAGGTGCGCTATTCAAGTACAGAGACTTTAGC TGAAGACCTCATGCAGTTACTCATTCGCCGCACTGAG FAP NM_004460 873 GTTGGCTCACGTG 874 GACAGGACCGAAACA 875 AGCCACTGCAAACATACTCG 876 GTTGGCTCACGTGGGTTACTGATGAACGAGTATGTTT FAS NM_000043 877 GGATTGCTCAACA ACCATGCT 878 GGCATTAACACTTTTG GACGATAA 879 TCTGGACCCTCCTACCTCTG GTTCTTACGT 880 GGATTGCTCAACAACCATGCTGGGCATCTGGACCCT CCTACCTCTGGTTCTTACGTCTGTTGCTAGATTATCG FASLG NM_000639 881 GCACTTTGGGATT CTTTCCATTAT 882 GCATGTAAGAAGACC CTCACTGAA 883 ACAACATTCTCGGTGCCTGT AACAAAGAA 884 GCACTTTGGGATTCTTTCCATTATGATTCTTTGTTAC AGGCACCGAGAATGTTGTATTCAGTGAGGGTCTTCTT FASN NM_004104 885 GCCTCTTCCTGTTC 886 GCTTTGCCCGGTAGC 887 TCGCCCACCTACGTACTGGC 888 GCCTCTTCCTGTTCGACGGCTCGCCCACCTACGTACT FCGR3A NM_000569 889 GTCTCCAGTGGAA 890 AGGAATGCAGCTACT 891 CCCATGATCTTCAAGCAGGG 892 GTCTCCAGTGGAAGGGAAAAGCCCATGATCTTCAAG FGF10 NM_004465 893 TCTTCCGTCCCTGT 894 AGAGTTGGTGGCCTC 895 ACACCATGTCCTGACCAAGG 896 TCTTCCGTCCCTGTCACCTGCCAAGCCCTTGGTCAGG FGF17 NM_003867 897 GGTGGCTGTCCTC 898 TCTAGCCAGGAGGAG 899 TTCTCGGATCTCCCTCAGTC 900 GGTGGCTGTCCTCAAAATCTGCTTCTCGGATCTCCCT FGF5 NM_004464 901 GCATCGGTTTCCA 902 AACATATTGGCTTCGT 903 CCATTGACTTTGCCATCCGG 904 GCATCGGTTTCCATCTGCAGATCTACCCGGATGGCAA FGF6 NM_020996 905 GGGCCATTAATTC TGACCAC 906 CCCGGGACATAGTGA TGAA 907 CATCCACCTTGCCTCTCAGG CAC 908 GGGCCATTAATTCTGACCACGTGCCTGAGAGGCAAG GTGGATGGCCCTGGGACAGAAACTGTTCATCACTAT FGF7 NM_002009 909 CCAGAGCAAATGG CTACAAA 910 TCCCCTCCTTCCATGT AATC 911 CAGCCCTGAGCGACACACAA GAAG 912 CCAGAGCAAATGGCTACAAATGTGAACTGTTCCAGC CCTGAGCGACACACAAGAAGTTATGATTACATGGAA FGFR2 NM_000141 913 GAGGGACTGTTGG CATGCA 914 GAGTGAGAATTCGAT CCAAGTCTTC 915 TCCCAGAGACCAACGTTCAA GCAGTTG 916 GAGGGACTGTTGGCATGCAGTGCCCTCCCAGAGACC AACGTTCAAGCAGTTGGTAGAAGACTTGGATCGAAT FGFR4 NM 002011 917 CTGGCTTAAGGAT GGACAGG 918 ACGAGACTCCAGTGC TGATG 919 CCTTTCATGGGGAGAACCGC ATT 920 CTGGCTTAAGGATGGACAGGCCTTTCATGGGGAGAA CCGCATTGGAGGCATTCGGCTGCGCCATCAGCACTG 2015227398 15 Sep 2015 127

«HUM 'Mmimf: f mm \o t .ir« afd Primer XtifUt-liOti 1 I.QT!> SO U rttrw Primw Swqufruwr: s P 111 Γι-ohi >ig Hi SO \m)>Ui>.n vψκιιΐΛ·: FKBP5 NM 004117 921 CCCACAGTAGAGG 922 GGTTCTGGCTTTCACG 923 TCTCCCCAGTTCCACAGCAG 924 XCACAGTAGAGGGGTCTCATGTCTCCCCAGTTCCAC FLNA NM 001456 925 GAACCTGCGGTGG 926 GAAGACACCCTGGCC 927 TACCAGGCCCATAGCACTGG 928 jAACCTGCGGTGGACACTTCCGGTGTCCAGTGCTAT FLNC NM 001458 929 CAGGACAATGGTG 930 TGATGGTGTACTCGC 931 ATGTGCTGTCAGCTACCTGC 932 :aggacaatggtgatggctcatgtgctgtcagctac FLT1 NM 002019 933 GGCTCCTGAATCT 934 TCCCACAGCAATACT 935 CTACAGCACCAAGAGCGAC 936 jGCTCCTGAATCTATCTTTGACAAAATCTACAGCACC FLT4 NM 002020 937 ACCAAGAAGCTGA 938 CCTGGAAGCTGTAGC 939 AGCCCGCTGACCATGGAAGA 940 i xcaagaagctgaggacctgtggctgagcccgctga FN1 NM 002026 941 GGAAGTGACAGAC 942 ACACGGTAGCCGGTC 943 ACTCTCAGGCGGTGTCCACA 944 jGAAGTGACAGACGTGAAGGTCACCATCATGTGGAC FOS NM 005252 945 CGAGCCCTTTGAT GACTTCCT 946 GGAGCGGGCTGTCTC AGA 947 TCCCAGCATCATCCAGGCCC AG 948 ( i :gagccctttgatgacttcctgttcccagcatcatcc ^GGCCCAGTGGCTCTGAGACAGCCCGCTCC FOXOl NM 002015 949 GTAAGCACCATGC 950 GGGGCAGAGGCACTT 951 TATGAACCGCCTGACCCAAG 952 3TAAGCACCATGCCCCACACCTCGGGTATGAACCGC FOXP3 NM 014009 953 CTGTTTGCTGTCCG 954 GTGGAGGAACTCTGG 955 TGTTTCCATGGCTACCCCAC 956 :tgtttgctgtccggaggcacctgtggggtagccat FOXQ1 NM 033260 957 TGTTTTTGTCGCAA 958 TGGAAAGGTTCCCTG 959 TGATTTATGTCCCTTCCCTC 960 rGTTTTTGTCGCAACTTCCATTGATTTATGTCCCTTCC FSD1 NM 024333 961 AGGCCTCCTGTCC 962 TGTGTGAACCTGGTC 963 CGCACCAAACAAGTGCTGCA 964 i VGGCCTCCTGTCCrrCTACAATGCCCGCACCAAACAA FYN NM 002037 965 GAAGCGCAGATCA 966 CTCCTCAGACACCAC 967 CTGAAGCACGACAAGCTGGT 968 jAAGCGCAGATCATGAAGAAGCTGAAGCACGACAAG G6PD NM 000402 969 AATCTGCCTGTGG 970 CGAGATGTTGCTGGT 971 CCAGCCTCAGTGCCACTTGA 972 i YYTCTGCCTGTGGCCTTGCCCGCCAGCCTCAGTGCCA GABRG2 NM_198904 973 CCACTGTCCTGAC AATGACC 974 GAGATCCATCGCTGT GACAT 975 CTCAGCACCATTGCCCGGAA AT 976 XACTGTCCTGACAATGACCACCCTCAGCACCATTGC XGGAAATCGCTCCCCAAGGTCTCCTATGTCACAGC GADD45 NM 001924 977 GTGCTGGTGACGA 978 CCCGGCAAAAACAAA 979 TTCATCTCAATGGAAGGATC 980 jTGCTGGTGACGAATCCACATTCATCTCAATGGAAG GADD45 NM 015675 981 ACCCTCGACAAGA 982 TGGGAGTTCATGGGT 983 TGGGAGTTCATGGGTACAGA 984 l UXCTCGACAAGACCACACTTTGGGACTTGGGAGCT GDF15 NM 004864 985 CGCTCCAGACCTA 986 ACAGTGGAAGGACCA 987 TGTTAGCCAAAGACTGCCAC 988 XrCTCCAGACCTATGATGACTTGTTAGCCAAAGACTG GHR NM 000163 989 CCACCTCCCACAG 990 GGTGCGTGCCTGTAG 991 CGTGCCTCAGCCTCCTGAGT 992 XACCTCCCACAGGTTCAGGCGATTCCCGTGCCTCAG GNPTAB NM 024312 993 GGATTCACATCGC 994 GTTCTTGCATAACAAT 995 CCCTGCTCACATGCCTCACA 996 3GATTCACATCGCGGAAAGTCCCTGCTCACATGCCTC GNRH1 NM_000825 997 AAGGGCTAAATCC AGGTGTG 998 CTGGATCTCTGTGGCT GOT 999 TCCTGTCCTTCACTGTCCTT GCCA 1000 1 \AGGGCTAAATCCAGGTGTGACGGTATCTAATGATG rCCTGTCCTTCACTGTCCTTGCCATCACCAGCCACAG GPM6B NM_0010019 94 1001 ATGTGCTTGGAGT GGCCT 1002 TGTAGAACATAAACA CGGGCA 1003 CGCTGAGAAACCAAACACAC CCAG 1004 l ^TGTGCTTGGAGTGGCCTGGCTGGGTGTGTTTGGTTT :tcagcggtgcccgtgtttatgttctaca GPNMB NM 0010053 40 1005 CAGCCTCGCCTTT AAGGAT 1006 TGACAAATATGGCCA AGCAG 1007 CAAACAGTGCCCTGATCTCC GTTG 1008 :agcctcgcctttaaggatggcaaacagtgccctga rCTCCGTTGGCTGCTTGGCCATATTTGTCA GPR68 NM 003485 1009 CAAGGACCAGATC 1010 GGT AGGGC AGGAAGC 1011 CTCAGCACCGTGGTCATCTT 1012 :AAGGACCAGATCCAGCGGCTGGTGCTCAGCACCGT GPS1 NM 004127 1013 AGT ACAAGCAGGC 1014 GCAGCTCAGGGAAGT 1015 CCTCCTGCTGGCTTCCTTTG 1016 ^GTACAAGCAGGCTGCCAAGTGCCTCCTGCTGGCTT GRB7 NM_005310 1017 CCATCTGCATCCA 1018 GGCCACCAGGGTATT 1019 CTCCCCACCCTTGAGAAGTG 1020 XATCTGCATCCATCTTGTTTGGGCTCCCCACCCTTG GREM1 NM_013372 1021 GTGTGGGCAAGGA 1022 GACCTGATTTGGCCT 1023 TCCACCCTCCCTTTCTCACT 1024 3TGTGGGCAAGGACAAGCAGGATAGTGGAGTGAGAA GSK3B NM_002093 1025 GACAAGGACGGCA 1026 TTGTGGCCTGTCTGG 1027 CCAGGAGTTGCCACCACTGT 1028 3ACAAGGACGGCAGCAAGGTGACAACAGTGGTGGCA GSN NM_000177 1029 CTTCTGCTAAGCG GTACATCGA 1030 GGCTCAAAGCCTTGC TTCAC 1031 ACCCAGCCAATCGGGATCGG C 1032 ( i :TTCTGCTAAGCGGTACATCGAGACGGACCCAGCCA VTCGGGATCGGCGGACGCCCATCACCGTGGl'GAAGC GSTM1 NM_000561 1033 AAGCTATGAGGAA AAGAAGTACACGA 1034 GGCCCAGCTTGAATT TTTCA 1035 TCAGCCACTGGCTTCTGTCA TAATCAGGAG 1036 ^AGCTATGAGGAAAAGAAGTACACGATGGGGGACGC rCCTGATTATGACAGAAGCCAGTGGCTGAATGAAAA GSTM2 NM_000848 1037 CTGCAGGCACTCC 1038 CCAAGAAACCATGGC 1039 CTGAAGCTCTACTCACAGTT 1040 :TGCAGGCACTCCCTGAAATGCTGAAGCTCTACTCAC HDAC1 NM_004964 1041 CAAGT ACCACAGC GATGACTACATTA 1042 GCTTGCTGTACTCCG ACATGTT 1043 TTCTTGCGCTCCATCCGTCC AGA 1044 :AAGTACCACAGCGATGACTACATTAAATTCTTGCGC rCCATCCGTCCAGATAACATGTCGGAGTACAGCAAG HDAC9 NM_178423 1045 AACCAGGCAGTCA CCTTGAG 1046 CTCTGTCTTCCTGCAT CGC 1047 CCCCCTGAAGCTCTTCCTCT GCTT 1048 l ^ACCAGGCAGTCACCTTGAGGAAGCAGAGGAAGAGC rTCAGGGGGACCAGGCGATGCAGGAAGACAGAG HGD NM_000187 1049 CTCAGGTCTGCCC 1050 TTATTGGTGCTCCGTG 1051 CTGAGCAGCTCTCAGGATCG 1052 7TCAGGTCTGCCCCTACAATCTCTATGCTGAGCAGCT HIP1 NM_005338 1053 CTCAGAGCCCCAC 1054 GGGTTTCCCTGCCAT 1055 CGACTCACTGACCGAGGCCT 1056 :TCAGAGCCCCACCTGAGCCTGCCGACTCACTGACC HIRIP3 NM 003609 1057 GGATGAGGAAAAG 1058 TCCCTAGCTGACTTTC 1059 CCATTGCTCCTGGTTCTGGG 1060 jGATGAGGAAAAGGGGGATTGGAAACCCAGAACCAG HK1 NM 000188 1061 TACGCACAGAGGC 1062 GAGAGAAGTGCTGGA 1063 TAAGAGTCCGGGATCCCCAG 1064 rACGCACAGAGGCAAGCAGCTAAGAGTCCGGGATCC HLA-G NM 002127 1065 CCATCCCCATCAT 1066 CCGCAGCTCCAGTGA 1067 CTGCAAGGACAACCAGGCC 1068 XTGCGCGGCTACTACAACCAGAGCGAGGCCAGTTC HLF NM 002126 1069 CACCCTGCAGGTG 1070 GGTACCTAGGAGCAG 1071 TAAGTGATCTGCCCTCCAGG 1072 7ACCCTGCAGGTGTCTGAGACTAAGTGATCTGCCCTC HNF1B NM 000458 1073 TCCCAGCATCTCA 1074 CGT ACCAGGTGT ACA 1075 CCCCTATGAAGACCCAGAAG 1076 rCCCAGCATCTCAACAAGGGCACCCCTATGAAGACC HPS1 NM 000195 1077 GCGGAAGCTGTAT 1078 TTCGGATAAGATGAC 1079 CAGTCACCAGCCCAAAGTGC 1080 jCGGAAGCTGTATGTGCTCAAGTACCTGTTTGAAGT HRAS NM 005343 1081 GGACGAATACGAC 1082 GCACGTCTCCCCATC 1083 ACCACCTGCTTCCGGTAGGA 1084 iGACGAAT ACGACCCCACT AT AG AGO ATT CCT ACCG 128 2015227398 15 Sep 2015

