AU2008202159B2 - Compositions of ezetimibe and methods for the treatment of cholesterol-associated benign and malignant tumors - Google Patents

Description

P/00/011 Regulation 3.2 AUSTRALIA Patents Act 1990 ORIGINAL COMPLETE SPECIFICATION STANDARD PATENT Invention Title: "COMPOSITIONS OF EZETIMIBE AND METHODS FOR THE TREATMENT OF CHOLESTEROL-ASSOCIATED BENIGN AND MALIGNANT TUMORS" The following statement is a full description of this invention, including the best method of performing it known to me/us: -1 COMPOSITIONS OF EZETIMIBE AND METHODS FOR THE TREATMENT OF CHOLESTEROL-ASSOCIATED BENIGN AND MALIGNANT TUMORS 5 FIELD OF THE INVENTION The invention relates to the prevention and/or treatment of cholesterol-associated tumors by means of administering azetidinone-based cholesterol absorption inhibitors. Particularly, hypertrophy of prostate tissues, breast tissues, endometrial tissues, and colon tissues are controlled by the oral administration of a therapeutically effective amount of 10 ezetimibe, for example, or its phenolic glucuronide. Combinations of ezetimibe, and/or at least one of its analogs efficacious in controlling the absorption of cholesterol, and at least one other anticancer agent selected from the group consisting of (a steroidal antiandrogen, a non steroidal antiandrogen, an estrogen, diethylstilbestrol, a conjugated estrogen, a selective estrogen receptor modulator (SERM), a taxane, and a LHRH analog) are 15 provided for the prevention or treatment of cholesterol-associated tumors. BACKGROUND OF THE INVENTION Although cholesterol metabolism has been studied extensively in the liver and intestinal tract of humans and experimental animals little attention has been directed to the 20 cholesterol metabolism in the male prostate gland and the female mammary gland in both their normal and pathologic diseased states. The etiology and progression for benign and malignant tumors of these glands still remains largely a mystery. Cholesterol-rich diets have had a significant epidemiological association with the variety of human cancer diseases. Particularly, cancers of the prostate and mammary glands and of the colon have 25 been linked to high-fat "western" diets including the intake of fat of animal origin. Kolonel, et aL., 1999, J. NatI. Cancer Inst. 91:414-428; Willett, 1989, Nature 338:389 394. The mechanisms, however, by which these cancers are initiated and progress, as related to the dietary fat, are poorly understood. The polyene macrolides and in particular, the aromatic heptaene macrolide, 30 candicidin, have been in clinical use for the treatment of human benign prostatic -- 2 hyperplasia for many years in several countries. Various other hypocholesterolemic drugs that interfere with cholesterol absorption and resorption in the gastrointestinal tract have also been in clinical use for the same human prostatic disease. Schaffner, 1983, in: "Benign Prostatic Hypertrophy", Frank Hinman, Jr. ed. Springer-Velag, New York, 5 pp.280-307 reviewed clinical studies with candicidin and other polyene macrolides. Candicidin in long-term rat studies has been shown to inhibit tumor initiation and progression as compared to untreated controls. Haditirto, 1974, Ph.D. Dissertation, Rutgers University. Other inhibitors of cholesterol absorption - resorption include the bile acid sequestering anionic exchange resins such as Cholestyramine@ and Colestipol@. 0 These have also been shown to alter the course of prostatic disease in animals and humans. Colestipol@ inhibited benign prostatic hypertrophy in hamsters. Wang. et al., 1976, Investigative Urol. 14:66-71. Cholestyramine@ has been shown to be effective in some patients with prostatic carcinoma. Addleman, 1972, N. England J. Med., 287:1047. As hypocholesterolemic drugs, the phytosterols, beta-sitosterol and stigmasterol, for example, .5 are also known for their ability to inhibit cholesterol absorption and resorption by a mass action effect requiring large doses. In a controlled double blind study beta-sitosterol was found to be effective in the treatment of benign prostatic hyperplasia. Ebbinghaus et al., 1977, Z. Allg.Med., 53:1054-1058. It has been approved for human use.in Europe. The phytosterols are also components of a variety of herbal medicines prescribed for the !0 treatment of prostate disease. Extracts of the berries of the plant, saw palmetto, and the bark of Pygeum africanum, also known as Tadenan, for example, contain significant quantities of beta-sitosterol. SUMMARY OF THE INVENTION 25 The present invention is directed to methods of prevention or treatment of a cholesterol-associated tumor comprising administering a therapeutically effective amount of an azetidinone-based cholesterol absorption inhibitor particularly ezetimibe, one of its analogs or its phenolic glucuronide to a patient wherein the patient is either at risk of developing a cholesterol-associated tumor or already exhibits a cholesterol-associated 30 tumor.

