NZ617418B2 - Antimicrobial/adjuvant compounds and methods - Google Patents

Abstract

Disclosed are antimicrobial and adjuvant compounds according to Formulae Ia and Ib where the substituents are as defined herein. Also disclosed are prodrugs and pharmaceutically acceptable salts thereof of the compounds and the use of the compounds as antimicrobials and in the manufacture of medicaments for the treatment of microbial infection. ents for the treatment of microbial infection.

Description

ANTIMICROBIAL / ADJUVANT COMPOUNDS AND METHODS CROSS—REFERENCE TO RELATED APPLICATION This application claims the benefit of U.S. Provisional Application Ser. No. 61/448,682, filed March 3, 2011, the t of which is hereby incorporated by reference in its entirety.

BACKGROUND Infection is an invasion of a host organism by a foreign organism, lly to the detriment of normal function in the host. In treating humans and other animals for infection and post-infective atory disease (e.g. septic shock), practitioners usually rely on chemical compounds known to have antibiotic affects, whether antiviral, cterial, antifungal, or the like.

Unfortunately, many ens have become resistant to current otic treatments. Antibiotic resistance is therefore an increasingly significant clinical issue, calling for novel antibiotics. ally valuable would be new antimicrobial adjuvant compounds, which, while not necessarily antimicrobial themselves, can se the potency, efficacy, and/or spectrum of activity of antibiotics when co—administered or given as combination therapy. There is a need for new antimicrobials and new antimicrobial adjuvant compounds.

SUMMARY The compounds ofthe present invention are of utility in antimicrobial infections.

In some cases, the compounds are themselves antimicrobial. In some cases, the nds (“antimicrobial adjuvants”) have beneficial effects in conjunction with an antimicrobial, reducing the dose of antibiotic required for antimicrobial activity when administered in combination. In some cases, the compounds are both antimicrobial and antimicrobial adjuvants.

In l, in an aspect, compounds of a Ia are provided: W0 2012/116452 R10 (la) in which E is ~CH2~ or is absent such that ene is directly connected to phenyl; R1, R2, R9, Rm, and R“ are each independently hydrogen, methyl, ethyl, propyl, cyclopropyl, butyl, cyclobutyl, pentyl, hexyl, isopropryl, isobutyl, neopentyl, methoxy, or ethoxy; additionally, R1 and R2 may connect to form a phenyl or benzofuran ring; additionally, R9 and Rm may connect to form a phenyl or benzofuran ring; R3 and R3 are each independently hydrogen, methyl, ethyl, , isopropyl, chlorine, fluorine, tert—butyl, methylsulfonyl, methoxy, or ethoxy; R4 and R7 are each independently en, chlorine, methyl ester, ethyl ester, methyl, ethyl, propyl, cyclopropyl, bury], cyclobutyl, isopropryl, yl, methoxy, or ethoxy; and R5 and R5 are each independently hydrogen, cyclopentyl, ropyl, furan, thiophene, trifluoromethyl, romethyl ether, lthiol, formaldehyde, chlorine, fluorine, bromine, phenyl, methyl, ethyl, isopropyl, , butyl, cyclobutyl, isobutyl, neOpentyl, pentyl, methoxy, or .

In general, in an aspect of the invention, a method, of treatment of a microbial infection is provided including administering an effective amount of an antimicrobial compound disclosed herein to a patient in need thereof.

In general, in an aspect of the invention, a method of treatment of a microbial infection is provided including administering an effective amount of an antimicrobial W0 2012/116452 adjuvant compound disclosed herein and an effective amount ofan antimicrobial compound to a patient in need thereof.

Compounds of other formulae are provided as described in the Detailed Description below.

BRIEF DESCRIPTION OF GS s thiophene- or thiazole~core compounds and ated compound identifiers of the present invention, along with their respective minimum inhibitory concentrations against MRSA generally following the procedure in Example 9 (“MRSA_MIC” ed in uM). depicts phenyl-core compounds and associated compound identifiers of the present ion.

DETAILED DESCRIPTION Definitions Unless otherwise defined, terms as used in the specification refer to the following definitions, as detailed below.

The term “acyl” as used herein means an alkyl group, as defined herein, ed to the parent molecular moiety through a carbonyl group, as defined herein.

Representative es of acyl include, but are not limited to, , 1~oxopropyl, 2,2- dimethyl—I-oxopropyl, l-oxobutyl, and 1-oxopentyl'.

The term “acyloxy” as used herein means an acyl group, as defined herein, appended to the parent molecular moiety through an oxygen atom. Representative examples of acyloxy include, but are not limited to, acetyioxy, propionyloxy, and isobutyryloxy.

The terms “administration” or “administering” compound should be understood to mean providing a compound of the present invention to an individual in a form that can be introduced into that individual’s body in an amount effective for prophylaxis, treatment, or diagnosis, as applicable. Such forms may include for e oral dosage forms, injectable dosage forms, transdermal dosage forms, inhalation dosage forms, and rectal dosage forms.

The term “alkenyl” as used herein means a straight chain, branched and/or cyclic hydrocarbon having fr0m 2 to 20 (tag, 2 to 10 or 2 to 6) carbon atoms, and including at least one carbOn-carbon double bond. Representative alkenyl moieties include vinyl, allyl, l-butenyl, 2-butenyi, isobutylenyl, 1-pentenyl, Z-pentenyl, 3-methyl-l-butenyl, 2- methyl-Z-butenyl, 2,3—dimethyl—2—butenyl, l—hexenyl, 2—hexeny1, 3~hexenyi, 1—heptenyl, 2-heptenyl, 3—heptenyl, nyl, 2—octenyl, 3-octenyl, 1-nonenyl, 2—nonenyl, 3—nonenyl, nyl, 2-deceny1 and 3-decenyl.

The term “alkoxy” as used herein means an alkyl group, as defined herein, appended to the parent molecular moiety through an oxygen atom. Representative examples of aikoxy include, but are not limited to, methoxy, ethoxy, propoxy, 2—propoxy, butoxy, tert—butoxy, pentyloxy, and hexyloxy.

The term “alkoxyalkoxy” as used herein means an alkoxy group, as defined herein, appended to the parent molecular moiety through another alkoxy group, as defined herein. Representative examples of alkoxyalkoxy include, but are not limited to, tert-butoxymethoxy, 2—ethoxyethoxy, 2-methoxyethoxy, and methoxymethoxy.

The term “alkoxyalkyl” as used herein means an alkoxy group, as defined , appended to the parent molecular moiety through an alkyl group, as defined herein.

Representative es of alkoxyalkyl include, but are not limited to, tert- butoxymethyl, 2-ethoxyethyl, oxyethy1, and methoxymethyl.

The term “alkoxycarbonyl” as used herein means an alkoxy group, as defined herein, appended to the parent molecular moiety through a carbonyl group, as defined herein. Representative examples of alkoxycarbonyl include, but are not limited to, methoxycarbonyl, carbonyl, and tert—butoxycarbonyl.

The term “alkoxyimino” as used herein means an alkoxy group, as defined herein, ed to the parent lar moiety through an imino group, as defined herein.

Representative examples of alkoxyimino include, but are not limited to, ethoxy(imino)methyl and methoxy(imino)methyl.

The term “alkoxysulfonyl” as used herein means an alkoxy group, as defined herein, appended to the parent molecular moiety h a yl group, as defined herein. Representative es of alkoxysulfonyl include, but are not limited to, methoxysulfonyl, ethoxysulfonyl, and propoxysulfonyl.

The term “alkyl” as used herein means a straight or branched chain arbOn containing from 1 to 20 carbon atoms, preferably from 1 to 10 carbon atoms, more W0 2012/116452 2012/050130 preferably 1, 2, 3, 4, 5, or 6 carbons. Representative examples of alkyl include, but are not limited to, methyl, ethyl, n—propyl, iso—propyl, nubutyl, sec-butyl, iso-butyl, tert~butyl, n~pentyl, isopentyl, neopentyl, l, 3-methylhexyl, 2,2—dimethy1pentyl, 2,3- dimethylpentyl, n—heptyl, n—octyl, n—nonyl, and n-decyl.

The term “alkylarnino” as used herein means an alkyl group, as defined , ed to the parent molecular moiety through a NH group. Representative examples of alkylamino include, but are not limited to, methylamino, ethylarnino, isopropylamino, and butylamino.

The term “alkylcarbony ” as used herein means an alkyl group, as defined herein, appended to the parent molecular moiety through a carbonyl group, as defined herein.

Representative examples of alkylcarbonyl include, but are not limited to, methylcarbonyl, ethylcarbonyl, isopropylcarbonyl, n-propylcarbonyl, and the like.

The term “alkylsulfonyl” as used herein means an alkyl group, as defined herein, appended to the parent molecular moiety through a sulfonyl group, as defined herein.

Representative es of alkylsulfonyl include, but are not limited to, sulfonyl and ethylsulfonyl.

The term “alkynyl” as used herein means a straight or branched chain hydrocarbon group containing from 2 to 10 carbon atoms, and preferably 2, 3, 4, or 5 carbons, and containing at least one carbon—carbon triple bond. Representative es of alkynyl include, but are not limited to, enyl, l-propynyl, 2—propynyl, nyl, 2—pentyny1, and l-butynyl.

The term “amido” as used herein means an amino, alkylamino, or dialkylarnino group appended to the parent molecular moiety through a carbonyl group, as defined herein. Representative examples of amide include, but are not d to, aminocarbonyl, methylaminocarbonyl, ylaminocarbonyl, and ethylrnethylarninocarbonyl.

The term “amino” as used herein means a —NH2 group.

The term “aryl” as used herein means a monocyclic hydrocarbon aromatic ring system. Representative es of aryl include, but are not limited to, phenyl.

The term “arylalkyl” as used herein means an aryl group, as defined herein, appended to the parent molecular moiety through an alkyl group, as defined herein.

Representative examples 'of arylalkyl include, but are not limited to, benzyl, 2- phenylethyl and 3-phenylpropyl.

The term “carbonyl” as used herein means a —C(=O)— group.

The term “carboxy” as used herein means a —COOH group, which may be protected as an ester group: alkyl.

The term “cyano” as used herein means a —CN group.

The term “cyanophenyl” as used herein means a —CN group appended to the parent molecular moiety h a phenyl group, including, but not limited to, 4- cyanophenyl, 3-cyanophenyl, and Z-cyanophenyl.

The term “cycloalkyl” as used herein means a saturated cyclic hydrocarbon gr0up containing from 3 to 8 carbons. Examples of lkyl include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl.

The term “cycloalkylcarbonyl” as used herein means a lkyl group, as defined herein, appended to the parent molecular moiety through a carbonyl group, as defined . Representative examples of cycloalkylcarbonyl include, but are not limited to, cyclopropylcarbonyl, cyclopentylcarbonyl, cyclohexylcarbonyl, and cycloheptylcarbonyl.

The term “dialkylamino” as used herein means two independent alkyl groups, as defined herein, appended to the parent molecular moiety through a nitrogen atom.

Representative examples of lamino include, but are not limited to, dimethylamino, lamino, ethylrnethylamino, and butylrnethylamino.

The term “fluoro” as used herein means —F.

The term “fluoroalkoxy” as used herein means at least one fluoroalkyl group, as defined herein, ed to the parent molecular moiety through an oxygen group, as defined herein. Representative examples of fluoroalkyl include, but are not limited to, tn'fluoromethoxy (CF30-), and difluoromethoxy (CHF20—).

The term “fluoroalkyl” as used herein means at least one fluoro group, as defined herein, appended to the parent molecular moiety h an alkyl' group, as defined herein. entative examples of fluoroalkyl include, but are not limited to, fluoromethyl, difluoromethyl, trifluoromethyl, pentafluoroethyl, and 2,2,2—trifluoroethyl.

The term “formyl” as used herein means a —C(O)H group.

The term “halo” or “halogen” as used herein means Cl, Br, I, or F.

The term “haloalkoxy” as used herein means at least one halogen, as defined herein, appended to the parent molecular moiety through an alkoxy group, as defined herein. Representative examples of haloalkoxy include, but are not limited to, 2— fluoroethoxy, trifluoromethoxy, and pentafluoroethoxy.

The term “haloalkyl” as used herein means at least one halogen, as defined herein, appended to the parent lar moiety h an alkyl group, as defined .

Representative examples of haloalkyl include, but are not limited to, chloromethyl, 2- fluoroethyl, romethyl, pentafluoroethyl, and 2-chloro-3 -fluoropentyl.

The term “heteroaryl”, as used herein, refers to an ic ring containing one or more heteroatoms independently selected from nitrogen, oxygen, or sulfur, or a tautomer thereof. Such rings can be monocyclic or bicyclic as further described herein. Heteroaryl rings are connected to the parent molecular moiety through a carbon or nitrogen atorn.

The terms “monocyciic heteroaryl” or “5— or 6-membered heteroaryl ring”, as used herein, refer to 5» or 6-membered ic rings containing 1, 2, 3, or 4 heteroatoms independently selected from nitrogen, oxygen, or sulfiir, or a tautomer thereof. es of such rings include, but are not limited to, a ring wherein one carbon is replaced with an O or atom; one, two, or three N atoms arranged in a suitable manner to provide an ic ring; or a ring wherein two carbon atoms in the ring are replaced with one 0 or S atom and one N atom. Such rings can include, but are not limited to, a six—membered aromatic ring wherein one to four of the ring carbon atoms are replaced by nitrogen atoms, mbered rings containing a sulfur, oxygen, or nitrogen in the ring; five membered rings containing one to four nitrogen atoms; and five membered rings containing an oxygen or sulfur and one to three nitrogen atoms. Representative examples of 5— to 6-membered heteroaryl rings include, but are not limited to, furyl, imidazolyl, isoxazolyl, isothiazolyl, oxazolyl, pyrazinyl, lyl, zinyl, pyridinyl, pyrimidinyl, pyrrolyl, tetrazolyl, {1,2,3]thiadiazolyl, [1,2,3]oxadiazolyl, thiazolyl, thienyl, [1,2,3]triazinyl, [l riazinyl, [1 riazinyl, [1 ,2,3]triazoly1, and {l,2,4]triazolyl.

The term lic heteroaryl” or “8- to 12-membered bicyclic heteroaryl ring”, as used herein, refers to an 8-, 9-, 10-, 11-, or lZ-membered bicyclic aromatic ring containing at least 3 double bonds, and wherein the atoms ofthe ring include one or more heteroatoms W0 2012/116452 independently selected fi'om oxygen, , and nitrogen. Representative examples of bicyclic aryl rings include indolyl, benzothienyl, benzofirranyl, indazolyl, benzimidazolyl, hiazolyl, benzoxazolyl, benzoisothiazolyl, benzoisoxazolyl, quinolinyl, isoquinolinyl, quinazolinyl, quinoxalinyl, phthalazinyl, pteridinyl, purinyl, yridinyl, inyl, thieno[2,3-d]imidazole, thieno[3,2-b]pyridinyl, and pyrrolopyrimidinyl.

The terms “heterocyclic ring” and “heterocycle”, as used , refer to a 4— to 12-membered monocyclic or bicyclic ring containing one, two, three, four, or five heteroatoms independently selected from the group consisting of nitrogen, oxygen, and sulfur and also containing either at least one carbon atom attached to four other atoms or one carbon atom substituted with an oxo group and attached to two other atoms. Four- and five-membered rings may have zero or one double bond. Six-membered rings have zero, one, or two double bonds. Seven— and eight-membered rings may have zero, one, two, or three double bonds. The non-aromatic heterocycie groups ofthe invention can be attached through a carbon atom or a nitrogen atom. The non-aromatic heterocycle groups may be present in tautomeric form. Representative examples of nitrogen- containing heterocycles e, but are not limited to, azepanyl, inyl, aziridinyl, azocanyl, dihydropyridazinyi, dihydropyridinyl, dihydropyrimidinyl, morphoiinyl, piperazinyl, piperidinyl, pyrrolidinyl, pyrrolinyl, dihydrothiazolyl, dihydropyridinyl, and thiomorpholinyl. Representative examples of non—nitrogen containing non—aromatic heterocycles include, but are not limited to, dioxanyl, dithianyl, tetrahydrofuryl, dihydropyranyl, tetrahydropyranyl, and [1,3]dioxolany1.

The term “hydroxy” as used herein means an ——OH group.

The term “hydroxyalkyi” as used herein means at least one y group, as defined herein, appended to the parent molecular moiety through an alkyl group, as defined herein. Representative examples oxyalkyl include, but are not limited to, hydroxymethyi, 2-hydroxyethyl, 2—methyl-2—hydroxyethyl, 3~hydroxypropyl, 2,3- dihydroxypentyl, and 2—ethyl-4—hydroxyheptyl.

The term “hydroxy-protecting group” means a tuent which ts hydroxyl groups against undesirable reactioris during synthetic procedures. Examples of hydroxy—protecting groups include, but are not limited to, methoxymethyl, W0 2012/116452 benzyloxymethyl, 2-methoxyethoxymethyl, methylsi1yl)ethoxymethyl, benzyl, triphenylmethyl, 2,2,2-trichloroethyl, tubutyl, trimethylsilyl, ldimethylsiiyl, tbutyldiphenylsilyl , methylene acetal, acetonide benzylidene acetal, cyclic ortho esters, methoxymethylene, cyclic carbonates, and cyclic boronates. Hydroxy-protecting groups are appended onto hydroxy groups by reaction of the compound that contains the hydroxy group with a base, such as triethylamine, and a reagent selected from an alkyl halide, alkyl trifilate, trialkylsilyl halide, triallcylsilyl triflate, alyldialkylsilyltriflate, or an hloroformatc, CHzlz, or a dihaloboronate ester, for example with methyliodide, benzyl , tn'ethylsilyltriflate, acetyl chloride, benzylchloride, or dimethylcarbonate.

A protecting group also may be appended onto a hydroxy group by reaction of the compound that contains the y group with acid and an alkyl acetal.

The term “imino” as defined herein means a—C(=NH)— group.

The term “mercapto” as used herein means a —SH group.

The term “nitro” as used herein means a—NOZ group.

The term “nitrogen protecting group" as used herein means those groups intended to protect a nitrogen atom against undesirable reactions during synthetic procedures.

Nitrogen protecting groups comprise carbamates, amides, Nubenzyl derivatives, and imine derivatives. Preferred nitrogen protecting groups are , benzoyl, benzyl, benzyloxycarbonyl (Cbz), formyI, phenylsulfonyl, pivaloyl, tert-butoxycarbonyl (Boo), tert-butylacetyl, trifluoroacetyl, and triphenylmethyl (trityl). Nitrogen-protecting groups are appended onto y or secondary amino groups by reacting the nd that contains the amine group with base, such as triethylarnine, and a reagent selected from an alkyl , an alkyl te, a dialkyl anhydride, for example as represented by (alkyl- 0) 2 C=O, a diaryl ide, for example as represented by (aryl-O) 2 CmO, an acyl halide, an hloroformate, or an alkylsulfonylhalide, an arylsulfonylhalide, or halo- CON(alkyl) 2 for example acetylchloride, benzoylchloride, benzylbromide, benzyloxycarbonylchloride, formylfluoride, phenylsulfonylchloride, pivaloylchloride, (tert—butyl~O—-C=O) 2 O, trifluoroacetic anhydride, and nylmethylchloride.

The term “oxo” as used herein means (w).

Unless ise indicated, the term “prodrug” encompasses pharmaceutically acceptable esters, carbonates, thiocarbonates, N-acyl derivatives, N—acyloxyalkyl W0 2012/116452 derivatives, quaternary derivatives of tertiary amines, N—Mannich bases, Schiff bases, aminoacid conjugates, phosphate esters, metal salts and ate esters of compounds disclosed herein. Examples ofprodrugs include compounds that comprise a biohydrolyzable moiety (e.g., a biohydrolyzable amide, biohydrolyzable carbamate, rolyzable carbonate, biohydrolyzable ester, biohydrolyzable phosphate, or biohydrolyzable ureide analog). Prodrugs ofcompounds disclosed herein are readily envisioned and ed by those of ordinary skill in the art. Sec, e.g., Design of Prodrugs , Bundgaard, A. Ed, Elseview, 1985; Bundgaard, hours, “Design and Application of Prodrugs,” A ok of Drug Design and Development , Krosgaard- Larsen and hours. Bundgaard, Ed., 1991, Chapter 5, p. 113-491; and Bundgaard, hours, ed Drug Delivery Review, 1992, 8, l-38.

Unless otherwise indicated, the term “protecting group” or “protective group,” when used to refer to part of a molecule ted to a chemical reaction, means a chemical moiety that is not reactive under the conditions of that chemical reaction, and which may be removed to provide a moiety that is reactive under those conditions.

Protecting groups are well known in the art. See, e.g., Greene, T. W. and Wuts, P.G.M., Protective Groups in Organic Synthesis (3 rd ed., John Wiley & Sons: 1999); Larock, R.

C., hensive Organic Transformations (2nd ed., John Wiley & Sons: 1999). Some examples include , diphenylmethyl, trityl, Cbz, Boc, Fmoc, methoxycarbonyl, ethoxycar‘oonyl, and pthalimido. ting groups include, for example, nitrogen protecting groups and hydroxy-protecting groups.

The term nyl” as used herein means a —S(O)2 — group.

The term “thioalkoxy” as used herein means an alkyl group, as defined herein, appended to the parent molecular moiety through a sulfur atom. Representative examples ofthioalkoxy include, but are no limited to, methylthio, ethylthio, and propylthio. n nds ofthe present invention may exist as stereoisomers wherein, asymmetric or chiral centers are present. These stereoisorners are “R” or “S” depending on the configuration of substituents around the chiral carbon atom. The terms “R” and “S” used herein are configurations as defined in IUPAC 1974 Recommendations for Section E, Fundamental Stereochemistry, in Pure Appl. Chem, 1976, 45: 13—30. The invention contemplates various stereoisomers and mixtures thereof and these are 2012/050130 cally included within the scope of this invention. Stereoisomers include enantiomers and diastereorners, and mixtures of enantiomers or diastereomers. Individual stereoisomers of compounds of the invention may be prepared synthetically from commercially available starting materials which contain asymmetric or chiral centers or by preparation of racemic mixtures followed by resolution well known to those of ordinary skill in the art. These methods of resolution are exemplified by (1) attachment of a mixture ofenantiomers to a chiral auxiliary, separation of the resulting mixture of diastereomers by recrystallization or chromatography and optional liberation of the Optically pure product from the auxiliary as described in s, Hannaford, Smith, and Tatchell, “Vogel's Textbook ofPractical Organic Chemisuy”, 5th edition (1989), Longman Scientific & Technical, Essex CM20 2JE, England, incorporated herein by reference for the disclosure of s for separation and purification of diastereomers or (2) direct tion of the mixture of optical enantiomers on chiral chromatographic columns or (3) fractional recrystallization s. n compounds of the present ion may exist as cis or trans isomers, wherein substituents on a ring may attached in such a manner that they are on the same side of the ring-(cis) relative to each other, or on opposite sides of the ring relative to each other (trans). Such methods are well known to those of ry skill in the art, and may include tion of s by recrystallization or chromatography. It should be understood that the compounds of the invention may possess tautomeric forms, as well as geometric isomers, and that these also constitute an aspect of the invention.