Of1u-i.il N\mM: NihiiImtt \o 1 urn.inj PHttKT Ν(·([ΙΗΊΗ(·: smtf* V) &amp;<Ailr><· I’mrn-r Stoui-ih*. SI O ίΐβ I'rohfrSfttiU'nctt IB NO Am plhw* Λ<·φ«·#κ·<·: HSD17B1 0 NM 004493 1085 CCAGCGAGTTCTT GATGTGA 1086 ATCTCACCAGCCACC AGG 1087 TCATGGGCACCTTCAATGTG ATCC 1088 CCACCAGACAAGACCGATTCGCTGGCCTCCATTTCTT CAACCCAGTGCCTGTCATGAAACTTGTGG HSD17B2 NM 002153 1089 GCTTTCCAAGTGG 1090 TGCCTGCGATATTTGT 1091 AGTTGCTTCCATCCAACCTG 1092 GCTTTCCAAGTGGGGAATTAAAGTTGCTTCCATCCAA HSD17B3 NM 000197 1093 GGGACGTCCTGGA ACAGT 1094 TGGAGAATCTCACGC ACTTC 1095 CTTCATCCTCACAGGGCTGC TGGT 1096 GGGACGTCCTGGAACAGTTCTTCATCCTCACAGGGCT GCTGGTGTGCCTGGCCTGCCTGGCGAAGTGCGTGAG HSD17B4 NM 000414 1097 CGGGAAGCTTCAG 1098 ACCTCAGGCCCAAT A 1099 AGGCGGCGTCCTATTTCCTC 1100 CGGGAAGCTTCAGAGTACCTTTGTATTTGAGGAAAT HSD3B2 NM 000198 1101 GCCTTCCTTTAACC 1102 GGAGTAAATTGGGCT 1103 ACTTCCAGCAGGAAGCCAAT 1104 GCCTTCCTTTAACCCTGATGTACTGGATTGGCTTCCT HSP90A B1 NM 007355 1105 GCATTGTGACCAG CACCTAC 1106 GAAGTGCCTGGGCTT TCAT 1107 ATCCGCTCCATATTGGCTGT CCAG 1108 GCATTGTGACCAGCACCTACGGCTGGACAGCCAATA TGGAGCGGATCATGAAAGCCCAGGCACTTC HSPA5 NM 005347 1109 GGCT AGT AGAACT GGATCCCAACA 1110 GGTCTGCCCAAATGC TTTTC 1111 TAATTAGACCTAGGCCTCAG CTGCACTGCC 1112 GGCTAGTAGAACTGGATCCCAACACCAAACTCTTAA TTAGACCTAGGCCTCAGCTGCACTGCCCGAAAAGCA HSPA8 NM 006597 1113 CCTCCCTCTGGTG GTGCTT 1114 GCTACATCTACACTTG GTTGGCTTAA 1115 CTCAGGGCCCACCATTGAAG AGGTTG 1116 CCTCCCTCTGGTGGTGCTTCCTCAGGGCCCACCATTG AAGAGGTTGATTAAGCCAACCAAGTGTAGATGTAGC HSPB1 NM 001540 1117 CCGACTGGAGGAG CAT AAA 1118 ATGCTGGCTGACTCT GCTC 1119 CGCACTTTTCTGAGCAGACG TCCA 1120 CCGACTGGAGGAGCATAAAAGCGCAGCCGAGCCCAG CGCCCCGCACTTTTCTGAGCAGACGTCCAGAGCAGA HSPB2 NM 001541 1121 CACCACTCCAGAG 1122 TGGGACCAAACCAT A 1123 CACCTTTCCCTTCCCCCAAG 1124 CACCACTCCAGAGGTAGCAGCATCCTTGGGGGAAGG HSPE1 NM 002157 1125 GCAAGCAACAGTA GTCGCTG 1126 CCAACTTTCACGCT A ACTGGT 1127 TCTCCACCCTTTCCTTTAGA ACCCG 1128 GCAAGCAACAGTAGTCGCTGTTGGATCGGGTTCTAA AGGAAAGGGTGGAGAGATTCAACCAGTTAGCGTGAA HSPG2 NM 005529 1129 GAGTACGTGTGCC 1130 CTCAATGGTGACCAG 1131 CAGCTCCGTGCCTCTAGAGG 1132 GAGTACGTGTGCCGAGTGTTGGGCAGCTCCGTGCCT ICAM1 NM 000201 1133 GCAGACAGTGACC ATCTACAGCTT 1134 CTTCTGAGACCTCTG GCTTCGT 1135 CCGGCGCCCAACGTGATTCT 1136 GCAGACAGTGACCATCTACAGCTTTCCGGCGCCCAA CGTGATTCTGACGAAGCCAGAGGTCTCAGAAG IER3 NM 003897 1137 GTACCTGGTGCGC GAGAG 1138 GCGTCTCCGCTGTAG TGTT 1139 TCAAGTTGCCTCGGAAGTCC CAGT 1140 GTACCTGGTGCGCGAGAGCGTATCCCCAACTGGGAC TTCCGAGGCAACTTGAACTCAGAACACTACAGCGGA IFI30 NM 006332 1141 ATCCCATGAAGCC 1142 GCACCATTCTTAGTG 1143 AAAATTCCACCCCATGATCA 1144 ATCCCATGAAGCCCAGATACACAAAATTCCACCCCA MT1 NM_001548 1145 TGACAACCAAGCA 1146 CAGTCTGCCCATGTG 1147 AAGTTGCCCCAGGTCACCAG 1148 TGACAACCAAGCAAATGTGAGGAGTCTGGTGACCTG IFNG NM_000619 1149 GCT AAAACAGGGA AGCGAAA 1150 CAACCATTACTGGGA TGCTC 1151 TCGACCTCGAAACAGCATCT GACTCC 1152 GCTAAAACAGGGAAGCGAAAAAGGAGTCAGATGCTG TTTCGAGGTCGAAGAGCATCCCAGTAATGGTTG IGF1 NM 000618 1153 TCCGGAGCTGTGA 1154 CGGACAGAGCGAGCT 1155 TGTATTGCGCACCCCTCAAG 1156 TCCGGAGCTGTGATCTAAGGAGGCTGGAGATGTATT IGF1R NM_000875 1157 GCATGGTAGCCGA AGATTTCA 1158 TTTCCGGTAATAGTCT GTCTCATAGATATC 1159 CGCGTCATACCAAAATCTCC GATTTTGA 1160 GCATGGTAGCCGAAGATTTCACAGTCAAAATCGGAG ATTTTGGTATGACGCGAGATATCTATGAGACAGACTA IGF2 NM_000612 1161 CCGTGCTTCCGGA 1162 TGGACTGCTTCCAGG 1163 TACCCCGTGGGCAAGTTCTT 1164 CCGTGCTTCCGGACAACTTCCCCAGATACCCCGTGGG IGFBP2 NM_000597 1165 GTGGACAGCACCA 1166 CCTTCATACCCGACTT 1167 CTTCCGGCCAGCACTGCCTC 1168 GTGGACAGCACCATGAACATGTTGGGCGGGGGAGGC IGFBP3 NM_000598 1169 ACATCCCAACGCA 1170 CCACGCCCTTGTTTCA 1171 ACACCACAGAAGGCTGTGA 1172 ACATCCCAACGCATGCTCCTGGAGCTCACAGCCTTCT IGFBP5 NM_000599 1173 TGGACAAGTACGG 1174 CGAAGGTGTGGCACT 1175 CCCGTCAACGTACTCCATGC 1176 TGGACAAGTACGGGATGAAGCTGCCAGGCATGGAGT IGFBP6 NM_002178 1177 TGAACCGCAGAGA CCAACAG 1178 GTCTTGGACACCCGC AGAAT 1179 ATCCAGGCACCTCTACCACG CCCTC 1180 TGAACCGCAGAGACCAACAGAGGAATCCAGGCACCT CTACCACGCCCTCCCAGCCCAATTCTGCGGGTGTCCA IL10 NM 000572 1181 CTGACCACGCTTT 1182 CCAAGCCCAGAGACA 1183 TTGAGCTGTTTTCCCTGACC 1184 CTGACCACGCTTTCTAGCTGTTGAGCTGTTTTCCCTG IL11 NM 000641 1185 TGGAAGGTTCCAC 1186 TCTTGACCTTGCAGCT 1187 CCTGTGATCAACAGTACCCG 1188 TGGAAGGTTCCACAAGTCACCCTGTGATCAACAGTA IL17A NM 002190 1189 TCAAGCAACACTC 1190 CAGCTCCTTTCTGGGT 1191 TGGCTTCTGTCTGATCAAGG 1192 TCAAGCAACACTCCTAGGGCCTGGCTTCTGTCTGATC ILIA NM 000575 1193 GGTCCTTGGTAGA 1194 GGATGGAGCTTCAGG 1195 TCTCCACCCTGGCCCTGTTA 1196 GGTCCTTGGTAGAGGGCTACTTTACTGTAACAGGGC IL1B NM 000576 1197 AGCTGAGGAAGAT 1198 GGAAAGAAGGTGCTC 1199 TGCCCACAGACCTTCCAGGA 1200 AGCTGAGGAAGATGCTGGTTCCCTGCCCACAGACCT IL2 NM_000586 1201 ACCTCAACTCCTG CCACAAT 1202 CACTGTTTGTGACAA GTGCAAG 1203 TGCAACTCCTGTCTTGCATT GCAC 1204 ACCTCAACTCCTGCCACAATGTACAGGATGCAACTCC TGTCTTGCATTGCACTAAGTCTTGCACTTGTCACAAA IL6 NM_000600 1205 CCTGAACCTTCCA 1206 ACCAGGCAAGTCTCC 1207 CCAGATTGGAAGCATCCATC 1208 CCTGAACCTTCCAAAGATGGCTGAAAAAGATGGATG IL6R NM_000565 1209 CCAGCTTATCTCA 1210 CTGGCGTAGAACCTT 1211 CCTTTGGCTTCACGGAAGAG 1212 CCAGCTTATCTCAGGGGTGTGCGGCCTTTGGCTTCAC IL6ST NM_002184 1213 GGCCTAATGTTCC 1214 AAAATTGTGCCTTGG 1215 CATATTGCCCAGTGGTCACC 1216 GGCCTAATGTTCCAGATCCTTCAAAGAGTCATATTGC IL8 NM_000584 1217 AAGGAACCATCTC ACTGTGTGTAAAC 1218 ATCAGGAAGGCTGCC AAGAG 1219 TGACTTCCAAGCTGGCCGTG GC 1220 AAGGAACCATCTCACTGTGTGTAAACATGACTTCCAA GCTGGCCGTGGCTCTCTTGGCAGCCTTCCTGAT ILF3 NM 004516 1221 GACACGCCAAGTG 1222 CTCAAGACCCGGATC 1223 ACACAAGACTTCAGCCCGTT 1224 GACACGCCAAGTGGTTCCAGGCCAGAGCCAACGGGC 129 2015227398 15 Sep 2015

Of1u-i.il Nwnfwl: An <-«<<»> NihiiImtt <SE.o m NO 1 »i-« .ii-d Prirm·»· Ncifih-ihc: si.g τι» Srttrw Prim» i M.Q Jl»NO i>n>be NetproiBie: Nl <J 11) NO \m))lR»n N<-c|m-nn·: V) Swqufruwr: ILK NM_0010147 94 1225 CTCAGGATTTTCTC GCATCC 1226 1 VGGAGCAGGTGGAGA :tgg 1227 ATGTGCTCCCAGTGCTAGGT GCCT 1228 CTC^ TAG \GGATTTTCTCGCATCCAAATGTGCTCCCAGTGC TTGCCTGCCAGTCTCCACCTGCTCCT IMMT NM 006839 1229 CTGCCTATGCCAG 1230 jCTTTTCTGGCTTCCT 1231 CAACTGCATGGCTCTGAACA 1232 CTG( 3CTATGCCAGACTCAGAGGAATCGAACAGGCTG ING5 NM 032329 1233 CCTACAGCAAGTG 1234 :atctcgtaggtctg 1235 CCAGCTGCACTTTGTCGTCA 1236 CCT^ \CAGCAAGTGCAAGGAATACAGTGACGACAAAG INHBA NM 002192 1237 GTGCCCGAGCCAT 1238 :ggtagtggttgatg 1239 ACGTCCGGGTCCTCACTGTC 1240 GTG CCCGAGCCATATAGCAGGCACGTCCGGGTCCTC INSL4 NM 002195 1241 CTGTCATATTGCCC 1242 :agattccagcagcc 1243 TGAGAAGACATTCACCACCA 1244 CTGr rCATATTGCCCCATGCCTGAGAAGACATTCACCA ITGA1 NM181501 1245 GCTTCTTCTGGAG 1246 XTGTAGATAATGAC 1247 TTGCTGGACAGCCTCGGTAC 1248 GCTr rCTTCTGGAGATGTGCTCTATATTGCTGGACAGC ITGA3 NM 002204 1249 CCATGATCCTCAC 1250 jAAGCTTTGTAGCCG 1251 CACTCCAGACCTCGCTTAGC 1252 CCA rGATCCTCACTCTGCTGGTGGACTATACACTCCA ITGA4 NM 000885 1253 CAACGCTTCAGTG 1254 jTCTGGCCGGGATTC 1255 CGATCCTGCATCTGTAAATC 1256 CAA CGCTTCAGTGATCAATCCCGGGGCGATTTACAG ITGA5 NM 002205 1257 AGGCCAGCCCTAC 1258 jTCTTCTCCACAGTCC 1259 TCTGAGCCTTGTCCTCTATC 1260 AGO CCAGCCCTACATTATCAGAGCAAGAGCCGGATA ITGA6 NM 000210 1261 CAGTGACAAACAG 1262 jTTTAGCCTCATGGG 1263 TCGCCATCTTTTGTGGGATT 1264 CAG TGACAAACAGCCCTTCCAACCCAAGGAATCCCA ITGA7 NM 002206 1265 GATATGATTGGTC GCTGCTTTG 1266 i i ^GAACTTCCATTCCCC VCCAT 1267 CAGCCAGGACCTGGCCATCC G 1268 GAT. TGG \TGATTGGTCGCTGCTTTGTGCTCAGCCAGGACC CCATCCGGGATGAGTTGGATGGTGGGGAATGGA ITGAD NM 005353 1269 GAGCCTGGTGGAT 1270 i VCT GT CAGGAT GCCC 1271 CAACTGAAAGGCCTGACGTT 1272 GAG CCTGGTGGATCCCATCGTCCAACTGAAAGGCCT ITGB3 NM 000212 1273 ACCGGGAGCCCTA CATGAC 1274 XTTAAGCTCTTTCAC rGACTCAATCT 1275 AAATACCTGCAACCGTTACT GCCGTGAC 1276 ACC GTTj TGGGAGCCCTACATGACGAAAATACCTGCAACC X rl'GACGAGATT GAGT C AGT GAAAGAGC ITGB4 NM 000213 1277 CAAGGTGCCCTCA 1278 jCGCACACCTTCATC 1279 CACCAACCTGTACCCGTATT 1280 CAA GGTGCCCTCAGTGGAGCTCACCAACCTGTACCC ITGB5 NM 002213 1281 TCGTGAAAGATGA 1282 jGT gaacatcatgac 1283 TGCTATGTTTCTACAAAACC 1284 TCGr rGAAAGATGACCAGGAGGCTGTGCTATGTTTCTA ITPR1 NM 002222 1285 GAGGAGGTGTGGG 1286 jTAATCCCATGTCCG 1287 CCATCCTAACGGAACGAGCT 1288 GAG GAGGTGTGGGTGTTCCGCTTCCATCCTAACGGA ITPR3 NM 002224 1289 TTGCCATCGTGTC 1290 i VT GGAGCTGGCGT CA 1291 TCCAGGTCTCGGATCTCAGA 1292 TTG< rCATCGTGTCAGTGCCCGTGTCTGAGATCCGAGA ITSN1 NM 003024 1293 TAACTGGGATGCA 1294 :tctgccttaactggc 1295 AGCCCTCTCTCACCGTTCCA 1296 TAA CTGGGATGCATGGGCAGCCCAGCCCTCTCTCAC JAG1 NM 000214 1297 TGGCTTACACTGG 1298 jCAT agctgtgagat 1299 ACTCGATTTCCCAGCCAACC 1300 TGG CTTACACTGGCAATGGTAGTTTCTGTGGTTGGCT JUN NM 002228 1301 GACTGCAAAGATG GAAACGA 1302 rAGCCATAAGGTCCG :tctc 1303 CTATGACGATGCCCTCAACG CCTC 1304 GAC CCT< TGCAAAGATGGAAACGACCTTCTATGACGATGC rAACGCCTCGTTCCTCCCGTCCGAGAGCGGACCT JUNB NM 002229 1305 CTGTCAGCTGCTG 1306 i \GGGGGTGTCCGT AA 1307 CAAGGGACACGCCTTCTGAA 1308 CTGr rCAGCTGCTGCTTGGGGTCAAGGGACACGCCTT KCNN2 NM 021614 1309 TGTGCTATTCATCC 1310 jGGCATAGGAGAAGG 1311 TTATACATTCACATGGACGG 1312 TGT( 3CTATTCATCCCATACCTGGGAATTATACATTCA KCTD12 NM 138444 1313 AGCAGTTACTGGC 1314 rGGAGACCTGAGCAG 1315 ACTCTTAGGCGGCAGCGTCC 1316 AGC AGTTACTGGCAAGAGGGAGAAAGGACGCTGCCG KHDRBS NM 006558 1317 CGGGCAAGAAGAG 1318 :tgtagacgcccttt 1319 CAAGACACAAGGCACCTTCA 1320 CGGi GCAAGAAGAGTGGACTAACTCAAGACACAAGGC KIAA019 NM 014846 1321 CAGACACCAGCTC 1322 l \ AC ATT GT G AGGCGG 1323 TCCCCAGTGTCCAGGCACAG 1324 CAG ACACCAGCTCTGAGGCCAGTTAATCATCCCCAG KIAA024 NM 014734 1325 CCGTGGGACATGG 1326 jAAGCAAGTCCGTCT 1327 TCCGCTAGTGATCCTTTGCA 1328 CCG TGGGACATGGAGTGTTCCTTCCGCTAGTGATCCT KIF4A NM 012310 1329 AGAGCTGGTCTCC 1330 3CTGGTCTTGCTCTGT 1331 CAGGTCAGCAAACTTGAAAG 1332 AGA GCTGGTCTCCTCCAAAATACAGGTCAGCAAACT KIT NM_000222 1333 GAGGCAACTGCTT ATGGCTT AATT A 1334 3GCACTCGGCTTGAG :at 1335 TTACAGCGACAGTCATGGCC GCAT 1336 GAG GCC GCAACTGCTTATGGCTTAATTAAGTCAGATGCG ATGACTGTCGCTGTAAAGATGCTCAAGCCGAGT KLC1 NM_ 182923 1337 AGTGGCTACGGGA 1338 rGAGCCACAGACTGC 1339 CAACACGCAGCAGAAACTG 1340 AGT 3GCTACGGGATGAACTGGCCAACACGCAGCAGA KLF6 NM_001300 1341 CACGAGACCGGCT 1342 3CTCTAGGCAGGTCT 1343 AGTACTCCTCCAGAGACGGC 1344 CAC 3AGACCGGCTACTTCTCGGCGCTGCCGTCTCTGG KLK1 NM_002257 1345 AACACAGCCCAGT TTGTTCA 1346 7CAGGAGGCTCATGT rGAAG 1347 TCAGTGAGAGCTTCCCACAC CCTG 1348 AAC CAC ACAGCCCAGTTTGTTCATGTCAGTGAGAGCTTCC \CCCTGGCTTCAACATGAGCCTCCTGG KLK10 NM 002776 1349 GCCCAGAGGCTCC 1350 :agaggtttgaacag 1351 CCTCTTCCTCCCCAGTCGGC 1352 GCC CAGAGGCTCCATCGTCCATCCTCTTCCTCCCCAG KLK11 NM 006853 1353 CACCCCGGCTTCA 1354 :atcttcaccagcat 1355 CCTCCCCAACAAAGACCACC 1356 CAC 3CCGGCTTCAACAACAGCCTCCCCAACAAAGAC KLK14 NM 022046 1357 CCCCTAAAATGTT 1358 :tcatcctcttggctc 1359 CAGCACTTCAAGTCCTGGCT 1360 CCC< 3TAAAATGTTCCTCCTGCTGACAGCACTTCAAGT KLK2 NM 005551 1361 AGTCTCGGATTGT 1362 rGTACACAGCCACCT 1363 TTGGGAATGCTTCTCACACT 1364 AGT 3TCGGATTGTGGGAGGCTGGGAGTGTGAGAAGC KLK3 NM 001648 1365 CCAAGCTTACCAC 1366 VGGGTGAGGAAGACA 1367 ACCCACATGGTGACACAGCT 1368 CCA \GCTTACCACCTGCACCCGGAGAGCTGTGTCAC KLRK1 NM 007360 1369 TGAGAGCCAGGCT 1370 ^TCCTGGTCCTCTTTG 1371 TGTCTCAAAATGCCAGCCTT 1372 TGA 3AGCCAGGCTTCTTGTATGTCTCAAAATGCCAGC KPNA2 NM 002266 1373 TGATGGTCCAAAT 1374 VAGCTT CACAAG1 I'G 1375 ACTCCTGTTTTCACCACCAT 1376 TGA rGGTCCAAATGAACGAATTGGCATGGTGGTGAA KRT1 NM 006121 1377 TGGACAACAACCG 1378 rATCCTCGTACTGGG 1379 CCTCAGCAATGATGCTGTCC 1380 TGG <\C AACAACCGC AGT CT CGACCT GGAC AGC AT C A KRT15 NM 002275 1381 GCCTGGTTCTTCA 1382 :ttgctggtctggatc 1383 TGAACAAAGAGGTGGCCTCC 1384 GCC TGGTTCTTCAGCAAGACTGAGGAGCTGAACAAA KRT18 NM 000224 1385 AGAGATCGAGGCT 1386 3GCCTTTTACTTCCTC 1387 TGGTTCTTCTTCATGAAGAG 1388 AGA GATCGAGGCTCTCAAGGAGGAGCTGCTCTTCAT KRT2 NM 000423 1389 CCAGTGACGCCTC 1390 3GGCATGGCTAGAAG 1391 ACCTAGACAGCACAGATTCC 1392 CCA 3TGACGCCTCTGTGTTCTGGGGCGGAATCTGTGC KRT5 NM 000424 1393 TCAGTGGAGAAGG 1394 rGCCATATCCAGAGG 1395 CCAGTCAACATCTCTGTTGT 1396 TCA< 3TGGAGAAGGAGTTGGACCAGTCAACATCTCTG 130 2015227398 15 Sep 2015