-3 In addition, the current invention is directed to methods of using ezetirnibe to prevent or treat a cholesterol-associated tumor selected from the group consisting of either benign or malignant tumors of the prostate, breast, endometrium and colon. 5 The invention is further directed to methods and compositions fog co-administering ezetimibe and at least one other anticancer agent selected from the group consisting of a steroidal antiandrogen, a non-steroidal antiandrogen, an estrogen, diethylstilbestrol, a conjugated estrogen, a selective estrogen receptor modulator (SERM), a taxane, and a t0 LHRH analog for the prevention or treatment of a cholesterol-associated tumor. Further the invention is directed to an article of manufacture comprising indication labeling; particularly, an article of manufacture comprising a container, instructions, and a composition, wherein the composition comprises a therapeutically effective amount of an 15 azetidinone-based cholesterol absorption inhibitor, preferably ezetimibe, and the instructions are for the administration of the composition for the prevention or treatment of a cholesterol-associated tumor. DETAILED DESCRIPTION OF THE INVENTION 20 Unless defined otherwise, all technical and scientific terms used herein have the same meaning as is commonly understood by one of skill in the art to which this invention belongs. All publications and patents referred to herein are incorporated by reference. cholesterol mediation of disease conditions 25 The epoxycholesterols, for example, have long been attributed to have mitogenic, mutagenic, carcinogenic and cyotoxic properties. The in vivo level of epoxycholesterols as oxidized metabolites of cholesterol in the male prostate and female mammary gland, for example, is the direct result of cholesterol content. Particularly, cholesterol and its metabolites including cholesterol epoxides (epoxycholesterols), e.g., cholesterol 59 and 30 6fl-epoxide, are related to and mediate disease processes, particularly benign and/or -4 malignant cholesterol-associated tumors or otherwise cholesterol-associated abnormal or cancerous cell growth or cell-mass including but not limited to tumors associated with prostate, breast, endometrial, and colon tissues. In these tissues the epoxycholesterols also serve as a marker of benign and malignant diseases. 5 prostate The cholesterol content of the human prostate gland tissues, for example, doubles with the appearance of prostate tumors. Swyer, 1942, Cancer Res., 2:372-375; Schaffner, C.P., et al., Cancer Detect. Prevent., 1980, vol.3, p 143. Further, the appearance of the cholesterol epoxides 10 has been reported in the tissues and secretions of diseased human prostate glands confirming the doubling of tissue cholesterol content and the significant content of the epoxycholesterols with the diagnosis of human prostatic hyperplasia and carcinoma. Sporer et al., 1982, Urology, 6:244 250. 15 breast and endometrial. Studies with the human female mammary gland also report significant increases of cholesterol in the breast fluid aspirates and the simultaneous appearance of the epoxycholesterols, for example, with the aging female human mammary gland. Particularly, the appearance of isometric epoxycholesterols, e.g., beta-epoxycholesterol, is correlated to benign 20 and malignant breast tumors. Petrakis, et al., 1981, Cancer Res., 41:2563-2565; Wrench et al., 1989, Cancer Res., 49:2168-2174. Elevated beta-epoxycholesterol is also detected, for example, in the plasma of endometrial cancer patients. Kucuk, et al., 1994, Can Epidemiol. Biomark. Prevention, 3:57 1-574. The appearance of the epoxycholesterols, e.g., beta-epoxycholesterol, is directly related to the increase of cholesterol in body fluids and tissues. 25 azetidinone-based cholesterol absorption inhibitors The present invention is directed toward compositions of azetidinone-based cholesterol absorption inhibitors, e.g., ezetimibe and its glucuronides and its analogs, for the reduction of cholesterol levels in vivo and reduces epoxycholesterol formation and the initiation and 30 progression of benign and malignant tumors and methods of use therefore. The compositions and -5 methods of the invention are particularly for the prevention or control or treatment of benign or malignant cholesterol-associated tumors or cholesterol-associated cell growth or cell-masses including but not limited to tumors associated with prostate, colon, endometrial, or breast tissues - or - prostate, colon, breast, or endometrial cancer. Methods are provided for the prevention and 5 treatment of cholesterol-associated tumors by the inhibition of their initiation and progression by the inhibition of cholesterol absorption and resorption in the gastrointestinal tract. Oral administration of ezetimibe compositions disclosed herein, for example, are preferred embodiments of the present invention for the treatment and/or prevention of benign prostatic hypertrophy or other cholesterol-related benign or malignant tumors, for example, associated .0 with prostate, breast, endometrial or colon tissues. Ezetimibe by the oral route, for example, has a significant effect on the initiation and progression of prostatic disease and other cholesterol-associated tumors, for example, in experimental animals. 'The BIO 87.20 male Syrian hamster is a well-known inbred line that .5 develops, spontaneously, an age-dependent and genetic related cystic prostatic hypertrophy and is recognized in the art as an excellent model for human prostate disease. See, Examples I-rn, infra. Ezetimibe by the oral route inhibits the development of cystic prostatic hypertrophy in the BIO 87.20 male Syrian hamster. Ezetimibe also inhibits other cholesterol-associated tumor formation in this animal disease model. Treatment of BIO 87.20 male Syrian hamsters with !0 ezetimibe, beginning at 6 months of age, inhibits prostatic enlargement. BIO 87.20 male Syrian hamsters treated with ezetimibe, beginning at 12 months of age (when the prostatic enlargement is already in progress), reduces the prostatic mass or volume. Higher doses of ezetimibe, for example, at the 1000ig/kg body weight have a greater effect on the inhibition and reversal of the prostate enlargement of the BIO 87.20 hamster. Further, histopathological examination of the 25 prostates, for example, of the ezetimibe treated BIO 87.20 hamsters show a more normal histology, as seen in prostate sections of the BIO 1.5 hamsters. The BIO 87.20 male Syrian hamster is noted for the emergence of tumors throughout the body at 18 months of age or older. At 18 months the BIO 87.20 hamsters demonstrate the presence of cholesterol-associated tumors in addition to that of the prostate gland. The BIO 87.20 control animals of Example II herein, for -6 example, exhibit several different tumors at the end of the experiment; whereas, the BIO 87.20 ezetimibe-treated animals of the same age exhibit no tumors. Prostate cancer is a leading cause of mortality in males in North America with between 5 30,000 to 40,000 deaths per year in the United States. While chemotherapy has not been very efficacious in the treatment of prostate cancer, an alternative approach has been to target prostate cancer cell survival pathways, particularly the P13 kinase/Akt/PTEN signaling axis, which has been identified as an important cell survival mechanism in PCa and other cancers. Recently it has been determined that membrane cholesterol is an important component of a mechanism that 10 transfers survival signals from the cell exterior to the Akt1 serine-threonine kinase. It has also been shown recently that elevated serum cholesterol accelerates the growth, lowers the extent of cellular apotosis, and increases the level of Akt activation in prostate tumors within a murine prostate cancer model. Findings indicate that cholesterol plays a critical role in the ability of prostate cancer cells to resist apoptotic stimuli. The role of cholesterol in prostate cancer cell 15 survival is so critical that drugs that bind cholesterol (polyene macrolides), extract cholesterol from membranes (cyclodextrin) or block cholesterol synthesis (statins) all alter essential membranes (lipid rafts) and reduce their capacity to regulate cell signaling. Cholesterol plays a crucial role in specifically regulating prostate cancer growth and survival. In these studies a unique prostate murine model system is used. This model features the human prostate 20 adenocarcinoma cell line, LNCaP transfected with HB-EGF. LNCaP cells resemble typical prostate tumor cells in their general morphology, production of PSA and PTEN null status. Stable transfection with HB-EGF, a physiologically relevant EGFR ligand that originates in the prostatic stroma, permits LNCaP cells to form tumors in vivo in an androgen -independent manner. Subcutaneous implantation of these LNCaP cells into 4 quadrants of SCID mice results 25 in rapid tumor growth in intact and castrated mice over an eight week period. Ezetimibe and related compounds SCH 48461 and SCH 58053 do not generally affect serum cholesterol levels in mice fed a low cholesterol diet, but do lower elevated serum cholesterol levels induced by high cholesterol diets. Mice placed on a high cholesterol diet 4 weeks prior to'tumor implantation are started on ezetimibe and SCH 48461 @ 30mg/kg of body weight 2 weeks after 30 tumor implantation. After 12 weeks the evaluation of tumor volumes in each mouse as -7 compared to the untreated controls revealed that drug treatment significantly inhibits the progression and growth of the implanted tumors. Cholesterol-associated tumor as used herein refers to benign or malignant tumors or otherwise cholesterol-associated abnormal or cancerous cell growth or cell-mass including but not limited to tumors associated with prostate (for example, prostatic hyperplasia) tissue, colon tissue, breast tissue, or endometrial tissue - or - prostate cancer, colon cancer, breast cancer, or endometrial cancer. ) Azetidinone-based cholesterol absorption inhibitors, for example, are described by Rosenblum, S.B., et al., J. Med. Chem., 41(6):973-80 (1998)). Azetidinone-based compounds are potent, orally active inhibitors of cholesterol absorption. Bioorg. Med. Chem., 7(10):2199-202 (1999). A particularly preferred azetidinone-based compound for use in compositions and methods of the present invention is ezetimibe (l-(4-fluorophenyl)-(3R)-[3 -(4-fluorophenyl)-(3 5 S)-hydroxypropyl]-(45)-(4-hydroxyphenyl)-2-azetidinone) (also referred to in the literature as SCH 58235 or ZETIA@) and its phenolic glucuronide, SCH60663. Br. J. Pharmacol., 129(8):1748-54 (2000). Two other ezetimibe related analogs and cholesterol absorption inhibitors for use in compositions and methods of the present invention, for example, are referred to in the literature as: 1) SCH 58053 or (+)-7-(4-chlorophenyl)-2-(4-flourophenyl)-7-hydroxy 3 3R-(4-hydroxyphenyl)-2-azaspiro[3,5] nonan-1-one) J. Lipid Res., 43:1864-1873(2002) and 2). SCH 48461 or (3R)-3Phenylpropyl)-1.(4S)-bis(4-methoxvphenyl)-2-azetidinone. J Med. Chem., 41:973-980 (1998) Ezetimibe's mode of action involves the inhibition of cholesterol absorption and Z5 resorption in the intestinal tract. This mechanism of action also involves the increased excretions of cholesterol and its intestinal generated metabolites with the feces. This effect of ezetinibe results in lowered body cholesterol levels, increased cholesterol synthesis, and decreased triglyceride synthesis. The increased cholesterol synthesis initially provides for the maintenance of cholesterol levels in the circulation, levels that eventually decline as the inhibition of -8 cholesterol absorption and resorption continues. The overall effect of drug action is the lowering of cholesterol levels in the circulation and tissues of the body. A preferred azetidinone-based cholesterol absorption inhibitors for use in compositions 5 and methods of the present invention is ezetimibe or a stereoisomeric mixture thereof, diastereomerically enriched, diastereomerically pure, enantiomerically enriched or enantiomerically pure isomer thereof, or a prodrug of such compound, mixture or isomer thereof, or a pharmaceutically acceptable salt of the compound, mixture, isomer or prodrug. Lo Another preferred azetidinone-based cholesterol absorption inhibitors is the phenolic glucuronide of ezetimibe (Br. J. Pharmacol., 129(8): 1748-54 (2000)) or a stereoisomeric mixture thereof, diastereomerically enriched, diastereomerically pure, enantiomerically enriched or enantiomerically pure isomer thereof, or a prodrug of such compound, mixture or isomer thereof, or a pharmaceutically acceptable salt of the compound, mixture, isomer or prodrug. 15 The expression "prodrug" as used herein refers to compounds that are drug precursors which following administration, release the drug in vivo via chemical or physiological process (e.g., a prodrug on being brought to the physiological pH is converted to the desired drug form). Exemplary prodrugs upon cleavage release the corresponding free acid. For example, by means 20 of hydrolyzable ester-forming residues of the compounds. Compositions of the invention basically comprise an effective dose or a pharmaceutically effective amount or a therapeutically effective amount of an azetidinone based cholesterol absorption inhibitor, preferably ezetimibe and/or its phenolic glucuronide or at least one 25 ezetimibe pharmacologically active analog, to prevent, or control the growth, or reduce the size of benign prostatic hypertrophy or other cholesterol-related benign or malignant tumors, for example, associated with prostate, breast, endometrial or colon tissues. Compositions described herein comprise azetidinone-based cholesterol absorption 30 inhibitors, preferably ezetimibe its phenolic glucuronide, or one of its analogs and may further -- 9 comprise at least one other anticancer agent. These compositions are preferably orally administered. Solid dosage forms for oral administration include capsules, tablets, pills, powders and granules and for companion animals the solid dosage forms include an admixture with food and chewable forms. In such solid dosage forms, the active compound is admixed with at least 5 one inert pharmaceutically acceptable carrier such as sucrose, lactose, or starch. Such dosage forms can also comprise, as is normal practice, additional substances other than such inert diluents, e.g., lubricating agents such as magnesium stearate. In the case of capsules, tablets and pills, the dosage forms may also comprise buffering agents. Tablets and pills can additionally be prepared with enteric coatings. In the case of chewable forms, the dosage form may comprise 10 flavoring agents and perfuming agents. The dosage of active ingredient in the compositions of this invention may be varied; however, it is necessary that the amount of the active ingredient be such that a suitable dosage form is obtained. The selected dosage depends upon the desired therapeutic effect, on the route 15 of administration, and on the duration of the treatment. Generally, dosage levels of between about 100pzg to about 200pg/kg of body weight daily are administered to humans and other animals, e.g., mammals, to obtain effective release of ezetimibe for methods described herein. Oral administration of ezetimibe, for example, inhibits both dietary and biliary cholesterol absorption and resorption in the intestinal tract, thereby lowering serum cholesterol levels as 20 associated with the reduced low density lipoprotein (LDL) levels and increased high density lipoprotein (HDL) levels. The preferred dosage range of ezetimibe in compositions for administration to a patient in need of prevention or treatment described herein is from about 5 mg to about 150 mg per day. A more preferred range is from about 5mg to about 100 mg per day. An even more preferred range is from about 8 mg to about 50 mg per day. A most preferred 25 range is from about 10mg to about 25 mg per day. A composition for oral administration which comprises about 10mg ezetimibe for a single daily dosage to prevent, or control the growth, or reduce the size of benign prostatic hypertrophy or other cholesterol-related benign or malignant tumors, for example, associated with prostate, breast, endometrial or colon tissues, is a particularly preferred embodiment of the present invention. A composition for oral 30 administration which comprises about 15mg ezetimibe for a single daily dosage to prevent, or -10 control the growth, or reduce the size of benign prostatic hypertrophy or other cholesterol-related benign or malignant tumors, for example, associated with prostate, breast, endometrial or colon tissues is another preferred embodiment of the present invention. A composition for oral administration which comprises about 20mg ezetimibe for a single daily dosage to prevent, or 5 control the growth, or reduce the size of benign prostatic hypertrophy or other cholesterol-related benign or malignant tumors, for example, associated with prostate, breast, endometrial or colon tissues is another preferred embodiment of the present invention. A composition for oral administration which comprises about 25mg ezetimibe for a single daily dosage to prevent, or control the growth, or reduce the size of benign prostatic hypertrophy or other cholesterol-related 10 benign or malignant tumors, for example, associated with prostate, breast, endometrial or colon tissues is another preferred embodiment of the present invention. A composition for oral administration which comprises about 30mg ezetimibe for a single daily dosage to prevent, or control the growth, or reduce the size of benign prostatic hypertrophy or other cholesterol-related benign or malignant tumors, for example, associated with prostate, breast, endometrial or colon 15 tissues is another preferred embodiment of the present invention. A composition for oral administration which comprises about 35mg ezetimibe for a single daily dosage to prevent, or control the growth, or reduce the size of benign prostatic hypertrophy or other cholesterol-related benign or malignant tumors, for example, associated with prostate, breast, endometrial or colon tissues is another preferred embodiment of the present invention. A composition for oral 20 administration which comprises about 40mg ezetimibe for a single daily dosage to prevent, or control the growth, or reduce the size of benign prostatic hypertrophy or other cholesterol-related benign or malignant tumors, for example, associated with prostate, breast, endometrial or colon tissues - is another preferred embodiment of the present invention. A composition for oral administration which comprises about 45mg ezetimibe for a single daily dosage to prevent, or 25 control the growth, or reduce the size of benign prostatic hypertrophy or other cholesterol-related benign or malignant tumors, for example, associated with prostate, breast, endometrial or colon tissues - is another preferred embodiment of the present invention. These compounds can be administered by any means known in the art. Such modes include oral, rectal, nasal, topical (including buccal and sublingual) or parenteral (including 30 subcutaneous, intramuscular, intravenous and intradermal) administration.