It should be noted that a chemical moiety that forms part of a larger compound may be described herein using a name commonly accorded it when it exists as a single molecule or a name commonly accorded its radical. For example, the terms “pyridine” and “pyridyl” are accorded the same meaning when used to describe a moiety attached to other chemical moieties. Thus, for example, the two phrases “XOH, wherein X is l” and “XOH, wherein X is pyridine” are accorded the same meaning, and encompass the compounds pyridin-Z-ol, pyridin—3-ol and pyridin—4~ol.

It should also be noted that names of compounds having one or more chiral centers that do not specify the stereochemistry of those centers encompass pure stereoisomers and mixtures thereof. Moreover, any atom shown in a drawing with WO 16452 unsatisfied valences is d to be attached to enough hydrogen atoms to satisfy the valences. In addition, chemical bonds depicted with one solid line parallel to one dashed line encompass both single and double (e.g., ic) bonds, if valences permit.

The term “phannaceutically acceptable excipient”, as used herein, means a non- toxic, inert solid, semi-solid or liquid filler, diluent, encapsulating material or formulation auxiliary of any type. Some examples of materials which can serve as pharmaceutically acceptable carriers are sugars such as lactose, glucose and sucrose; starches such as corn starch and potato starch; ose and its tives such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; powdered tragacanth; malt; gelatin; talc; cocoa butter and suppository waxes; oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; glycols; such a propylene glycol; esters such as ethyl oleate and ethyl laurate; agar; buffering agents such as magnesium ide and aluminum hydroxide; alginic acid; pyrogen—i’ree water; isotonic saline; Ringer's on; ethyl alcohol, and phosphate buffer solutions, as well as other non— toxic compatible lubricants such as sodium lauryl e and magnesium stearate, as well as coloring agents, releasing agents, coating agents, sweetening, flavoring and perfuming agents, preservatives and antioxidants can also be present in the composition, according to the judgment of one d in the art of formulations.

Unless otherwise indicated, a “therapeutically effective amount” of a compound is an amount sufficient to treat a disease or condition, or one or more ms associated with the disease or condition. In some embodiments, “treatment” may be determined by comparison to an untreated control.

- The term “subject” is intended to include living organisms in which disease may occur. Examples of subjects include humans, s, cows, sheep, goats, dogs, cats, mice, rats, and transgenic Species thereof.

The present invention is based, at least in part, on the design of nds that inhibit acyl carrier protein (ACP) synthase (AcpS), the enzyme responsible for converting apo—ACP into holo-ACP. AcpS is present not only in Gram-positive and -negative bacteria, but also in acid-fast bacteria such as cterium tuberculosis, and even in protozoa such as dz‘umfalciparum. Accordingly, AcpS inhibitors are expected to have deleterious effects on the viability of many microbes. AcPS inhibitors are be expected to have deleterious effects on the maintenance ofmicrobial cell function, including, for example, increased porosity of cell membranes and dysfunction of efflux pumps in such membranes. The spectrum of activity demonstrated for compounds of the present invention includes many Gram-positive bacteria including Methicillinwresistant S. aureus, with selected compounds retaining activity against Gram-negative pathogens such as PS. aeruginosa, Ac. ii, and S. maltophilz'a, and the like. Compounds of the t invention with low or no antimicrobial potency against, for example, PS. aeruginosa, may have antimicrobial adjuvant (“adjuvant” or “pro-antibiotic”) s when co—administered with antimicrobial nds such as, for example, azithromycin, erythromycin, or ampiciliin, or with crobial compounds ofthe present ion.

Though the utility of the compounds disclosed herein does not dependper se on their being AcpS tors, the design effects toward that goal resulted, at least in part, in the discovery of the compounds ofthe present invention.

According to an embodiment, a compound, or prodrug or pharmaceutically acceptable salt f, is provided according to one of Formulas I or has as structure as set forth in one of Formulas I: 2012/050130 R1 R3 //N‘“‘~NH R4 \N/ S \ / R? N R3 R10 (1b) in which E is ~CH2— or is absent whereby thiophene is directly connected to phenyl, R1, R2, R9, R10, and Rn (ifpresent) are each independently selected from the group consisting ofhydrogen, methyl, ethyl, propyl, cyclopropyi, butyl, cyciobutyl, pentyl, hexyl,isopropry1, isobutyl, neopentyl, methoxy, and ethcxy; additionally, R1 and R2 may connect to form a phenyi or benzofuran ring; onally, R9 and R10 may connect to form a phenyl or benzofuran ring; R3 and R3 are each independently selected from the group consisting ofhydrogen, methyl, ethyl, propyl, isopropyl, chlorine, fluorine, tert-butyl, methylsulfonyl, methoxy, and ethoxy; R4 and R7 are each independently selected from the group consisting ofhydrogen, chlorine, methyl ester, ethyl ester, methyl, ethyl, prOpyl, cyclopropyl, butyl, cycIobutyl, isopropryl, yl, methoxy, and ethoxy; and R5 and R5 are each independently selected from the group consisting of en, cyclopentyl, cyclopropyl, furan, thiophene, romethyl, tn'fluoromethyl ether, methylthiol, formaldehyde, ne, e, bromine, phenyl, methyl, ethyl, isopropyl, propyl, butyl, cyclobutyl, isobutyl, neopentyl, pentyl, methoxy, and ethoxy.

WO 16452 2012/050130 In some embodiments, R3 and R2 are either independently selected from the group consisting ofhydrogen and , or form a phenyl ring whereby the ring system is naphthyl; R9 and R10 are either independently selected from the group consisting of hydrogen and methyl, or form a phenyl ring whereby the ring system is naphthyl; R11 is hydrogen; R3 and R3 are each independently selected fi'om the group consisting of hydrogen, methyl, chlorine, fluorine, isopropyl, tert-butyl, methoxy, and methylsulfonyl; R4 and R7 are each independently selected from the group consisting of hydrogen, methyl, chlorine, and ethyl ester; and R5 and R6 are each independently selected from the group consisting of methyl, ethyl, phenyl, hydrogen, chlorine, isopropyl, entyl, bromine, ropyl, trifluorornethyl, trifluoromethyl ether, methylthiol, formaldehyde, furan, and thiophene. In some embodiments, E is absent. In some embodiments, R1, R2, R4, R7, R9, and Rm are each hydrogen. In some embodiments, R3 and R3 are each chlorine; and R5 and R5 are selected from the group consisting of hydrogen, methyl, and isopropyl. In a preferred ment, the compound is DNM0488. In a preferred embodiment, the compound is DNM0548. In a preferred embodiment, the compound is DNMO606. In a preferred ment, the compound is DNMO631. (These compound identifiers are given ing to ) In a preferred embodiment, the compound is selected from the group consisting of those compounds listed in According to an embodiment, a compound, or prodrug or pharmaceutically acceptable salt thereof, is provided according to one of Formulas II: W0 2012/116452 2012/050130 (IIb) in which R1, R2, R3, R5, and R12 are each independently selected from the group consisting of hydrogen, methyl, ethyl, propyl, cyclopropyl, butyl, Cyclobutyl, pentyl, hexyl, isopmpryl, isobutyl, neopentyl, methoxy, and ; R4, and R13 are each ndently selected frorn the group consisting of hydrogen, methyl, ethyl, propyl, cyclopropyl, butyl, cyclobutyl, pcntyl, hexyl, isopropryl, isobutyl, neopentyl, methoxy, ethoxy, and dialkylamine; additionally, R4 and R5 may connect to form a phenyl ring; additionally, R12 and R13 may connect to form a phenyl ring; R6 and R“ are each independently selected from the group consisting of hydrogen, chlorine, e, hydroxy, phenyl ether, methyl, ethyl, propyl, i30propyl, tert-butyl, methoxy, and ethoxy; R1 and R19 are each independently selected from the group consisting ofhydrogen, chlorine, methyl, ethyl, propyl, isopropyl, tert—butyl, methoxy, and ethoxy; and R8 and R9 are each independently selected from the group consisting of en, halogen, methyl, ethyl, propyl, isopropyl, tert-butyl, methoxy, ethoxy, yl optionally tuted with methyl, and phenyl optionally substituted with chlorine, bromine, carboxylic acid, and alkyl.

In some embodiments, R5 and R11 are each independently selected from the group consisting ofhydrogen, chlorine, fluorine, hydroxy, phenyl ether, and alkyl; R7 and R10 are each independently ed from the group consisting ofhydrogen, chlorine, and alkyl; and R3 and R9 are each independently selected from the group consisting of hydrogen, halogen, methyl, isopropyl, naphthyl ally substituted with methyl, and phenyl optionally substituted with chlorine, bromine, carboxylic acid, and alkyl. In some embodiments, the compound is according to a 1121. In some embodiments, the compound is a compound according to Formula 111). In a preferred embodiment, the compound is selected from the group consisting of those compounds listed in In a preferred embodiment of the invention, a compound is selected from the group consisting of 5-(4,5-Bis(4-chloro-2—methylpheny1)thiophen-Z—yl)-lH-tetrazole (DNM0488), 5-(4,5—Bis(4—methy1naphthalenyl)thiophen~2-yl)—1 H—tetrazole (DNMO486), 5~(4,5—Bis(4-chlorophenyl)thiophen-Z—yl)-lH—tetrazole (DNM0487), 5—(4,5- Di(biphenyl-2~yl)thiophen—2-yl)-1H~tetrazole (DNMO489), 5—(4,5-Bis(3-chloro methylphenyl)thiophen-2—yl)-1H-tetrazole (DNM0504), 5-(4,5—Bis(5-chloro methylphenyl)thiophenyl)-lH—tetrazole (DNMOSOS), 5—(4,5—Bis(3,4- dimethylphenyl)thiophen—2~yl)-lH—tetrazole (DNM0509), -Bis(4- pylphenyl)thiophen-2~yl)~1H—tetrazole (DNMOSIZ), 5-(4,5—Bis(2- methylphenyl)thiophen-Z-yl)—1H—tetrazole (DNM0531), 5-(4,S-Bis(2- isopropylphenyl)thiophen—2-yl)-1I-I-tetrazole (DNM0534), 5-(4,5-Bis(2» phenoxyphenyl)thiophen-2—yl)—lH—tetrazole (DNM0536), 5—(4,5~Bis(4—fluoronaphthalen- l—yl)thiophen—2—yl)—lH-tetrazole (DNM0537), 5-(4,5-Bis(2-chlorophenyl)thiophen~2-yl)- lH—tetrazole (DNM053 8), —Bis(2—ethyl~phenyl)thiophen—2-yl)—lH-tetrazole . (DNMOS41), 5-(4,5-Bis(dibenzo[b,d]fiiranyl)thiophen-2—yl)~lH-tetrazole 42), -(4,5-Di(benzofuranyl)thiophenyl)-lH-tetrazole 43), -Bis(2- yphenyl)thiophenyl)-1H—tetrazole (DNM0544), 5-(4,5-Bis{2,3- dimethoxyphenyl)thiophen—2-yl)-1H—tetrazole (DNM0545), 5-(4,5-Bis(4-tert- butylphenyl)thi0pheny1)— l H—tetrazole (DNM0546), 544,5~Bis(4-chloro—2-iso- propylphenyl)thiophen—2-yl)-lH—tetrazole (DNM0548), 5—(4,5—Bis(2— (trifluoromethyl)phenyl)thi0phen-2—yl)~lH-tetrazole (DNM0549), 5-(4,5-Bis(2- (trifluoromethoxy)phenyl)thiophen—2-yl)—lI-I-tetrazole (DNMOSSO), 5—(4,5-Bis(2,4— W0 2012/116452 dimethoxyphenyl)thiophen—2-yl)—lH-tetrazole (DNMOSSZ), 5—(4,5-Bis(2,6- oxyphenyl)thiophen—2-yl)-1H—tetrazole (DNMOSSB), (2,2'-(5-(1H—tetrazoI—5- y1)thiophene-2,3—diyl)bis(2,1-phenylene))dimethanol (DNMOSSS), 2,2’-(5-(1H—tetrazoI—5— yl)thiophene-2,3—diyl)dibenza1dehyde (DNM0556), S—(4,5-Di(furan—3—yI)thiophen—2-yl)— lH—tetrazole (DNM0557), 5-(4,5—Di(thiophen—3-yI)thiophen—2-yl)~1H—tetrazole (DNMOSSS), 5-(4,5—Bis(2—(thiophen—3-y1)pheny1)thiophen—2—yl)-1H~tetrazole (DNM0559), 5-(4,5~Bis(2-( furan~3—yl)phenyl)thiophen~2—yl)-1H—tetrazolc (DNMOS60), -(4,5—Bis(2-chloro-4~methy1pheny1)thiophen—2~y1)—lH-tetrazole (DNM0563), 5-(4,5- Bis(4—methoxy-3,5-dimethylphenyl)thiophen-Z-yl)-1H—tetrazole (DNM0564), 5-(4,5— Bis(3~chloromethoxyphenyl)thiophen—2-yl)—lH—tetrazole 6S), Diethyl 3,3'-(5- (1H—tetrazol-5—yl)thiophene~2,3-diy1)dibenzoate (DNM0593), 1,1 '-(3,3'-(5-(1H—tetrazol- -y1)thiophene—2,3~diyl)bis(3,1-phenylene))dibutanone (DNM0599), Diethyl 5,5'-(5~ (1H~tetrazol~S-yl)thiophene-2,3-diyl)bis(3-hydroxybenzoate), 5-(4,5—Bis(3— butylphenyl)thiophen—2—y1)—1H—tetrazole (DNM0608), 5—(4,5-bis(3— (cyclopentylmethyl)phenyl)thiophen—2-y1)-lH—tetrazole (DNM0612), 5-(4-(4-Chloro—2— methylphenyl)(4-methylnaphthalen—1-y1)thiophenyl)—1H—tetrazole (DNM0576), 5- (4—(4-Chlorophenyl)(4-methy1naph’rhalen-1—yl)thiopheny1)—lH-tetrazole (DNM0572), 5—(4—(5—Chlorophenyl)—4-(4-methy1naphthalen- I-y1)thiophenyl)-1H- tetrazole CDNMOS75), 5—(4—(4—Fluorophenyl)-5—(4—(methylsu1fony1)phenyl)thiophen yl)—1H—tetrazole (DNM0592), Ethyl 3—(2~(4~methylnaphthalen—1-yl)-S—(1H-tetrazol—S- yl)thiophen—3-y1)benzoate 96), Ethyi 3—(2-(4-chloro~2-methylphenyI)—5—(1H— oI-S—yl)thiopheny1)benzoate (DNM0597), s(4-chloromethylpheny1) (1H-tetrazol~5—yl)thiazole (DNM0574), 4,5-Bis(5-chloromethylphenyl)—2-(1H— tetrazoI-S—yl)thiazole (DNM0567), 4,5~Bis(3~chloro—4—met'hylphenyl)~2~(IH—tetrazol—S— yl)thiazole (DNM0568), 4,5-Bis(4—methylnaphthalen—1—y1)—2-(1H-tetrazolyl)thiazole (DNM0569), 4,5~Bis(4-chlorophenyl)—2-(1H-tetrazol—5-yi)thiazoie (DNM0573), 4,5- Bis(4-fluoronaphthalen—1-y1)—2—(1H—tetrazolLS—ylfihiazoie (DNM0578), 4,5—Bis(4— yl)-2~(1H—tetrazol~5-yl)thiazole (DNM0581), 4,5-Bis(4-t—butylphenyl)-2—(1H- ol-S-yl)thiazole (DNM0582), 4,5-Bis(3,4—dimethy1phenyl)~2~(1H-tetrazol-S- y1)thiazole (DNMOS 83), 4,5-Bis(4-chloro-2—isopropylphenyl)—2—( lH-tetrazol—S- azole (DNM0584), 5-(5-(4-Chlorobenzy1)—4—(4—methy1naphthalen—1-y1)thiophen-2— W0 16452 yl)—1H~tetrazole (DNM0577), 5-(5-(4-Chlorobenzyl)-4—(4-chlorophenyl)thjopheny1)—1H— tetrazole (DNM0579), 5-(5~(4-Chlorobenzyl)-4~(4-chloro~2-iso- propylphenylfihiophenyl}1H-tetrazole SO), 5-(5-(4-Chlorobenzyl)—4-(4_ fluoronaphthalen—1—y1)thiophen—2-yl)-IH—tetrazole (DNM0587), S-(4—(Biphenylyl)—5— (4—chlorobenzyi)thiophen-Z-yl)~lH-tetrazole (DNM0588), Ethyl 3-(2—(4-chlorobenzyl)—5- (1H—tetrazol-S-yl)thiophen—3~yl)benzoate (DNM0595), 1-(3r(2-(4—Ch10robenzy1)—S-(1H- tetrazol-S-yl)thiophen—3-y1)phenyl)butan—1-one (DNM0600), 5-(4—(3-Buty1phenyl)—5-(4- chIorobenzyl)thiophen-2«yl)-1H—tetrazoie (DNM0606), Ethyl 3—(2—(4—chlorobenzyl)—5- (1H—tetrazol—5-yl)thiophen—3-yl)hydroxybenzoate (DNM0609), 5-(5-(4- Chlorobenzyl)~4~(3-(cyc1opentyhnethyl)phenyl)thiophen—2—yl)-1H—tetrazole (DNM06 1 0), 3-(2-(4—Chlorobenzyl)(1H-tetrazoi—S—y1)thiophen—3-yl)phcnol 1 3), 5—(5-(4- benzyI)—4-(3—methoxyphenyl)thiophen-2~y1)~1I-I-tetrazole 15), 5-(5-(4— Chlorobenzyl)~4—(3-butoxyphenyl)thiopheny1)— 1 H—tetrazole (DNM06 I 6), 5-(5—(4— Chlorobenzyl)~4—(3-ethoxyphenyl)thiopheny1)-1H—tetrazole (DNM0617), 5-(5—(4- benzyl)(3-propoxyphenyl)thiophen—2-yl)~1 H-tetrazole (DNM06 l 8), 5—(3 ,5- Bis(4-methylnaphthalenyl)pheny1)~2H—tetrazole (DNM0461), 5~(2,5-Bis(4- methylnaphthalen—l-y1)pheny1)-IH—tetrazole (DNM0446), 5-(2,5-Bis(4—chloro methylphenyl)phenyl)-lH—tetrazole (DNM0447), 5—(3,5~Bis(4—chlorOpheny1)phenyl)—2H- tetrazole (DNM0470), 5-(4'-Chloro~5-(4-methy1naphthalen—l—y1)biphenyl~3—y1)—1H— tetrazole (DNM0480), 5-(3,5-Bis(4-fluoronaphthalen—1-y1)phenyl)-2H-tetrazole (DNM0539), 4,5-Bis(4-ch10roisopropylphenyl)thiophene-Z—carboxylic acid (DNM0566), 4,5-Bis(4-biphenyl)thiophene-Z-carboxylic acid (DNM0497), 4,5-Bis(4- chlorophenyl)thiophene—2-carboxylic acid (DNM0498), 4,5-Bis(5-chloro-2— methylphenyl)thiophene—2-carboxy1ic acid OI), 4,5—Bis(3— chlorophenyl)thiophene-2~carboxylic acid (DNMOSOZ), 4,5—Bis(2,4— ylphenyl)thiophene-2ncarboxylic acid (DNM0503), 4,5-Bis(4-chlor02- methylphenyl)thiophene—2—carboxylic acid (DNM0561), S-(4,5—Bis(4—chloro—2— isopropylphenyl)-3 -methylthiophen—2-yl)—lH-tetrazole (DNM0631), 5-(4,5-bis(4-chloro— 2~methylphenyl)methylthiophen—2-yl)-1H—tetrazole 14), 5—(4,5-Bis(4- methylnaphthalen—l~y1)-3~methy1thiophenyl)-1H-tetrazole (DNM0620), 5-(4,5-Bis(3- WO 16452 henyl)—3-methylthiophen—2—yl)-lH-tetrazole (DNM0627), and 5-(4,5-Bis(4~ fluoronaphthalen- l -yl)-3 lthiophen—2—yl)— l H—tetrazo1e (DNMO628).

According to an embodiment of the invention, a method of treatment of a microbial ion is provided sing administering an effective amount of an antimicrobial compound as bed herein to a patient in need thereof. Such a patient may be, for example, a human or other mammal that is infected with a pathogenic bacterium. In some embodiments, the microbial infection is substantially caused by Gram-positive bacteria. In some embodiments, the microbial infection is substantially caused by Enterococcus sp. In some embodiments, the microbial infection is substantially caused by Staphylococcus sp. In some embodiments, the microbial infection is substantially caused by Bacillus sp. In some embodiments, the microbial ion is substantially caused by Staphylococcus epidermidis, Staphylococcus aureus (including methicillin-resistant Staphylococcus aureus [MRSA]), Enterococcus sp. (including vancomycin—resistant Enterococcus [VRE]), Streptococcus sp., or Bacillus sp.

To say that a microbial infection is “substantially caused” by a particular bacterium or species thereof indicates that the given ial genus or species is thought to be primau'ly responsible for the pathology, epidemiology, or symptom profile of the microbial infection and does not rule out the presence of other ial species (e.g., benignly zing bacteria). In some embodiments, the microbial infection is skin and skin structure infection of , wound infections, diabetic foot infections, osteomyelitis, pneumonia, anthrax infection, impetigo or secondarily-acquired traumatic lesions, gasteroenteritis, meningitis, pneumonia, gonorrhea, peptic ulcers, nosocomial infections, or blood-stream infection, among others. In some embodiments, the described antimicrobial agents are used as medicinal compounds, for example, for treating humans, or as veterinary compounds, for e, for treating s, poultry, livestock and the like, as well as in aquaculture and agricultural ations.