Offui.il Svrahnl: NiiiiiImtt <sr.o n> \o In» ir.1 I’nniii StfEUcliOtS \o irtcfsi- I’riim-r N 11» it! m Γι-ohi Hi NO Ampiii»h Ήη<η-ικ<·: KRT75 NM 004693 1397 TCAAAGTCAGGTA CGAAGATGAAATT 1398 ACGTCCTTTTTCAGGG CTACAA 1399 TTCATTCTCAGCAGCTGTGC GCTTGT 1400 TCAA CAGC AGTCAGGTACGAAGATGAAATTAACAAGCGCA TGCTGAGAATGAATTTGTAGCCCTGAAAAAGG KRT76 NM 015848 1401 ATCTCCAGACTGC TGGTTCC 1402 TCAGGGAATTAGGGG ACAGA 1403 TCTGGGCTTCAGATCCTGAC rccc 1404 ATCT ATCT CCAGACTGCTGGTTCCCAGGGAACCCTCCCTAC GGGCTTCAGATCCTGACTCCCTTCTGTCCCCTA KRT8 NM 002273 1405 GGATGAAGCTTAC ATGAACAAGGTAG 1406 CATATAGCTGCCTGA GGAAGTTGAT 1407 CGTCGGTCAGCCCTTCCAGG c 1408 GGAI TCGC GAAGCTT ACATGAACAAGGTAGAGCTGGAGTC CTGGAAGGGCTGACCGACGAGATCAACTTCCT L1CAM NM 000425 1409 CTTGCTGGCCAAT 1410 TGATTGTCCGCAGTC 1411 ATCTACGTTGTCCAGCTGCC 1412 CTTG CTGGCCAATGCCTACATCTACGTTGTCCAGCTG LAG3 NM 002286 1413 GCCTTAGAGCAAG 1414 CGGTTCTTGCTCCAGC 1415 TCTATCTTGCTCTGAGCCTG 1416 GCCT TAGAGCAAGGGATTCACCCTCCGCAGGCTCAG LAMA3 NM 000227 1417 CCTGTCACTGAAG 1418 TGGGTTACTGGTCAG 1419 ATTCAGACTGACAGGCCCCT 1420 CCTG TCACTGAAGCCTTGGAAGTCCAGGGGCCTGTC LAMA4 NM_002290 1421 GATGCACTGCGGT 1422 CAGAGGATACGCTCA 1423 CTCTCCATCGAGGAAGGCAA 1424 GATC rCACTGCGGTTAGCAGCGCTCTCCATCGAGGAA LAMA5 NM_005560 1425 CTCCTGGCCAACA 1426 ACACAAGGCCCAGCC 1427 CTGTTCCTGGAGCATGGCCT 1428 CTCC TGGCCAACAGCACTGCACTAGAAGAGGCCATG LAMB1 NM_002291 1429 CAAGGAGACTGGG 1430 CGGCAGAACTGACAG 1431 CAAGTGCCTGTACCACACGG 1432 CAAC iGAGACTGGGAGGTGTCTCAAGTGCCTGTACCA LAMB3 NM_000228 1433 ACTGACCAAGCCT 1434 GTCACACTTGCAGCA 1435 CCACTCGCCATACTGGGTGC 1436 ACTG ACCAAGCCTGAGACCTACTGCACCCAGTATGG LAMC1 NM_002293 1437 GCCGTGATCTCAG 1438 ACCTGCTTGCCCAAG 1439 CCTCGGTACTTCATTGCTCC 1440 GCCC iTGATCTCAGACAGCTACTTTCCTCGGTACTTCA LAMC2 NM_005562 1441 ACTCAAGCGGAAA TTGAAGCA 1442 ACTCCCTGAAGCCGA GACACT 1443 AGGTCTTATCAGCACAGTCT CCGCCTCC 1444 ACTC CA CA AAGCGGAAATTGAAGCAGATAGGTCTTATCAG lGTCTCCGCCTCCTGGATTCAGTGTCTCGGCTTC LAPTM5 NM 006762 1445 TGCTGGACTTCTG 1446 TGAGATAGGTGGGCA 1447 TCCTGACCCTCTGCAGCTCC 1448 TGCT GGACTTCTGCCTGAGCATCCTGACCCTCTGCAG LGALS3 NM 002306 1449 AGCGGAAAATGGC 1450 CTTGAGGGTTTGGGT 1451 ACCCAGATAACGCATCATGG 1452 AGCC IGAAAATGGCAGACAATTTTTCGCTCCATGATG LIG3 NM 002311 1453 GGAGGTGGAGAAG 1454 ACAGGTGTCATCAGC 1455 CTGGACGCTCAGAGCTCGTC 1456 GGAC jGTGGAGAAGGAGCCGGGCCAGAGACGAGCTCT LIMS1 NM 004987 1457 TGAACAGTAATGG 1458 TTCTGGGAACTGCTG 1459 ACTGAGCGCACACGAAACA 1460 TGA4 lCAGTAATGGGGAGCTGTACCATGAGCAGTGTT LOX NM 002317 1461 CCAATGGGAGAAC 1462 CGCTGAGGCTGGTAC 1463 CAGGCTCAGCAAGCTGAACA 1464 CCA4 lTGGGAGAACAACGGGCAGGTGTTCAGCTTGCT LRP1 NM_002332 1465 TTTGGCCCAATGG GCTAAG 1466 GTCTCGATGCGGTCG TAGAAG 1467 TCCCGGCTGGGCGCCTCTAC Γ 1468 TTTG GGCC GCCCAATGGGCTAAGCCTGGACATCCCGGCTG iCCTCTACTGGGTGGATGCCTTCTACGACCGCAT LTBP2 NM_000428 1469 GCACACCCATCCT 1470 GATGGCTGGCCACGT 1471 CTTTGCAGCCCTCAGAACTC 1472 GCAC 'ACCCATCCTTGAGTCTCCTTTGCAGCCCTCAGA LUM NM 002345 1473 GGCTCTTTTGAAG GATTGGTAA 1474 AAAAGCAGCTGAAAC AGCATC 1475 CCTGACCTTCATCCATCTCC AGCA 1476 GGCI ATCT CTTTTGAAGGATTGGTAAACCTGACCTTCATCC CCAGCACAATCGGCTGAAAGAGGATGCTGTTT MAGEA4 NM 002362 1477 GCATCTAACAGCC 1478 CAGAGTGAAGAATGG 1479 CAGCTTCCCTTGCCTCGTGT 1480 GCAT CTAACAGCCCTGTGCAGCAGCTTCCCTTGCCTC MANF NM 006010 1481 CAGATGTGAAGCC 1482 AAGGGAATCCCCTCA 1483 TTCCTGATGATGCTGGCCCT 1484 CAG/ lTGTGAAGCCTGGAGCTTTCCTGATGATGCTGG MAOA NM 000240 1485 GTGTCAGCCAAAG 1486 CGACTACGTCGAACA 1487 CCGCGATACTCGCCTTCTCT 1488 GTGT CAGCCAAAGCATGGAGAATCAAGAGAAGGCGA MAP3K5 NM 005923 1489 AGGACCAAGAGGC 1490 CCTGTGGCCATTTCA 1491 CAGCCCAGAGACCAGATGTC 1492 AGO/ ^CCAAGAGGCTACGGAAAAGCAGCAGACATCTG MAP3K7 NM_145333 1493 CAGGCAAGAACTA GTTGCAGAA 1494 CCTGTACCAGGCGAG ATGTAT 1495 TGCTGGTCCTTTTCATCCTG GTCC 1496 CAGC AAAC 1CAAGAACT AGTTGCAGAACTGGACCAGGATGA IGACCAGCAAAATACATCTCGCCTGGTACAGG MAP4K4 NM_004834 1497 TCGCCGAGATTTC 1498 CTGTTGTCTCCGAAG 1499 AACGTTCCTTGTTCTCCTGC 1500 TCGC CGAGATTTCCTGAGACTGCAGCAGGAGAACAA MAP7 NM_003980 1501 GAGGAACAGAGGT 1502 CTGCCAACTGGCTTTC 1503 CATGTACAACAAACGCTCCG 1504 GAGC jAACAGAGGTGTCTGCACTTCCATGTACAACAA MAPKAP K3 NM_004635 1505 AAGCTGCAGAGAT AATGCGG 1506 GTGGGCAATGTTATG GCTG 1507 ATTGGCACTGCCATCCAGTT TCTG 1508 AAGC TCCA TGCAGAGATAATGCGGGATATTGGCACTGCCA GTTTCTGCACAGCCATAACATTGCCCAC MCM2 NM_004526 1509 GACTTTTGCCCGC TACCTTTC 1510 GCCACTAACTGCTTC AGTATGAAGAG 1511 ACAGCTCATTGTTGTCACGC CGGA 1512 GACT AATC TTTGCCCGCTACCTTTCATTCCGGCGTGACAAC rAGCTGTTGCTCTTCATACTGAAGCAGTTAGTGG MCM3 NM_002388 1513 GGAGAACAATCCC 1514 ATCTCCTGGATGGTG 1515 TGGCCTTTCTGTCTACAAGG 1516 GGAC jAACAATCCCCTTGAGACAGAATATGGCCTTTC MCM6 NM_005915 1517 TGATGGTCCTATG TGTCACATTCA 1518 TGGGACAGGAAACAC ACCAA 1519 CAGGTTTCATACCAACACAG GCTTCAGCAC 1520 TGAT ACCA GGTCCTATGTGTCACATTCATCACAGGTTTCAT ACACAGGCTTCAGCACTTCCTTTGGTGTGTTTC MDK NM 002391 1521 GGAGCCGACTGCA 1522 GACTTTGGTGCCTGT 1523 ATCACACGCACCCCAGTTCT 1524 GGAC jCCGACTGCAAGTACAAGTTTGAGAACTGGGGT MDM2 NM 002392 1525 CTACAGGGACGCC 1526 ATCCAACCAATCACC 1527 CTTACACCAGCATCAAGATC 1528 CTAC AGGGACGCCATCGAATCCGGATCTTGATGCTG MEEK NM 014791 1529 AGGATCGCCTGTC 1530 TGCACATAAGCAACA 1531 CCCGGGTTGTCTTCCGTCAG 1532 AOOA lTCGCCTGTCAGAAGAGGAGACCCGGGTTGTCT MET NM 000245 1533 GACATTTCCAGTC CTGCAGTCA 1534 CTCCGATCGCACACA TTTGT 1535 TGCCTCTCTGCCCCACCCTT TGT 1536 GAC/ ACCC OTTCCAGTCCTGCAGTCAATGCCTCTCTGCCCC TTTGTTCAGTGTGGCTGGTGCCACGACAAATGT MGMT NM 002412 1537 GTGAAATGAAACG 1538 GACCCTGCTCACAAC 1539 CAGCCCTTTGGGGAAGCTGG 1540 GTG/ lAATGAAACGCACCACACTGGACAGCCCTTTGG MGST1 NM 020300 1541 ACGGATCTACCAC ACCATTGC 1542 TCCATATCCAACAAA AAAACTCAAAG 1543 TTTGACACCCCTTCCCCAGC CA 1544 ACGC CCCC rATCTACCACACCATTGCATATTTGACACCCCTT AGCCAAATAGAGCTTTGAGTTTTTTTGTTGGAT MICA NM 000247 1545 ATGGTGAATGTCA 1546 AAGCCAGAAGCCCTG 1547 CGAGGCCTCAGAGGGCAAC 1548 ATGC rTGAATGTCACCCGCAGCGAGGCCTCAGAGGGC 131 2015227398 15 Sep 2015