-11 For ease to the patient oral administration is preferred. However, as practiced by those skilled in the art other routes of administration may be necessary. Thus, depending upon the situation - the skilled artisan must determine which form of administration is best in a particular 5 case - balancing dose needed versus the number of times per month administration is necessary. combination therapy Compositions of the invention comprise an effective dose or a pharmaceutically effective amount or a therapeutically effective amount of an azetidinone-based cholesterol absorption 10 inhibitor, preferably ezetimibe or its phenolic glucuronide, and at least one other anticancer agent, for the treatment or prevention of benign prostatic hypertrophy or other cholesterol-related benign or malignant tumors, for example, associated with prostate, breast, endometrial or colon tissues. Examples of agents for use in compositions and methods of the invention described herein include but are not limited to steroidal or non steroidal antiandrogens (e.g., finasteride 15 (PROSCAR@), cyproterone acetate (CPA), flutamide (4'-nitro-3'-trifluorormethyl isobutyranilide), bicalutamide (CASODEX@), and nilutamide), estrogens, diethylstilbestrol (DES), conjugated estrogens (e.g., PREMARIN@), selective estrogen receptor modulator (SERM) compounds (e.g., tamoxifen, raloxifene, droloxifene, idoxifene), Taxanes (e.g., paclitaxel (TAXOL@), docetaxel (TAXOTERE@)), LHRII analogs (e.g., goserelin acetate 20 (ZOLADEX@), leuprolide acetate (LUPRON@)). taxanes Docetaxel (TAXOTERE@) based regimens, for example, are reported to be treatment options for the management of patients with advanced, androgen-independent prostate cancer. 25 Docetaxel in combination with ezetimibe, for example, should achieve a significant response in treatment or prevention of benign prostatic hypertrophy or other cholesterol-related benign or malignant tumors, for example, associated with prostate, breast, endometrial or colon tissues in patients with measurable disease. See, e.g., Oncology (Huntingt.), 16(6 Suppl. 6):63-72 (2002). Any taxane may be used as an anticancer agent for use in the compositions and methods of this 30 invention.