According to an embodiment ofthe invention, a method oftreatment of a microbial infection is ed comprising administering an effective amount of an antimicrobial adjuvant compound as described herein and an effective amount of an antimicrobial nd to a patient in need thereof. Such a patient may be, for example, a human or other mammal that is infected with a pathogenic bacterium. In some W0 16452 embodiments, the antimicrobial adjuvant compound is DNM0487. In some embodiments, the antimicrobial adjuvant compound is DNM0488. In some embodiments, the antimicrobial adjuvant compound is DNMOS48. In some embodiments, the antimicrobial adjuvant compound is itself antimicrobial to some extent.

Thus combinations are possible, not only of compounds of the present invention with each other, but also between a compound of the present invention and a known antibacterial nd. In some embodiments, the microbial ion is skin and skin structure infection of ulcers, wound infections, diabetic foot infections, osteomyelitis, pneumonia, impetigo or secondarily-acquired traumatic lesions, gasteroenteritis, meningitis, pneumonia, aemia, urinary tract infections, gonorrhea, peptic ulcers, nosocomial infections, blood-stream infection, brucellosis, campylobacteriosis, Cat Scratch fever, cholera, legionellosis, leptospirosis, Lyme disease, dosis, meningitis, pertussis, plague, salmonellosis, shigellosis, is, tuiaremia, typhoid fever, or urinary tract infection. An effective amount of one or more of the above—described antimicrobials may be used in the preparation of a medicament as described above for the treatment of a disease, disorder or condition caused by a pathogenic bacteria selected from the group including but by no means limited to Escherichia, Salmonella, Pseudomonas, Neisseria, Legionella, Haemophiius, Campylobacter, bacter and In another ment ofthe invention, there is provided a method of manufacturing a medicament for treating a microbial infection comprising admixing an antimicrobial compound as described herein with a suitable cxcipient.

In other embodiments, there is provided the use of an antimicrobial compound as described herein for ng a microbial infection.

In another embodiment of the invention, there is provided a method of manufacturing a medicament for treating a ial infection comprising admixing an antimicrobial adjuvant as described herein with a suitable antimicrobial compound.

In other embodiments, there is ed the use of an antimicrobial adjuvant as described herein for treating a microbial infection. In some embodiments, the antimicrobial adjuvant is used with or coadministered with an antimicrobial compound.

WO 16452 According to an embodiment, a pharmaceutical composition is ed comprising a compound ofthe present invention and a pharmaceutically acceptable excipient.

The microbial infection may be substantially caused by one or more Gram— positive ia.

The microbial infection may se a Staphylococcal infection.

The microbial infection may comprise Enterococcal infection.

The microbial ion may comprise Bacillus infection.

The microbial infection may besubstantially caused by a bacterial species selected from the group ting of Staphylococcus aureus, Staphylococcus epidermidis, Enterococcus is, Enterococcus faecium, Bacillus cereus, and Streptococcus sp.

Alternatively, the microbial infection is substantially caused by Gram—negative bacteria.

The microbial ion may be crobial.

The microbial infection may be substantially caused by Pseudomonas aeruginosa.

The antimicrobial adjuvant compound may be selected from the group consisting ofDNMO487, DNM0488, and DNM0548.

It is of note that the antimicrobials or antimicrobial adjuvants may be prepared to be administered in a variety of ways, for example, topically, orally, intravenously, intramuscularly, subcutaneously, intraperitoneally, asally or by local or systemic intravascular infusion using means known in the art and as discussed below. Compounds ofthe present invention may be used, for example, to treat infections caused by Gram negative bacteria such as E. 0011' and/or Gram positive bacteria such as .S'. aureus. In some embodiments, compounds have antimicrobial effects against antibiotic-resistant strains such as, for example, methicillin—resistant S. aureus (MRSA). In some embodiments, compounds have antimicrobial effects against Mycobacterium tuberculosis.

The crobial or antimicrobial adjuvant compounds may be arranged to be delivered at a tration of about 1 nM to about 50 mM; or 10 nM to about 50 mM; or 100 nM to about SOmM; or 1 M to about 50 mM; or 10 uM to 50 mM or 100 uM to 50 mM. As will be appreciated by one of skill in the art, this may be the “effective amoun ,” that is, a sufficient dosage is administered such that a concentration within one of the envisioned ranges is attained at the required site. In some antimicrobial embodiments, the effective amount will be informed at least in part by the minimum inhibitory concentration of the compound required to t bacteriostatic or bacteriocidal effects against the en of st. In some antimicrobial adjuvant embodiments, the effective amount will be informed at least in part by the approximate minimum concentration required to produce the desired adjuvant effect with a ermined known antibiotic against the pathogen of interest. In some embodiments, the effective amount will be calibrated so as to produce a serum level ofover ten times the MIC, or over five times the MIC, or over three times the MIC, or at the MIC in a subject suffering from an infection. In some ments, the effective amount will be calibrated so as to produce an in situ concentration of over ten times the MIC, or over five time the MIC, or over time times the MIC, or at the MIC.

An effective amount of one of the compounds of the invention can be employed in pure form or, where such forms exist, in pharmaceutically able salt form.

Alternatively, the compound can be administered as a ceutical compositiOn containing the compound of interest in combination with one or more pharmaceutically acceptable carriers. It will be understood, however, that the total daily usage ofthe compounds and compositions ofthe invention will be decided by the attending physician within the scope of sound medical judgment. The specific ive dose level for any ular patient will depend upon a variety offactors including the disorder being treated and the severity of the disorder; activity of the specific compound employed; the specific composition employed; the age, body weight, general health, sex and diet of the patient; the time of administration, route of administration, and rate of excretion of the specific compound employed; the duration of the treatment; the risk/benefit ratio; drugs used in combination or coincidental with the specific compound employed; and like factors well known in the medical arts. For example, it is well within the skill ofthe art to start doses of the compound at levels lower than required to e the desired therapeutic effect and to lly increase the dosage until the desired effect is achieved.

The total daily dose of the compounds of the present invention as administered to a human or lower animal may range from about 0.0003 to about 30 mg/kg ofbody weight. For purposes of oral administration, more preferable doses can be in the range of from about 0.0003 to about 1 mg/kg body weight. If desired, the effective daily dose can be divided into multiple doses for es of administration; consequently, single dose compositions may n such amounts or submultiples f to make up the daily close. For oral administration, the compositions of the invention are ably provided in the form oftablets ning about 1.0, about 5.0, about 10.0, about 15.0, about 25.0, about 50.0, about 100, about 250, or about 500 milligrams of the active ingredient.

For best results, whether a particular compound is antimicrobial or an antimicrobial adjuvant with respect to the pathogen of interest (and in the case of adjuvants, with respect to the antimicrobial compound being used) would lly be ascertained by a number of microbiological methods known in the art, including for e, the methods disclosed in the Examples. It is well understood in the art that while every “antimicrobial” compound has an antimicrobial effect against at least one pathogen at one or more effective amounts, antimicrobial compounds vary as to their spectrum and/or potency of activity. Moreover, while every “antimicrobial adjuvant” nd has an adjuvant effect in conjunction with at least one antimicrobial compound against at least one pathogen at one or more effective s ofboth crobial compound and adjuvant, antimicrobial nt compounds vary as to their spectrum, potency of activity, and/or compatibility with particular antimicrobial compounds. As will be apparent to one knowledgeable in the art, the total dosage will vary according to many factors, including but by no means limited to the weight, age and condition of the individual or patient.

In some embodiments, one or more of the crobial compounds may be cc— administered with one or more known antibiotics. In some embodiments, one or more of the antimicrobial adjuvant compounds my be co-administered with one or more antimicrobial compounds; in which case the total effective amount of antimicrobial compound may be less than would otherwise be required in the absence of antimicrobial adjuvant, e.g. ~8 times less, or ~16 times less, or ~32 times less, or ~64 times less, or ~125 times less, or ~250 times less. in some embodiments, the antimicrobial adjuvant compounds are not themselves antimicrobial. In some embodiments, the antimicrobial adjuvant compounds are themselves antimicrobial. In some embodiments, one or more W0 2012/116452 of the antimicrobial adjuvant compounds may be combined with one or more antimicrobial compounds in a single dosage form. In some embodiments, the antimicrobial compound is an antimicrobial compound ofthe t invention. In some embodiments, the crobial compound is a known antimicrobial compound such as, for example, almecillin, amdinocillin, amikacin, amoxicillin, amphomycin, amphotericin B, llin, azacitidine, azaserine, azithromycin, azlociilin, aztreonam, sinin, allopurinol, amicacin, aminoglycosides, amphotericin B, ampicillin, ansamycins, anthracyclines, antimycotics, azithromycin, bacampiciliin, bacitracin, benzyl penicilloylpoiylysine , bleomycin, din A, butoconazole, candicidin, capreomycin, carbenicillin, cefacior, cefadroxil, cefamandole, cefazoline, cefdinir, cefepime, cefixime, oxime, cefinetazole, zime, cefonicid, cefoperazone, ceforanide, cefotaxime, cefotetan, cefotiam, cefoxitin, cefpiramide, cefpodoxime, cefprozil, cefsuiodin, cefiazidime, cefiibuten, ceftizoxime, ceftriaxone, cefuroxime, cephacetrile, cephalexin, cephaloglycin, cephaloridine, cephalothin, irin, cephradine, chloramphenicoi, cilastatin, cinnamycin, ciprofloxacin, clarithromycin, clavulanic acid, clindamycin, inol, cloxacillin, colistimethate, colistin, cyclacillin, cycloserine, cyclospon'ne, (Leu- Pro), camptothecin, cefataxime, cephalexin, cephaiosporins, chalcomycin, chartreusin, chlorotetracyciines, thricin, chrymutasins, chrysomicin M, chrysomicin V, clomocyclines, dactinomycin, dalbavancin, dalfopristin, ycin, daunorubicin, demeclocycline, detorubicin, dicloxacillin, dihydrostreptomycin, dirithromycin, doxorubicin, doxycycline, ellipticines, eisamicin, epirubicin, erythromycin, eveminomycin, filipins, fluconazoles, fungichromins, fusidic acid, floxacillin, fosfomycin, gentamycin, gilvocarin, fulvin, viridin, guamecyclines, gemifloxacin, gramicidin, hetacillin, icin, imipenem, an, ivermectin, ilosamides, itraconazoles, kanamycin, laspartomycin, linezolid, loracarbef, lankamycin, lincomycin, magainin, meclocycline, meropenem, methacycline, mezlocillin, minocycline, mitomycin, moenomycin, moxalactam, moxifloxacin, enolic acid, macrolides, methicillins, mitoxantrone, nafcillin, cin, neomycin, netilmicin, niphimycin, nitrofurantoin, ocin, nalidixic acid, norfloxin, nystatin, nystatins, ofloxacin, oieanomycin, oxytetracyline, paromomycin, penicillamine, phenethicillin, piperacillin, plicamycin, pristinamycin, peciiocin, penicillins, pesticides, phosphomycin, W0 2012/116452 pimarcin, siinycin, polyenes, polymyxin B, polymyxin E, quinupristin, quinolones, ravidomycin, reserpines, rifamycin, ristocetins A and B, rifabutin, in, rifamycin, rolitetmcycline, sisomycin, spiramycin, spironolactone, sulfacetamide sodium, sulphonamide, spectrinornycin, streptomycin, streptozocin, sulbactam, sultamicillin, tacrolimus, tazobactam, lanin, teiithromycin, cins, tetracyclines, thiamphenicols, thiolutins, tobramycin, tyrothricin, ticarcillin, tigecycline, tobramycin, troleandomycin, tunicamycin, tyrthricin, vancomycin, vidarabine, Viomycin, virginiamycin, and wortmannins; the presence of a plural item in the foregoing list g to refer to one or more members of a family of antibiotics known in the art by that name. Which compound or compounds should be co-administered or compounded in combination with compounds ofthe present invention depends on a number of factors, including but not necessarily limited to the efficacy ofthe agent or agents in the absence of antimicrobial adjuvant compounds, the ism of action ofthe compound(s), the identity of the pathogen causing or potentiating the sick ion, and/or the severity of the sick condition in the subject.

The pharmaceutical compositions can be formulated for oral administration in solid or liquid form, for parenteral intravenous, subcutaneous, uscular, eritoneal, intra—arterial, or ermal injection, for or for vaginal, nasal, topical, or rectal administration. Pharmaceutical compositions of the present iOn suitable for Oral administration can be presented as discrete dosage forms, e.g., tablets, chewable tablets, caplets, capsules, liquids, and flavored . Such dosage forms n predetermined amounts of active ingredients, and may be prepared by methods of pharmaCy well known to those skilled in the mt. See generally, ton's Pharmaceutical Sciences, 18th ed., Mack Publishing, Easton Pa. (1990).

Parenteral dosage forms can be administered to ts by various routes including subcutaneous, intravenous (including bolus injection), intramuscular, and rterial. Because their administration typically bypasses patients' natural defenses against contaminants, parenteral dosage forms are specifically sterile or capable of being sterilized prior to administration to a patient. Examples ofparenteral dosage forms include solutions ready for injection, dry products ready to be dissolved or suspended in a pharmaceutically acceptable vehicle for injection, suspensions ready for injection, and ons. Pharmaceutical compositions for parenteral injection comprise pharrnaceutically acceptable sterile aqueous or nonaqueous solutions, dispersions, suspensions or emulsions and sterile powders for reconstitution into sterile injectable solutions or dispersions. Examples of suitable s and nonaqueous rs, ts, solvents or vehicles include water, ethanol, polyols (propylene glycol, hylene glycol, glycerol, and the like, and suitable mixtures thereof), vegetable oils (such as olive oil) and injectable organic esters such as ethyl oleate, or suitable mixtures thereof.

Suitable fluidity ofthe composition may be maintained, for e, by the use of a coating such as in, by the maintenance ofthe required particle size in the case of diSpersions, and by the use of surfactants. These itions may also contain adjuvants such as preservative agents, wetting agents, emulsifying agents, and dispersing agents. Prevention ofthe action ofmicroorganisms may be ensured by various antibacterial and anfifiingal agents, for example, parabens, butanol, phenol, sorbic acid, and the like. It may also be desirable to e isotonic agents, for example, sugars, sodium chloride and the like. Prolonged absorption of the injectable pharmaceutical form may be brought about by the use of agents delaying absorption, for example, aluminum monostearate and gelatin.

In some cases, in order to prolong the effect of a drug, it is often desirable to slow the absorption of the drug from subcutaneous or intramuscular injection. This may be lished by the use of a liquid suspension of crystalline or amorphous material with poor water solubility. The rate of absorption of the drug then depends up0n its rate of dissolution which, in turn, may depend upon crystal size and crystalline form.

Alternatively, delayed absorption of a parenterally administered drug form is lished by dissolving or ding the drug in an oil vehicle.

Suspensions, in addition to the active compounds, may contain suspending agents, for example, ethoxylated isostearyl alcohols, polyoxyethyiene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar-agar, tragacanth, and mixtures f. If desired, and for more effective distribution, the compounds ofthe invention can be incorporated into slow-release or targeted—delivery systems such as r matrices, liposomes, and microspheres. They may be sterilized, for example, by ion through a bacteria-retaining filter or by incorporation of W0 2012/116452 sterilizing agents in the form of sterile solid compositions, which may be dissolved in sterile water or some other sterile injectable medium ately before use.

Injectable depot forms are made by g microencapsulated matrices of the drug in biodegradable polymers such as polylactide-polyglycolide. ing upon the ratio of drug to polymer and the nature of the particular polymer employed, the rate of drug e can be controlled. es of other biodegradable polymers include poly(orthoesters) and poly(anhydrides). Depot injectable formulations also are prepared by entrapping the drug in liposomes or microemulsions which are ible with body tissues. The injectable formulations can be sterilized, for example, by filtration through a ial-retaining filter or by incorporating sterilizing agents in the form of sterile solid compositions which can be dissolved or dispersed in sterile water or other sterile injectable medium just prior to use.

Injectable preparations, for e, sterile injectable aqueous or nous suspensions may be formulated according to the known art using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation may also be a sterile able solution, suspension or emulsion in a nontoxic, parenterally acceptable diluent or solvent such as a solution in 1,3-butanedioi. Among the acceptable vehicles and solvents that may be employed are water, Ringer's solution, U.S.P. and isotonic sodium chloride on. In on, sterile, fixed oils are conventionally employed as a solVent or suspending medium. For this purpose any bland fixed oil can be employed including synthetic mono- or diglycerides. In addition, fatty acids such as oleic acid are used in the preparation of injectables.

Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In such solid dosage forms, one or more compounds of the invention is mixed with at least one inert pharmaceutically acceptable carrier such as sodium e or iurn phosPhate and/or a) fillers or extenders such as starches, lactose, sucrose, glucose, mannitol, and salicylic acid; b) binders such as carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidinone, e, and acacia; c) humectants such as glycerol; d) disintegrating agents such as agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate; e) solution retarding agents such as paraffin; t) absorption accelerators such as quaternary W0 2012/116452 ammonium compounds; g) wetting agents such as cetyl alcohol and glycerol monostearate; h) absorbents such as kaolin and bentonite clay; and i) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, and es thereof. In the case of capsules, tablets and pills, the dosage form may also se buffering agents.

Solid compositions of a similar type may also be employed as fillers in soft and hard-filled gelatin capsules using lactose or milk sugar as well as high molecular weight polyethylene glycols. The solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings and other coatings well known in the pharmaceutical formulating art. They may optionally contain opacifying agents and can also be of a compositiou that they release the active ingredient(s) only, or preferentially, in a certain part of the intestinal tract in a delayed . Examples of materials which can be useful for delaying release of the active agent can include polymeric substances and waxes.

Dosage forms for topical administration may include s, sprays, ointments and inhalants. A compound of the present ion can be mixed under sterile conditions with a pharmaceutically acceptable carrier and any needed preservatives, buffers or propellants which may be required. Opthalmic formulations, eye ointments, powders and solutions are plated as being within the scope of this invention. Aqueous liquid compositions comprising compounds ofthe invention also are contemplated.

Liquid dosage forms for oral administration include ceutically acceptable emulsions, microemulsions, solutions, sions, syrups and elixirs. In addition to the active compounds, the liquid dosage forms may n inert diluents commonly used in the art such as, for example, water or other solvents, solubilizing agents and emulsifiers such as ethyl alcohol, isopmpyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene , 1,3-butylene , dimethylformamide, oils (in particular, cottonseed, nut, corn, germ, olive, castor, and sesame oils), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols and fatty acid esters of an, and mixtures thereof.

Besides inert diluents, the oral compositions can also include adjuvants such as wetting agents, emulsifying and suspending , sweetening, flavoring, and perfuming agents. Dosage forms for topical or transdermal administration of a compound of this invention include ointments, pastes, creams, lotions, gels, powders, solutions, , inhalants or patches. A desired compound ofthe invention is admixed under sterile conditions with a pharmaceutically acceptable r and any needed vatives or buffers as may be required. Ophthalmic formulation, ear drops, eye ointments, powders and solutions are also contemplated as being within the scope of this invention. The ointments, , creams and gels may contain, in addition to an active nd of this invention, animal and vegetable fats, oils, waxes, paraffins, starch, tragacanth, cellulose derivatives, polyethylene glycols, silicones, bentonites, silicic acid, talc and zinc oxide, or mixtures thereof.

Powders and sprays can contain, in addition to the nds of this invention, lactose, talc, silicic acid, aluminum hydroxide, calcium silicates and ide powder, or ‘es of these substances. Sprays can additionally contain customary propellants such as chlorofluorohydrocarbons. nds ofthe invention may also be administered in the form of liposomes.

As is known in the art, liposomes are generally derived from phospholipids or other lipid substances. Liposomes are formed by mono-- or lamellar hydrated liquid crystals that are dispersed in an aqueous medium. Any non-toxic, physiologically acceptable and metabolizable lipid capable of forming liposomes may be used. The present cempositions in liposome form may contain, in on to the compounds of the invention, stabilizers, preservatives, and the like. The preferred lipids are the natural and synthetic phospholipids and phosphatidylcholines hins) used separately or together. Methods to form liposomes are known in the art. See, for example, Prescott, Ed, Methods in Cell Biology, Volume XIV, Academic Press, New York, N.Y., (1976), p 33 et seq.

In some embodiments, one or more of the antimicrobials or antimicrobial adjuvants at concentrations or dosages discussed above may be combined with a pharmaceutically or pharmacologically acceptable r, ent or diluent, either biodegradable or non-biodegradable. Exemplary es of carriers include, but are by no means limited to, for example, poly(ethylene~vinyl acetate), copolymers of lactic acid and glycolic acid, poly(lactic acid), gelatin, collagen matrices, polysaccharides, poly(D,L lactide), poly(malic acid), poly(caprolactone), celluloses, albumin, starch, casein, dextran, wo 2012/116452 2012/050130 polyesters, ethanol, mathacrylate, polyurethane, polyethylene, vinyl polymers, glycols, mixtures thereof and the like. Standard excipients include n, casein, lecithin, gum acacia, cholesterol, tragacanth, c acid, benzalkonium chloride, calcium stearate, yl monostearate, cetostearyl alcohol, crogol emulsifying wax, sorbitan , polyoxyethylene alkyl , polyoxyethylene castor oil derivatives, polyoxyethylene sorbitan fatty acid esters, polyethylene glycols, polyoxyethylene stearates, colloidol silicon dioxide, phosphates, sodium dodecylsulfate, carboxymethylcellulose calcium, ymethylcellulose sodium, methylcellulose, hydroxyethylcellulose, hydroxypropylcellulose, hydroxypropylmethycellulose phthalate, noncrystalline cellulose, magnesium aluminum silicate, triethanolamine, polyvinyl alcohol, polyvinylpyrrolidone, sugars and es.

As will be apparent to one dgeable in the art, specific carriers and carrier combinations known in the art may be selected based on their preperties and release characteristics in view of the intended use. Specifically, the carrier may be pH-sensitive, thermo—sensitive, thermo~gelling, arranged for sustained release or a quick burst. In some embodiments, rs of different classes may be used in combination for multiple effects, for example, a quick burst followed by sustained release.