Offluil !»νίιιΕκ>Ι. VtWftfoft Nmnlii-lt smm 1 i.i« ir.1 Pnrmr ScTiUrtR-r: wmm V) Primer Smm'm'ri ϋϋ! lYohr SHI it) NO Antplhw* MKI67 NM_002417 1549 GATTGCACCAGGG 1550 TCCAAAGTGCCTCTG 1551 CCACTCTTCCTTGAACACCC 1552 GATTGCACCAGGGCAGAACAGGGGAGGGTGTTCAAG MLXIP NM_014938 1553 TGCTTAGCTGGCA 1554 CAGCCTACTCTCCAT 1555 CATGAGATGCCAGGAGACCC 1556 TGCTTAGCTGGCATGTGGCCGCATGAGATGCCAGGA MMP11 NM_005940 1557 CCTGGAGGCTGCA ACATACC 1558 TACAATGGCTTTGGA GGATAGCA 1559 ATCCTCCTGAAGCCCTTTTC GCAGC 1560 CCTGGAGGCTGCAACATACCTCAATCCTGTCCCAGG CCGGATCCTCCTGAAGCCCTTTTCGCAGCACTGCTAT MMP2 NM_004530 1561 CAGCCAGAAGCGG 1562 AGACACCATCACCTG 1563 AAGTCCGAATCTCTGCTCCC 1564 CAGCCAGAAGCGGAAACTTAAAAAGTCCGAATCTCT MMP7 NM_002423 1565 GGATGGTAGCAGT CTAGGGATTAACT 1566 GGAATGTCCCATACC CAAAGAA 1567 CCTGTATGCTGCAACTCATG AACTTGGC 1568 GGATGGTAGCAGTCTAGGGATTAACTTCCTGTATGCT GCAACTCATGAACTTGGCCATTCTTTGGGTATGGGAC MMP9 NM 004994 1569 GAGAACCAATCTC 1570 CACCCGAGTGTAACC 1571 ACAGGTATTCCTCTGCCAGC 1572 GAGAACCAATCTCACCGACAGGCAGCTGGCAGAGGA MPPED2 NM 001584 1573 CCGACCAACCCTC 1574 AGGGCATTTAGAGCT 1575 ATTTGACCTTCCAAACCCAC 1576 CCGACCAACCCTCCAATTATATTTGACCTTCCAAACC MRC1 NM 002438 1577 CTTGACCTCAGGA 1578 GGACTGCGGTCACTC 1579 CCAACCGCTGTTGAAGCTCA 1580 CTTGACCTCAGGACTCTGGATTGGACTTAACAGTCTG MRPL13 NM 014078 1581 TCCGGTTCCCTTCG 1582 GTGGAAAAACTGCGG 1583 CGGCTGGAAATTATGTCCTC 1584 TCCGGTTCCCTTCGTTTAGGTCGGCTGGAAATTATGT MSH2 NM 000251 1585 GATGCAGAATTGA 1586 TCTTGGCAAGTCGGT 1587 CAAGAAGATTTACTTCGTCG 1588 GATGCAGAATTGAGGCAGACTTTACAAGAAGATTTA MSH3 NM_002439 1589 TGATTACCATCAT GGCTCAGA 1590 CTTGTGAAAATGCCA TCCAC 1591 TCCCAATTGTCGCTTCTTCT GCAG 1592 TGATTACCATCATGGCTCAGATTGGCTCCTATGTTCC TGCAGAAGAAGCGACAATTGGGATTGTGGATGGCAT MSH6 NM 000179 1593 TCTATTGGGGGAT 1594 CAAATTGCGAGTGGT 1595 CCGTTACCAGCTGGAAATTC 1596 TCTATTGGGGGATTGGTAGGAACCGTTACCAGCTGG MTA1 NM 004689 1597 CCGCCCTCACCTG AAGAGA 1598 GGAATAAGTTAGCCG CGCTTCT 1599 CCCAGTGTCCGCCAAGGAGC G 1600 CCGCCCTCACCTGCAGAGAAACGCGCTCCTTGGCGG ACACTGGGGGAGGAGAGGAAGAAGCGCGGCTAACTT MTPN NM 145808 1601 GGTGGAAGGAAAC 1602 CAGCAGCAGAAATTC 1603 AAGCTGCCCACAATCTGCTG 1604 GGTGGAAGGAAACCTCTTCATTATGCAGCAGATTGT MTSS1 NM 014751 1605 TTCGACAAGTCCT 1606 CTTGGAACATCCGTC 1607 CCAAGAAACAGCGACATCA 1608 TTCGACAAGTCCTCCACCATTCCAAGAAACAGCGAC MUC1 NM 002456 1609 GGCCAGGATCTGT GGTGGTA 1610 CTCCACGTCGTGGAC ATTGA 1611 CTCTGGCCTTCCGAGAAGGT ACC 1612 GGCCAGGATCTGTGGTGGTACAATTGACTCTGGCCTT CCGAGAAGGTACCATCAATGTCCACGACGTGGAG MVP NM 017458 1613 ACGAGAACGAGGG CATCTATGT 1614 GCATGTAGGTGCTTC CAATCAC 1615 CGCACCTTTCCGGTCTTGAC ATCCT 1616 ACGAGAACGAGGGCATCTATGTGCAGGATGTCAAGA CCGGAAAGGTGCGCGCTGTGATTGGAAGCACCTACA MYBL2 NM_002466 1617 GCCGAGATCGCCA AGATG 1618 CTTTTGATGGTAGAG TTCCAGTGATTC 1619 CAGCATTGTCTGTCCTCCCT GGCA 1620 GCCGAGATCGCCAAGATGTTGCCAGGGAGGACAGAC AATGCTGTGAAGAATCACTGGAACTCTACCATCAAA MYBPC1 NM_002465 1621 CAGCAACCAGGGA 1622 C AGC AGT AAGTGCCT 1623 AAATTCGCAAGCCCAGCCCC 1624 CAGCAACCAGGGAGTCTGTACCCTGGAAATTCGCAA MYC NM 002467 1625 TCCCTCCACTCGG AAGGACTA 1626 CGGTTGTTGCTGATCT GTCTCA 1627 TCTGACACTGTCCAACTTGA CCCTCTT 1628 TCCCTCCACTCGGAAGGACTATCCTGCTGCCAAGAG GGTCAAGTTGGACAGTGTCAGAGTCCTGAGACAGAT MYLK3 NM 182493 1629 CACCTGACTGAGC TGGATGT 1630 GATGTAGTGCTGGTG CAGGT 1631 CACACCCTCACAGATCTGCC TGGT 1632 CACCTGACTGAGCTGGATGTGGTCCTGTTCACCAGGC AGATCTGTGAGGGTGTGCATTACCTGCACCAGCACT MY06 NM 004999 1633 AAGCAGTTCTGGA 1634 GATGAGCTCGGCTTC 1635 CAATCCTCAGGGCCAGCTCC 1636 AAGCAGTTCTGGAGCAGGAGCGCAGGGACCGGGAGC NCAM1 NM 000615 1637 TAGTTCCCAGCTG 1638 CAGCCTTGTTCTCAGC 1639 CTCAGCCTCGTCGTTCTTAT 1640 TAGTTCCCAGCTGACCATCAAAAAGGTGGATAAGAA NCAPD3 NM 015261 1641 TCGTTGCTTAGAC 1642 CTCCAGACAGTGTGC 1643 CTACTGTCCGCAGCAAGGCA 1644 TCGTTGCTTAGACAAGGCGCCTACTGTCCGCAGCAA NCOR1 NM 006311 1645 AACCGTTACAGCC 1646 TCTGGAGAGACCCTT 1647 CCAGGCTCAGTCTGTCCATC 1648 AACCGTTACAGCCCAGAATCCCAGGCTCAGTCTGTCC NCOR2 NM 006312 1649 CGTCATCTACGAA 1650 GAGCACTGGGTCACA 1651 CCTCATAGGACAAGACGTGG 1652 CGTCATCTACGAAGGCAAGAAGGGCCACGTCTTGTC NDRG1 NM 006096 1653 AGGGCAACATTCC 1654 CAGTGCTCCTACTCC 1655 CTGCAAGGACACTCATCACA 1656 AGGGCAACATTCCACAGCTGCCCTGGCTGTGATGAG NDUFS5 NM 004552 1657 AGAAGAGTCAAGG 1658 AGGCCGAACCTTTTC 1659 TGTCCAAGAAAGGCATGGCT 1660 AGAAGAGTCAAGGGCACGAGCATCGGGTAGCCATGC NEK2 NM 002497 1661 GTGAGGCAGCGCG ACTCT 1662 TGCCAATGGTGTACA ACACTTCA 1663 TGCCTTCCCGGGCTGAGGAC T 1664 GTGAGGCAGCGCGACTCTGGCGACTGGCCGGCCATG CCTTCCCGGGCTGAGGACTATGAAGTGTTGTACACC NET02 NM 018092 1665 CCAGGGCACCATA 1666 AACGGTAAATCAAGG 1667 AGCCAACCCTTTTCTCCCAT 1668 CCAGGGCACCATACTGTTTCCAGCAGCCAACCCTTTT NEXN NM 144573 1669 AGGAGGAGGAAGA 1670 GAGCTCCTGATCTGG 1671 TCATCTTCAGCAGTGGAGCC 1672 AGGAGGAGGAAGAAGGTAGCATCATGAATGGCTCCA NFAT5 NM 006599 1673 CTGAACCCCTCTC 1674 AGGAAACGATGGCGA 1675 CGAGAATCAGTCCCCGTGGA 1676 CTGAACCCCTCTCCTGGTCACCGAGAATCAGTCCCCG NFATC2 NM 173091 1677 CAGTCAAGGTCAG 1678 CTTTGGCTCGTGGCAT 1679 CGGGTTCCTACCCCACAGTC 1680 CAGTCAAGGTCAGAGGCTGAGCCCGGGTTCCTACCC NFKB1 NM 003998 1681 CAGACCAAGGAGA 1682 AGCTGCCAGTGCTAT 1683 AAGCTGTAAACATGAGCCGC 1684 CAGACCAAGGAGATGGACCTCAGCGTGGTGCGGCTC NFKBIA NM 020529 1685 CT ACTGGACGACC 1686 CCTTGACCATCTGCTC 1687 CTCGTCTTTCATGGAGTCCA 1688 CTACTGGACGACCGCCACGACAGCGGCCTGGACTCC NME1 NM_000269 1689 CCAACCCTGCAGA CTCCAA 1690 ATGTATAATGTTCCTG CCAACTTGTATG 1691 CCTGGGACCATCCGTGGAGA CTTCT 1692 CCAACCCTGCAGACTCCAAGCCTGGGACCATCCGTG GAGACTTCTGCATACAAGTTGGCAGGAACATTATAC NNMT NM 006169 1693 CCT AGGGCAGGGA 1694 CTAGTCCAGCCAAAC 1695 CCCTCTCCTCATGCCCAGAC 1696 CCTAGGGCAGGGATGGAGAGAGAGTCTGGGCATGAG NOS3 NM_000603 1697 ATCTCCGCCTCGC 1698 TCGGAGCCATACAGG 1699 TTCACTCGCTTCGCCATCAC 1700 ATCTCCGCCTCGCTCATGGGCACGGTGATGGCGAAG 132 2015227398 15 Sep 2015

Ofl1cl.il Sv ni hoi. Numbers iipn e forward Printer JimHefler: wmm V) Srtfrw Primer Srctunwr; lie ϊ»80 Pr<4>e ?><.< (lu-itce: 111 Ampllcon .SeijuetKe: NOX4 NM_016931 1701 CCTCAACTGCAGC CTTATCC 1702 TGCTTGGAACCTTCTG TGAT 1703 CCGAACACTCTTGGCTTACC TCCG 1704 CCTCAACTGCAGCCTTATCCTTTTACCCATGTGCCGA ACACTCTTGGCTTACCTCCGAGGATCACAGAAGGTTC NPBWR1 NM 005285 1705 TCACCAACCTGTT 1706 GATGTTGATGGGCAG 1707 ATCGCCGACGAGCTCTTCAC 1708 TCACCAACCTGTTCATCCTCAACCTGGCCATCGCCGA NPM1 NM_002520 1709 AATGTTGTCCAGG TTCTATTGC 1710 CAAGCAAAGGGTGGA GTTC 1711 AACAGGCATTTTGGACAACA CATTCTTG 1712 AATGTTGTCCAGGTTCTATTGCCAAGAATGTGTTGTC CAAAATGCCTGTTTAGTTTTTAAAGATGGAACTCCAC NRG1 NM_013957 1713 CGAGACTCTCCTC ATAGTGAAAGGTA 1714 CTTGGCGTGTGGAAA TCTACAG 1715 ATGACCACCCCGGCTCGTAT GTCA 1716 CGAGACTCTCCTCATAGTGAAAGGTATGTGTCAGCC ATGACCACCCCGGCTCGTATGTCACCTGTAGATTTCC NRIP3 NM_020645 1717 CCCACAAGCATGA 1718 TGCTCAATCTGGCCC 1719 AGCTTTCTCTACCCCGGCAT 1720 CCCACAAGCATGAAGGAGAAAAGCTTTCTCTACCCC NRP1 NM_003873 1721 CAGCTCTCTCCAC GCGATTC 1722 CCCAGCAGCTCCATT CTGA 1723 CAGGATCTACCCCGAGAGAG CCACTCAT 1724 CAGCTCTCTCCACGCGATTCATCAGGATCTACCCCGA GAGAGCCACTCATGGCGGACTGGGGCTCAGAATGGA NUP62 NM 153719 1725 AGCCTCTTTGCGT CAATAGC 1726 CTGTGGTCACAGGGG TACAG 1727 TCATCTGCCACCACTGGACT CTCC 1728 AGCCTCTTTGCGTCAATAGCAACTGCTCCAACCTCAT CTGCCACCACTGGACTCTCCCTCTGTACCCCTGTGAC OAZ1 NM 004152 1729 AGCAAGGACAGCT 1730 GAAGACATGGTCGGC 1731 CTGCTCCTCAGCGAACTCCA 1732 AGCAAGGACAGCTTTGCAGTTCTCCTGGAGTTCGCTG OCLN NM 002538 1733 CCCTCCCATCCGA 1734 GACGCGGGAGTGTAG 1735 CTCCTCCCTCGGTGACCAAT 1736 CCCTCCCATCCGAGTTTCAGGTGAATTGGTCACCGAG ODC1 NM 002539 1737 AGAGATCACCGGC GTAATCAA 1738 CGGGCTCAGCTATGA TTCTCA 1739 CCAGCGTTGGACAAATACTT TCCGTCA 1740 AGAGATCACCGGCGTAATCAACCCAGCGTTGGACAA ATACTTTCCGTCAGACTCTGGAGTGAGAATCATAGCT OLFML2 NM 015441 1741 CATGTTGGAAGGA 1742 CACCAGTTTGGTGGT 1743 TGGCCTGGATCTCCTGAAGC 1744 CATGTTGGAAGGAGCGTTCTATGGCCTGGATCTCCTG OLFML3 NM 020190 1745 TCAGAACTGAGGC 1746 CCAGATAGTCTACCT 1747 CAGACGATCCACTCTCCCGG 1748 TCAGAACTGAGGCCGACACCATCTCCGGGAGAGTGG OMD NM 005014 1749 CGCAAACTCAAGA CTATCCCA 1750 CAGTCACAGCCTCAA TTTCATT 1751 TCCGATGCACATTCAGCAAC TCTACC 1752 CGCAAACTCAAGACTATCCCAAATATTCCGATGCAC ATTCAGCAACTCTACCTTCAGTTCAATGAAATTGAGG OR51E1 NM_152430 1753 GCATGCTTTCAGG 1754 AGAAGATGGCCAGCA 1755 TCCTCATCTCCACCTCATCC 1756 GCATGCTTTCAGGCATTGACATCCTCATCTCCACCTC OR51E2 NM_030774 1757 TATGGTGCCAAAA 1758 GTCCTTGTCACAGCT 1759 ACATAGCCAGCACCCGTGTT 1760 TATGGTGCCAAAACCAAACAGATCAGAACACGGGTG OSM NM_020530 1761 GTTTCTGAAGGGG 1762 AGGTGTCTGGTTTGG 1763 CTGAGCTGGCCTCCTATGCC 1764 GTTTCTGAAGGGGAGGTCACAGCCTGAGCTGGCCTC PAGE1 NM 003785 1765 CAACCTGACGAAG TGGAATC 1766 CAGATGCTCCCTCAT CCTCT 1767 CCAACTCAAAGTCAGGATTC TACACCTGC 1768 CAACCTGACGAAGTGGAATCACCAACTCAAAGTCAG GATTCTACACCTGCTGAAGAGAGAGAGGATGAGGGA PAGE4 NM 007003 1769 GAATCTCAGCAAG AGGAACCA 1770 GTTCTTCGATCGGAG GTGTT 1771 CCAACTGACAATCAGGATAT TGAACCTGG 1772 GAATCTCAGCAAGAGGAACCACCAACTGACAATCAG GATATTGAACCTGGACAAGAGAGAGAAGGAACACCT PAK6 NM 020168 1773 CCTCCAGGTCACC 1774 GTCCCTTCAGGCCAG 1775 AGTTTCAGGAAGGCTGCCCC 1776 CCTCCAGGTCACCCACAGCCAGTTTCAGGAAGGCTG PATE1 NM 138294 1777 TGGTAATCCCTGG 1778 TCCACCTTATGCCTTT 1779 CAGCACAGTTCTTTAGGCAG 1780 TGGTAATCCCTGGTTAACCTTCATGGGCTGCCTAAAG PC A3 NR 015342 1781 CGTGATTGTCAGG 1782 AGAAAGGGGAGATGC 1783 CTGAGATGCTCCCTGCCTTC 1784 CGTGATTGTCAGGAGCAAGACCTGAGATGCTCCCTG PCDHGB NM_018927 1785 CCCAGCGTTGAAG 1786 GAAACGCCAGTCCGT 1787 ATTCTTAAACAGCAAGCCCC 1788 CCCAGCGTTGAAGCAGATAAGAAGATTCTTAAACAG PCNA NM_002592 1789 GAAGGTGTTGGAG 1790 GGTTTACACCGCTGG 1791 ATCCCAGCAGGCCTCGTTGA 1792 GAAGGTGTTGGAGGCACTCAAGGACCTCATCAACGA PDE9A NM_0010015 70 1793 TTCCACAACTTCC GGCAC 1794 AGACTGCAGAGCCAG ACCA 1795 TACATCATCTGGGCCACGCA GAAG 1796 TTCCACAACTTCCGGCACTGCTTCTGCGTGGCCCAGA TGATGTACAGCATGGTCTGGCTCTGCAGTCT PDGFRB NM 002609 1797 CCAGCTCTCCTTCC 1798 GGGTGGCTCTCACTT 1799 ATCAATGTCCCTGTCCGAGT 1800 CCAGCTCTCCTTCCAGCTACAGATCAATGTCCCTGTC PECAM1 NM_000442 1801 TGTATTTCAAGAC CTCTGTGCACTT 1802 TTAGCCTGAGGAATT GCTGTGTT 1803 TTTATGAACCTGCCCTGCTC CCACA 1804 TGTATTTCAAGACCTCTGTGCACTTATTTATGAACCT GCCCTGCTCCCACAGAACACAGCAATTCCTCAGGCT PEX10 NM_153818 1805 GGAGAAGTTCCCT CCCCAG 1806 ATCTGTGTCCAGGCC CAC 1807 CTACCTTCGGCACTACCGCT GAGC 1808 GGAGAAGTTCCCTCCCCAGAAGCTCATCTACCTTCGG CACTACCGCTGAGCCGGCGCCCGGGTGGGCCTGGAC PGD NM 002631 1809 ATTCCCATGCCCT 1810 CTGGCTGGAAGCATC 1811 ACTGCCCTCTCCTTCTATGA 1812 ATTCCCATGCCCTGTTTTACCACTGCCCTCTCCTTCT PGF NM 002632 1813 GTGGTTTTCCCTCG 1814 AGCAAGGGAACAGCC 1815 ATCTTCTCAGACGTCCCGAG 1816 GTGGTTTTCCCTCGGAGCCCCCTGGCTCGGGACGTCT PGK1 NM_000291 1817 AGAGCCAGTTGCT GTAGAACTCAA 1818 CTGGGCCTACACAGT CCTTCA 1819 TCTCTGCTGGGCAAGGATGT TCTGTTC 1820 AGAGCCAGTTGCTGTAGAACTCAAATCTCTGCTGGG CAAGGATGTTCTGTTCTTGAAGGACTGTGTAGGCCCA PGR NM 000926 1821 GATAAAGGAGCCG 1822 TCACAAGTCCGGCAC 1823 TAAATTGCCGTCGCAGCCGC 1824 GATAAAGGAGCCGCGTGTCACTAAATTGCCGTCGCA PHTF2 NM 020432 1825 GATATGGCTGATG 1826 GGTTTGGGTGTTCTTG 1827 ACAATCTGGCAATGCACAGT 1828 GATATGGCTGATGCTGCTCCTGGGAACTGTGCATTGC PTK3C2A NM 002645 1829 ATACCAATCACCG CACAAACC 1830 CACACTAGCATTTTCT CCGCATA 1831 TGTGCTGTGACTGGACTTAA CAAATAGCCT 1832 ATACCAATCACCGCACAAACCCAGGCTATTTGTTAAG TCCAGTCACAGCACAAAGAAACATATGCGGAGAAAA PTK3CA NM 006218 1833 GTGATTGAAGAGC 1834 GTCCTGCGTGGGAAT 1835 TCCTGCTTCTCGGGATACAG 1836 GTGATTGAAGAGCATGCCAATTGGTCTGTATCCCGA PTK3CG NM 002649 1837 GGAGAACTCAATG 1838 TGATGCTTAGGCAGG 1839 TTCTGGACAATTACTGCCAC 1840 GGAGAACTCAATGTCCATCTCCATTCTTCTGGACAAT PIM1 NM 002648 1841 CTGCTCAAGGACA 1842 GGATCCACTCTGGAG 1843 TACACTCGGGTCCCATCGAA 1844 CTGCTCAAGGACACCGTCTACACGGACTTCGATGGG 133 2015227398 15 Sep 2015