-12 A preferred method of the invention accordingly comprises orally co-administering to a patient in need of treatment a therapeutically effective amount of an azetidinone-based cholesterol absorption inhibitor, preferably ezetimibe, and a taxane preferably selected from the 5 group consisting essentially of (paclitaxel and docetaxel) or an effective derivative or analog thereof for the treatment or prevention of benign prostatic hypertrophy or other cholesterol related benign or malignant tumors, for example, associated with prostate, breast, endometrial or colon tissues. See, e.g., U.S. Patent No. 6,395,770 Method and compositions for administering taxanes orally to human patients, May 28, 2002; U.S. Patent No. 6,380,405 Taxane Prodrugs, 0 April 30, 2002; U.S. Patent No. 6,239,167 Antitumor compositions containing taxane derivatives, May 29, 2001. SERMs An azetidinone-based cholesterol absorption inhibitor, preferably ezetimibe, may be 5 combined with a mammalian selective estrogen receptor modulator (SERM) to prevent, or control the growth, or reduce the size of benign pro static hypertrophy or other cholesterol related benign or malignant tumors, for example, associated with prostate, breast, endometrial or colon tissues. Any SERM may be used as an anticancer agent for use in the compositions and methods of this invention. The term selective estrogen receptor modulator includes both estrogen 0 agonist and estrogen antagonists and refers to compounds that bind with the estrogen receptor, inhibit bone turnover and prevent bone loss. In particular, estrogen agonists are herein defined as chemical compounds capable of binding to the estrogen receptor sites in mammalian tissue, and mimicking the actions of estrogen in one or more tissue. Estrogen antagonists are herein defined as chemical compounds capable of binding to the estrogen receptor sites in mammalian tissue, 25 and blocking the actions of estrogen in one or more tissues. A preferred SERM is tamoxifen: (ethanamine,2-[-4-(1,2-diphenyl-l-butenyl)phenoxy]-N,N-dimethyl, (Z)-2,2-hydroxy- 1 ,2,3 propanetri-carboxylate (1:1)) and associated compounds which are disclosed in U.S. Pat. No. 4,536,516, the disclosure of which is hereby incorporated by reference. Another related compound is 4-hydroxy tamoxifen which is disclosed in U.S. Pat. No. 4,623,660, the disclosure 30 of which is hereby incorporated by reference. Another preferred SERM is raloxifene: -13 (methanone, [6-hydroxy-2-(4-hydroxyphenyl) benzo[b]thien-3-yl] [4-[2-( 1 piperidinyl)ethoxy]phenyl]-, hydrochloride) and associated compounds which are disclosed in U.S. Pat. No. 4,418,068, the disclosure of which is hereby incorporated by reference. Another preferred SERM is idoxifene: Pyrrolidine, 1-1-[4-[-1 -(4-iodophenyi)-2-phenyl- 1 5 Butenyl]phenoxy]ethyl] and associated compounds which are disclosed in U.S. Pat. No. 4,839,155, the disclosure of which is hereby incorporated by reference. In particular, an effective dosage for droloxifene is in the range of 0.1 to 40 mg/kg/day, preferably 0.1 to 5 mg/kg/day. In particular, an effective dosage for raloxifene is in the range of 10 0.1 to 100 mg/kg/day, preferably 0.1 to 10 mg/kg/day. In particular, an effective dosage for tamoxifen is in the range of 0.1 to 100 mg/kg/day, preferably 0.1 to 5 mg/kg/day. In particular, an effective dosage for 4-hydroxy tamoxifen is in the range of 0.0001 to 100 mg/kg/day, preferably 0.001 to 10 mg/kg/day. 15 A preferred method of the invention accordingly comprises orally co-administering to a patient in need of treatment a therapeutically effective amount of an azetidinone-based cholesterol absorption inhibitor, preferably ezetimibe, and a SERM selected from the group consisting essentially of(tamoxifen, raloxifene, droloxifene, and idoxifene) or an effective derivative or analog thereof for the treatment or prevention of benign prostatic hypertrophy or 20 other cholesterol-related benign or malignant tumors, for example, associated with prostate, breast, endometrial or colon tissues. See, e.g., U.S. Pat. No. 5,047,431, U.S. Pat. No. 6,245,352 and U.S. Pat. No. 5,972,383 the disclosures of which are hereby incorporated by reference. steroidal or non steroidal antiandrogens 25 An azetidinone-based cholesterol absorption inhibitor, preferably ezetimibe, may be combined with a steroidal or non steroidal antiandrogen to prevent, or control the growth, or reduce the size of benign prostatic hypertrophy or other cholesterol-related benign or malignant tumors, for example, associated with prostate, breast, endometrial or colon tissues. Any steroidal or non-steroidal antiandrogen may be used as the second compound of this invention. See, e.g., 30 U.S. Patent No. 5,610,150, and U.S. Patent No. 6,015,806.