In other embodiments, one or more of the antimicrobials or crobial adjuvants at coucentrations or dosages described above may be encapsulated for delivery.

Specifically, the compounds may be encapsulated in biodegradable microspheres, microcapsules, microparticles, or nanospheres. The delivery vehicles may be composed of, for example, hyaluronic acid, polyethylene glycol, poly(lactic acid), gelatin, poly(E- caprolactone), or a poly(lactic-glycolic) acid polymer. Combinations may also be used, as, for example, gelatin nanospheres may be coated with a polymer of poly(lactic~ glycolic) acid. As will be apparent to one knowledgeable in the art, these and other suitable delivery vehicles may be prepared according to protocols known in the art and utilized for delivery of the compounds.

It is of note that the above described antimicrobials may be combined with permeation enhancers known in the art for improving ry. Examples of permeation enhancers include, but are by no means limited to those compounds described in US. Pat.

Nos. 3,472,931; 3,527,864; 3,896,238; 256; 3,952,099; 4,046,886; 4,130,643; 4,130,667; 4,299,826; 4,335,115; 4,343,798; 4,379,454; 4,405,616; 4,746,515; 4,788,062; 4,820,720; 4,863,73 8; 970; and 5,378,730; British Pat. No. 1,011,949; and Idson, 1975, J. Pharm. Sci. —924.

A “pharmaceutically acceptable salt” includes a salt that retains the desired biological activity of the parent antimicrobial or antimicrobial adjuvant compound and does not impart any undesired toxicological effects. es of such salts are salts of acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, nitric acid, and the like; acetic acid, oxalic acid, tartaric acid, succinic acid, malic acid, benzoic acid, pamoic acid, alginic acid, methanesulfonic acid, naphthalenesulfonic acid, and the like. Also included are salts of cations such as sodium, potassium, lithium, zinc, c0pper, barium, bismuth, calcium, and the like; or organic s such as trialkylammonium.

Combinations of the above salts are also useful. It is to be understood that a compound disclosed herein in a salt-free form and analogous compound in a phannaceutically acceptable salt form are both compounds of the t invention. Additionally, gs, e.g. esters of the compounds disclosed herein, are compounds ofthe present invention. The preparation and use of acid on salts, carboxylate salts, amino acid additiOn salts, and zwitterion salts of compOunds ofthe present inventiOn may also be considered pharmaceutically acceptable ifthey are, within the scope of sound medical nt, le for use in contact with the tissues ofhumans and lower animals without undue toxicity, irritation, allergic response, and the like, are commensurate with a reasonable benefit/risk ratio, and are effective for their intended use. Such salts may also include various es and hydrates ofthe compound of the present invention.

In some embodiments, the bed antimicrobial compounds used as medicinal compounds, for example, for treating humans, or as veterinary compounds, for example, for treating animals, poultry, livestock and the like, as well as in aquaculture and agricultural ations.

While various embodiments ofthe invention have been described above, it will be recognized and understood that modifications may be made therein, and the appended claims are ed to cover all such modifications which may fall within the spirit and scope ofthe invention.

Certain embodiments ofthe invention will now be illustrated using examples.

W0 2012/116452 Example 1 Synthesis ofDNM0488 and analogues NBS :91N U c: TFA+HZSO4 / \ “2 ON Br 3 3 wrong, PMPP diamhala-rfizcos ”he" NINSW1213:”his:IFHN—N 4,5-Dibromothiophenecarboxamide Thiophene—Z—carbonitrile (3.27 g, 30.0 mmol) was dissolved in a mixture of H2804 (10 mL) and TFA (20 mL). NBS (11.75 g, 66.0 mmol) was added in portion in 20 min. After complete addition, the reaction mixture was further stirred for 4h, and then poured onto 200 g of crushed ice. A white solid was formed, which was collected, washed with water, and dried together with orous pentoxide under vacuum to afford 8.50 g (99 %) of product. bromothiophene-Z-carbonitrile An oven-dried round-bottomed flask was charged with bromofl1i0phenecarboxamide (4.28g, 15.0 mmol) and 20 mL of DMF under an argon here. The solution was cooled on an ice—water bath, and cyauuric chloride (1.81 g, 9.8 mmol) was then added in one portion. After stirring at 0 °C for 1 h, the reaction mixture was warmed to room temperature, and stirred for a further 3h. 100 mL ofwater was added. A white solid was formed, which was collected through suction ion, washed with water, and dried together with phosphorous pentoxide under vacuum to afford 3.70 g (92 %) ofproduct. 4,5-Bis(4-chloromethylpheny1)thiophenc—Z-carbonitrile A round-bottomed flask was charged with 4,5-dibromothi0phene»2—carbonitrile (534 mg, 2.00 11111101), 4- chloro~2~methylphenylboronic acid (818 mg, 4.80 mmol) and Pd(PPh3)4 (136 mg, 0.10 mmol). After degassed, dioxane (10 mL) and aqueous sodium carbonate (5 mL, 2M, 10 mmol) was added. The reaction mixture was heated to 90 °C. The ss of the reaction was monitored by TLC. After the reaction was te, 50 mL of water was added, and the reaction mixture was extracted with ethyl acetate. The organic phase was dried over anhydrous sodium sulphate and then coucentrated. The residue was purified by flash ch-ornatography (hexane: CH2C12 = 2: 1). 0.60 g (84 %) of product was obtained as clear oil. —(4,5-Bis(4—chloromethylphenyl)thiophenyl)—1H-tetrazole (DNM0488) A round-bottomed flask was charged with 4,5-bis(4-chloromethylphenyl)thi0phene—2- carbonitriie (600 mg, 1.67 mmol), zinc bromide (945 mg, 4.20 mmol) and sodium azide (273 mg, 4.20 mmol). After degassed, DMF (5 mL) was added. The reaction mixture was heated to 110 °C and stirred at this temperature until complete. The reaction was cooled to rt, and 30 mL of 0.1 N aqueous HCi was added. The reaction mixture was extracted with ethyl acetate. The organic phase was dried over anhydrous sodium te and then concentrated. The e was purified by flash tography (hexane: EtOAc: AcOH = 30: 10: 1). 603 mg (90 %) uct was obtained as a white solid, 1H NMR (DMSO, 500 MHz) 5 7.82 (s, 1H), 7.36 (d, J= 2.04 Hz, 1H), 7.34 (d, J= 1.94 Hz, 1H), 7.31 (d, J = 8.25 Hz, 1H), 7.28 (dd, J1= 8.26 Hz, J2 = 1.99 Hz, 1H), 7.17 (dd, J1= 8.20 Hz, J2 = 2.05 Hz, 1H), 7.02 (d, J= 8.28 Hz, 1H), 2.17 (s, 3H), 2.03 (5,311); 13c NMR (DMSO, WO 16452 125 MHZ)8140.58, 139.14, 139.12, 138.19, 133.59, 133.39, , 132.47, 131.71, 131.17, 130.57, 130.17, 130.11, 126.00, 125.80, 19.69, 19.57.

The following compounds were also prepared using an analogous method: -(4,5—Bis(4—methylnaphthalen-l-yl)thiophen-Z—yI)—1H-tetrazole (DNM0486) 1H NMR (DMSO, 500 MHZ.) 6 8.00 (d, J= 8.49 Hz, 1H), 7.98 - 7.90 (m, 3H), 7.88 (d, J = 8.49 Hz, 1H), 7.57 - 7.50 (m, 2H), 7.50 — 7.42 (m, 2H), 7.33 (t, J= 7.47 Hz, 1H), 7.29 (d, J2 7.36 Hz, 1H), 7.19 (d, J= 7.22 Hz, 1H), 7.13 (d, J= 7.22 Hz, 1H), 2.57 (s, 3H), 2.52 (s, 3H); l3C NMR (DMSO, 125 MHz) 6 , 135.65, 134.20, 132.18, 132.13, 131.98, 131.68, 131.30, 130.95, 129.21, 127.85, 127.50, 126.30, 126.22, , 125.95, 125.89, 125.81, 125.75, , 124.50, 19.07, 18.97. -(4,5-Bis(4-ch10rophenyl)thiophen-2—yl)—1H-tetrazole (DNM0487) IH NMR (DMSO, 500 MHz) 5 7.90 (s, 1H), 7.50 — 7.45 (111,411), 7.38 - 7.31 (m, 4H); ”c NMR (DMSO, 125 MHz) 5 140.14, 137.85, , , 132.63, 131.58, 131.15, 130.74, 130.54, 129.19, 128.94, 124.74. -(4,5-Di(biphenyI-Z-yl)thiophenyl)-1H—tetrazole (DNM0489) 1H NMR (DMSO, 500 MHz).8 7.48 (s, 1H), 7.35 (td, J; = 7.55 Hz, J2 = 1.00 Hz, 1H), 7.30 (td, J; m 7.55 Hz, J2 = 1.00 Hz, 1H), 7.21 — 7.03 (m, 10H), 6.72 - 6.65 (m, 2H), 6.65 - 6.58 (m, 3H), 6.55 (d, J: 7.45 Hz, 111); 13C NMR (DMSO, 125 MHz) 8 141.91, 140.83, 140.46, 140.19, 139.33, , 132.02, 131.21, 130.32, 130.23, 130.20, 129.88, 128.63, 128.46, 128.45, 127.95, 127.88, 127.70, 127.45, 127.34, 126.61, 126.34. -(4,5—Bis(3—chloromethylphenyl)thiophen-2—y1)-lH-tetrazole (DNM0504) lH NMR (DMSO, 500 MHZ) 5 7.89 (s, 1H), 7.41 (d, J: 1.70 Hz, 1H), 7.39 (d, J= 1.75 Hz, 1H), 7.37 (d, J= 8.05 Hz, 1H), 7.35 (d, J= 8.00 Hz, 1H), 7.18 (dd, J; = 7.85 HZ, J2 = 1.80 Hz, 1H), 7.12 (dd, J, = 7.83 Hz, J2 = 1.73 Hz, 1H), 2.34 (s, 6H); 130 NMR (DMSO, 125 MHz)8139.77, , 136.19, 135.06, 134.02, , 133.55, 131.80, 131.68, , 131.54, 128.79, 128.69, ,127.43,19.38, 19.32.

S-(4,S-Bis(S—chloro-Z-methylphenyl)thiophenyl)-1H-tetrazole 08) ]H NMR (DMSO, 500 MHz) 8 7.84 (s, 1H), 7.41 (d, J= 2.25 Hz, 1H), 7.36 (dd, J, = 8.23 Hz, J2 = 2.28 Hz, 1H), 7.31 - 7.23 (m, 3H), 7.11 (s, 1H), 2.13 (s, 3H), 2.01 (s, 3H); 13C NMR(DMSO, 125 MHz) 8 140.36, 139.01, 136.27, 135.74, 134.71,133.43, 132.26, 132.23, 131.07, 130.59, 130.15, 130.01,129.56, 128.95, 19.26, 19.16. -(4,S-Bis(3,4-dimethylphenyl)thiophenyl)-1H—tetrazole (DNM0509) 1H NMR (DMSO, 500 MHz) 8 7.84 (s, 1H), 7.21 (s, 1H), 7.19 (s, 1H), 7.12 — 7.06 (m, 2H), 6.98 (dd, J, = 8.28 Hz, J2 a 1.63 Hz, 1H), 6.94 (dd, J, = 7.67 Hz, J2 = 1.48 Hz, 1H), 2.22 (s, 6H), 2.20 (s, 3H), 2.19 (s, 3H); 13C NMR (DMSO, 125 MHz) 8 141.17, 138.26, 136.90, 136.80,136.57, 135.65, 132.51, 131.95, 130.14, 129.86, 129.68, 129.65,129.54, 126.21, 126.06, 19.41, 19.33,19.14, 19.11. -(4,5-Bis(4-isopr0pylphenyl)thiophenyl)-lH—tetrazole (DNM0512) ]H NMR (DMSO, 500 MHz) 8 7.84 (s, 1H), 7.30 - 7.23 (m, 8H), 2.90 (8613, J= 6.88 Hz, 1H), 1.24 - 1.18 (m, 12H); 13C NMR (DMSO, 125 MHZ)5148.87, 147.77, 141.11, 138.25, 132.48, 132.01, 130.15, 128.75, 128.55, 126.93, 126.67, 33.11, 33.08, 23.76, 23.65. -(4,S-Bis(2-methylphenyl)thiophen—Z-yl)-1H-tetrazole (DNM0531) 1H NMR (DMSO, 500 MHz) 8 7.81 (s, 1H), 7.33 - 7.16 (m, 6H), 7.02 (t, J= 7.35 Hz, 1H), 7.02 (t, J= 7.44 Hz, 1H), 2.16 (s, 3H), 2.02 (s, 3H); 130 NMR (DMSO, 125 MHz) 8 141.77, WO 16452 140.03, 136.57, 135.51, 134.73, 131.87, 131.48, 131.26,130.42, 130.39, 130.10, 128.93, 127.89, 125.89, 125.72, 19.91, 19.75. ~(4,S-Bis(2-isopropylphenyl)thiophen-2—yl)—lH—tetrazole 34) 1H NMR (CDC13, 500 MHz) 8 7.75 (s, 1H), 7.29 — 7.19 (m, 5H), 7.14 — 7.08 (m, 1H), 7.07 ~ 7.00 (m, 2H), 3.10 - 2.95 (m, 2H), 1.04 (s, 6H), 0.95 (d, J= 6.80 Hz, 6H); 13c NMR (CDC13, 125 MHz) 6 148.48, 147.51, , 132.28, 131.09, 129.46, 128.43, , 126.15, 125.72, 125.66, 30.36, 10.15, 24.46. -(4,5—Bis(2—phenoxyphenyl)thiophen-2—yl)—1H—tetrazole (DNM0536) 1H NMR (DMSO, 500 MHz) 5 7.91 (s, 1H), 7.39 - 7.22 (m, 8H), 7.16 - 7.05 (m, 4H), 6.79 (d, J = 7.90 Hz, 1H), 6.75 (d, J: 7.65 Hz, 1H), 6.66 - 6.61 (m, 4H); ”c NMR (DMSO, 125 MHz)5155.83, 155.76, 154.04, 153.87, 138.02, 136.48, 131.75, ,131.33, 130.42, 129.91, 129.86, 129.50, 126.43, 124.08, 123.68, 123.64, 123.46, , 118.95, 118.70, 118.37,117.81. -(4,5-Bis(4-fluoronaphthalen—1-yl)thiophen—2—yl)-1H-tetrazole (DNM0537) IH NMR (DMSO, 500 MHz) 6 8.05 — 7.95 (m, 4H), 7.85 (d, J= 8.50 Hz, 1H), 7.66 — 7.56 (m, 3H), 7.53 (t, J= 7.53 Hz, 1H), 7.40 (t, J= 7.55 Hz, 1H), 7.34 - 7.26 (m, 2H), 7.15 (dd, J17» 10.55 Hz, J2 a 8.00 Hz, 1H); 13C NMR (DMSO, 125 MHz) 5 159.20, 158.48, 157.19, 156.48, 140.42, 139.24, 132.83, 132.79, 132.44, 132.40, 132.00, , 129.84, 128.92, 128.89, 127.97, 127.90, 127.80, 127.75, 127.02, 126.85, 125.89, 125.85, 125.53, 125.51, 125.41, , 122.86, 122.75, 122.73, 122.62, 120.25, 120.23, , 120.19, 109.42, 109.28, 109.26, 109.12.