Of1u-i.il 'Minimi: Vin-vAm \iniil«-r: «eoo) \o 1 urn.inj Prir ν·([ΐΗΊΗ(·: if ipi \o K»-'c-rs«· PriffM-T ΙΙϋΙ iiiiii Probe Ill Amplhw* Λ<·φ«·#κ·<·: PLA2G7 NM 005084 1845 CCTGGCTGTGGTT 1846 TGACCCATGCTGATG 1847 TGGCAATACATAAATCCTGT 1848 CCTGGCTGTGGTTTATCCTTTTGACTGGCAATACATA PLAU NM 002658 1849 GTGGATGTGCCCT 1850 CTGCGGATCCAGGGT 1851 AAGCCAGGCGTCTACACGAG 1852 GTGGATGTGCCCTGAAGGACAAGCCAGGCGTCTACA PLAUR NM 002659 1853 CCCATGGATGCTC 1854 CCGGTGGCTACCAGA 1855 CATTGACTGCCGAGGCCCCA 1856 CCCATGGATGCTCCTCTGAAGAGACTTTCCTCATTGA PLG NM 000301 1857 GGCAAAATTTCCA 1858 ATGTATCCATGAGCG 1859 TGCCAGGCCTGGGACTCTCA 1860 GGCAAAATTTCCAAGACCATGTCTGGACTGGAATGC PLK1 NM 005030 1861 AATGAATACAGTA TTCCCAAGCACAT 1862 TGTCTGAAGCATCTTC TGGATGA 1863 AACCCCGTGGCCGCCTCC 1864 AATGAATACAGTATTCCCAAGCACATCAACCCCGTG GCCGCCTCCCTCATCCAGAAGATGCTTCAGACA PLOD2 NM_000935 1865 CAGGGAGGTGGTT GCAAAT 1866 TCTCCCAGGATGCAT GAAG 1867 TCCAGCCTTTTCGTGGTGAC TCAA 1868 CAGGGAGGTGGTTGCAAATTTCTAAGGTACAATTGC TCTATTGAGTCACCACGAAAAGGCTGGAGCTTCATG PLP2 NM 002668 1869 CCTGATCTGCTTCA 1870 GCAGCAAGGATCATC 1871 ACACCAGGCTACTCCTCCCT 1872 CCTGATCTGCTTCAGTGCCTCCACACCAGGCTACTCC PNLIPRP NM 005396 1873 TGGAGAAGGTGAA 1874 CACGGCTTGGGTGTA 1875 ACCCGTGCCTCCAGTCCACA 1876 TGGAGAAGGTGAACTGCATCTGTGTGGACTGGAGGC POSTN NM 006475 1877 GTGGCCCAATTAG 1878 TCACAGGTGCCAGCA 1879 TTCTCCATCTGGCCTCAGAG 1880 GTGGCCCAATTAGGCTTGGCATCTGCTCTGAGGCCA PPAP2B NM 003713 1881 ACAAGCACCATCC 1882 CACGAAGAAAACTAT 1883 ACCAGGGCTCCTTGAGCAAA 1884 ACAAGCACCATCCCAGTGATGTTCTGGCAGGATTTGC PPFIA3 NM 003660 1885 CCTGGAGCTCCGT 1886 AGCCACATAGGGATC 1887 CACCCACTTTACCTTCTGGT 1888 CCTGGAGCTCCGTTACTCTCAGGCACCCACTTTACCT PPP1R12 A NM_002480 1889 CGGCAAGGGGTTG ATATAGA 1890 TGCCTGGCATCTCTA AGCA 1891 CCGTTCTTCTTCCTTTCGAG CTGC 1892 CGGCAAGGGGTTGATATAGAAGCAGCTCGAAAGGAA GAAGAACGGATCATGCTTAGAGATGCCAGGCA PPP3CA NM_000944 1893 ATACTCCGAGCCC 1894 GGAAGCCTGTTGTTT 1895 TACATGCGGTACCCTGCATC 1896 ATACTCCGAGCCCACGAAGCCCAAGATGCAGGGTAC PRIMA1 NM_178013 1897 ATCCTCTTCCCTGA 1898 CCCAGCTGAGAGGGA 1899 TGACGCATCCAGGGCTCTAG 1900 ATCCTCTTCCCTGAGCCGCTGACGCATCCAGGGCTCT PRKAR1 NM_002735 1901 ACAAAACCATGAC 1902 TGTCATCCAGGTGAG 1903 AAGGCCATCTCCAAGAACGT 1904 ACAAAACCATGACTGCGCTGGCCAAGGCCATCTCCA PRKAR2 B NM_002736 1905 TGATAATCGTGGG AGTTTCG 1906 GC ACCAGGAGAGGT A GCAGT 1907 CGAACTGGCCTTAATGTACA ATACACCCA 1908 TGATAATCGTGGGAGTTTCGGCGAACTGGCCTTAAT GTACAATACACCCAGAGCAGCTACAATCACTGCTAC PRKCA NM 002737 1909 CAAGCAATGCGTC 1910 GT AAATCCGCCCCCT 1911 CAGCCTCTGCGGAATGGATC 1912 CAAGCAATGCGTCATCAATGTCCCCAGCCTCTGCGG PRKCB NM 002738 1913 GACCCAGCTCCAC 1914 CCCATTCACGTACTCC 1915 CCAGACCATGGACCGCCTGT 1916 GACCCAGCTCCACTCCTGCTTCCAGACCATGGACCGC PROM1 NM 006017 1917 CTATGACAGGCAT 1918 CTCCAACCATGAGGA 1919 ACCCGAGGCTGTGTCTCCAA 1920 CT ATGACAGGCATGCCACCCCGACCACCCGAGGCTG PROS1 NM 000313 1921 GCAGCACAGGAAT 1922 CCCACCTATCCAACCT 1923 CTCATCCTGACAGACTGCAG 1924 GCAGCACAGGAATCTTCTTCTTGGCAGCTGCAGTCTG PSCA NM_005672 1925 ACCGTCATCAGCA AAGGCT 1926 CGTGATGTTCTTCTTG CCC 1927 CCTGTGAGTCATCCACGCAG TTCA 1928 ACCGTCATCAGCAAAGGCTGCAGCTTGAACTGCGTG GATGACTCACAGGACTACTACGTGGGCAAGAAGAAC PSMD13 NM_002817 1929 GGAGGAGCTCTAC ACGAAGAAG 1930 CGGATCCTGCACAAA ATCA 1931 CCTGAAGTGTCAGCTGATGC CACA 1932 GGAGGAGCTCTACACGAAGAAGTTGTGGCATCAGCT GACACTTCAGGTGCTTGATTTTGTGCAGGATCCG PTCH1 NM 000264 1933 CCACGACAAAGCC 1934 TACTCGATGGGCTCT 1935 CCTGAAACAAGGCTGAGAAT 1936 CCACGACAAAGCCGACTACATGCCTGAAACAAGGCT PTEN NM_000314 1937 TGGCTAAGTGAAG ATGACAATCATG 1938 TGCACATATCATTAC ACCAGTTCGT 1939 CCTTTCCAGCTTTACAGTGA ATTGCTGCA 1940 TGGCTAAGTGAAGATGACAATCATGTTGCAGCAATT CACTGTAAAGCTGGAAAGGGACGAACTGGTGTAATG PTGER3 NM_000957 1941 TAACTGGGGCAAC 1942 TTGCAGGAAAAGGTG 1943 CCTTTGCCTTCCTGGGGCTC 1944 TAACTGGGGCAACCTTTTCTTCGCCTCTGCCTTTGCC PTGS2 NM_000963 1945 GAATCATTCACCA GGCAAATTG 1946 CTGTACTGCGGGTGG AACAT 1947 CCTACCACCAGCAACCCTGC CA 1948 GAATCATTCACCAGGCAAATTGCTGGCAGGGTTGCT GGTGGTAGGAATGTTCCACCCGCAGTACAG PTH1R NM_000316 1949 CGAGGT AC AAGCT GAGATCAAGAA 1950 GCGTGCCTTTCGCTTG AA 1951 CCAGTGCCAGTGTCCAGCGG CT 1952 CGAGGTACAAGCTGAGATCAAGAAATCTTGGAGCCG CTGGACACTGGCACTGGACTTCAAGCGAAAGGCACG PTHLH NM 002820 1953 AGTGACTGGGAGT GGGCTAGAA 1954 AAGCCTGTTACCGTG AATCGA 1955 TGACACCTCCACAACGTCGC TGGA 1956 AGTGACTGGGAGTGGGCTAGAAGGGGACCACCTGTC TGACACCTCCACAACGTCGCTGGAGCTCGATTCACG PTK2 NM 005607 1957 GACCGGTCGAATG 1958 CTGGACATCTCGATG 1959 ACCAGGCCCGTCACATTCTC 1960 GACCGGTCGAATGATAAGGTGTACGAGAATGTGACG PTK2B NM 004103 1961 CAAGCCCAGCCGA 1962 GAACCTGGAACTGCA 1963 CTCCGCAAACCAACCTCCTG 1964 CAAGCCCAGCCGACCTAAGTACAGACCCCCTCCGCA PTK6 NM 005975 1965 GTGCAGGAAAGGT TCACAAA 1966 GCACACACGATGGAG TAAGG 1967 AGTGTCTGCGTCCAATACAC GCGT 1968 GTGCAGGAAAGGTTCACAAATGTGGAGTGTCTGCGT CCAATACACGCGTGTGCTCCTCTCCTTACTCCATCGT PTK7 NM 002821 1969 TCAGAGGACTCAC 1970 CATACACCTCCACGC 1971 CGCAAGGTCCCATTCTTGAA 1972 TCAGAGGACTCACGGTTCGAGGTCTTCAAGAATGGG PTPN1 NM 002827 1973 AATGAGGAAGTTT 1974 CTTCGATCACAGCCA 1975 CTGATCCAGACAGCCGACCA 1976 AATGAGGAAGTTTCGGATGGGGCTGATCCAGACAGC PTPRK NM 002844 1977 TCAAACCCTCCCA 1978 AGCAGCCAGTTCGTC 1979 CCCCATCGTTGTACATTGCA 1980 TCAAACCCTCCCAGTGCTGGCCCCATCGTTGTACATT PTTG1 NM 004219 1981 GGCTACTCTGATC TATGTTGATAAGG 1982 GCTTCAGCCCATCCTT AGCA 1983 CACACGGGTGCCTGGTTCTC CA 1984 GGCTACTCTGATCTATGTTGATAAGGAAAATGGAGA ACCAGGCACCCGTGTGGTTGCTAAGGATGGGCTGAA PYCARD NM 013258 1985 CTTT AT AGACC AG 1986 AGCATCCAGCAGCCA 1987 ACGTTTGTGACCCTCGCGAT 1988 CTTT AT AGACCAGCACCGGGCTGCGCTT ATCGCGAG RAB27A NM 004580 1989 TGAGAGATTAATG 1990 CCGGATGCTTTATTCG 1991 ACAAATTGCTTCTCACCATC 1992 TGAGAGATTAATGGGCATTGTGTACAAATTGCTTCTC RAB30 NM 014488 1993 TAAAGGCTGAGGC 1994 CTCCCCAGCATCTCAT 1995 CCATCAGGGCAGTTGCTGAT 1996 TAAAGGCTGAGGCACGGAGAAGAAAAGGAATCAGCA 134 2015227398 15 Sep 2015