-14 A preferred method of the invention accordingly comprises orally co-administering to a patient in need of treatment a therapeutically effective amount of an azetidinone-based cholesterol absorption inhibitor, preferably ezetimibe, and a steroidal or non steroidal antiandrogen selected from the group consisting essentially of (finasteride (PROSCAR@)), 5 cyproterone acetate (CPA), flutamide (4'-nitro-3 '-trifluorormethyl isobutyranilide), bicalutamide (CASODEX@), and nilutamide) or an effective derivative or analog thereof for the treatment or prevention of benign pro static hypertrophy or other cholesterol-related benign or malignant tumors, for example, associated with prostate, breast, endometrial or colon tissues. 10 Finasteride (PROSCAR@), in an amount of between about 1mg to about 10mg, preferably about 5mg, may be orally co-administered in a pharmaceutical composition which further comprises about 10mg ezetimibe, for example, for a single daily dosage, to prevent or control the growth, or reduce the size of benign prostatic hypertrophy or other cholesterol related benign or malignant tumors, for example, associated with prostate. 15 luteinizing hormone releasing hormone (LHRH) analog or agonist An azetidinone-based cholesterol absorption inhibitor, preferably ezetimibe, may be co administered with a luteinizing hormone releasing hormone (LHRH) analog or agonist to prevent, or control the growth, or reduce the size of benign prostatic hypertrophy or other 20 cholesterol-related benign or malignant tumors, for example, associated with prostate, breast, endometrial or colon tissues. Any LHRH analog or agonist may be used as the second compound of this invention. A preferred method of the invention accordingly comprises orally co-administering to a 25 patient in need of treatment a therapeutically effective amount of an azetidinone-based cholesterol absorption inhibitor, preferably ezetimibe, and a LHRH analog or agonist selected from the group consisting essentially of (goserelin acetate (ZOLADEX@) and leuprolide acetate (LUPRON@)) or an effective derivative or analog thereof for the treatment or prevention of benign prostatic hypertrophy or other cholesterol-related benign or malignant tumors, for 30 example, associated with prostate, breast, endometrial or colon tissues.