S-(4,5-Bis(2-chlorophenyl)thiophenyl)-1H-tetrazole (DNM0538) 1H NMR (CDC13, 500 MHZ) 5 8.07 (s, 1H), 7.36 (d, J= 8.05 Hz, 1H), 7.31 (d, J= 7.91 Hz, 1H), W0 2012/116452 7.29 - 7.22 (m, 2H), 7.18 (dt,J1 = 1.06 Hz, J; = 7.50 Hz, 1H), 7.15 — 7.09 (m, 1H), 7.08 - 7.01 (m, 2H); 13C NMR (CDC13, 125 MHz) 5 142.02, 139.64, 134.55, 134.27, 133.64, 133.21, 132.59, 132.11, 131.78, 130.56,130.40, 130.23, 129.57, , ,124.30. -(4,5-Bis(2—ethylphenyl)thi0phenyl)-lI-I-tetrazole (DNM0541) lH NMR (DMSO, 500 MHz) 5 7.77 (s, 1H), 7.33 - 7.20 (m, 5H), 7.17 (t, J= 7.43 Hz, 1H), 7.08 (dt, J, = 1.10 Hz, J2 = 7.33 Hz, 1H), 7.03 (d, J= 7.50 Hz, 1H), 2.50 (q, J= 7.50 Hz, 2H), 2.43 (q, J= 7.50 Hz, 2H), 1.06 (1,.1= 7.50 Hz, 3H), 0.96 (1, J= 7.50 Hz, 3H); 13(2 NMR (DMSO, 125 MHz) 5 142.74, , 141.58,139.90, 134.00, 131.53, 131.48, 130.97, 130.33, 129.16, 128.69, 128.45, 128.11, 125.71, 125.56, 123.74, 25.51, 25.38, 15.09, .01. -(4,5-Bis(dibenz0 [b,d]furanyl)thiophen—Z-yl)-1H—tetrazole (DNM0542) 1H NMR (DMSO, 500 MHz) 5 8.26 (s, 1H), 8.15 - 8.07 (m, 3H), 8.04 (dd, J, = 7.63 Hz, J2 = 0.98 Hz, 1H), 7.50 — 7.40 (m, 4H), 7.40 -7.34 (m, 2H), 7.34 — 7.22 (m, 4H), ”C NMR (DMSO, 125 MHz)5155.30, 155.22,152.65, 152.36, 137.03, 135.52, 131.54, 128.15, , 127.75,127.68, 124.22, 124.05, ,123.34, 123.20, 123.16, 123.12, 121.82, 121.36, ,120.90, 119.86, 117.22,111.38, 111.25. -(4,5-Di(benzofuran-Z—yl)thi0phen~2~yl)—1H—tetrazole (DNM0543) 1H NMR (DMSO, 500 MHz) 5 8.23 (s, 1H), 7.74 (d, J= 7.75 Hz, 2H), 7.67 (1, J= 7.28 Hz, 2H), 7.49 (s, 1H), 7.45 - 7.37 (m, 3H), 7.33 (dt, J; = 2.20 Hz, J; = 7.40 Hz, 2H); 13C NMR (DMSO, 125 MHz) 5 154.12, 153.91, 149.67, 147.67,130.34, 130.05, 128.48, 128.36, 128.21, 125.92, 125.38, 123.75, 123.53,121.93, 121.68, 111.27,111.26, 107.06, 106.01. -(4,S—Bis(2-methoxyphenyl)thiophenyl)~lH-tetrazole (DNM0544) 1H NMR (DMSO, 500 MHZ) 8 7.82 (s, 1H), 7.35 - 7.28 (m, 2H), 7.10 (dd, J1= 7.55 Hz, J2 = 1.50 W0 2012/116452 Hz, 1H), 7.04 (d, J= 8.35 Hz, 2H), 6.98 (dd, J, = 7.50 Hz, J; = 1.35 Hz, 1H), 6.90 _ 6.80 (m, 2H), 3.62 (s, 3H), 3.59 (s, 3H); 13C NMR (DMSO, 125 MHz) 5 156.29, 156.25, 138.75, 137.01, 131.72, 130.91, 130.30, 130.09, 129.16, 124.59, 121.73, 120.37, 120.29, 111.85, 111.60, 55.28, 55.22. —(4,5—Bis(2,3-dimethoxyphenyl)thiophen—2-yl)-1H-tetrazole (DNM0545) lH NMR (DMSO, 500 MHz) 8 7.82 (s, 1H), 7.03 (dt, .1; = 1.20 Hz, J; = 8.00 Hz, 2H), 6.98 (d, J = 7.70 Hz, 1H), 6.94 (d, J = 8.00 Hz, 1H), 6.67 (dd, J; = 7.75 Hz, J2 = 1.40 Hz, 1H), 6.63 (dd, J, = 7.58 Hz, J; = 1.58 Hz, 1H), 3.82 (s, 6H), 3.63 (s, 3H), 3.57 (s, 3H); 13C NMR (DMSO, 125 MHz) 5 152.73, 146.13, 146.07, 138.01, 136.68, 131.51, 129.53, 126.64, 123.88, 122.59, 122.43, , 112.63, 60.12, 59.96, 55.72, 55.67. —(4,5—Bis(4-tert~butylphenyl)tl1iophen~2-yl)-lH-tetrazole 46) 1H NMR (DMSO, 500 MHz) 6 7.84 (s, 1H), 7.53 - 7.37 (m, 4H), 7.30 (d, J = 8.41 Hz, 2H), 7.27 (d, J= 8.41 Hz, 2H), 1.29 (s, 9H), 1.28 (s, 9H); 13C NMR GDMSO, 125 MHz) 5 151.15, 150.07, , 132.17, 132.12, 129.84, 128.46, 128.28, 125.80, 125.53, 34.46, 34.37, 31.09, 30.99. -(4,5-Bis(4-chloro—2-iso-propylphenyl)thiophen-2—yl)-1H-tetrazole (DNM0548) 1H NMR (DMSO, 500 MHz) 5 7.75 (s, 1H), 7.43 - 7.39 (m, 2H), 7.31 - 7.26 (m, 2H), 7.30 (dd, J; = 2.24 Hz, J2 = 8.30 Hz, 1H), 7.06 (d, J= 8.30 Hz, 1H), 2.97 — 2.82 (m, 2H), 1.04 (broad, 6H), 0.95 (d, J = 6.80 HZ, 6H); 13C NMR (DMSO, 125 MHz) 8 150.00, 149.20,140.39, 138.88, 134.47, , 133.30, 132.20, 131.71, , 128.76, 126.16, , 125.97, 125.76, 29.95, 29.77, 23.54 (broad). -(4,5-Bis(2-(trifluoromethyl)phenyl)thiophen-2—yl)-1H-tetrazole (DNM0549) 1H NMR (DMSO, 500 MHZ) 5 7.88 - 7.78 (m, 3H), 7.66 - 7.58 (m, 2H), 7.56 - 7.49 (m, 2H), 7.41 (broad, 1H), 7.23 - 7.15 (m, 1H); l3c NMR (DMSO, 125 MHz) 8 -(4,5-Bis(2—(trifluoromethoxy)phenyl)thiophen-Z-y1)-1H-tetrazole (DNMOSSO) 1H NMR (DMSO, 500 MHZ) 6 7.90 (s, 1H), 7.57 - 7.51 (m, 1H), 7.51 — 7.46 (m, 1H), 7.45 — 7.34 (m, 5H), 7.32 (dd, J; = 7.50 Hz, J2 = 1.70 Hz, 1H); 13C NMR (DMSO, 125 MHz) 8 145.44, 136.94, 135.8, 132.48, 131.7, 131.17, 130.98, 130.22, 127.82, 127.62, 127.4, 125.37, 120.88, 120.54, 118.85, 118.81. —(4,5-Bis(2,4-dimethoxyphenyl)thiophen-Z-yl)~1H—tetrazole (DNMOSSZ) IH NMR (DMSO, 500 MHz) 5 7.75 (s, 1H), 6.99 (d, J= 8.45 Hz, 1H), 6.87 (d, J= 8.45 Hz, 1H), 6.64 — 6.58 (m, 2H), 6.46 (dd, J1= 8.50 Hz, J; = 2.25 Hz, 1H), 6.43 (dd, J1= 8.50 Hz, J; = 2.25 Hz, 1H), 3.764 (s, 3H), 3.756 (s, 3H), 3.66 (s, 3H), 3.65 (s, 3H); 13(3 NMR (DMSO, 125 MHZ) 5 , 160.13, 157.52, 157.46, 138.46, 136.36, 131.91, 131.62, , , 114.29, 105.25, 104.89, 98.82, 98.79, 55.49, 55.40, 55.29, 55.19. -(4,S-Bis(2,6—dimethoxyphenyl)thiophen-Z—yl)—1H-tetrazoie (DNM0553) 1H NMR (DMSO, 500 MHz) 8 7.68 (s, 1H), 7.22 (t, J= 8.38 HZ, 1H), 7.16 (d, J= 8.33 Hz, 1H), 6.57 (d, J= 8.40 Hz, 2H), 6.55 (d, J= 8.35 HZ, 1H), 3.49 (s, 6H), 3.47 (s, 6H); 13C NMR (DMSO, 125 MHZ) 8 157.67, 157.29, 134.19, 132.23, 130.01, 128.89, , 110.53,103.80, 103.73, 55.26, 55.10. (2,2'-(5-(lH-tetrazol-S-yl)thiophene-Z,3-diyl)bis(2,1—phenylene))dimethanol (DNMDSSS) 1H NMR (DMSO, 500 MHZ) 8 7.85 (s, 1H), 7.51 (d, J= 7.60 Hz, 1H), 7.48 (d, J= 7.70 Hz, 1H), 7.37 (dt, J1= 1.47 Hz, J3 = 7.45 Hz, 1H), 7.33 ~ 7.21 (m, 3H), 7.13 (dt, .1; = 1.00 Hz, J2 = 7.49 HZ, 1H), 7.02 (dd, J,- = 1.00 Hz, J2 = 7.60 Hz, 1H), 4.36 (s, W0 2012/116452 2H), 4.23 (s, 2H); 13C NMR (DMSO, 125 MHz) 5 141.19, 140.84, 140.12, 139.48, 133.01, 131.45, 131.19, 130.04, 129.89, 128.87, 127.85, 127.65, ,126.65, 126.59, 60.71, 60.44. 2,2'-(S-(lH-tetrazol—S-yl)thiophene-Z,3—diyl)dibenzaldehyde (DNM0556) iH NMR(CDC13, 500 MHz) 5 9.91 (s, 1H), 9.80 (s, 1H), 7.93 (s, 1H), 7.75 (d, J= 7.80 Hz, 2H), 7.59 (dt, J, = 1.35 Hz, J; = 7.55 Hz, 1H), 7.49 - 7.41 (m, 3H), 7.34 (t, J= 7.53 Hz, 1H), 7.18 (d, J: 7.50 Hz, 1H); 13C NMR(CDC13, 125 MHz) 5 192.30, 191.20, 153.22, 140.91, 139.47,137.51, 134.79, 134.72, , 134.40,134.04, 133.00, 132.44,132.21, 130.28, , ,129.32, 126.53. -(4,5-Di(furanyl)thiophen-Z-yl)—1H~tetrazole (DNM0557) ]H NMR (DMSO, 500 MHz) 5 8.03 (dd, J, = 0.93 Hz, J, = 1.38 Hz, 1H), 7.93 (dd, J, = 0.90 Hz, J; = 1.30 Hz, 1H), 7.87 (s, 1H), 7.83 (14,]; 1.60 Hz, 1H), 7.77 (t, J= 1.70 Hz, 1H), 6.63 (dd, J, = 0.83 Hz, J2 = 1.83 Hz, 1H), 6.59 (dd, J, ~—~ 0.80 Hz, J2 = 1.80 Hz, 1H); 13(3 NMR (DMSO, 125 MHz) 5 144.46,143.86, 141.52, 140.76, 131.73, 130.93, 130.30, 119.66, , 110.91, 110.38. -(4,5-Di(thi0phen—3—yl)thiophen-2—yl)-1H—tetrazole (DNMOSSS) ‘H NMR (DMSO, 500 MHz) 5 7.89 (s, 1H), 7.68 (dd, J, = 1.33 Hz, J, = 2.88 Hz, 1H), 7.65 (dd, J, = 2.95 Hz, J2 = 4.95 Hz, 1H), 7.61 (dd, J, = 2.95 Hz, J, = 4.90 Hz, 1H), 7.59 (dd, J, = 1.28 Hz, J2 = 2.88 Hz, 1H), 7.03 (dd, J, = 1.18 Hz, J2 a 4.93 Hz, 1H); 13C NMR (DMSO, 125 MH2)813S.88, 135.23, 133.66, 132.73, 131.40, 127.61,127.59, 127.52, 126.80, 125.14,123.88. -(4,5-Bis(2-(thiophenyl)phenyl)thiophen—2—yl)—lH-tetrazole (DNM0559) 1H NMR (DMSO, 500 MHz) 5 7.65 (s, 1H), 7.40 — 7.35 (m, 2H), 7.32 (dt, J, = 1.15 Hz, W0 2012/116452 J; = 7.51 Hz, 1H), 7.29 - 7.23 (m, 3H), 7.16 - 7.00 (m, 2H), 6.90 (dd, J; = 1.23 Hz, J2 a 2.88 Hz, 1H), 6.85 (dd, J1= 1.23 Hz, J2 = 2.88 Hz, 1H), 6.73 (d, J= 7.65 Hz, 1H), 6.64 (d, J= 7.65 Hz, 1H), 6.58 (dd, J} = 1.20 Hz, J2 : 4.95 Hz, 1H), 6.53 (dd, J]: 1.20 Hz, J; 24.95 Hz, 1H); l3CNMR (DMSO, 125 MHz) 5 142.01, 140.99, 140.62, 139.74, 135.76, 135.02, 132.86, 131.72, 130.96, , 129.91, 129.73, 129.37, 128.68, 127.94, , 127.73,'127.16, 127.06, 125.56, 125.39, 123.16, 122.78. -(4,5—Bis(2-( furanyl)pheny1)thiophen—Z-yl)-1H-tetrazole (DNM0560) 1H NMR (DMSO, 500 MHz) 5 7.61 (s, 1H), 7.59 - 7.56 (m, 2H), 7.38 - 7.31 (m, 3H), 7.30 - 7.25 (m, 2H), 7.25 - 7.22 (m, 1H), 7.19 — 7.14 (m, 1H), 7.12 ((11, J; = 1.30 Hz, J; = 7.58 Hz, 1H), 6.96 (d, J= 7.54 Hz, 1H), 6.85 (dd, J; = 0.95 Hz, J2 = 7.66 Hz, 1H), 6.20 (dd, J; = 0.83 Hz, J2 = 1.78 Hz, 1H), 6.12 (dd, .71 = 0.80 Hz, J2 = 1.75 Hz, 1H); 130 NMR (DMSO, 125 MHZ) 5 142.01, 140.99, 140.62, 139.74, 135.76, , 132.86, 131.72, 130.96, 130.16, 129.91, 129.73, 129.37, 128.68, 127.94, 127.81, 127.73,127.16,127.06, 125.56, 125.39, , 122.78. -(4,5—Bis(2-chloromethylphenyl)thiophen-Z—yl)—1H—tetrazole (DNM0563) 1H NMR (DMSO, 500 MHz) 5 7.85 (s, 1H), 7.36 - 7.32 (m, 2H), 7.29 (d, J= 7.80 Hz, 1H), 7.06 (d, J= 7.74 Hz, 1H), 7.09-7.02 (m, 2H), 2.30 (s, 3H), 2.28 (s, SE); 13C NMR (DMSO, 125 MHz)6141.00, 139.73, , 132.59, 132.54, 131.88, 131.38, , 130.57, 130.19, 130.03, 128.06, 127.87, 127.82, 20.39, 10.37. -(4,5-Bis(4-methoxy-3,S-dimethylphenyl)thiophenyl)—lH—tetrazole (DNM0564) 1H NMR (DMSO, 500 MHz) 8 7.83 (s, 1H), 7.03 (s, 2H), 6.99 (5, 21-1), 3.67 (s, 6H), 2.172 (s, 6H), 2.166 (s, 6H); ”C NMR (DMSO, 125 MHz) 8 156.89, 156.12, W0 16452 140.69, 137.89, 131.78, 130.86, 130.45, 130.27, 129.19, 128.99, 127.91, 59.43, 59.42, .78, 15.74. -(4,5—Bis(3-chloromethoxyphenyl)thiophen—2—yl)—1H—tetrazoie 65) IH NMR (DMSO, 500 MHz) 5 7.87 (s, 1H), 7.41 (d, J= 2.05 Hz, 1H), 7.39 (d, J= 2.00 Hz, 1H), 7.30 (dd, J1= 2.15 Hz, J2 = 8.65 Hz, 1H), 7.22 (dd, J; = 2.08 Hz, J; = 8.58 Hz, 1H), 7.20 (d, J= 8.70 Hz, 1H), 7.17 (d, J= 8.65 Hz, 1H), 3.89 (s, 3H), 3.88 (s, 3H); l3C NMR (DMSO, 125 MHz) 8 172.03, , 154.03, 139.42, 136.95, 131.63, , 129.84, 129.19, 128.70, 127.88, 125.48, 121.39, 121.18, 113.24, 113.01, 56.29, 56.18.

Diethyl 3,3’-(5—(1H—tetrazol-S-y1)thiophene-2,3-d1yl)dibenzoate (DNM0593) lH NMR (CDC13, 500 MHZ) 5 8.17 (s, 1H), 8.14 (s, 1H), 8.06 (s, 1H), 8.03 (d, J= 7.75 Hz, 1H), 7.98 (d, J= 7.75 Hz, 1H), 7.48 (d, J= 7.90 Hz, 1H), 7.44 - 7.37 (m, 3H), 7.34 (t, J= 7.75 Hz, 1H), 4.44 - 4.35 (m, 4H), 1.42 — 1.35 (m, 6H); 13C NMR , 125 MHz) 167.33, 166.35, , 138.70, 135.47, 133.75, 133.73, 133.17, 132.49, 131.28, 131.27, , 130.44, 130.20, 129.85, 129.20, 128.99, 128.97, 124.48, 61.88, 61.64, 14.39. 1,1'—(3,3'-(5-(1H—tetrazol-S-ylfihiophene—2,3-diyl)bis(3,1-phenylene))dibutan- 1-0ne(DNM0599) lH NMR(CDC13, 500 MHz) 5 8.13 (s, 1H), 8.08 (s, 1H), 7.98 — 7.89 (m, 3H), 7.54 (d, J= 7.80 Hz, 1H), 7.46 (t, J= 7.60 Hz, 1H), 7.40 (t, J= 7.70 Hz, 1H), 2.94 (t, J"—' 7.28 Hz, 2H), 2.85 (t, J= 7.30 Hz, 2H), 1.80 — 1.67 (m, 4H), 0.99 (t, J: 7.35 Hz, 3H), 0.98 (t, J= 7.35 Hz, 3H); 13C NMR (CDC13, 125 MHz) 5 200.52, 139.04, 137.96, 137.80, 136.06, 134.09, 134.00, 133.71, 132.47, 129.80, 129.51, 129.44, 129.04, 128.58, 128.04, 41.20, 41.06, 18.23, 18.18, 14.28.

W0 2012/116452 2012/050130 Diethyl 5,5‘-(S-(1H—tetrazol-S—yl)thiophene-Zfi—diyl)bis(3-hydroxybenzoate) (DNM0607) 1H NMR (DMSO, 500 MHz) 5 10.12 (s, 1H), 9.99 (s, 1H), 7.87 (s, 1H), 7.43 — 7.29 (m, 4H), 6.97 (s, 1H), 6.93 (s, 1H), 4.35 - 4.17 (m, 4H), 1.33 — 1.20 (m, 6H); 13C NMR (DMSO, 125 MHZ) 5 , 165.07, 157.89, , 139.98, 138.09, 136.36, 133.97,131.78,131.60,131.02,120.17,120.10,120.01,115.86, 115.11, 60.91, 60.81, 14.07, 14.00. —(4,5—Bis(3—butylphenyl)thiophen-Z-yl)-IH-tetrazole (DNM0608) 1H NMR (CDClg, 500 MHz) 6 7.44 (s, 1H), 6.91 — 6.56 (m, 8H), 8.06 (s, 1H), 2.31 - 2.05 (m, 4H), 1.29 — 1.16 (m, 4H), 1.16 — 1.00 (m, 4H), 0.81 - 0.67 (m, 6H); 13C NMR (CDC13, 125 MHz) 5 142.80, 142.62, 139.91, 138.91, 136.26,134.07,129.86, 129.58, 129.54, 128.54, 127.63, 127.02, 126.88, 126.61, 35.70, 35.68, 33.79, 33.68, 22.66, 22.63, 14.33. -(4,5-bis(3-(cyclopcntylmethyl)phenyl)thiophen-Z-yl)~1H-tetrazole (DNM0612) II-I NMR (CDC13, 500 MHz) 5 8.00 (s, 1H), 7.24 - 7.17 (m, 2H), 7.17 - 7.09 (m, 4H), 7.09 - 7.04 (m, 2H), 2.54 - 2.48 (m, 4H), 2.00 — 1.87 (111, 21-1), 1.69 — 1.54 (m, 8H), 1.54 — 1.41 (m, 4H), 1.17 - 1.00 (m, 4H); 131: NMR (CDC13, 125 MHz) 5 152.34, 144.09, 143.03, 142.82, 139.94, , 133.02, 132.94, 129.97, 129.77, 129.12, , 128.57, 128.14, 126.76, 126.41, 122.42, 42.08, 41.99, 32.56, 25.06.

Example 2 sis of DNM0576 and analogues Br Br sz(dba)3, man-1),, KF Pd,(dba),. P(B 11-03. KF Br CN ”1 N dioxanc S dioxanc flNaNJ, mar, ”2.2—- / N DMF. Ar1 An CN If HN"‘N W0 2012/116452 4-Bromo-S—(4—methylnaphthalen-l-yl)thiophene—2—carbonitrile Around- bottomed flask was charged with 4,S—dibromothiophene-Z—carbonitrile (536 mg, 2.00 mmol), 4—methylnaphthalen—I-ylboronic acid (409 mg, 2.20 mmol), Pd2(dba)3 (18.3 mg, 0.020 mmol) and KF (383 mg, 6.60). After degassed, e (5 mL) and P(Bu—t)3 (0.24 mL, 0.2M, 0.048 mmol) was added. The reaction mixture was stirred at rt until te. mL of water was added, and the reaction mixture was extracted with ethyl acetate. The organic phase was dried over anhydrous sodium sulphate and then concentrated. The residue was purified by flash chromatography (hexane: CHZClz = 3: l). 0.58 g (88 %) of product was obtained as a white solid. 4—(4~Chloromethylphenyl)(4-methylnaphthalen-l-yl)thiophene—2— carbonitrile A round—bottomed flask was charged with 4-bromo—5-(4- methylnaphthalen—l-yl)thiophene—2—carbonitrile (203 mg, 0.62 mmol), ro methylphenylboronic acid (119 mg, 0.70 mmol), Pd2(dba)3 (9.2 mg, 0.010 mmol) and KF (126 mg, 2.17). After degassed, dioxane (2.0 mL) and P(Bu—t)3 (0.15 mL, 0.2M, 0.03 mmol) was added. The reaction mixture was stirred at rt until complete. 20 mL of water was added, and the on mixture was extracted with ethyl e. The organic phase was dried over anhydrous sodium sulphate and then concentrated. The residue was purified by flash tography (hexane: CH2C12 = 3: 1). 0.21 g (90.6 %) of product was obtained as a white solid. -(4—(4-Chloromethylphenyl)(4-methylnaphthalen-l-yl)thiophen-2—yl)- 1H-tetrazole (DNM0576) A round-bottomed flask was charged with 4-(4—ehloro~2— methylphenyi)-5—(4-methylnaphthalen—l-y1)thiophenecarbonitrile (209 mg, 0.56 mmol), zinc bromide (338 mg, 1.50 mmol) and sodium azide (97.5 mg, 1.50 mmol).

W0 2012/116452 After degassed, DMF (3 mL) was added. The reaction mixture was heated to 110 °C and stirred at this temperature until complete. The on was cooled to rt, and 30 mL of 0.1 N aqueous HCl was added. The reaction e was extracted with ethyl acetate. The organic phase was dried over anhydrous sodium sulphate and then concentrated. The residue was purified by flash chromatography (hexane: EtOAc: AcOH = 30: 10: 1). 196 mg (84 %) of product was obtained as a white solid, 1H NMR (DMSO, 500 MHz) 5 8.03 (d, J74 8.40 Hz, 1H), 7.88 (s, 1H), 7.80 (d, J= 8.05 Hz, 1H), 7.55 (t, J= 7.68 Hz, 1H), 7.48 (t, J: 7.65 Hz, 1H), 7.45 (d, J= 7.19 Hz, 1H), 7.37(d, J= 7.30 Hz, 1H), 7.23 (d, J‘2 1.95 Hz, 1H), 7.09 (d, J= 8.25 Hz, 1H), 7.04 (dd, J1= 8.25 Hz, J; = 2.06 Hz, 1H), 2.65 (s, 3H), 2.17 (s, 3H); 13C NMR (DMSO, 125 MHz) 6 172.05, 141.01, 139.65, 138.35, , 133.73, 132.18,132.10,131.64, ,131.26,129.80, 129.23, 127.62, 126.45, 126.26, 126.02, 125.53, 125.51, 124.67, 21.08, 19.82.

The following compounds were also ed using an analogous method: -(4-(4-Chlorophenyl)—5—(4—methylnaphthalen—1—y1)thi0phen—2~yl)—1H~ tetrazole (DNM05‘72) 1H NMR (CDC13, 500 MHz) 5 8.09 (d, J= 8.39 Hz, 1H), 7.84 (s, 1H), 7.82 (d, J= 8.46 Hz, 1H), 7.56 (t, J= 7.32 Hz, 1H), 7.44 (t, J= 7.84 Hz, 1H), 7.31 (d, J= 7.25 Hz, 1H), 7.2301, J= 7.24 Hz, 1H), 7.17—7.09 (m, 4H), 2.76 (s, 311); ”c NMR (CDCI3, 125 MHz) 6 152.64, 142.65, 138.24, 135.00, 134.09, 133.65, , 132.16, 131.91, 131.87, 129.76, 128.93, 127.95, 126.54, 126.37, 126.30, 126.11, 124.78, , 19.80. —(4—(5-Chlor0phenyl)—4—(4-methylnaphthalen—l-yl)thiophen-2—yl)—1H— tetrazole (DNM0575) 1H NMR (DMSO, 500 MHz) 8 8.11 (s, 1H), 8.09 (d, J= 8.44 Hz, 1H), 7.65 (d, J= 8.11 Hz, 1H), 7.58 — 7.53 (111,211), 7.49 - 7.42 (m, 2H), 7.28 — 7.20 (m, W0 2012/116452 4H), 2.72 (s, 3H); ”C NMR (DMSO, 125 MHz) 5 139.74, 139.15, 136.18, 133.66, 132.29, 132.15, 131.14, 130.40, 129.40, 129.23,128.61, 127.69, 126.69, 126.36, 126.28, 125.47,124.77, 19.23. 4-Fluorophenyl)-5—(4—(methylsulfonyl)phenyl)thi0phen-2—yl)—1H- tetrazole (DNM0592) 1H NMR (DMSO, 500 MHz) 5 7.96 (s, 1H), 7.94 (d, J= 8.45 Hz, 2H), 7.57 (d, J= 8.45 Hz, 2H), 7.45 _ 7.38 (m, 2H), 7.28 (t, J: 8.80 Hz, 2H), 3.27 (s, 3H); l3c NMR(DMSO, 125 MHz) 5 , , 141.61, 139.83, 139.71, 137.14, 131.45,131.38,131.32, 129.58, 129.42, , 128.42,127.46, 124.88,116.37, 116.19, 43.36.

Ethyl 3—(2-(4-methylnththalen-1—yl)-5—(1H-tetrazol-S-yl)thiophen yl)benzoate (DNM0596) 1H NMR (01301;, 500 MHz) 5 8.00 (d, J= 8.40 Hz, 1H), 7.96 (1, J= 1.64 Hz, 1H), 7.87 (s, 1H), 7.83 (dt, J; = 7.85 Hz, .72 = 1.35 Hz, 1H), 7.73 (d, J: 8.36 Hz, 1H), 7.35 (1, J= 7.60 Hz, 1H), 7.28 (d, J= 7.79 Hz, 1H), 7.22(d, J= 7.74 Hz, 1H), 7.17 (d, J= 7.10 Hz, 1H), 7.14 (1, J2 7.85 Hz, 1H), 4.24 (q, J= 7.07 Hz, 2H), 2.68 (s, 3H), 1.24 (t, J= 7.15 Hz, 3H); 13C NMR (CDC13, 125 MHz) 6166.77, 143.43, 138.88, 135.13, 134.17, 133.51, 133.05, ,132.03, 131.41,130.72, 129.59,129.31,128.91, 127.95,126.46, 126.36, 126.14, 124.71, 123.55, 61.60, 19.76, 14.21.