Offlclul wm hoi. WcVslod Niiinhcri HI \o fforttanJ Primer Jimtienec: V) Reverse Primer Seniienee. ϋϋI Prnhr Sequence: IB WO Ampllcon .Sequence: RAB31 NM_006868 1997 CTGAAGGACCCT A 1998 ATGCAAAGCCAGTGT 1999 CTTCTCAAAGTGAGGTGCCA 2000 CTGAAGGACCCTACGCTCGGTGGCCTGGCACCTCAC RAD21 NM_006265 2001 TAGGGATGGTATC TGAAACAACA 2002 TCGCGTACACCTCTG CTC 2003 CACTTAAAACGAATCTCAAG AGGGTGACCA 2004 TAGGGATGGTATCTGAAACAACAATGGTCACCCTCTT GAGATTCGTTTTAAGTGTAATTCCATAATGAGCAGAG RAD51 NM 002875 2005 AGACTACTCGGGT 2006 AGCATCCGCAGAAAC 2007 CTTTCAGCCAGGCAGATGCA 2008 AGACTACTCGGGTCGAGGTGAGCTTTCAGCCAGGCA RAD9A NM_004584 2009 GCCATCTTCACCA 2010 CGGTGTCTGAGAGTG 2011 CTTTGCTGGACGGCCACTTT 2012 GCCATCTTCACCATCAAGGACTCTTTGCTGGACGGCC RAF1 NM_002880 2013 CGTCGTATGCGAG 2014 TGAAGGCGTGAGGTG 2015 TCCAGGATGCCTGTTAGTTC 2016 CGTCGTATGCGAGAGTCTGTTTCCAGGATGCCTGTTA RAGE NM_014226 2017 ATTAGGGGACTTT 2018 GGGTGGAGATGTATT 2019 CCGGAGTGTCTATTCCAAGC 2020 ATTAGGGGACTTTGGCTCCTGCCGGAGTGTCTATTCC RALA NM_005402 2021 TGGTCCTGAATGT 2022 CCCCATTTCACCTCTT 2023 TTGTGTTTCTTGGGCAGTCT 2024 TGGTCCTGAATGTAGCGTGTAAGCTTGTGTTTCTTGG RALBP1 NM_006788 2025 GGTGTCAGATATA AATGTGCAAATGC 2026 TTCGATATTGCCAGC AGCTATAAA 2027 TGCTGTCCTGTCGGTCTCAG TACGTTCA 2028 GGTGTCAGATATAAATGTGCAAATGCCTTCTTGCTGT CCTGTCGGTCTCAGTACGTTCACTTTATAGCTGCTGG RAP1B NM_0010109 42 2029 TGACAGCGTGAGA GGTACTAGG 2030 CTGAGCCAAGAACGA CTAGCTT 2031 CACGCATGATGCAAGCTTGT CAAA 2032 TGACAGCGTGAGAGGTACTAGGTTTTGACAAGCTTG CATCATGCGTGAGTATAAGCTAGTCGTTCTTGGCTCA RARB NM 000965 2033 ATGAACCCTTGAC CCCAAGT 2034 GAGCTGGGTGAGATG CTAGG 2035 TGTGCTCTGCTGTGTTCCCA CTTG 2036 ATGAACCCTTGACCCCAAGTTCAAGTGGGAACACAG CAGAGCACAGTCCTAGCATCTCACCCAGCTC RASSF1 NM 007182 2037 AGGGCACGTGAAG TCATTG 2038 AAAGAGTGCAAACTT GCGG 2039 CACCACCAAGAACTTTCGCA GCAG 2040 AGGGCACGTGAAGTCATTGAGGCCCTGCTGCGAAAG TTCTTGGTGGTGGATGACCCCCGCAAGTTTGCACTCT RBI NM 000321 2041 CGAAGCCCTTACA 2042 GGACTCTTCAGGGGT 2043 CCCTTACGGATTCCTGGAGG 2044 CGAAGCCCTTACAAGTTTCCTAGTTCACCCTTACGGA RECK NM 021111 2045 GTCGCCGAGTGTG 2046 GTGGGATGATGGGTT 2047 TCAAGTGTCCTTCGCTCTTG 2048 GTCGCCGAGTGTGCTTCTGTCAAGTGTCCTTCGCTCT REG4 NM 032044 2049 TGCTAACTCCTGC ACAGCC 2050 TGCTAGGTTTCCCCTC TGAA 2051 TCCTCTTCCTTTCTGCTAGCC TGGC 2052 TGCTAACTCCTGCACAGCCCCGTCCTCTTCCTTTCTG CTAGCCTGGCTAAATCTGCTCATTATTTCAGAGGGGA RET, A NM 021975 2053 CTGCCGGGATGGC 2054 CCAGGTTCTGGAAAC 2055 CTGAGCTCTGCCCGGACCGC 2056 CTGCCGGGATGGCTTCTATGAGGCTGAGCTCTGCCC RFX1 NM 002918 2057 TCCTCTCCAAGTTC 2058 CAGGCCCTGGT AC AG 2059 TCCAATGGACCAAGCACTGT 2060 TCCTCTCCAAGTTCGAGCCCGTGCTCCAATGGACCAA RGS10 NM 0010053 39 2061 AGACATCCACGAC AGCGAT 2062 CCATTTGGCTGTGCTC TTG 2063 AGTTCCAGCAGCAGCCACCA GAG 2064 AGACATCCACGACAGCGATGGCAGTTCCAGCAGCAG CCACCAGAGCCTCAAGAGCACAGCCAAATGG RGS7 NM_002924 2065 CAGGCTGCAGAGA GCATTT 2066 TTTGCTTGTGCTTCTG CTTG 2067 TGAAAATGAACTCCCACTTC CGGG 2068 CAGGCTGCAGAGAGCATTTGCCCGGAAGTGGGAGTT CATTTTCATGCAAGCAGAAGCACAAGCAAA RHOA NM 001664 2069 TGGCATAGCTCTG 2070 TGCCACAGCTGCATG 2071 AAATGGGCTCAACCAGAAA 2072 TGGCATAGCTCTGGGGTGGGCAGTTTTTTGAAAATG RHOB NM 004040 2073 AAGCATGAACAGG 2074 CCTCCCCAAGTCAGT 2075 CTTTCCAACCCCTGGGGAAG 2076 AAGCATGAACAGGACTTGACCATCTTTCCAACCCCTG RHOC NM 175744 2077 CCCGTTCGGTCTG 2078 GAGCACTCAAGGTAG 2079 TCCGGTTCGCCATGTCCCG 2080 CCCGTTCGGTCTGAGGAAGGCCGGGACATGGCGAAC RLN1 NM_006911 2081 AGCTGAAGGCAGC CCTATC 2082 TTGGAATCCTTTAATG CAGGT 2083 TGAGAGGCAACCATCATTAC CAGAGC 2084 AGCTGAAGGCAGCCCTATCTGAGAGGCAACCATCAT TACCAGAGCTACAGCAGTATGTACCTGCATTAAAGG RND3 NM 005168 2085 TCGGAATTGGACT 2086 CTGGTTACTCCCCTCC 2087 TTTTAAGCCTGACTCCTCAC 2088 TCGGAATTGGACTTGGGAGGCGCGGTGAGGAGTCAG RNF114 NM 018683 2089 TGACAGGGGAAGT 2090 GGAAGACAGCTTTGG 2091 CCAGGTCAGCCCTTCTCTTC 2092 TGACAGGGGAAGTGGGTCCCCAGGTCAGCCCTTCTC R0B02 NM 002942 2093 CTACAAGGCCCAG 2094 CACCAGTGGCTTTAC 2095 CTGTACCATCCACTGCCAGC 2096 CTACAAGGCCCAGCCAACCAAACGCTGGCAGTGGAT RRM1 NM 001033 2097 GGGCTACTGGCAG 2098 CTCTCAGCATCGGTA 2099 CATTGGAATTGCCATTAGTC 2100 GGGCTACTGGCAGCTACATTGCTGGGACTAATGGCA RRM2 NM 001034 2101 CAGCGGGATT AAA 2102 ATCTGCGTTGAAGCA 2103 CCAGCACAGCCAGTTAAAAG 2104 CAGCGGGATTAAACAGTCCTTTAACCAGCACAGCCA S100P NM 005980 2105 AGACAAGGATGCC 2106 GAAGTCCACCTGGGC 2107 TTGCTCAAGGACCTGGACGC 2108 AGACAAGGATGCCGTGGATAAATTGCTCAAGGACCT SAT1 NM 002970 2109 CCTTTTACCACTGC 2110 ACAATGCTGTGTCCTT 2111 TCCAGTGCTCTTTCGGCACT 2112 CCTTTTACCACTGCCTGGTTGCAGAAGTGCCGAAAGA SCUBE2 NM_020974 2113 TGACAATCAGCAC ACCTGCAT 2114 TGTGACTACAGCCGT GATCCTTA 2115 CAGGCCCTCTTCCGAGCGGT 2116 TGACAATCAGCACACCTGCATTCACCGCTCGGAAGA GGGCCTGAGCTGCATGAATAAGGATCACGGCTGTAG SDC1 NM 002997 2117 GAAATTGACGAGG 2118 AGGAGCT AACGGAGA 2119 CTCTGAGCGCCTCCATCCAA 2120 GAAATTGACGAGGGGTGTCTTGGGCAGAGCTGGCTC SDC2 NM 002998 2121 GGATTGAAGTGGC 2122 ACCAGCCACAGTACC 2123 AACTCCATCTCCTTCCCCAG 2124 GGATTGAAGTGGCTGGAAAGAGTGATGCCTGGGGAA SDHC NM 003001 2125 CTTCCCTCGGGTCT 2126 TTCCCTCCTGGTAAA 2127 TTACATCCTCCCTCTCCCCG 2128 CTTCCCTCGGGTCTCAGGCATTTACATCCTCCCTCTC SEC14L1 NM_0010395 73 2129 AGGGTTCCCATGT GACCAG 2130 GCAGGCATGCTGTGG AAT 2131 CGGGCTTCTACATCCTGCAG TGG 2132 AGGGTTCCCATGTGACCAGGTGGCCGGGCTTCTACA TCCTGCAGTGGAAATTCCACAGCATGCCTGC SEC23A NM 006364 2133 CGTGTGCATTAGA 2134 CCCATTACCATGTATC 2135 TCCTGGAGATGAAATGCTGT 2136 CGTGTGCATTAGATCAGACAGGTCTCCTGGAGATGA SEMA3A NM 006080 2137 TTGGAATGCAGTC 2138 CTCTTCATTTCGCCTC 2139 TTGCCAATAGACCAGCGCTC 2140 TTGGAATGCAGTCCGAAGTCGCAGAGAGCGCTGGTC SEPT9 NM 006640 2141 CAGTGACCACGAG 2142 CTTCGATGGTACCCC 2143 TTGCCAATAGACCAGCGCTC 2144 CAGTGACCACGAGTACCAGGTCAACGGCAAGAGGAT SERPINA 3 NM 001085 2145 GTGTGGCCCTGTC TGCTTA 2146 CCCTGTGCATGTGAG AGCTAC 2147 AGGGAATCGCTGTCACCTTC CAAG 2148 GTGTGGCCCTGTCTGCTTATCCTTGGAAGGTGACAGC GATTCCCTGTGTAGCTCTCACATGCACAGGG 135 2015227398 15 Sep 2015

«H1cl.il !»νίηΕκ»Ι. VctCWoft Niiinhcrs mom iHfc forward Miner .SmUefler: smm V) Reverse Primer Sen lienee; $80 ise Probe Sequence: 111 Ampftren A«i|Wemv; SERPINB 5 NM_002639 2149 CAGATGGCCACTT TGAGAACATT 2150 GGCAGCATTAACCAC AAGGATT 2151 AGCTGACAACAGTGTGAACG ACCAGACC 2152 CAGATGGCCACTTTGAGAACATTTTAGCTGACAACA GTGTGAACGACCAGACCAAAATCCTTGTGGTTAATG SESN3 NM 144665 2153 GACCCTGGTTTTG 2154 GAGCTCGGAATGTTG 2155 TGCTCTTCTCCTCGTCTGGC 2156 GACCCTGGTTTTGGGTATGAAGACTTTGCCAGACGA SFRP4 NM 003014 2157 TACAGGATGAGGC 2158 GTTGTTAGGGCAAGG 2159 CCTGGGACAGCCTATGTAAG 2160 TACAGGATGAGGCTGGGCATTGCCTGGGACAGCCTA SH3RF2 NM_152550 2161 CCATCACAACAGC CTTGAAC 2162 CACTGGGGTGCTGAT CTCTA 2163 AACCGGATGGTCCATTCTCC TTCA 2164 CCATCACAACAGCCTTGAACACTCTCAACCGGATGG TCCATTCTCCTTCAGGGCGCCATATGGTAGAGATCAG SH3YL1 NM 015677 2165 CCTCCAAAGCCAT 2166 CTTTGAGAGCCAGAG 2167 CACAGCAGTCATCTGCACCA 2168 CCTCCAAAGCCATTGTCAAGACCACAGCAGTCATCT SHH NM 000193 2169 GTCCAAGGCACAT 2170 GAAGCAGCCTCCCGA 2171 CACCGAGTTCTCTGCTTTCA 2172 GTCCAAGGCACATATCCACTGCTCGGTGAAAGCAGA SHMT2 NM 005412 2173 AGCGGGTGCTAGA 2174 ATGGCACTTCGGTCT 2175 CCATCACTGCCAACAAGAAC 2176 AGCGGGTGCTAGAGCTTGTATCCATCACTGCCAACA SIM2 NM 005069 2177 GATGGTAGGAAGG 2178 CACAAGGAGCTGTGA 2179 CGCCTCTCCACGCACTCAGC 2180 GATGGTAGGAAGGGATGTGCCCGCCTCTCCACGCAC SIPA1L1 NM 015556 2181 CT AGGACAGCTTG 2182 CATAACCGTAGGGCT 2183 CGCCACAATGCCCTCAT AGT 2184 CTAGGACAGCTTGGCTTCCATGTCAACTATGAGGGC SKIL NM 005414 2185 AGAGGCTGAATAT 2186 CTATCGGCCTCAGCA 2187 CCAATCTCTGCCTCAGTTCT 2188 AGAGGCTGAATATGCAGGACAGTTGGCAGAACTGAG SLC22A3 NM 021977 2189 ATCGTCAGCGAGT 2190 CAGGATGGCTTGGGT 2191 CAGCATCCACGCATTGACAC 2192 ATCGTCAGCGAGTTTGACCTTGTCTGTGTCAATGCGT SLC25A2 1 NM_030631 2193 AAGTGTTTTTCCCC CTTGAGAT 2194 GGCCGATCGATAGTC TCTCTT 2195 TCATGGTGCTGCATAGCAAA TATCCA 2196 AAGTGTTTTTCCCCCTTGAGATAATGGATATTTGCTA TGCAGCACCATGAAGAAGAGAGACTATCGATCGGCC SLC44A1 NM_080546 2197 AGGACCGTAGCTG 2198 ATCCCATCCCAATGC 2199 TACCATGGCTGCTGCTCTTC 2200 AGGACCGTAGCTGCACAGACATACCATGGCTGCTGC SMAD4 NM_005359 2201 GGACATTACTGGC 2202 ACCAATACTCAGGAG 2203 TGCATTCCAGCCTCCCATTT 2204 GGACATTACTGGCCTGTTCACAATGAGCTTGCATTCC SMARCC 2 NM 003075 2205 TACCGACTGAACC CCCAA 2206 GACATCACCCGCTAG GTTTC 2207 TATCTTACCTCTACCGCCTG CCGC 2208 TACCGACTGAACCCCCAAGAGTATCTTACCTCTACCG CCTGCCGCCGAAACCTAGCGGGTGATGTC SMARC D1 NM 003076 2209 CCGAGTTAGCATA TCCCAGG 2210 CCTTTGTGCCCAGCTG TC 2211 CCCACCCTTGCTGTGTTGAG TCTG 2212 CCGAGTTAGCATATCCCAGGCTCGCAGACTCAACAC AGCAAGGGTGGGAGACAGCTGGGCACAAAGG SMO NM 005631 2213 GGCATCCAGTGCC 2214 CGCGATGTAGCTGTG 2215 CTTCACAGAGGCTGAGCACC 2216 GGCATCCAGTGCCAGAACCCGCTCTTCACAGAGGCT SNAI1 NM 005985 2217 CCCAATCGGAAGC 2218 GT AGGGCTGCTGGAA 2219 TCTGGATTAGAGTCCTGCAG 2220 CCCAATCGGAAGCCTAACTACAGCGAGCTGCAGGAC SNRPB2 NM 003092 2221 CGTTTCCTGCTTTT 2222 AGGT AGAAGGCGCAC 2223 CCCACCTAAGGCCTACGCCG 2224 CGTTTCCTGCTTTTGGTTCTTACAGTAGTCGGCGTAG SOD1 NM_000454 2225 TGAAGAGAGGCAT 2226 AATAGACACATCGGC 2227 TTTGTCAGCAGTCACATTGC 2228 TGAAGAGAGGCATGTTGGAGACTTGGGCAATGTGAC SORBS 1 NM_015385 2229 GCAGATGAGTGGA 2230 AGCGAGTGAAGAGGG 2231 ATTTCCATTGGCATCAGCAC 2232 GCAGATGAGTGGAGGCTTTCTTCCAGTGCTGATGCC SOX4 NM_003107 2233 AGATGATCTCGGG 2234 GCGCCCTTCAGTAGG 2235 CGAGTCCAGCATCTCCAACC 2236 AGATGATCTCGGGAGACTGGCTCGAGTCCAGCATCT SPARC NM_003118 2237 TCTTCCCTGTACAC TGGCAGTTC 2238 AGCTCGGTGTGGGAG AGGT A 2239 TGGACCAGCACCCCATTGAC GG 2240 TCTTCCCTGTACACTGGCAGTTCGGCCAGCTGGACCA GCACCCCATTGACGGGTACCTCTCCCACACCGAGCT SPARCL NM 004684 2241 GGCACAGTGCAAG 2242 GATTGAGCTCTCTCG 2243 ACTTCATCCCAAGCCAGGCC 2244 GGCACAGTGCAAGTGATGACTACTTCATCCCAAGCC SPDEF NM_012391 2245 CCATCCGCCAGTA TTACAAG 2246 GGGTGCACGAACTGG TAGA 2247 ATCATCCGGAAGCCAGACAT CTCC 2248 CCATCCGCCAGTATTACAAGAAGGGCATCATCCGGA AGCCAGACATCTCCCAGCGCCTCGTCTACCAGTTCGT SPINK1 NM_003122 2249 CTGCCATATGACC 2250 GTTGAAAACTGCACC 2251 ACCACGTCTCTTCAGAAGCC 2252 CTGCCATATGACCCTTCCAGTCCCAGGCTTCTGAAGA SPINT1 NM_003710 2253 ATTCCCAGCACAG 2254 AGATGGCTACCACCA 2255 CTGTCGCAGTGTTCCTGGTC 2256 ATTCCCAGCACAGGCTCTGTGGAGATGGCTGTCGCA SPP1 NM_0010400 58 2257 TCACACATGGAAA GCGAGG 2258 GTTCAGGTCCTGGGC AAC 2259 TGAATGGTGCATACAAGGCC ATCC 2260 TCACACATGGAAAGCGAGGAGTTGAATGGTGCATAC AAGGCCATCCCCGTTGCCCAGGACCTGAAC SQLE NM_003129 2261 ATTTTCGAGGCCA AAAAATC 2262 CCTGAGCAAGGATAT TCACG 2263 TGGGCAAGAAAAACATCTCA TTCCTTTG 2264 ATTTTCGAGGCCAAAAAATCATTTTACTGGGCAAGA AAAACATCTCATTCCTTTGTCGTGAATATCCTTGCTC SRC NM_005417 2265 TGAGGAGTGGTAT TTTGGCAAGA 2266 CTCTCGGGTTCTCTGC ATTGA 2267 AACCGCTCTGACTCCCGTCT GGTG 2268 TGAGGAGTGGTATTTTGGCAAGATCACCAGACGGGA GTCAGAGCGGTTACTGCTCAATGCAGAGAACCCGAG SRD5A1 NM 001047 2269 GGGCTGGAATCTG 2270 CCATGACTGCACAAT 2271 CCTCTCTCGGAGGCCACAGA 2272 GGGCTGGAATCTGTCTAGGAGCCCTCTCTCGGAGGC SRD5A2 NM_000348 2273 GTAGGTCTCCTGG CGTTCTG 2274 TCCCTGGAAGGGTAG GAGTAA 2275 AGACACCACTCAGAATCCCC AGGC 2276 GTAGGTCTCCTGGCGTTCTGCCAGCTGGCCTGGGGAT TCTGAGTGGTGTCTGCTTAGAGTTTACTCCTACCCTT ST5 NM_005418 2277 CCTGTCCTGCCAG 2278 CAGCTGCACAAAACT 2279 AGTCACGAGCACCCAGCGA 2280 CCTGTCCTGCCAGAGCATGGATGAAGTTTCGCTGGGT STAT1 NM_007315 2281 GGGCTCAGCTTTC AGAAGTG 2282 ACATGTTCAGCTGGT CCACA 2283 TGGCAGTTTTCTTCTGTCAC CAAAA 2284 GGGCTCAGCTTTCAGAAGTGCTGAGTTGGCAGTTTTC TTCTGTCACCAAAAGAGGTCTCAATGTGGACCAGCT STAT3 NM 003150 2285 TCACATGCCACTTT 2286 CTTGCAGGAAGCGGC 2287 TCCTGGGAGAGATTGACCAG 2288 TCACATGCCACTTTGGTGTTTCATAATCTCCTGGGAG STAT5A NM 003152 2289 GAGGCGCTCAACA TGAAATTC 2290 GCCAGGAACACGAGG TTCTC 2291 CGGTTGCTCTGCACTTCGGC CT 2292 GAGGCGCTCAACATGAAATTCAAGGCCGAAGTGCAG AGCAACCGGGGCCTGACCAAGGAGAACCTCGTGTTC STAT5B NM 012448 2293 CCAGTGGTGGTGA 2294 GCAAAAGCATTGTCC 2295 CAGCCAGGACAACAATGCG 2296 CCAGTGGTGGTGATCGTTCATGGCAGCCAGGACAAC 136 2015227398 15 Sep 2015