-t5 estrogens, diethylstilbestrol (DES), conjugated estrogens (e.g., PREMARIN@) An azetidinone-based cholesterol absorption inhibitor, preferably ezetimibe, may be co administered with an estrogen, diethylstilbestrol (DES), or conjugated estrogen to prevent, 5 or control the growth, or reduce the size of benign prostatic hypertrophy or other cholesterol related benign or malignant tumors, for example, associated with prostate, breast, endometrial or colon tissues. A preferred method of the invention accordingly comprises orally co-administering to a 10 patient in need of treatment a therapeutically effective amount of an azetidinone-based cholesterol absorption inhibitor, preferably ezetimibe, and a estrogen, diethyistilbestrol (DES), or conjugated estrogen e.g., PREMARIN@ or an effective derivative or analog thereof for the treatment or prevention of benign prostatic hypertrophy or other cholesterol- related benign or malignant tumors, for example, associated with prostate, breast, endometrial or colon tissues. 15 Article of Manufacture An article of manufacture is provided which comprises a container, e.g., a vial, written instructions, and a formulated composition, wherein the composition comprises a therapeutically effective amount of an azetidinone-based cholesterol absorption inhibitor, and the instructions 20 are for - or - indicate the administration of the composition for the prevention or treatment of a cholesterol-associated tumor, e.g., prostate tumor, breast tumor, endometrial tumor, and/or colon tumor. A preferred article of manufacture comprises ezetimibe as the azetidinone-based cholesterol absorption inhibitor. Another preferred article of manufacture so described further comprises at least one other anticancer agent, e.g., a steroidal antiandrogen, a non-steroidal 25 antiandrogen, an estrogen, diethylstilbestrol, a conjugated estrogen, a selective estrogen receptor modulator (SERM), a taxane, and/or a LHRH analog. 30 -16 EXAMPLES EXAMPLE I Experimental animal groups 5 The BIO 87.20 male Syrian hamster is a well-known inbred line that develops, spontaneously, an age-dependent and genetic related cystic prostatic hypertrophy and is recognized in the art as an excellent model for human prostate disease. See, e.g., Homburger et al., 1970, Proc. Soc. Exptl. Biol. Med., 134:284-286; Homburger, 1972, Health Lab Sci., 9:103 111; Wang et al., 1976, Invest. Urol., 14:66-71. The BIO 87.20 strain of hamster develops 10 multiple tumors after 12 months of age. Histopathological examination of the enlarged prostate of the BIO 87.20 male Syrian hamster generally reveals a cystic dilation of the prostatic acini, which are filled with eosinophilic amorphous material. The stromal hyperplasia and changes in the epithelial cells are 15 also observed upon microscopic examination. Control male Syrian hamster such as the BIO 1.5 strain do not generally develop tumors at 12 months or older. The BIO 87,20 and BIO 1.5 strains of male Syrian hamster are employed in the ezetimibe studies presented herein. BIO 87.20 hamsters maintained on a cholesterol-rich diet revealed on autopsy a marked accumulation of cholesterol in the liver and severe hypercholesterolemia that led to animal death. The study 20 further clearly revealed that whereas hepatic cholesterol synthesis in the normal hamster is under negative feedback control with dietary cholesterol, hepatic cholesterol synthesis in the BIO 87.20 hamster is under no such feedback control. Schaffner et al., Lipids 16:835-840 (1981). The defect in cholesterol synthesis and the development of cystic prostatic hypertrophy in the BIO 87.20 male hamster is related. 25 16 BIO 1.5 and 48 BIO 87.20 male Syrian hamsters are obtained from Bio Breeders, Inc. Boston, Massachusetts. Among these animals, 8 BIO 1.5 and 24 BIO 87.20 hamsters are 6 months of age, while the remainder, consisting of 6 BIO 1.5 and 24 BIO 87.20 hamsters, are 12 months of age. The animals are housed in individual cages. All animals are given water ad 30 libitum and are housed under automatic regime of 12 hours artificial light and 12 hours darkness.