Ethyl 3-(2—(4-chloro—2—methylphenyl)—5—(lH-tetrazol—S-yl)thiophen yl)benzoate (DNM0597) 1H NMR (CD013, 500 MHz) 5 8.01 — 7.95 (m, 2H), 7.78 (s, 1H), 7.41 = 8.22 Hz, J2 = 1.50 Hz, 1H), 7.14 (d, — 7.33 (m, 2H), 7.23 (s, 1H), 7.19 (dd, J, J= 8.22 Hz, 1H), 4.38 (q, J= 7.15 Hz, 2H), 2.04 (s, 3H), 1.40 (t, J= 7.14 Hz, 3H); 131: NMR (CDC13, 125 MHz) 5166.58, 142.77, 138.62, 138.53, 134.15, , 133.52, 132.76, 132.50, 131.59, , , 129.59, 129.35, 129.24, 126.57, 124.34, 61.74, .16, 14.39.

Example 3 Synthesis ofDNM0574 and analogues / l) ammo] N ArB(Gl-l)z ska; / ) Br 2)McOH ”(WW4"“2003 s dioxanc + H10 C)mAcOH+NaOAc 4,5—Dibromothiazole To a solution of 2,4,5-triibromothiazole (3.12 g, 9.67 mmol) in anhydrous THF (25 mL) was added i-PngCl (4.84 mL, 2M in THF, 9.67 mmol) at ice-sait bath temperature under argon. After te addition, the on was stiired for 1h at ice-salt bath temperature, and then quenched with methanol (2 mL). The reaction was worked up with a typical procedure, and the crude product was purified by flash chromatography (30 % ofDCM in hexane) to afford 1.45 g of product. 4,5-Bis(4-chloro-Z-methylphenyl)thiazole A round-bottomed flask was charged with 4,5-dibromothiazole (160 mg, 0.66 mmoi), ro-2—methylphenylboronic acid (269 mg, 1.58 rnmol) and Pd(PPh3)4 (38.1 mg, 0.033 mmol). Afier degassed, dioxane (5 mL) and aqueous sodium carbonate (3 mL, 2M, 6 mmol) was added. The reaction mixture was heated to 90 °C. The progress of the on was monitored by TLC. After the reaction was complete, 50 mL of water was added, and the reaction mixture was extracted with ethyl acetate. The organic phase was dried over anhydrous sodium sulphate and then trated. The residue was purified by flash chromatography to afford 170 mg ofproduct as clear oil.

Z-Bromo—4,S-bis(4-ch10ro-Z-methylphenyl)thiazole A solution of 4,5—Bis(4- chloro-2—methy1phenyl)thiazole (170 mg, 0.51 mmol), NBS (100 mg, 0.56 mmol) and NaOAc (82 mg, 1.0 mmol ) in AeOH (5 mL) and DCM (3 mL) was stirred overnight at rt, and then quenched with water (25 mL) and extracted with ethyl acetate. The organic phase was dried over anhydrous sodium sulphate and then concentrated. The residue was purified by flash chromatography to afford 196 mg of product. 4,5-Bis(4-chloromethylphenyl)thiazole—Z—carbonitrile A round~bottomed flask was charged with 2—Bromo—4,5~bis(4-chloromethylphenyl)thiazola (196 mg, 0.47 mmol) and CuCN (84.2 mg, 0.94 mmol). After degassed, 3 mL ofDMF was added. The reaction mixture was heated to 150 °C ght. After cooled to room temperature, the reaction was quenched with 25 mL of water, and extracted with ethyl acetate. The organic phase was dried over anhydrous sodium sulphate and then concentrated. The residue was purified by flash chromatography to afford 150 mg of t. 4,5—Bis(4—chloro—2—methylphenyl)—2-(lH-tetrazoI—S-yl)thiazole (DNM0574) A round-bottomed flask was charged with 4,5-bis(4-chloromethylphenyl)thiazole—2— itrile (150 mg, 0.42 mmol), zinc bromide (189 mg, 0.84 mmol) and sodium azide (45.5 mg, 0.84 mmol). After degassed, DMF (3 mL) was added. The reaction e was heated to 150 °C overnight. After cooled to rt, the reaction was quenched with 30 mL of 0.1 N aqueous HCl, and extracted with ethyl e. The organic phase was dried over anhydrous sodium sulphate and then concentrated. The residue was purified by flash tography (hexane: EtOAc: AcOH = 30: 10: l). 120 mg of product was obtained as a white solid, lH NMR (DMSO, 500 MHz) 8 7.93 (d, J= 8.41 Hz, 11-1), 7.57 (d, J= 1.90 Hz, 1H), 7.49 - 7.44 (m, 2H), 7.33 — 7.28 (m, 2H), 2.63 (s, 3H), 2.18 (s, 3H); ”C NMR (DMSO, 125 MHz) 5 166.07, 153.61, 139.50,138.74,134.92, 133.48, 132.53, 131.87, 131.42, 130.27, 129.98, 126.64, 125.67, 21.13, 19.51.

The following compounds were also prepared using an analogous method: 4,5~Bis(5-chloromethylpheny1)(1H—tetrazol—S-yl)thiazole (DNM0567) 1H NMR (DMSO, 500 MHz) 5 7.95 (d, J= 2.17 Hz, 1H), 7.54 (dd, J; = 8.25 Hz, J2 = 1.80 Hz, 11-1), 7.48 (d, J»?- 8.25 Hz, 1H), 7.45 (dd, J; = 8.23 Hz, J2 = 2.33 Hz, 1H), 7.41 — 7.36 (111, 21-1), 2.61 (s, 3H), 2.11 (s, 3H); 13C NMR (DMSO, 125 MHz) 5 165.31, 152.99, , 135.54, 135.31, 133.72, 133.07, 132.04, 130.99, 130.09, 129.94, 129.64, 128.79, 128.72, 20.82, 19.03. 4,5-Bis(3—chloromethylphenyl)(1H-tetrazol-S—yl)thiazole (DNM0568) 1H NMR (DMSO, 500 MHz) 8 8.09 (d, J= 1.30 Hz, 1H), 7.92 (dd, J; = 7.93 Hz, J; = 1.38 Hz, 1H), 7.79 (d, J= 1.20 Hz, 1H), 7.57 - 7.50 (m, 2H), 7.43 (d, J= 7.95 Hz, 1H), 2.42 (8, 31-1), 2.39 (5, 31-1); 13(3 NMR (DMSO, 125 MHz) 5 166.30, 153.18, , 136.51, 134.29, 133.22, 132.68, 132.20, 131.49, 131.20, 129.01, 127.53, 126.22, 125.36, 19.68, 19.50. 4,5-Bis(4-methylnaphthalen—l—yl)—2—(1H-tetrazol—S-yl)thiazole (DNM0569) 1H NMR (DMSO, 500 MHz) 8 9.99 (d, J= 7.91 Hz, 1H), 8.16 (t, Jfi 9.52 Hz, 2H), 8.04 (d, J= 7.35 Hz, 1H), 7.86 (d, J= 8.41 Hz, 1H), 7.47 - 7.49 (m, 6H), 7.47 (d, J= 7.30 Hz, 1H), 2.764 (s, 3H), 2.761 (s, 3H); 13'C NMR (DMSO, 125 MHz) 5 167.71, 154.55, , 135.74, 132.61, 132.41, , 129.79, 129.58, 129.06, 128.26, , 127.41, 126.69, 126.34, 126.23, 126.13, 126.04, 125.93, 125.83, , 124.59, 19.52, 19.23. s(4-chloropheny1)(1H-tetrazol-5—yl)thiazole (DNM0573) 1I-INMR (DMSO, 500 MHZ) 6 8.10 (d, J= 8.43 Hz, 2H), 7.74 (d, J= 8.44 Hz, 2H), 7.64 (d, J= 7.50 Hz, 2H), 53 (d, J= 8.50 Hz, 2H); ”C NMR (DMSO, 125 MHz) 5 166.61, 153.57, 135.94, 133.87, 132.11, 130.85, , 129.52, 128.50, 128.29. 4,5-Bis(4-f1uomnaphthalen—l-yl)—2-(1H-tetrazol-5~yl)thiazo1e(DNM0578) lI-I NMR (DMSO, 500 MHZ) 6 9.02-8.95 (m, 1H), 8.24-8.16 (m, 3H), 7.88 (d, J= 8.49 Hz, 1H), 7.79-7.73 (m, 1H), 7.73—7.67 (m, 2H), 7.62 (t, J= 7.80 Hz, 1H), 7.58 (dd, J1: 8.21 Hz, J2 =10.14 Hz, 1H), 7.47 (dd, J, = 7.96 Hz, J; =10.s4 Hz, 1H); 13C NMR (DMSO, 125 MHZ) 6 166.76, 160.47, 159.60, , 157.59, 153.30, 132.99, , 131.08, 130.07, 129.99, 129.08, 128.99, 128.92, 127.82, 127.58, 126.92, 125.63, 123.39, 123.26, 123.23, , 120.54, 120.49, 120.35, 120.30, 109.92, , 109.55, 109.40. 4,5-Bis(4-biphenyl)—2—(lH—tetrazol—S-yl)thiazole (DNM0581) 1H NMR (DMSO, 500 MHz) 6 8.17 (d, .1: 8.32 Hz, 2H), 7.89 (d, J= 8.32 Hz, 2H), 7.84-7.72 (m, 8H), 7.56-7.47 (m, 4H), .38 (m, 2H); 13C NMR (DMSO, 125 MHz) 6 167.64, 154.70, 142.75, 140.67, 139.37, 138.93, 132.38, 131.10, 129.43, 129.12, 129.05, 128.23, 127.84, 127.60, 127.17, 126.79, 126.70, 126.68. 4,5-Bis(4ni-butylphenyl)—2—(1H—tetrazol—5—yl)thiazole (DNM0582) 'H NMR (DMSO, 500 MHz) 6 7.99 (d, J = 8.25 Hz, 2H), 7.60 (d, J= 8.25 Hz, 4H), 7.48 (d, .13 8.30 Hz, 2H), 1.34 (s, 9H), 1.32 (s, 9H); 13C NMR (DMSO, 125 MHz) 6 167.93, 155.03, 154.21, 151.65, 130.58, 129.56, 128.52, 126.35, , 125.27, 34.77, 34.51, 31.02, .87. 4,5-Bis(3,4—dimethylphenyl)-2—(1H-tetrazolyl)thiazole (DNM0583) 1H NMR (DMSO, 500 MHz) 6‘ 7.35 (s, 1H), 7.79 (dd, J, = 1.65, J2 = 7.80 Hz, 1H), 7.47 (s, W0 2012/116452 1H), 7.34 (d, J= 7.95 Hz, 1H), 7.29 (d, J= 7.70 Hz, 1H), 7.19 (d, J= 7.35 Hz, 1H), 2.34 (s, 3H), 2.31 (s, 3H), 2.28 (s, 3H), 2.25 (s, 3H); ”c NMR (DMSO, 125 MHz) 5 168.65, 155.73, 140.64, 138.00, 137.85, , 131.33, 130.85, 130.32, 130.17, 129.93, 127.70, , 124.49, 19.91, 19.87, 19.75, 19.71. 4,5-Bis(4-chloro-Z—isopropylphenyl)(lH—tetrazol-S-yl)thiazole (DNM0584) II-I NMR (DMSO, 500 MHz) 5 7.73 (d, J= 8.35 Hz, 1H), 7.62 (d, J= 2.20 Hz, 1H), 7.52 (d, J= 2.10 Hz, 1H), 7.47 (dd, J; = 2.18, J; = 8.33 Hz,1H), 7.30 (dd, J; = 2.13, J2 = 8.23 Hz, 1H), 7.25 (d, J= 8.20 Hz, 1H), 3.77 (sep, J= 6.85 Hz, 1H), 2.93 (sep, J= 6.83 Hz, 1H), 1.21 (d, J= 6.85 Hz, 6H), 1.11 (d, J= 6.80 Hz, 6H); 13c NMR (DMSO, 125 MHz) 6 166.64, 154.25, 150.51, 150.06, 136.15, 134.62, 132.72, 132.50, 131.94, 129.84, 127.13, 126.93, 126.22, 126.16, 30.57, 29.36, 23.72, 23.59.

Example 4 Synthesis of DNM0577 and analogues 1 1' N | N15 or“NRC, . \ 1) 1 / \ F TFA + H250, DMF / ON 2) ArICHO s | CN 0 S l I H / 5551“ y / 10,13th __ on TFA + 01,121., on P11216611), Pam-1),. KF ”1 An N Ara / \ NaN], 2113!: b / \ s s \f'I An N1 HN—N 4,5-Diiodothiophene—Z-carboxamide Thiophene—Z-cm‘bonitrile (5.39 g, 49.39 mmol) was dissolved in a mixture 01112804 (10 mL) and TFA (30 mL). NIS (23.34 g, 103.74 mmol) was added in n in 30 min. After 00mplete addition, the reaction e was further stirred for 4h, and then poured onto 300 g of crushed ice. A white solid was formed, which was collected, washed with water, and dried together with phosphorous pentoxide under vacuum to afi'ord 17.8 g (95 %) ofproduct. 4,5-Diiodothiophene—Z—carbonitrile An oven-dried round~bottomed flask was charged with iodothiophene-2—carboxamide (5.68 g, 15.0 mmol) and 20 mL of DMF under an argon atmosphere. The solution was cooled on an ice-water bath, and cyanuric chloride (1.81 g, 9.8 mmol) was then added in one portion. After stirring at 0 °C for 1 h, the reaction mixture was warmed to room temperature, and stirred for a further 3h. 100 mL of water was added. A white solid was formed, which was collected h n filtration, washed with water, and dried together with orous pentoxide under vacuum to afford 4.87 g (90 %) of product. —((4-Chlor0phenyl)(hydroxy)methyl)—4-iodothiophene-Z-carbonitrile An oven-dried round—bottomed flask was charged with 4,5-diiodothiophene-Z—carbonitrilc (1.80 g, 5.0 mmol). After degassed, anhydrous THF (15 mL) was added h syringe.

After cooled to ~78 °C, i-PngCl (3 mL, 2.0 M, 6.0 mmol) was added dropwise. The reaction was further stirred for 30 min at —78 °C after complete addition. 4— chlorobenzaldehyde (0.98 g, 7.0 mmol) in 5 mL THF was then added. After stirring for min at -78 °C for 1 h, the reaction mixture was warmed to room temperature. 50 mL of saturated s NH4C1 was added. The reaction mixture was extracted with ethyl acetate. The organic phase was dried over anhydrous sodium sulphate and then concentrated. The residue was purified by flash chromatography (hexane: EtOAc: CH2CL2 = 30: 3: 10). 1.39 g (74 %) of product was obtained as a white solid. .5-(4-Chlorobenzyl)-4—iodothiophenecarbonitrile A round-bottomed flask was charged with 5~((4—chlorophenyl)(hydroxy)methyl)iodothiophene—2-carbonitrile (1.67 g, 4.45 mmol). After degassed, dichloromethane (8 mL), TFA (4 mL) and triethylsilane (1.1 mL, 6.9 mmol) was added sequentially through e. The reaction was further stirred at rt until complete, and then concentrated. The residue was purified by flash chromatography (hexane: CH2CL2 = 2: 1). 1.36 g (85 %) of product was obtained as a white solid.

S-(4-Chlorobenzyl)—4~(4—methylnaphthalen—l-yl)thiophene-Z-carbonitrile A bottomed flask was d with 5-(4-chlorobenzyl)-4~iodothiophene—2— carbonitrile (180 mg, 0.50 mmol), 4-methylnaphthalen-l-ylboronic acid (102 mg, 0.55 mmol), Pd2(dba)3 (9.2 mg, 0.010 mmol) and KF (126 mg, 2.17). After degassed, dioxane (2.0 mL) and P(Bu-t)3 (0.15 mL, 0.2M, 0.03 mmol) was added. The reaction mixture was stirred at rt until complete. 20 mL of water was added, and the reaction mixture was extracted with ethyl acetate. The organic phase was dried over anhydrous sodium te and then concentrated. The residue was purified by flaSh chromatography (5 % ethyl acetate in hexane). 0.17 g (91 %) of product was obtained as a white solid. —(5-(4—Chlorobenzyl)-4—(4-methylnaphthalen—1-yl)thiophen—2—yl)—1H— ole (DNM0577) A round-bottOmed flask was charged with 5-(4-chlorobenzy1) (4-methylnaphthaleny1)thiophene-2~carbonitrile (170 mg, 0.45 mmol), zinc bromide (338 mg, 1.50 mmol) and sodium azide (97.5 mg, 1.50 mmol). After ed, DMF (3 mL) was added. The reaction mixture was heated to 110 °C and stirred at this temperature until complete. The reaction was cooled to It and 30 mL of 0.1 N aqueous HCl was added. The on mixture was extracted with ethyl acetate. The organic phase was dried over anhydrous sodium sulphate and then concentrated. The residue was purified by flash chromatography e: EtOAc: AcOH = 30: 10: 1). 161 mg (85 %) of product was obtained as a white solid, lH NMR(CDC13, 500 MHZ) 5 8.05 (d, J= 8.46 Hz, 1H), 7.73 (s, 1H), 7.63 (d, J= 8.36 Hz, 1H), 7.53 (t, J= 7.58 Hz, 1H), 7.42 (t, J= 7.53 Hz, 1H), 7.33 (d, J= 7.11 Hz, 1H), 7.23 (d, J= 7.08 Hz, 11-1), 7.14 (d, J= 8.25 Hz, 2H), 6.92 (d, J= 8.14 Hz, 2H), 3.86 (ABq, 2H, MAB = 0.06, JAB = 15.90 Hz), 2.70 (s, 3H); 13C NMR (CDC13, 125 MHz) 8 152.61, 145.54, 139.59, 137.96, 135.22, 132.97, 132.76, 132.69, 132.27, , 130.05, 128.81, 127.55, 126.33, 126.25, 126.20, 126.16, 124.75, 122.29, 34.24, 19.77.

The following compounds were also prepared using an analogous method: -(5-(4—Chlorobenzyl)—4—(4—chlorophenyl)thiophenyI)-1H-tetrazole 79) 1H NMR (DMSO, 500 MHz) 8 7.80 (s, 1H), 7.57 (d, Ja 8.70 Hz, 2H), 7.53 (d, J “—" 8.70 Hz, 2H), 7.40 (d, J= 8.34 Hz, 21-1), 7.26 (d, J= 8.34 Hz, 2H), 4.30 (s, 2H); 13C NMR (DMSO, 125 MHz)6143.21, 138.47, , 133.64, 132.51, 131.51, 130.65, 130.49, 130.24, 128.89, 128.65, 122.97, 32.99. -(5—(4—Chlorobenzyl)—4~(4—chloro—2-isa—propylpheny1)thiophenyl)—1H- ole (DNM0580) 1H NMR , 500 MHZ) 5 7.66 (s, 1H), 7.31 (d, J= 2.05 Hz, 1H), 7.19 (d, J= 8.35 Hz, 2H), 7.16 (dd, J, = 8.12 Hz, J2 = 2.10 Hz, 1H), 6.99 (d, J= 8.20 Hz, 1H), 7.23 (d, J= 78.35 Hz, 2H), 3.89 (ABq, 2H, ASAB = 0.04, JAB = 14.13 Hz), 2.76 (sep, J= 6.85 Hz, 1H), 1.10 - 0.97 (m, 6H); ”c NMR (CD013, 125 MHz) 5 152.60, 149.93, 144.93, 139.52, 137.57, 134.85, 132.95, , 131.93, 131.70, 130.02, 129.01, 126.41, 126.15, 122.28, 30.02, 30.85, 24.57, 23.42. 4-Chlorobenzyl)—4-(4-fluoronaphthalen-l-yl)thiophen-2—y1)—1H- tetrazole (DNM0587) 1H NMR (DMSO, 500 MHz) 5 8.17 (d, J'= 8.25 Hz, 1H), 7.75 ~ 7.69 (m, 2H), 7.69 - 7.62 (m, 2H), 7.53 - 7.42 (m, 2H), 7.30 - 7.25 (m, 2H), 7.11 - 7.06 (111,211), 4.10 - 3.91 (m, 2H); 13C NMR (CDC13, 125 MHz) 5 158.86, 156.86, 144.75, 138.33, 137.42, 132.85, 132.81, 131.60, 131.28, 130.30, 128.78, 128.75, 128.41, 127.93, 127.85, 127.78, 126.99, 125.40, 125.38, 123.10, 122.97, 120.43, 120.39, 109.53, 109.37, 33.01, 21.07. -(4-(BiphenyI-Z-yl)(4—chlorobenzyl)thiophen-2~yl)-1H—tetrazole (DNM0588) 1H NMR (DMSO, 500 MHz) 6 7.58 — 7.52 (m, 1H), 7.52 - 7.46 (m, 3H), 7.43 (d, J= 7.60 Hz, 21-1), 7.33 - 7.24 (m, 5H), 7.20 — 7.13 (m, 2H), 6.89 (d, J= 8.33 Hz, 2H), 3.73 (s, 2H); 130 NMR (DMSO, 125 MHz) 6 143.34, 140.80, 140.48, 139.40, 138.13, 133.20,131.66, , 130.76, 130,42, 130.30,129.03, 128.69, , 128.23, 127.71, 127.03, 122.21, 33.01.