official wm hoi. Wiluhew ϋϋϋ iltlli forward Prbmr Knewflcrs wmm V) Srtfrw Primer Sntnmm ise I’pfthv WMjuwttv; Ill Ampllcon .Sequence: STMN1 NM 005563 2297 AATACCCAACGCA 2298 GGAGACAATGCAAAC 2299 CACGTTCTCTGCCCCGTTTC 2300 AATACCCAACGCACAAATGACCGCACGTTCTCTGCC STS NM 000351 2301 GAAGATCCCTTTC CTCCTACTGTTC 2302 GGATGATGTTCGGCC TTGAT 2303 CTGCGTGGCTCTCGGCTTCC CA 2304 GAAGATCCCTTTCCTCCTACTGTTCTTTCTGTGGGAA GCCGAGAGCCACGCAGCATCAAGGCCGAACATCATC SULF1 NM 015170 2305 TGCAGTTGTAGGG AGTCTGG 2306 TCTCAAGAATTGCCG TTGAC 2307 TACCGTGCCAGCAGAAGCCA AAG 2308 TGCAGTTGTAGGGAGTCTGGTTACCGTGCCAGCAGA AGCCAAAGAAAGAGTCAACGGCAATTCTTGAGA SUMOl NM 003352 2309 GTGAAGCCACCGT 2310 CCTTCCTTCTTATCCC 2311 CTGACCAGGAGGCAAAACCT 2312 GTGAAGCCACCGTCATCATGTCTGACCAGGAGGCAA SVIL NM 003174 2313 ACTTGCCCAGCAC 2314 GACACCATCCGTGTC 2315 ACCCCAGGACTGATGTCAAG 2316 ACTTGCCCAGCACAAGGAAGACCCCAGGACTGATGT TAF2 NM_003184 2317 GCGCTCCACTCTC AGTCTTT 2318 CTTGTGCTCATGGTG ATGGT 2319 AGCCTCCAAACACAGTGACC ACCA 2320 GCGCTCCACTCTCAGTCTTTACTAAGGAATCTACAGC CTCCAAACACAGTGACCACCATCACCACCATCACCAT TARP NM_0010037 99 2321 GAGCAACACGATT CTGGGA 2322 GGCACCGTTAACCAG CTAAAT 2323 TCTTCATGGTGTTCCCCTCC TGG 2324 GAGCAACACGATTCTGGGATCCCAGGAGGGGAACAC CATGAAGACTAACGACACATACATGAAATTTAGCTG TBP NM 003194 2325 GCCCGAAACGCCG 2326 CGTGGCTCTCTTATCC 2327 TACCGCAGCAAACCGCTTGG 2328 GCCCGAAACGCCGAATATAATCCCAAGCGGTTTGCT TFDP1 NM_007 111 2329 TGCGAAGTGCTTT TGTTTGT 2330 GCCTTCCAGACAGTC TCCAT 2331 CGCACCAGCATGGCAATAAG CTTT 2332 TGCGAAGTGCTTTTGTTTGTTTGTTTTCGTTTGGTTAA AGCTTATTGCCATGCTGGTGCGGCTATGGAGACTGTC TFF1 NM_003225 2333 GCCCTCCCAGTGT GCAAAT 2334 CGTCGATGGTATTAG GATAGAAGCA 2335 TGCTGTTTCGACGACACCGT TCG 2336 GCCCTCCCAGTGTGC AAAT AAGGGCTGCTGTTTCGAC GACACCGTTCGTGGGGTCCCCTGGTGCTTCTATCCTA TFF3 NM_003226 2337 AGGCACTGTTCAT CTCAGTTTTTCT 2338 CATCAGGCTCCAGAT ATGAACTTTC 2339 CAGAAGCGCTTGCCGGGAG CAAAGG 2340 AGGCACTGTTCATCTCAGCTTTTCTGTCCCTTTGCTC CCGGCAAGCGCTTCTGCTGAAAGTTCATATCTGGAG TGFA NM_003236 2341 GGTGTGCCACAGA CCTTCCT 2342 ACGGAGTTCTTGACA GAGTTTTGA 2343 TTGGCCTGTAATCACCTGTG CAGCCTT 2344 GGTGTGCCACAGACCTTCCTACTTGGCCTGTAATCAC CTGTGCAGCCTTTTGTGGGCCTTCAAAACTCTGTCAA TGFB1I1 NM_0010424 54 2345 GCTACTTTGAGCG CTTCTCG 2346 GGTCACCATCTTGTGT CGG 2347 CAAGATGTGGCTTCTGCAAC CAGC 2348 GCTACTTTGAGCGCTTCTCGCCAAGATGTGGCTTCTG CAACCAGCCCATCCGACACAAGATGGTGACC TGFB2 NM_003238 2349 ACCAGTCCCCCAG 2350 CCTGGTGCTGTTGTA 2351 TCCTGAGCCCGAGGAAGTCC 2352 ACCAGTCCCCCAGAAGACTATCCTGAGCCCGAGGAA TGFB3 NM_003239 2353 GGATCGAGCTCTT 2354 GCCACCGATATAGCG 2355 CGGCCAGATGAGCACATTGC 2356 GGATCGAGCTCTTCCAGATCCTTCGGCCAGATGAGC TGFBR2 NM_003242 2357 AACACCAATGGGT 2358 CCTCTTCATCAGGCC 2359 TTCTGGGCTCCTGATTGCTC 2360 AACACCAATGGGTTCCATCTTTCTGGGCTCCTGATTG THBS2 NM_003247 2361 CAAGACTGGCTAC ATCAGAGTCTTAG 2362 CAGCGTAGGTTTGGT CATAGATAGG 2363 TGAGTCTGCCATGACCTGTT TTCCTTCAT 2364 CAAGACTGGCTACATCAGAGTCTTAGTGCATGAAGG AAAACAGGTCATGGCAGACTCAGGACCTATCTATGA THY1 NM 006288 2365 GGACAAGACCCTC 2366 TTGGAGGCTGTGGGT 2367 CAAGCTCCCAAGAGCTTCCA 2368 GGACAAGACCCTCTCAGGCTGTCCCAAGCTCCCAAG TIAM1 NM 003253 2369 GTCCCTGGCTGAA 2370 GGGCTCCCGAAGTCT 2371 TGGAGCCCTTCTCCCAAGAT 2372 GTCCCTGGCTGAAAATGGCCTGGAGCCCTTCTCCCAA TIMP2 NM 003255 2373 TCACCCTCTGTGA 2374 TGTGGTTCAGGCTCTT 2375 CCCTGGGACACCCTGAGCAC 2376 TCACCCTCTGTGACTTCATCGTGCCCTGGGACACCCT TIMP3 NM 000362 2377 CTACCTGCCTTGCT 2378 ACCGAAATTGGAGAG 2379 CCAAGAACGAGTGTCTCTGG 2380 CTACCTGCCTTGCTTTGTGACTTCCAAGAACGAGTGT TK1 NM_003258 2381 GCCGGGAAGACCG TAATTGT 2382 CAGCGGCACCAGGTT CAG 2383 CAAATGGCTTCCTCTGGAAG GTCCCA 2384 GCCGGGAAGACCGTAATTGTGGCTGCACTGGATGGG ACCTTCCAGAGGAAGCCATTTGGGGCCATCCTGAAC TMPRSS NM_005656 2385 GGACAGTGTGCAC 2386 CTCCCACGAGGAAGG 2387 AAGCACTGTGCATCACCTTG 2388 GGACAGTGTGCACCTCAAAGACTAAGAAAGCACTGT TMPRSS 2ERGA DQ204772 2389 GAGGCGGAGGGCG AG 2390 ACTGGTCCTCACTCA CAACT 2391 TAAGGCTTCCTGCCGCGCTC CA 2392 GAGGCGGAGGCGGAGGGCGAGGGGCGGGGAGCGCC GCCTGGAGCGCGGCAGGAAGCCTTATCAGTTGTGAG TMPRSS 2ERGB DQ204773 2393 GAGGCGGAGGGCG AG 2394 TTCCTCGGGTCTCCAA AGAT 2395 CCTGGAATAACCTGCCGCGC 2396 GAGGCGGAGGGCGAGGGGCGGGGAGCGCCGCCTGG AGCGCGGCAGGTTATTCCAGGATCTTTGGAGACCCG TNF NM_000594 2397 GGAGAAGGGTGAC 2398 TGCCCAGACTCGGCA 2399 CGCTGAGATCAATCGGCCCG 2400 GGAGAAGGGTGACCGACTCAGCGCTGAGATCAATCG TNFRSF1 0A NM_003844 2401 TGCACAGAGGGTG TGGGTTAC 2402 TCTTCATCTGATTTAC AAGCTGTACATG 2403 CAATGCTTCCAACAATTTGT TTGCTTGCC 2404 TGCACAGAGGGTGTGGGTTACACCAATGCTTCCAAC AATTTGTTTGCTTGCCTCCCATGTACAGCTTGTAAAT TNFRSF1 OB NM_003842 2405 CTCTGAGACAGTG CTTCGATGACT 2406 CCATGAGGCCCAACT TCCT 2407 CAGACTTGGTGCCCTTTGAC TCC 2408 CTCTGAGACAGTGCTTCGATGACTTTGCAGACTTGGT GCCCTTTGACTCCTGGGAGCCGCTCATGAGGAAGTT TNFRSF1 8 NM 148901 2409 CAGAAGCTGCCAG TTCCC 2410 CACCCACAGGTCTCC CAG 2411 CCTTCTCCTCTGCCGATCGC TC 2412 CAGAAGCTGCCAGTTCCCCGAGGAAGAGCGGGGCGA GCGATCGGCAGAGGAGAAGGGGCGGCTGGGAGACCT TNFSF10 NM 003810 2413 CTTCACAGTGCTC 2414 CATCTGCTTCAGCTCG 2415 AAGTACACGTAAGTTACAGC 2416 CTTCACAGTGCTCCTGCAGTCTCTCTGTGTGGCTGTA TNFSF11 NM 003701 2417 AACTGCATGTGGG 2418 TGACACCCTCTCCACT 2419 ACATGACCAGGGACCAACCC 2420 AACTGCATGTGGGCTATGGGAGGGGTTGGTCCCTGG TOP2A NM 001067 2421 AATCCAAGGGGGA 2422 GT ACAGATTTTGCCC 2423 CATATGGACTTTGACTCAGC 2424 AATCCAAGGGGGAGAGTGATGACTTCCATATGGACT TP53 NM 000546 2425 CTTTGAACCCTTGC 2426 CCCGGGACAAAGCAA 2427 AAGTCCTGGGTGCTTCTGAC 2428 CTTTGAACCCTTGCTTGCAATAGGTGTGCGTCAGAAG TP63 NM 003722 2429 CCCCAAGCAGTGC 2430 GAATCGCACAGCATC 2431 CCCGGGTCTCACTGGAGCCC 2432 CCCCAAGCAGTGCCTCTACAGTCAGTGTGGGCTCCA 137 2015227398 15 Sep 2015