-17 Food consumption is be monitored to ensure equivalent intake in all animals. The drug, ezetimibe, is powdered and mixed into ground PURINA@ hamster chow. Concentrations of ezetimibe, in the food, is adjusted according to quantity of food intake per hamster per day and expressed as average sg of ezetimibe per kg body weight per day. 5 The animals are divided into 8 experimental groups containing 8 animals each. The groups may be characterized as follows: Group I BIO 1.5 Controls, 6 months of age 10 Group II BIO 1.5 Controls, 12 months of age Group III BIO 87.20 Controls, 6 months of age 15 Group IV BIO 87.20 Controls, 12 months of age Group V BIO 87.20 Treated, 6 months of age, ezetimibe @ 100 pg/kg body weight Group VI BIO 87.20 Treated, 6 months of age, ezetimibe @ 1000 pg/kg body weight 20 Group VII BIO 87.20 Treated, 12 months of age, ezetimibe @ 100 pg/kg body weight Group VIII BIO 87.20 Treated, 12 months of age, ezetimibe @ 1000 pg/kg body weight 25 Control BIO 1.5 and control and treated BIO 87.20 male Syrian hamsters, both 6 and 12 months of age, are used as follows. In the treated group of BIO 87.20 male Syrian hamsters, ezetimibe is administered as a powder in the diet. Treatment with doses of ezetimibe of 100 and 1000 micrograms per kilogram body weight of the BIO 87.20 animals is continued for 6 months. Control BIO 1.5 and BIO 87.20 animals do not receive ezetimibe. The food consumption of all 30 groups is monitored, in order to ensure comparable food intake. After 6 months of treatment the -18 animals are sacrificed and the prostate glands excised, weighed and preserved for histopathological examination. At the termination of the experiment, after 6 months, it is evident that all control BIO 5 87.20 hamsters have enlarged prostate glands. This is further evident when the ventral prostate glands are excised and weighed. By sharp contrast none of the BIO 1.5 hamsters, at 12 or 18 months of age show an enlargement of the prostate gland. In the BIO 87.20 control groups the enlargement of the gland is more pronounced at 18 months than at 12 months. The progression of prostatic enlargement is enhanced with time in this control group. Examination of the 10 prepared sections of the BIO 87.20 hamsters, at 12 and 18 months of age, reveal histologically the cystic prostatic hypertrophy that is well-known for this hamster strain. Further, microscopic examination reveals a distended acinus, which is absent in the examination of BIO 1.5 hamster prostate sections. L5 EXAMPLE II Ezetimibe treatment of BIO 87.20 hamsters 6 months of age Six months of ezetimibe treatment of the BIO 87.20 male Syrian hamsters, beginning at 6 months of age shows a marked inhibitory effect on the prostate gland volume as compared to the untreated control BIO 87.20 hamsters. The effect of 1OOOg ezetimibe /kg-body weight !0 demonstrates a greater inhibitory effect on size increase of the prostate gland than with the drug ezetimibe /kg-body weight dose. Upon histopathological examination it is further evident that after 6 months of ezetimibe treatment a marked reduction in the cystic prostatic hypertrophy is seen in treated animals when compared to the untreated control BIO 87.20 animals. After 6 months of ezetimibe treatment the ventral prostate gland weights of the BIO 87.20 animals are 25 similar to those of the BIO 1.5 (control) hamsters (where the development of cystic prostatic hypertrophy is not seen) of the same age. 30 -19 EXAMPLE m Ezetimibe treatment of BIO 87.20 hamsters 12 months of age Cystic prostatic hypertrophy is generally well established in BIO 87.20 male Syrian hamster at 12 months of age. All of the male Syrian hamsters, i.e., control and treated BIO 87.20, 5 and control BIO 1.5 are 12 months old at the beginning of the experiment. Treatment of the BIO 87.20 animals with doses of ezetimibe of 100 and 1000 micrograms per kilogram body weight is continued for.6 months. After the treatment, the excised ventral prostate weights are markedly reduced as compared to untreated BIO 87.20 controls. Further, the level of effect is again significantly greater at the higher dosage of ezetimibe. These results therefore indicate that 10 ezetimibe also reverses the prostatic enlargement once it has taken place. Miscellaneous tumors other than that of the prostate gland are observed in the examination of the untreated BIO 87.20 control hamsters. By contrast, very few tumors are observed in the ezetimibe treated BIO 87.20 animals, which are administered an oral dose at 1000pg/kg-body 15 weight. EXAMPLE IV Ezetimibe treatment of SCID mice transplanted with LNCaP cells transfected with HB-EGF.. 20 Subcutaneous implantations of LNCaP cells into 4 quadrants of SCID mice (2x10' cells in150 ul Matrigel per injection site) results in rapid tumor growth in intact and castrated hosts on a high cholesterol diet over an 8 week period. Two weeks after tumor cell implantation, the animals are given ezetimibe in their diet at 10 mg/kg body weight. After 6-12 weeks of treatment, examination reveals a marked inhibition of tumor growth and progression compared 25 to untreated controls in both intact and castrated animals. 30 -20 EXAMPLE V SCH 48461 treatment of SCID mice transplanted with LNCaP cells transfected with HB-EGF. In a study similar in design as in example IV animals are given SCH 48461 in their diet at 5 10mg/kg body weight. After 6-12 weeks of treatment examination reveal a significant inhibition of tumor growth and progression as compared to untreated controls in both intact and castrated animals. 10 All publications and patents mentioned in the above specification are herein incorporated by reference. Various modifications and variations of the described compositions and methods of the invention will be apparent to those skilled in the art without departing from the scope and spirit of the invention. Although the invention has been described in connection with specific preferred embodiments, it should be understood that the invention as claimed should not be 15 unduly limited to such specific embodiments. Indeed, various modifications of the described compositions and modes for carrying out the invention which are obvious to those skilled in the art or related fields are intended to be within the scope of the following claims.

Claims (20)

1. A method of inhibiting the initiation of a cholesterol-associated tumor comprising administering a therapeutically effective amount of an azetidinone-based cholesterol absorption inhibitor to a patient wherein the 5 patient is either at risk of developing a cholesterol-associated tumor or already exhibits a cholesterol-associated tumor wherein the azetidinone based cholesterol absorption inhibitor is selected from the group consisting of ezetimibe, a glucuronide of ezetimibe, SCH 48461 and SCH 58053.