Ethyl 3-(2-(4-01110robenzyI)(1H—tetrazol—S—yl)thiophen—3—yl)benzoate (DNM0595) IH NMR (DMSO, 500 MHz) 5 8.09-7.93 (m, 2H), 7.87 (s, 1H), 7.80 (d, J= 7.24 Hz, 1H), 7.66 (t, J= 7.71 Hz, 1H), 7.40 (d, J= 7.86 Hz, 2H), 7.26 (d, J= 7.86 Hz, 2H), 4.52 — 4.14 (m, 4H), 1.31 (1, J= 7.07 Hz, 3H); 13c NMR (DMSO, 125 MHz) 5 , 143.15, , 138.45, 135.17, 133.07, 131.49, 130.65, 130, 48,130.40,129.46, , 128.64, 128.37, 123.12, 60.94, 32.96, 14.11. 1~(3-(2-(4—Chlorobenzyl)—5-(1H—tetrazol-S-yl)thiophen-3—yl)phenyl)butan one (DNM0600) 1H NMR (DMSO, 500 MHz) 5 8.00 — 7.97 (m, 2H), 7.88 (s, 1H), 7.77 (d, J= 7.70 Hz, 1H), 7.65 (t, J= 7.95 Hz, 1H), 7.39 (d, J= 8.40 Hz, 2H), 7.26 (d, J= 8.40 Hz, 2H), 4.32 (s, 2H), 2.98 (t, J= 7.15 Hz, 2H), 1.63 (sex, J= 7.30 Hz, 1H), 0.93 (t, J= 7.35 Hz, 3H); l3c NMR (DMSO, 125 MHz) 8 199.80, , 138.94, 138.55, 137.20, 135.19, 132.80, , 130.68, 130.42, , 128.66, 127.75, 127.08, 123.27, 39.85, 32.99, 17.17, 13.61. -(4-(3~Butylphenyl)(4—chlorobenzyl)thiophen-Z-yl)-1H—tetrazole 06) 1H NMR (DMSO, 500 MHz) 5 7.81 (s, 1H), 7.42 - 7.34 (m, 3H), 7.32 — 7.26 (m, 2H), 7.26 ~ 7.21 (m, 3H), 4.30 (s, 2H), 2.62 (t, J= 7.65 Hz, 2H), 1.56 (qi, J= 7.55 Hz, 2H), 1.31 (sex, J= 7.54 Hz, 2H), 0.90 (t, .1: 7.37 Hz, 3H); ‘30 NMR (DMSO, 125 42.95, 139.99, 138.72, 134.71, 131.41, 130.87, 130.39, 128.77,128.59, 128.58, 128.34, 127.76, 125.68, 122.74, 34.73, 33.04, 21.75, 13.79.

Ethyl 3-(2—(4-chlorobenzyl)~5~(1H—tetrazolyl)thiophenyl)-5— ybenzoate (DNM0609) 1H NMR (DMSO, 500 MHz) 5 10.12 (s, 1H), 7.81 (s, 1H), 7.45 (s, 1H), 7.43 - 7.38 (m, 3H), 7.27 (d, J= 8.35 Hz, 2H), 7.14 (s, 1H), 4.34 - 4.25 (m, 4H), 1.30 (t, J= 7.10 Hz, 3H); 13C NMR (DMSO, 125 MHz) 8 165.41, 157.89, 150.98, 142.84, 138.79, 138.53,136.37, 131.64, 131.47, 130.45, 130.38, 128.64, 123.18, 119.84, 119.74, 114.99, 60.87, 32.99, 14.11. -(5-(4-Chlorobenzyl)—4—(3-(cyclopentylmethyl)phenyl)thiophen—2-yl)-IH- tetrazole (DNM0610) ]H NMR (DMSO, 500 MHz) 5 7.81 (s, 1H), 7.41 —7.43 (m, 3H), 7.30 (d, J= 7.70 Hz, 1H), 7.26 (s, 1H), 7.25 - 7.19 (m, 3H), 4.30 (s, 2H), 2.61 (d, J= 7.45 Hz, 2H), 2.10 — 2.01 (m, 1H), 1.67 — 1.54 (m, 4H), 1.54 - 1.41 (m, 2H), 1.21 - 1.09 (m, 2H); [31: NMR (DMSO, 125 MHz) 8 150.80, 142.42, 142.22, 140.05, 138.70, 134.61, 131.41, 130.92, 130.34, 128.70, 128.67,128.58, 128.11, 125.68, 122.65, 41.32, 41.17, 33.01, 31.90, 24.48. 4—Chlorobenzyl)(1H—tetrazolyl)thiophenyl)phenol (DNM0613) 1H NMR (DMSO, 500 MHz) 5 9.65 (s, 1H), 7.65 (s, 1H), 7.40 (d, J = 8.35 Hz, 2H), 7.29 (t, J: 7.88 Hz, 1H), 7.26 (d, J= 8.40 Hz, 2H), 6.91 (d, J= 7.90 Hz, 1H), 6.87 (s, 1H), 6.82 (dd, J; = 1.79 Hz, J; = 8.10 Hz, 1H), 4.29 (s, 2H); 13C NMR (DMSO, 125 MHz) 5 157.66, 142.45, 139.94, 138.72, 136.01, 131.46, 130.76, 130.44, 129.94, 128.64, , 115.20, 114.78, 33.07.

~(S-(4-Chlorobenzyl)—4-(3-methoxyphenyl)thiophenyl)—1H—tetrazole (DNM0615) 1H NMR (DMSO, 500 MHz) 8 7.84 (s, 1H), 7.47 - 7.40 (m, 3H), 7.29 (d, J = 8.45 Hz, 2H), 7.09 (d, J= 7.80 Hz, 1H), 7.06 — 7.04 (m, 1H), 7.02 (dd, J; = 2.00 Hz, J2 = 8.20 Hz, 1H), 4.34 (s, 2H), 3.81 (s, 3H); 13C NMR (DMSO, 125 MHZ) 5 159.49, 142.69, 139.68, 138.70, 136.13, 131.45, 130.87, , 129.99, , 120.68, 113.83, 113.50, 55.14, 33.06. -(5-(4-Chl0robenzyl)-4—(3—butoxyphenyl)thiophenyl)-1H-tetrazole (DNM0616) 1H NMR (DMSO, 500 MHz) 5 7.81 (s, 1H), 7.44 - 7.36 (m, 3H), 7.26 (d, J = 8.35 Hz, 2H), 7.05 (d, J= 7.60 Hz, 1H), 6.99 — 6.95 (m, 2H), 4.31 (s, 2H), 3.95 (t, J: 6.50 Hz, 2H), 1.69 (pen, J= 6.96 Hz, 2H), 1.43 (sex, J= 7.50 Hz, 2H), 0.94 (t, J= 7.40 Hz, 2H); 13C NMR (DMSO, 125 MHz) 5 158.90, 150.96, 142.40, 139.75, 133.74, 136.10, 131.43, 130.85, 130.39, 129.96, 128.63, 122.80, 120.51, 114.13, 67.14, 33.04, 30.73, 18.72, 13.72. -(5—(4~Chiorobenzyl)—4—(3~ethoxyphenyl)thiophen—yl)—lI-I—tetrazole (DNM0617) lH NMR (DMSO, 500 MHz) 5 7.81 (s, 1H), 7.44 - 7.37 (m, 3H), 7.26 (d, J = 8.40 Hz, 2H), 7.05 (d, J= 7.65 Hz, 1H), 7.01 - 6.95 (m, 2H), 4.31 (s, 2H), 4.04 (q, J= 7.00 1), 1.33 (t, J= 7.00 Hz, 3H); 130 NMR (DMSO, 125 MHz) 6 158.74, 142.67, WO 16452 139.70, 138.71, 136.11, 131.44, 130.89, 130.43, 130.01, 128.63, 120.54, 114.20, 114.01, 63.05, 33.05, 14.63. -(5-(4-Chlorobenzyl)—4-(3-propoxyphenyl)thiophen—2—yl)-lH-tetrazole 18) 1H NMR (DMSO, 500 MHz) 5 7.82 (s, 1H), 7.43 - 7.36 (m, 3H), 7.26 (d, J = 8.40 Hz, 2H), 7.05 (d, J= 7.65 Hz, 1H), 7.00 - 6.95 (m, 2H), 4.31 (s, 2H), 3.91 (1:, J= 6.60 Hz, 2H), 1.73 (sex, 7.06, 2H), 0.98 (t, J= 7.40 Hz, 3H); 13(1 NMR (DMSO, 125 MHz)6158.88,142.50, 139.74, 138.73, 136.09, 131.43, 130.91, 130.39, 129.97,128.63, 122.72, , 114.14, 68.92, 33.04, 22.02, 10.37.

Example 5 Synthesis ofDNM0461 and analogues /DPM PPM H i1:» N NM.

N\ NH _ 1.leq.Ar,13(0H), TM N130“); CHZCIl \ A:2Pd(PP}li‘))_i‘°::n§azCO] \ figfiyzco: NI l / | / r1", N 1=1 ,“F1f N\ NH I MN, 2,3,2 PhICI-IOI-LTEOH """B Wr \ / DMF \ Mme N "H 221:4- Brlrimmon), CH2C12 \ meringfixmragco, i —'N |\ ' / / -(3,5-Dibromophenyl)—1H—tetrazole A round—bottomed flask was charged with 3,5-dibromobenzonitrile (15.65 g, 60.00 mmol), sodium azide (7.80 g, 120.00 mmol) and zinc bromide (27.00 g, 120.00 mmol). Afier degassed, DMF (100 mL) was added. The reaction mixture was heated to 120 °C and stirred at this temperature until complete. The reaction was cooled to rt and then in an ice-water bath. 300 mL of 1N aqueous HCl was added. The white itate formed was collected by suction filtration, washed with water, and dried together with phosphorous pentoxide under vacuum to afford 17.32 g (95 %) of t. 2—Benzhydryl-5—(3,5—dibromophenyl)-2H-tetrazole A suspension of 5-(3,5- dibromophenyl)—lH-tetrazole (6.08 g, 20.00 mm01), diphenylmethanol (3.68 g, 20.00 mmol) and TsOH.I-I20 (0.38 g, 2.00. mmol) in toluene (50.0 mL) was heated to 100 °C until a clear solution was obtained. The solution was cooled to rt, and filtered. The filtration was concentrated, and the residue was purified by recrystallization in ether and hexane to afford 8.00 g (85 %) of product as a white solid. hydryl-5—(3,5-bis(4-methylnaphthalen-l-yl)phenyl)-2H-tetrazole A round—bottomed flask was charged with 2-benzhydryl(3,5-dibromophenyl)-2H— tetrazole (282 mg, 0.60 mmoi), 4-methylnaphthalen-l-ylboronic acid (251 mg, 1.32 mmol) and Pd(PPh3)4 (35 mg, 0.03 nunol). After degassed, dioxane (5 mL) and aqueous sodium carbonate (3 mL, 2M, 6.0 mmol) was added. The reaction mixture was heated to 90 °C until the reaction was complete. 30 mL of water was added, and the on mixture was ted with ethyl acetate. The organic phase was dried over anhydrous sodium sulphate and then concentrated. The residue was purified by flash chromatography (hexane: CHZCIZ = 2: 1) to afford 0.32 g (90 %) of product as a white solid. -(3,5-Bis(4-methylnaphthalen—l~yl)phenyl)-2H-tetrazole (DNM0461) 2— Benzhydryl—S-(3,5-bis(4—methylnaphthalen~1—y1)phenyl)-2H-tetrazole (0.32 g, 0.54 mmol) was dissolved in dichloromethane (2 mL). Anisole (0.3 mL, 2.76 mmol) and TFA wo 2012/116452 (1 mL) were added sequentially. The on was stirred overnight, and then concentrated. The e was d by flash chromatography (hexane: ethyl acetate: AcOH = 30: 10: 1) to afford 0.22 g (95.5 %) of product as a white solid, 1H NMR (DMSO, 500 MHz) 5 8.24 (d, J= 1.55 Hz, 2H), 8.25 (d, J: 8.15 Hz, 2H), 8.03 (d, J= 8.20 Hz, 2H), 7.72 (s, 1H), 7.65 (t, J= 7.58 Hz, 2H), 7.60 (t, J= 7.44 Hz, 2H), 7.56 (d, J = 7.15 Hz, 2H), 7.51 (d, J = 7.29 Hz, 2H), 2.74 (s, 6H); 13c NMR (DMSO, 125 MHz) 5 141.58, 136.38, 134.51, , 132.43, 130.72,127.15, 127.03, 126.51, 126.32, 126.11, , 124.76.19.21.

The following compounds were also prepared using an analogous method: —(2,5~Bis(4—methylnaphthalen-1—yl)phenyl)—1H-tetrazole(DNM0446) 1H NMR (DMSO, 500 MHz) 5 8.16 (d, J= 8.45 Hz, 1H), 8.06 (d, J= 8.41 Hz, 2H), 7.97 (s, 1H), 7.80 (dd, J, = 7.84 Hz, J; = 1.80 Hz, 1H), 7.69 — 7.49 (m, 7H), 7.42 (t, J= 7.56 Hz, 1H), 7.39 (d, J= 7.09 Hz, 1H), 7.28 (d, J= 7.09 Hz, 1H), 2.73 (s, 3H), 2.70 (s, 3H); 13C NMR (DMSO, 125 MHz)5139.96, 138.93, 136.37, 135.51, 134.38, 133.98,132.46, 132.21, 132.06, 132.03, 131.37, 130.98, 130.80, 127.11, 127.00, 126.43, 126.34, 126.10, 126.04, 125.89, 125.76, 125.73, 124.73, 124.45, 19.23, 19.14. -(2,5-Bis(4-chloromethylpheny1)phenyl)—lH-tetrazole (DNM0447) 1H NMR (DMSO, 500 MHZ) 5 7.79 (s, 1H), 7.65 (d, J= 7.16 Hz, 1H), 7.51 - 7.43 (m, 2H), 7.38 (s, 2H), 7.33 (s, 1H), 7.22 (d, J= 8.09 Hz, 11-1), 7.05 (d, J= 8.14 Hz, 1H), 2.33 (s, 3H), 1.97 (s, 311); 13C NMR (DMSO, 125 MHz) 5 139.71, 138.72, 132.58, 138.20, 138.13, 137.59,132.41, 132.10, 131.39, 131.20, 131.08, 130.96, 130.10, , 129.00, 126.06, 125.39, 20.05, 19.56.

W0 2012/116452 S—(3,5-Bis(4-chlorophenyl)phenyl)-2H-tetrazole (DNM0470) 1H NMR (DMSO, 500 MHz) 8 8.34 (d, J= 1.69 Hz, 2H), 8.16 (d, .1: 1.69 Hz, 2H), 7.95 — 7.91 (m, 4H), 7.65 — 7.61 (m, 4H); 13(3 NMR (DMSO, 125 MHz) 8 140.76, 137.76, 133.22, 129.07,128.94, 127.58, . -(4'—Chloro—S—(4—methylnaphthalen-I-yl)biphenylyl)-1H-tetrazole (DNM0480) 1H NMR (DMSO, 500 MHz) 8 8.42 (t, J= 1.60 Hz, 1H), 8.16 (d, J= 7.98 Hz, 1H), 8.14 (t, J= 1.58 Hz, 1H), 7.95 (t, J= 1.62 Hz, 1H), 7.92 (d,J= 7.90 Hz, 1H), 7.90 (d, J= 8.70 Hz, 2H), 7.67 — 7.63 (m, 1H), 7.61 (d, J= 8.70 Hz, 2H), 7,594.55 (m, 1H), 7.54 - 7.50 (m, 2H), 2.76 (s, 3H); 13C NMR(CDC13, 125 MHz) 8 143.14, 141.32, 138.28, ,134.98, 134.46, 133.04, 131.81, 131.53,129.36, 128.72, 128.11, 127.06, , 126.31,126.26, 126.08, 125.13, 124.79, 19.84. ~(3,5_Bis(4-fluoronaphthalen-I~yl)phenyl)—2H-tetrazole (DNM0539) 1H NMR (DMSO, 500 MHz) 5 8.26 (d, J= 1.55 Hz, 2H), 8.19 (dd, J; = 8.15 Hz, J2 = 1.87 Hz, 2H), 8.05 (d, J= 7.87 Hz, 2H), 7.77 (t, J= 1.55 Hz, 1H), 7.75-7.65 (m, 6H), 7.49 (dd, J, = 10.60 Hz, .12 = 7.95 Hz, 2H); 130 NMR (DMSO, 125 MHz) 8 158.81, 156.81, 140.71, 134.46, , 133.80, 132.00, 131.97,128.67,127.55, 127.48, 127.46, 126.96, 125.44, 125.42, 123.15, 123.02, 120.46, 120.42, 109.60, 109.44.

Example 6 Synthesis ofDNM0566 and analogues W0 2012/116452 U _NBL... n mm), —-—-————1— coca TFA +]{ 50 1 4 s Br COOEt Hymn”:‘Na co,mane+ kn Ar LiOI-l ”‘0 + “‘0“ coca Ar COOH Ethyl 4,5-dibromothiophene—Z-carboxylate To a stirred solution of ethyl thiophene-Z-caroxylate (12.62g, 80.8mmol) in 12 mL of sulfuric acid and 40 mL of TFA was added NBS (32.00g, 177.8mmol) in portions during 2~3 hours. After stirred overnight at room ature, the reaction e was poured onto ice. The white precipitate formed was collected by suction filtration, and purified by tallization in methanol. 23.38g (92%) ofproduct was obtained as a white solid, mp: 47.0 - 48.0 (lit. mp 48.0 m 49.0 °C, Bull. Chem. Soc. Jpn. 1991, 64 (8), 2566—8).

Ethyl 4,5-bis(4-chloroisopropylphenyl)thiophene—2—carhoxylate A round- bottomed flask was charged with ethyl bromothiophene-Z-carboxyiate (235.5 mg, 0.75 mmol), 4-chloroisopropylphenyboronic acid (327.5 mg, 1.65 mmol) and Pd(PPh3)4 (43 mg, 0.0375 mmol). After degassed, dioxane (5 mL) and aqueous sodium carbonate (3 mL, 2M, 6.0 mmol) was added. The on mixture was heated to 90 °C until the reaction was complete. After cooled to room temperature, the reaction e was diluted with 30 mL of water and then extracted with ethyl acetate. The organic phase was dried over anhydrous sodium sulphate and then concentrated. The residue was purified by flash chromatography (hexane: CI-12C12 m 2: 1) to afford 0.28 g (81 %) of product as a white solid. 4,5-Bis(4-chloro~2-isopropylphenyl)thiophene-Z—carboxylic acid (DNM0566) To a solution of ethyl 4,5-bis(4—chloro—2-isopropylphenyl)thiophene—2-carboxylate (280 mg, 0.61 mmol) in THF (3 mL) and MeOH (3 mL) was added a soiution of LiOH (72 mg, 3.0 mmol) in water (2 mL). The reaction was d at rt until complete, and then concentrated. The residue was re-dissolved in 20 mL r and acidified with 1N aq.

HCl to pH 2. The white precipitate was extracted with ethyl acetate. The c phase was dried over anhydrous sodium sulphate and then concentrated. The e was purified by recrystallization in a mixture t of ethyl acetate and hexane to afford 0.24 g (91 %) of product as a white solid, II-l NMR(CDC13, 500 MHz) 5 7.78 (s, 1H), 7.23 (d, J= 2.15 Hz, 1H), 7.22 (d, J= 1.88 Hz, 1H), 7.13 - 7.07 (m, 2H), 7.03 (dd, J1= 8.23 Hz, J2 = 2.20 Hz, 1H), 6.89 (d, J= 8.26 Hz, 1H), 2.95 (m, 2H), 1.04'(d, J= 4.94 Hz, 6H), 0.98 (d, J= 6.95 Hz, 6H); ”C NMR (013013, 125 MHz) 6 167.13, 149.87, 149.06, 146.58, 139.45, 137.21, 135.53, 134.39,132.79, 131.84, 131.57, 131.14, 129.01, 126.43, 126.28, 125.86, 125.77, 30.27, 30.23, 23.93.

The following compounds were also prepared using an analogous method: 4,5-Bis(4-biphenyl)thiophene—Z-carboxylic acid (DNM0497) 1H NMR (CDClg, 500 MHz) 6 13.34 (s, 1H), 7.86 (s, 1H), 7.72 - 7.65 (m, 8H), 7.50 - 7.32 (m, 10H); 13(3 NMR (CDCI3, 125 MHZ) 5 162.69, 143.94, 140.16, 139.30, 139.09, , 138.17, 135.56, 134.08, 133.00, 131.98, 129.46, 129.38, 129.02, 128.98, 127.87, 127.63, 127.12, 126.86, 126.61, 126.53. s(4-chlorophenyl)thiophene—Z-carboxylic acid 98) 1H NMR (CDC13, 500 MHz) 8 13.39 (s, 1H), 7.81 (s, 1H), 7.46 (d, J= 8.40 Hz, 2H), 7.42 (d, J= 8.40 Hz, 2H), 7.32 (d, J= 8.50 Hz, 2H), 7.29 (d, J4 8.42 Hz, 2H); 13(J NMR(CDC13, 125 MHz) 6 162.53, 143.06, 137.68, 135.30, 133.60, 133.47, 133.44, 132.47, 131.46, 130.74, 130.64, 129.13, 128.78. 4,5—Bis(5-ch10ro-Z-methylphenyl)thiophene-Z—carboxylic acid (DNM0501) lH NMR(CDC13, 500 MHz) 6 13.38 (s, 1H), 7.77 (s, 1H), 7.36 (d, J= 2.20 Hz, 1H), 7.34 (dd, J; = 2.20 Hz, J; = 8.15 Hz, 1H), 7.26 — 7.22 (m, 3H), 7.05 (s, 1H), 2.01 (s, 3H), 1.96 (s, 3H); 13C NMR (CDC13, 125 MHz) 5 162.63, 143.31, 138.84, 136.36, 135.58, 134.85, 134.75, 133.77, 133.76, 132.18, 132.08, 130.47, 130.07, 129.86, 129.60, 128.88, 127.74, 19.22, 19.09. 4,5-Bis(3—chlorophenyl)thiophene—Z-carboxylic acid (DNMOSOZ) 1H NMR (CDC13, 500 MHz) 5 13.34 (s, 1H), 7.85 (s, 1H), 7.47 (d, J; 8.10 Hz, 1H), 7.44 - 7.34 (m, 5H), 7.26 (d, J= 7.65 Hz, 1H), 7.20 (d, J= 7.45 Hz, 1H); 130 NMR (CDC13, 125 MHz) 5 162.50, 142.92, 137.70, , 135.23, 134.51, 133.77, 133.50, , 130.87, 130.48, 128.75, 128.56, , , , 127.63. 4,5-Bis(2,4-dimethylpheny1)thiophene-Z-carboxylic acid (DNM0503) 1H NMR (CDC13, 500 MHz) 5 13.19 (s, 1H), 7.66 (s, 1H), 7.12 (d, J= 8.25 Hz, 1H), 7.03 - 6.95 (m, 3H), 6.87 - 6.81 (m, 2H), 2.24 (s, 3H), 2.22 (s, 3H), 2.07 (s, 3H), 1.93 (s, 3H); 13C NMR (CDC13, 125 MHz) 5 162.82, 144.83, 139.77, 138.16, , , 135.28, 135.22, 132.49, , 130.98, 130.91, 130.02, 129.43, 126.45, 126.27, 20.66, 20.61, 19.78, 19.64. 4,5-Bis(4-chlor02-methylphenyl)thiophene-Z-carboxylic acid (DNM0561) 1H NMR (CDC13, 500 MHZ) 8 7.81 (s, 1H), 7.17 (d, J= 2.02 Hz, 1H), 7.14 (s, 1H), 7.13 - 7.11 (m, 2H), 7.03 (dd, J1= 8.24 Hz, J2 = 2.02 Hz, 1H), 6.87 (d, J= 8.24 Hz, 1H), 2.12 (s, 3H), 2.03 (s, 3H); 13C NMR (CD013, 125 MHz) 5 167.74, 147.16, 140.15, 139.20, 138.26, 137.56, 135.29, 134.06, 133.61, 132.85, 131.97, 131.74, 131.13, 131.03, 130.88, 126.50, 126.43, 20.61, 20.59.