official Nt in hul Ntnuftm smm >o ri>rttar<} PrtttKT Kndwfit-e: HHR NO Srtfrw I'nmtr Swtumcr. $80 ϋϋI Pr<4>e ?><.< (lu-nce: Ill Ampltaa* <S«4|Wvmv; TPD52 NM 005079 2433 GCCTGTGAGATTC 2434 ATGTGCTTGGACCTC 2435 TCTGCTACCCACTGCCAGAT 2436 GCCTGTGAGATTCCTACCTTTGTTCTGCTACCCACTG TPM1 NM0010180 05 2437 TCTCTGAGCTCTG CATTTGTC 2438 GGCTCTAAGGCAGGA TGCTA 2439 TTCTCCAGCTGACCCTGGTT CTCTC 2440 TCTCTGAGCTCTGCATTTGTCTATTCTCCAGCTGACC CTGGTTCTCTCTCTTAGCATCCTGCCTTAGAGCC TPM2 NM 213674 2441 AGGAGATGCAGCT 2442 CCACCTCTTCATATTT 2443 CCAAGCACATCGCTGAGGAT 2444 AGGAGATGCAGCTGAAGGAGGCCAAGCACATCGCTG TPP2 NM 003291 2445 TAACCGTGGCATC 2446 ATGCCAACGCCATGA 2447 ATCCTGTTCAGGTGGCTGCA 2448 TAACCGTGGCATCTACCTCCGAGATCCTGTTCAGGTG TPX2 NM012112 2449 TCAGCTGTGAGCT GCGGATA 2450 ACGGTCCTAGGTTTG AGGTTAAGA 2451 CAGGTCCCATTGCCGGGCG 2452 TCAGCTGTGAGCTGCGGATACCGCCCGGCAATGGGA CCTGCTCTTAACCTCAAACCTAGGACCGT TRA2A NM 013293 2453 GCAAATCCAGATC 2454 CTTCACGAAGATCCC 2455 AACTGAGGCCAAACACTCCA 2456 GCAAATCCAGATCCCAACACTTGCCTTGGAGTGTTTG TRAF3IP NM 147200 2457 CCTCACAGGAACC 2458 CTGGGGCTGGGAATC 2459 TGGATCTGCCAACCATAGAC 2460 CCTCACAGGAACCGAGCAGGCCTGGATCTGCCAACC TRAM1 NM 014294 2461 CAAGAAAAGCACC 2462 ATGTCCGCGTGATTCT 2463 AGTGCTGAGCCACGAATTCG 2464 CAAGAAAAGCACCAAGAGCCCCCCAGTGCTGAGCCA TRAP1 NM 016292 2465 TTACCAGTGGCTTT 2466 TGTCCCGGTTCTAACT 2467 TTCGGCGATTTCAAACACTC 2468 TTACCAGTGGCTTTCAGATGGTTCTGGAGTGTTTGAA TRIM14 NM 033220 2469 CATTCGCCTTAAG 2470 CAAGGTACCTGGCTT 2471 AACTGCCAGCTCTCAGACCC 2472 CATTCGCCTTAAGGAAAGCATAAACTGCCAGCTCTCA TRO NM 177556 2473 GCAACTGCCACCC 2474 TGGTGTGGATACTGG 2475 CCACCCAAGGCCAAATTACC 2476 GCAACTGCCACCCATACAGCTACCACCCAAGGCCAA TRPC6 NM_004621 2477 CGAGAGCCAGGAC TATCTGC 2478 TAGCCGTAGCAAGGC AGC 2479 CTTCTCCCAGCTCCGAGTCC ATG 2480 CGAGAGCCAGGACTATCTGCTCATGGACTCGGAGCT GGGAGAAGACGGCTGCCCGCAAGCCCCGCTGCCTTG TRPV6 NM_018646 2481 CCGTAGTCCCTGC AACCTC 2482 TCCTCACTGTTCACAC AGGC 2483 ACTTTGGGGAGCACCCTTTG TCCT 2484 CCGTAGTCCCTGCAACCTCATCTACTTTGGGGAGCAC CCTTTGTCCTTTGCTGCCTGTGTGAACAGTGAGGA TSTA3 NM 003313 2485 CAATTTGGACTTCT 2486 CACCTCAAAGGCCGA 2487 AACGTGCACATGAACGACAA 2488 CAATTTGGACTTCTGGAGGAAAAACGTGCACATGAA TUBB2A NM 001069 2489 CGAGGACGAGGCT 2490 ACCATGCTTGAGGAC 2491 TCTCAGATCAATCGTGCATC 2492 CGAGGACGAGGCTTAAAAACTTCTCAGATCAATCGT TYMP NM_001953 2493 CTATATGCAGCCA GAGATGTGACA 2494 CCACGAGTTTCTTACT GAGAATGG 2495 ACAGCCTGCCACTCATCACA GCC 2496 CTATATGCAGCCAGAGATGTGACAGCCACCGTGGAC AGCCTGCCACTCATCACAGCCTCCATTCTCAGTAAGA TYMS NM_001071 2497 GCCTCGGTGTGCC 2498 CGTGATGTGCGCAAT 2499 CATCGCCAGCTACGCCCTGC 2500 GCCTCGGTGTGCCTTTCAACATCGCCAGCTACGCCCT UAP1 NM_003115 2501 CTGGAGACGGTCG TAGCTG 2502 GCCAAGCTTTGTAGA AATAGGG 2503 TACCTGTAAACCTTTCTCGG CGCG 2504 CTGGAGACGGTCGTAGCTGCGGTCGCGCCGAGAAAG GTTTACAGGTACATACATTACACCCCTATTTCTACAA UBE2C NM 007019 2505 TGTCTGGCGATAA 2506 ATGGTCCCTACCCATT 2507 TCTGCCTTCCCTGAATCAGA 2508 TGTCTGGCGATAAAGGGATTTCTGCCTTCCCTGAATC UBE2G1 NM 003342 2509 TGACACTGAACGA 2510 AAGCAGAGAGGAATC 2511 TTGTCCCACCAGTGCCTCAT 2512 TGACACTGAACGAGGTGGCTTTTGTCCCACCAGTGCC UBE2T NM 014176 2513 TGTTCTCAAATTGC 2514 AGAGGTCAACACAGT 2515 AGGTGCTTGGAGACCATCCC 2516 TGTTCTCAAATTGCCACCAAAAGGTGCTTGGAGACC UGDH NM 003359 2517 GAAACTCCAGAGG 2518 CTCTGGGAACCCAGT 2519 TATACAGCACACAGGGCCTG 2520 GAAACTCCAGAGGGCCAGAGAGCTGTGCAGGCCCTG UGT2B1 NM 001076 2521 AAGCCTGAAGTGG 2522 CCTCCATTTAAAACCC 2523 AAAGATGGGACTCCTCCTTT 2524 AAGCCTGAAGTGGAATGACTGAAAGATGGGACTCCT UGT2B1 NM 001077 2525 TTGAGTTTGTCATG 2526 TCCAGGTGAGGTTGT 2527 ACCCGAAGGTGCTTGGCTCC 2528 TTGAGTTTGTCATGCGCCATAAAGGAGCCAAGCACC UHRF1 NM 013282 2529 CT ACAGGGGCAAA 2530 GGTGTCATTCAGGCG 2531 CGGCCATACCCTCTTCGACT 2532 CTACAGGGGCAAACAGATGGAGGACGGCCATACCCT UTP23 NM 032334 2533 GATTGCACAAAAA 2534 GGAAAGCAGACATTC 2535 TCGAAATTGTCCTCATTTCA 2536 GATTGCACAAAAATGCCAAGTTCGAAATTGTCCTCAT VCAM1 NM 001078 2537 TGGCTTCAGGAGC TGAATACC 2538 TGCTGTCGTGATGAG AAAATAGTG 2539 CAGGCACACACAGGTGGGA CACAAAT 2540 TGGCTTCAGGAGCTGAATACCCTCCCAGGCACACAC AGGTGGGACACAAATAAGGGTTTTGGAACCACTATT VCL NM 003373 2541 GATACCACAACTC CCATCAAGCT 2542 TCCCTGTTAGGCGCA TCAG 2543 AGTGGCAGCCACGGCGCC 2544 GATACCACAACTCCCATCAAGCTGTTGGCAGTGGCA GCCACGGCGCCTCCTGATGCGCCTAACAGGGA VCPIP1 NM 025054 2545 TTTCTCCCAGTACC 2546 TGAATAGGGAGCCTT 2547 TGGTCCATCCTCTGCACCTG 2548 TTTCTCCCAGTACCATTCGTGATGGTCCATCCTCTGC VDR NM 000376 2549 CCTCTCCTTCCAGC 2550 TCATTGCCAAACACTT 2551 CAGCATGAAGCTAACGCCCC 2552 CCTCTCCTTCCAGCCTGAGTGCAGCATGAAGCTAACG VEGFA NM 003376 2553 CTGCTGTCTTGGG 2554 GCAGCCTGGGACCAC 2555 TTGCCTTGCTGCTCTACCTC 2556 CTGCTGTCTTGGGTGCATTGGAGCCTTGCCTTGCTGC VEGFB NM 003377 2557 TGACGATGGCCTG 2558 GGTACCGGATCATGA 2559 CTGGGCAGCACCAAGTCCGG 2560 TGACGATGGCCTGGAGTGTGTGCCCACTGGGCAGCA VEGFC NM 005429 2561 CCTCAGCAAGACG TTATTTGAAATT 2562 AAGTGTGATTGGCAA AACTGATTG 2563 CCTCTCTCTCAAGGCCCCAA ACCAGT 2564 CCTCAGCAAGACGTTATTTGAAATTACAGTGCCTCTC TCTCAAGGCCCCAAACCAGTAACAATCAGTTTTGCCA VIM NM 003380 2565 TGCCCTTAAAGGA 2566 GCTTCAACGGCAAAG 2567 ATTTCACGCATCTGGCGTTC 2568 TGCCCTTAAAGGAACCAATGAGTCCCTGGAACGCCA VTI1B NM_006370 2569 ACGTTATGCACCC CTGTCTT 2570 CCGATGGAGTTTAGC AAGGT 2571 CGAAACCCCATGATGTCTAA GCTTCG 2572 ACGTTATGCACCCCTGTCTTTCCGAAACCCCATGATG TCTAAGCTTCGAAACTACCGGAAGGACCTTGCTAAA WDR19 NM 025132 2573 GAGTGGCCCAGAT 2574 GATGCTTGAGGGCTT 2575 CCCCTCGACGTATGTCTCCC 2576 GAGTGGCCCAGATGTCCATAAGAATGGGAGACATAC WFDC1 NM 021197 2577 ACCCCTGCTCTGT 2578 ATACCTTCGGCCACG 2579 CTATGAGTGCCACATCCTGA 2580 ACCCCTGCTCTGTCCCTCGGGCTATGAGTGCCACATC WISP1 NM_003882 2581 AGAGGCATCCATG AACTTCACA 2582 CAAACTCCACAGTAC TTGGGTTGA 2583 CGGGCTGCATCAGCACACGC 2584 AGAGGCATCCATGAACTTCACACTTGCGGGCTGCAT CAGCACACGCTCCTATCAACCCAAGTACTGTGGAGTT

OfHciai 'nfnirof. Van tort* w$m. V» In» ir.11’nni.r StifUt-liOti SIQlli NO K< Vtrw Primer Swquturr: M.Q iilli |>r<>hi Sequence: Ill Antplhw* AetpHiKe: WNT5A NM_003392 2585 GTATCAGGACCAC ATGCAGTACATC 2586 TGTCGGAATTGATAC TGGCATT 2587 TTGATGCCTGTCTTCGCGCC TTCT 2588 GTATCAGGACCACATGCAGTACATCGGAGAAGGCGC GAAGACAGGCATCAAAGAATGCCAGTATCAATTCCG wwox NM 016373 2589 ATCGCAGCTGGTG 2590 AGCTCCCTGTTGCAT 2591 CTGCTGTTTACCTTGGCGAG 2592 ATCGCAGCTGGTGGGTGTACACACTGCTGTTTACCTT XIAP NM_001167 2593 GCAGTTGGAAGAC ACAGGAAAGT 2594 TGCGTGGCACTATTTT CAAGA 2595 TCCCCAAATTGCAGATTTAT CAACGGC 2596 GCAGTTGGAAGACACAGGAAAGTATCCCCAAATTGC AGATTTATCAACGGCTTTTATCTTGAAAATAGTGCCA XRCC5 NM 021141 2597 AGCCCACTTCAGC 2598 AGCAGGATTCACACT 2599 TCTGGCTGAAGGCAGTGTCA 2600 AGCCCACTTCAGCGTCTCCAGTCTGGCTGAAGGCAG YY1 NM 003403 2601 ACCCGGGCAACAA 2602 GACCGAGAACTCGCC 2603 TTGATCTGCACCTGCTTCTG 2604 ACCCGGGCAACAAGAAGTGGGAGCAGAAGCAGGTGC ZFHX3 NM 006885 2605 CTGTGGAGCCTCT 2606 GGAGCAGGGTTGGAT 2607 ACCTGGCCCAACTCTACCAG 2608 CTGTGGAGCCTCTGCCTGCGGACCTGGCCCAACTCTA ZFP36 NM 003407 2609 CATTAACCCACTC 2610 CCCCCACCATCATGA 2611 CAGGTCCCCAAGTGTGCAAG 2612 CATTAACCCACTCCCCTGACCTCACGCTGGGGCAGGT ZMYND8 NM 183047 2613 GGTCTGGGCCAAA 2614 TGCCCGTCTTTATCCC 2615 CTTTTGCAGGCCAGAATGGA 2616 GGTCTGGGCCAAACTGAAGGGGTTTCCATTCTGGCCT ZNF3 NM 017715 2617 CGAAGGGACTCTG 2618 GCAGGAGGTCCTCAG 2619 AGGAGGTTCCACACTCGCCA 2620 CGAAGGGACTCTGCTCCAGTGAACTGGCGAGTGTGG ZNF827 NM 178835 2621 TGCCTGAGGACCC 2622 GAGGTGGCGGAGTGA 2623 CCCGCCTTCAGAGAAGAAAC 2624 TGCCTGAGGACCCTCTACCGCCCCCGCCTTCAGAGA ZWINT NM 007057 2625 TAGAGGCCATCAA 2626 TCCGTTTCCTCTGGGC 2627 ACCAAGGCCCTGACTCAGAT 2628 TAGAGGCCATCAAAATTGGCCTCACCAAGGCCCTGA 2015227398 15 Sep 2015

TABLEB 2015227398 15 Sep 2015 nicroRNA_! isa-miR-1 isa-miR-103 isa-miR-106b isa-miR-10a isa-miR-133a isa-miR-141 isa-miR-145 isa-miR-146b-5p isa-miR-150 isa-miR-152 isa-miR-155 isa-miR-182 isa-miR-191 isa-miR-19b isa-miR-200c isa-miR-205 isa-miR-206 isa-miR-21 isa-miR-210 isa-miR-22 isa-miR-222 isa-miR-26a isa-miR-27a isa-miR-27b isa-miR-29b isa-miR-30a isa-miR-30e-5p isa-miR-31 isa-miR-331 isa-miR-425 hsa-miR-449a hsa-miR-486-5p hsa-miR-92a hsa-miR-93 hsa-miR-99a

Sequence_ UGGAAUGUAAAGAAGUAUGUAU GCAGCAUUGUACAGGGCUAUGA UAAAGUGCUGACAGUGCAGAU UACCCUGUAGAUCCGAAUUUGUG UUUGGUCCCCUUCAACCAGCUG UAACACUGUCUGGUAAAGAUGG GUCCAGUUUUCCCAGGAAUCCCU UGAGAACUGAAUUCCAUAGGCU UCUCCCAACCCUUGUACCAGUG UCAGUGCAUGACAGAACUUGG UUAAUGCUAAUCGUGAUAGGGGU UUUGGCAAUGGUAGAACUCACACU CAACGGAAUCCCAAAAGCAGCUG UGUAAACAUCCUCGACUGGAAG UAAUACUGCCGGGUAAUGAUGGA UCCUUCAUUCCACCGGAGUCUG UGGAAUGUAAGGAAGUGUGUGG UAGCUUAUCAGACUGAUGUUGA CUGUGCGUGUGACAGCGGCUGA AAGCUGCCAGUUGAAGAACUGU AGCUACAUCUGGCUACUGGGU UUCAAGUAAUCCAGGAUAGGCU UUCACAGUGGCUAAGUUCCGC UUCACAGUGGCUAAGUUCUGC UAGCACCAUUUGAAAUCAGUGUU CUUUCAGUCGGAUGUUUGCAGC CUUUCAGUCGGAUGUUUACAGC AGGCAAGAUGCUGGCAUAGCU GCCCCUGGGCCUAUCCUAGAA AAUGACACGAUCACUCCCGUUGA UGGCAGUGUAUUGUUAGCUGGU UCCUGUACUGAGCUGCCCCGAG UAUUGCACUUGUCCCGGCCUGU CAAAGUGCUGUUCGUGCAGGUAG AACCCGUAGAUCCGAUCUUGUG SEQID NO 2629 2630 2631 2632 2633 2634 2635 2636 2637 2638 2639 2640 2641 2642 2643 2644 2645 2646 2647 2648 2649 2650 2651 2652 2653 2654 2655 2656 2657 2658 2659 2660 2661 2662 2663 139 [00137] Throughout this specification, unless the context requires otherwise, the word "comprise", or variations such as "comprises" or "comprising", will be understood to imply the inclusion of a stated step or element or integer or group of steps or elements or integers but not the exclusion of any other step or element or integer or group of elements or integers. 2015227398 06 Μ 2017 [00138] Any discussion of documents, acts, materials, devices, articles or the like which has been included in the present specification is not to be taken as an admission that any or all of these matters form part of the prior art base or were common general knowledge in the field relevant to the present disclosure as it existed before the priority date of each of the appended claims.

139A

Claims (16)

1. A method for determining a likelihood of cancer recurrence in a human patient with prostate cancer, comprising: measuring an expression level of an RNA transcript of BGN in a biological sample comprising prostate tissue obtained from the patient: predicting a likelihood of cancer recurrence for the patient based on the measured expression level of the RNA transcript of BGN; wherein an increased expression level of BGN RNA is positively associated with a increased risk of recurrence.

2. The method of claim 1, further comprising normalizing said RNA expression level to obtain a normalized BGN RNA expression level, wherein an increased expression level of normalized BGN RNA is positively associated with a increased risk of recurrence.

3. The method of any one of claims 1 or 2, further comprising generating a report based on a Recurrence Score (RS).

4. The method of any one of claims 1-3, wherein the likelihood of cancer recurrence is based on clinical recurrence-free interval (cRFI).

5. The method of any one of claims 1-4, wherein the likelihood of cancer recurrence is based on biochemical recurrence-free interval (bRFl).

6. The method of any one of claims 1-5, wherein the biological sample has a positive TMPRSS2 fusion status.

7. The method of any one of claims 1-5, wherein the biological sample has a negative TMPRSS2 fusion status.

8. The method of any one of claims 1-7, wherein the patient has early-stage prostate cancer.

9. The method of any one of claims 1-8, wherein the biological sample comprises prostate tumor tissue with the primary Gleason pattern for said prostate tumor.

10. The method of any one of claims 1-8, wherein the biological samples comprises prostate tumor tissue with the highest Gleason pattern for said prostate tumor.

11. The method of any one of claims 1-9, wherein the biological sample is prostate tumor tissue.

12. The method of any one of claims 1-6, wherein the biological sample is non-tumor prostate tissue.

13. The method of any one of claims 1-5, further comprising classifying the patient as TMPRSS2 fusion positive or negative, wherein an increased expression level of AZGPl is associated with a negative TMPRSS2 fusion status.

14. The method of any one of claims 1-5, wherein the biological sample comprises non-tumor prostate tissue, and further comprising measuring an expression level of INHBA, wherein an increased expression level of 1NHBA is negatively associated with good prognosis.

15. The method of any one of claims 1-14, further comprising determining a likelihood of upgrading or upstaging in the patient with prostate cancer, wherein an increased expression level of BGN RNA is positively associated with a increased risk of upgrading/upstaging.

16. The method of any one of claims 1-15, wherein the biological sample is a fixed, paraffin-embedded tissue sample.