2. A method according to claim 1 wherein the azetidinone-based 10 cholesterol absorption inhibitor is ezetimibe or a stereoisomeric mixture thereof, diastereomerically enriched, diastereomerically pure, enantiomerically enriched or enantiomerically pure isomer thereof, or a prodrug of such compound, mixture or isomer thereof, or a pharmaceutically acceptable salt of the compound, mixture, isomer or prodrug. 15

3. A method according to claim 1 wherein the cholesterol-associated tumor is selected from the group consisting of benign prostatic hypertrophy, benign breast tumor, benign endometrial tumor, and benign colon tumor.

4. A method according to claim 1 wherein the cholesterol-associated tumor is selected from the group consisting of malignant prostate tumor, 20 breast cancer tumor, endometrial cancer tumor, and colon cancer tumor.

5. A method according to claim 1 wherein a therapeutically effective amount is between 0.1 to 30 mg/kg of body weight daily.

6. A method according to claim 2 wherein a therapeutically effective amount is between 0.1 to 30 mg/kg of body weight daily. 25

7. A method of inhibiting the initiation of a cholesterol-associated tumor comprising co-administering a therapeutically effective amount of an azetidinone-based cholesterol absorption inhibitor and at least one other anticancer agent to a patient wherein the patient is either at risk of developing a cholesterol-associated tumor or already exhibits a cholesterol 30 associated tumor wherein the azetidinone-based cholesterol absorption inhibitor is selected from the group consisting of ezetimibe, a glucuronide of ezetimibe, SCH 48461 and SCH 58053 and at least one other anticancer 22 agent is selected from the group consisting of a steroidal antiandrogen, a non steroidal antiandrogen, an estrogen, diethylstilbestrol, a conjugated estrogen, a selective estrogen receptor modulator (SERM), a taxane, goserelin acetate (ZOLADEX@), and leuprolide acetate (LUPRON@). 5

8. A method according to claim 7 wherein the azetidinone-based cholesterol absorption inhibitor is ezetimibe.

9. A method according to claim 8 wherein at least one other anticancer agent is selected from the group consisting of a steroidal antiandrogen, a non steroidal antiandrogen, an estrogen, diethylstilbestrol, a conjugated 10 estrogen, a selective estrogen receptor modulator (SERM), a taxane, goserelin acetate (ZOLADEX@), and leuprolide acetate LUPRON@).

10. A method according to claim 9 wherein the non steroidal antiandrogen is selected from the group consisting of finasteride (PROSCAR®), flutamide (4'-nitro-3'-trifluorormethyl isobutyranilide), bicalutamide (CASODEX®), and 15 nilutamide.

11. A method according to claim 9 wherein the SERM is selected from the group consisting of tamoxifen, raloxifene, droloxifene, and idoxifene.

12. A method according to claim 9 wherein the taxane is selected from the group consisting of paclitaxel (TAXOL@), and docetaxel (TAXOTERE@). 20

13. A composition for inhibiting the initiation of a cholesterol-associated tumor comprising a therapeutically effective amount of an azetidinone-based cholesterol absorption inhibitor and at least one other anticancer agent wherein the azetidinone-based cholesterol absorption inhibitor is selected from the group consisting of ezetimibe, a glucuronide of ezetimibe, SCH 25 48461 and SCH 58053 and at least one other anticancer agent is selected from the group consisting of a steroidal antiandrogen, a non steroidal antiandrogen, an estrogen, diethylstilbestrol, a conjugated estrogen, a selective estrogen receptor modulator (SERM), a taxane, goserelin acetate (ZOLADEX®), and leuprolide acetate (LUPRON®). 30

14. A composition according to claim 13 wherein the azetidinone-based cholesterol absorption inhibitor is ezetimibe and/or a stereoisomeric mixture thereof, diastereomerically enriched, diastereomerically pure, 23 enantiomerically enriched or enantiomerically pure isomer thereof, or a prodrug of such compound, mixture or isomer thereof, or a pharmaceutically acceptable salt of the compound, mixture, isomer or prodrug.

15. A composition according to claim 14 wherein the non-steroidal 5 antiandrogen is selected from the group consisting of finasteride (PROSCAR@), flutamide (4'-nitro-3'-trifluorormethyl isobutyranilide), bicalutamide (CASODEX@), and nilutamide.

16. A composition according to claim 14 wherein the SERM is selected from the group consisting of tamoxifen, raloxifene, droloxifene, and idoxifene. 10

17. A composition according to claim 14 wherein the taxane is selected from the group consisting of paclitaxel (TAXOL@), and docetaxel (TAXOTERE@).

18. An article of manufacture comprising a container, instructions, and a composition, wherein the composition comprises a therapeutically effective 15 amount of an azetidinone-based cholesterol absorption inhibitor, when used for the administration of the composition to inhibit the initiation of a cholesterol-associated tumor or the control of a malignant cholesterol associated tumor wherein the azetidinone-based cholesterol absorption inhibitor is ezetimibe and/or a stereoisomeric mixture thereof, 20 diastereomerically enriched, diastereomerically pure, enantiomerically enriched or enantiomerically inure isomer thereof, or a prodrug of such compound, mixture or isomer thereof, or a pharmaceutically acceptable salt of the compound, mixture, isomer or prodrug.

19. An article of manufacture according to claim 18 wherein the tumor is 25 selected from the group consisting of a prostatic hypertrophy, a breast tumor, an endometrial tumor, and a colon tumor.

20. An article of manufacture according to claim 18 wherein the composition further comprises at least one other anticancer agent selected from the group consisting of a steroidal antiandrogen, a non-steroidal 30 antiandrogen, an estrogen, diethylstilbestrol, a conjugated estrogen, selective estrogen receptor modulator (SERM@), a taxane, goserelin acetate (ZOLADEX@), and leuprolide acetate (LUPRON®).