Example 7 Synthesis ofDNM0631 and ues ['L Br Br NJ”.

NBS N’L‘u / \ SOCI; A :I “‘0“ OOH Br COOH ”HI-”=0 Br com-I, 13”“ S S S Br Ar Ar ); NaN,, my, \ / / \ n Pd may, N: no, DMF B \N r on 8 ne -|- P 2{0 Ar ON s \ l/ ’ 015:?8r m NaN3, 2113:, d ‘:i d on DMF ““ 2-Bromo—3-methylthiophene NBS (8.90 g, 50.0 mmol) was added portion wise to a stirred solution of 3—mehy1thiophene (4.90 g, 50.0 mmol) in acetic acid (20 mL) at rt.

After complete addition, the reaction was stirred at rt until it was te. The reaction mixture was poured into ice-water, and then extracted with a 3:1 mixture t of hexane and ether. The organic layer was washed with 1N aq. NaOH and brine. After dried over anhydrous sodium sulphate, the organic phase was concentrated on vacuum to afford 2-bromo—3~methylthiphene (8.20 g, 92.7 %), which was used directly in the next step without further purification. 3-Methylthiophene-2—carbonitrile A round-bottomed flask was charged with 2- brornoInethylthiphene (8.20 g, 46.3 mmol)‘and CuCN (8.29 g, 92.6 mmol). After degassed, 50 mL ofDMF was added. The reaction mixture was heated to 150 °C W0 2012/116452 overnight. After cooled to room temperature, the reaction was quenched with 250 mL of water, and extracted with ethyl acetate. The c phase was dried over anhydrous sodium sulphate and then concentrated. The residue was passed through a short and fat silicon column, and the column was eluted with hexane to afford 5.07g of product. 4,5-Dibromo-S-methylthiophene~2-carbonitrile Bromine (4.25 mL, 82.6 mmol) was added drop wise to a stirred on of 3—mehylthiophene—Z-carbonitrile (5.07 g, 41.2 mmol) in DMF (20 mL) at rt. After te addition, the on was heated to 60 °C until it was complete. After cooled to rt, the reaction mixture was poured into ice- water. The light yellow solid formed was collected through suction filtration, and dried together with P205 under vacuum to afford 9.84 g of 4,5~dib1‘omo-3—methylthiophene—2— carbonitrile. 4,5—Bis(4—chloroisopropylphenyl)—3-methylthiophene—Z-carbonitrile A round-bottomed flask was charged with 4,5~dibromo—3-methylthiophene—2-carbonitrile (140.5 mg, 0.50 mmol), 2-(4-chlor0—2—isopropylphenyl)—4,4,S,S-tetramethyl-l,3,2- dioxaborolane (3 08.7 mg, 1.10 mmol) and Pd(PPh3)4 (29 mg, 0.025 mmol). After degassed, dioxane (5 mL) and aqueous sodium carbonate (3 mL, 2M, 6.0 mmol) was added. The on mixture was heated to 95 °C until the on was complete. After cooled to room temperature, the reaction mixture was diluted with 30 mL of water and then extracted with ethyl acetate. The organic phase was dried over anhydrous sodium sulphate and then concentrated. The residue was purified by flash chromatography (hexane: CH2C12 = 4: 1) to afford 190.5 mg (89 %) of product as a white solid.

-Bis(4-chloro-2—isopropylphenyl)~3-methylthiophenyl)-1H—tetrazole (DNM0631) A round-bottomed flask was charged with 4,5—bis(4—chloro-2— wo 2012/116452 isopropylphenyl)—3 -methy11hiOphene-Z-carbonitrile (190.5 mg, 0.44 mmoi), zinc bromide (338 mg, 1.50 mmol) and sodium azide (97.5 mg, 1.50 mmol). After degassed, DMF (3 mL) was added. The reactiOn mixture was heated to 130 ”C and stirred at this temperature until complete. The reaction was cooled to It and 3 0 mL of 0.1 N aqueous HCl was added. The reaction mixture was extracted with ethyl acetate. The organic phase was dried over anhydrous sodium sulphate and then concentrated. The residue was purified by flash chromatography (hexane: EtOAc: AcOI—I 2 30: 10: 1). 188.7 mg (90 %) of product was obtained as a white solid, 1H C13, 500 MHZ) 5 7.25 - 7.22 (m, 2H), 7.18 (dd, J1= 8.18 Hz, J2 = 2.13 Hz, 1H), 7.09 - 7.06 (m, 2H), 7.02 (d, J= 8.20 Hz, 1H), 3.04 (sep, J= 6.75 Hz, 1H), 2.74 (sep, J= 6.80 Hz, 1H), 2.37 (s, 3H), 1.12 (d, J2 6.80 Hz, 3H), 1.05 (d, J= 6.80 Hz, 6H), 0.90 (d, J= 6.80 Hz, 3H); l3C NMR (CDCI3, 125 MHZ) 5 , 149.87, 141.11, 140.81, 140.72, 135.33, 134.47,132.72, 132.02, 131.51, 129.01, 126.30, 126.20, 125.84, 125.52, 30.35, 30.10, 24.76, 23.60, 23.13, 15.78.

The following compounds were also ed using an analogous : —(4,5—bis(4-chloro-2—methylphenyl)~3-methylthiophen-Z-yl)-1H-tetrazole (DNM0614) lH NMR(CDC13, 500 MHz) 5 7.15 (s, 13), 7.11 (s, 1H), 7.08 (d, J= 8.20 Hz, 11-1), 7.05 ~ 7.01 (m, 2H), 6.93 (d, J= 8.30 Hz, 1H), 2.31 (s, 3H), 2.17 (s, 3H), 2.04 (s, 3H); 13C NMR (CDC13, 125 MHz) 5 , 140.34, 139.95, 139.41, 139.16, 134.57, 133.83, 133.81, , 132.20, 131.43, 130.52, 130.42, , 126.00, 20.81, 20.03, .56. -(4,5-Bis(4—methylnaphthalen-l-yi)-3—methylthiophenyl)-1H-tetrazole (DNM0620) 1H NMR (CDC13, 500 MHz) 6 8.04 (d, J= 8.30 Hz, 1H), 7.91 (d, J= 8.30 Hz,1H), 7.86 (d, J= 8.40 Hz, 1H), 7.77 (dd, J; = 8.30 Hz, J; = 1.10 Hz, 1H), 7.48 — 7.37 WO 16452 (m, 3H), 7.32 (t, J= 7.65 Hz, 1H), 7.18 (d, J= 7.20 Hz, 1H), 7.06 - 7.00 (m, 3H), 2.55 (s, 3H), 2.54 (s, 3H), 2.28 (s, 3H); l3c NMR(CDC13, 125 MHz) 5 , 141.77, 141.39, 135.21,134.23, 132.73, 132.52, 132.44, 132.35, 131.52, , 128.72, 127.94, 126.61, 126.30, 126.07, 125.82, 125.74, 125.70, 125.64, 125.52,124.44, 124.10, 119.63, 19.49, 19.42, 15.54. —(4,S-Bis(3-butylphenyl)—3-methylthiophen—2—yl)—lH—tetrazole (DNM0627) ‘H NMR (CD013, 500 MHz) 5 7.30 (1, J= 7.53 Hz, 1H), 7.17 (d, Jm 7.70 Hz, 1H), 7.15 (t, J= 7.60 Hz, 1H), 7.10 - 6.97 (m, 5H), 2.59 (t, J= 7.58 Hz, 2H), 2.48 (s, 3H), 2.46 (1., J = 7.75 Hz, 2H),1.57 - 1.49 (111,211), 1.45 - 137011.211), 1.33 - 1.18 (m, 4H), 0.90 (t, J = 7.35 Hz, 3H), 0.89 (1, J= 7.30 Hz, 3H); 131: NMR (CDC13, 125 MHz) 8 143.22, 143.19, ,141.28, 141.17, 135.74, 133.10, 130.46, 129.16,128.47, 128.30, 128.15, 127.67, 127.45, 126.23, 125.52, 35.42, 35.36, 33.56, 33.26, 30.31, 22.15, 22.07, 15.64, 13.92,. —(4,S-Bis(4-fluoronaphthalenyl)~3-methylthiophen-Z—yl)-1H-tetrazole (DNM0628) 1H NMR(CDC13, 500 MHz) 6 8.10 - 8.06 (m, 1H), 8.03 (d, J= 8.30 Hz, 1H), 7.96 (d, J= 8.50 Hz, 1H), 7.76 - 7.71 (m, 1H), 7.56 - 7.44 (m, 3H), 7.39 (1, J= 7.50 Hz, 1H), 7.22 (dd, J, = 5.40 Hz, J2 = 7.80 Hz, 1H), 7.08 (dd, J, = 5.38 Hz, .12 = 7.83 Hz, 1H), 6.95 - 6.87 (m, 2H), 2.34 (s, 3H); ”0 NMR (CD013, 125 MHz) 5 159.96, 159.45, 157.93, 157.44, 141.79,141.44, 133.85, 133.81,133.52, 133.49, 128.95, 128.89, 128.71, 128.67, 128.08, 128.02, 127.36,127.24, ,126.20, 125.67, , 125.39, , 123.59, 123.56, 123.43,121.05,121.01,120.70,120.66, 109.03, 108.87, 108.77, 108.61, .53.

Example 8 Measurement ofAcpS inhibition Materials W0 2012/116452 [3H]Acetyl-C0A was mixed as 1 volume ofPerkin Elmer NET290 radiolabeled stock (e. g. 0.1 rnCi/ml, 3.7 Ci/mmol in Na—acetate, pH 4.5-5.0 = 27 uM) with 1.2 volumes of 1 mM unlabelled acetyl—CoA (Sigma). Trichioroacetic acid was prepared as a 10% w/v solution. Bovine serum albumin was ed as a 25 mg/ml solution in water.

DTT was prepared as a 50 mM solution in water.

Procedure 1. A standard reaction contained the following in a total volume of 10 til: Stock reagents Volume (LL13 Final concentration 1 M Na—phosphate, pH 7 0.5 50 mM 0.1 M MgClz 1 10 mM 50 mM DTT 1 5 mM yl carrier n (ACP) 4.4 pg 50 uM AcpS enzyme ~0.06 pg 570 uM [3H]Acetyl-C0A 1 57 11M Test compound e.g. DNM0488 0.5 predetermined uM e.g. 500 or 50 ddeo to 10 pl 2. Tubes were incubated at RT. [3H]acetyi—C0A was added last to start the reaction.

To stop the reaction, 2 ul was removed to a 1.5»ml microfuge tube containing 800 nl cold 10% TCA. Up to four time points were collected for each on, e.g. at min, 10 min, 30 min, and 60 min. 3. 20 pl of 25 mg/ml BSA was added to each tube, mixed and incubated on ice for min, then fuged at 12,000 g for 5 min to form a pellet. Supernatant was removed using P1000. 4. Each pellet was washed twice with 800 pl of cold 10% TCA each time, then centrifuged at 12,000 g for 5 min after each wash. The supernatant was discarded.

Each pellet was ended in 50 ul of formic acid. The suspension was transferred to a scintillation vial and radioactivity was measured in 2 ml of liquid scintillation cocktail.

The random (stochastic) error in scintillation counting is proportional to the square root of the counts (not dpm) accumulated in the counting period; i.e. 10% for 100, 3.3% for 1,000, 1% for 10,000, etc. ed radioactive counts (dpm) are in the thousands, the assay provides useful data. s are reported in terms of% activity with respect to the substracted counts for working substrate.

A % activity of less than about 40% is indicative ofAcpS enzyme inhibition by the test compound at that concentration, For results, see the table in the Example below.

Example 9 Measurement ofminimum inhibitory concentrations (MICs) Susceptibilities to the compounds were determined using the National Committee for Clinical Laboratory Standards (NCCLS) M7-A6 broth microdilution method. Cation- adjustcd Mueller~Hinton broth (Ca2+, 25 pg/mL; Mg“, 12.5 pg/mL) microdilution panels were ed to n antimicrobial doubling dilution concentrations of an appropriate range. DMSO (diinethylsulfoxide) controls were orated into the panel to mimic the quantity ofDMSO used in dissolving some of the compounds at the higher concentrations. Each final panel well volume was 100 uL with a bacterial inoculum of 5 x 105 CPU (colony forming units) / mL of the relevant bacterium. Panels were read following 16 to 20 h of tion at 35 degrees Celsius in ambient air. The MIC (minimum inhibitory concentration) was defined as the lowest concentration of antimicrobial inhibiting e growth.

The following table, as well as indicates results of experiments of both Examples 8 and 9 where ble, demonstrating AcpS inhibition and antimicrobial effects on Methicillin-Resistant Staph. aureus (MRSA) and E. coli D22: % AcpS activity % AcpS MIC MIC E Compound @ 500 activity MRSA coii D22 ID uM at 50 mm (uM) (uM) DNMO486 0.16 0.82 2 >125 DNM0487 0.08 3.34 8 >125 DNM0488 0.32 11.49 4 125 DNM0489 0 17.89 2 >125 DNM0504 0.16 7.33 125 125 DNM0508 0.1 44.92 4 >125 DNM0509 0.2 31.07 15 >125 DNM0512 0.7 37.29 8 >125 DNM0531 4.38 96.76 60 >125 DNM0534 0 33.98 8 >125 DNM0535 3.48 92.62 125 >125 DNM0536 0.2 8.25 8 125 DNM0537 0.14 1.28 2 125 DNM0538 0.58 82.18 >125 DNM0541 0.3 63.12 mmmmmoocnmoo4>4>moo >125 2 0.7 9.58 >125 DNM0546 5.16 37.21 >125 DNM0548 0.1 3.1 60 9 0.48 71.31 >125 DNM0550 0.39 47.83 mm >125 9 0.5 7.01 >125 DNM0560 0.06 63.58 _|. >125 DNM0563 0.56 15.12 125 DNM0564 0.75 47.61 >125 DNM0565 0.11 22.91 >125 DNM0570 0.19 33.7 AAA 125 DNM0571 0.11 8.19 >125 DNM0575 0.23 5.86 >125 DNM0572 0.51 4.41 >125 6 0.15 5.88 >125 DNM0577 0.4 5.54 >125 DNM0579 3.02 15.74 125 DNM0580 0.85 1.62 125 DNM0587 2.13 DNM0588 2.67 DNM0446 25.36 >125 DNM0447 0.47 32.56 ._|. >125 DNM0453 23.76 >125 DNM0457 1.56 60 DNM0461 0.85 13.39 >125 DNM0464 0.28 9.03 >125 DNM0470 27.69 125 DNM0474 0.3 15.08 ,>125 DNMO479 0.1 12.79 >125 2.04 DNM0480 7.35 125 2.7 DNM0493 25.28 >125 DNM0510 AAnNN-bA—xoommbcoooouoom-pmoaoo 0.12 8.4 >125 DNM0511 14.36 >125 DNM0514 0.14 5.73 >125 DNM0515 3.44 30.26 >125 DNM0516 32.01 >125 DNM0520 20.86 >125 DNM0529 0.16 91.56 >125 DNM0532 0.22 30.66 0300 mOOl’QNN-Dtm >125 DNM0539 39.39 >125 DNM0648 60.1 Example 10 ement of combination MICS for evaluation of antibiotic adjuvant potential in Ps. aeruginosa Generally following the procedure ofExample 9, MICs of ampicillin, erythromycin, and azithromycin were determined against Ps. aeruginosa. Parallel to this effort, solutions were ed which were identical. To each of these were added 0.5 uL of stock solution of a given compound of the present invention, then diluted to produce a final concentration of 125 HM, 60 uM, and/or 30 11M for determination of adjuvant activity. Compounds were determined to be antibiotic adjuvants if the mixture produced a lower MIC than that of the antibiotic agent alone. Results were as follows, demonstrating antimicrobial nt effects: 0 DNM0487 at 125 uM (standalone MIC >2 mM) produced an up to 60-fold lower MIC for erythrornycin. v DNM0487 at 125 M produced an up to d lower MIC for llin.

- DNMO488 at 125 M produced an up to 60—fold lower MIC for ampicillin.

- DNM0501 at 60 uM produced an up to 60-fold lower MIC for eryttuomycin. o DNM0501 at 60 uM produced an up to 250—fold lower MIC for ampicillin. o DNM0537 at 60 uM produced an up to 60—fold lower MIC for erythromycin.

- DNM0537 at 30 MM produced an up to ~7-fold lower MIC for erythromycin.

- DNM0537 at 30 MM produced an up to 250-fold lower MIC for llin. 0 DNM0548 at 125 14M produced an up to 125-fold lower MIC for omycin. 0 DNM0548 at 125 uM produced an up to 125-fold lower MIC for ampicillin.

Example 11 Measurement of MICs for selected compounds against a panel of challenge pathogens The inocula for each ial strain were ed by g 5-10 distinct colonies frorn the culture plates and suspending them in the appropriate broth as per Clinical and Laboratory Standards Institute (CLSI) guidelines M07-A8, M24-A or M11— A7 as appropriate. The inoculum was resuspended by vigorous shaking on a vortex mixer for 155. The turbidity was then adjusted to McFarland rd 0.5 (1-5 x 106 CFU/ml).

The inoculum was fiu'ther diluted in the appropriate media for MIC tests to give a final W0 2012/116452 inoculum in each well of 2—8 x 105 CPU/m1. MICs were tested in the appropriate broth in accordance with the appropriate CLSI guidelines.

A stock solution of each compound was prepared at a concentration of 1.28 g/L in DMSO. The stock was further diluted in the appropriate media to give a top starting concentration of 128 mg/L in the assay. 100uL of the appropriate media, as per CLSI guidelines, was dispensed into each well in columns 2—12. 200uL of the each compound solution (at L) was sed into each well in column I. IOOpL aliquots were pipetted fi'om column 1 wells and dispensed into column 2 with a multichannel pipette (i 2% coefficient of variation) thus diluting two-fold. 100 uL samples were then pipetted from column 2 wells and diSpensed into column 3. The process was repeated through to column 10. The final 100 pl. of diluted drug from column 10 was then discarded. Row 11 acted as a positive control (no compound, but organisms added), Row 12 acted as a negative control (no compound, and no organisms . 100pL ofthe appropriate inoculurn suspension in the riate media, as per CLSI guidelines, was added to the appropriate wells. This ed in a well containing ZOOuL final volume (made up of IOOpL d compound or diluents and IOOnL of inoculum or broth alone). All plates were ted in the dark under aerobic or anaerobic conditions at 30-370C for 24-48 hours, according to the appropriate CLSI ine. Plates were read visually 24-48 hours post inoculation. Endpoints of 100% inhibition were determined (CLSI interpretation nts following visual examination).

MICso, MICgo, and 100% inhibition results are reported below in pig/ml. in the following Tables, indicating each compound tested against a given s and strain: Enterococcusfaecalis Bacillus cereus ATCC29212 NCTC6349 DNM0610 as -- DNM0488 DNMO487 n— DNMO477 -- DNM0566 DNM0576 DNMO466 W0 2012/116452 DNM0465 8 $0.25 SO 25. $0 25.

Vancomycin 2 2 1 Staplwhaoccus epidermidis Staphylocaccus epidermidis NR5122 NRS7 g H(D 00 ”#HH Staphylococcus aureus NRSZ (ATCC700698) DNM0466 1 a DNMOSOS ‘ DNM0547 .5 -. _O.5 DNM0537 -1 DNM0548 - - <0 25 Vancomycm - n Staphylococcus eus Staphylococcus aureus NRSI (ATCC700699) NR$382 : 100% so - DNM0610 $0.25 $0.25 $0.25 m DNM0488 $0.25 $0.25 $0.25 _ 7 O. Ln 0.5 0.25 1 H 50.25 50-25 O Ln 0.5 — 1 H _ 1 H 1 H O D. 0'! 0.5 - 0.5 0.25 0.5 0.5 0.5 0.5 0.5 0.5 $0.25 $0.25 $0.25 $0.25 50.25 $0.25 O .

Staphylococcus aureus Staphylococcus aureus NR5383 NR5384 ———— 0.5 0.5 2 1 —2 2 .5 1 Staphylococcus aureus Staphylococcus aureus EUP SAU04D EUP MRSA004 % 100% . 4I . -so.25-so.25-so.2s é-Eso. 5 $0.25 1 III-- “In0-25 -‘-“ Staphylococcus aureus Staphylococcus aureus EUP MRSAOOZ EUP MRSAOOZ DNMOSlO - DNMO488 I“ .— 2 l DNMOSGG so 25 W0 2012/116452 DNM0466 Compound DNM0510 DNM0488 DNMD487 DNM0477 DNMDSGG DNM0575 ' DNM0466 II DNM0465 DNMOSOB DNM0547 DNM0474 0.5 0.5 0.5 DNM0606 0.5 0.5 0.5 DNM0537 $0.25 50.25 $0.25 DNM0548 0.5 0.5 ycin 0 Staphylococus eus cs" AUR59 7005 IAUR58 7005 DNM0510 DNM0488 DNM0487 DNM0477 DNM0576 DN M0466 DNMD465 1 1 1 —-——11 —-—“-. _———--m DNM0610 0.5 0.5 0.5 -1 --1 DNMD488 0.25 llkE0.25 IE 0.5 H H -2 2 -4 4 DNM0566 0 5 0.25 0.25 -0.5 0.5 0 U1 DNM0576 0.25 0.25 -1 1 — 2:-i 2 2 - ! 0.5 0.5 1 1 H H 1 1 DNM0537 0.5 0.5 as 0 5 05 1 DNM0548 $0.25 $0.25 50.25 50 25 50.25 50.25 >16 >16 VHCh >16 >15 These s indicate that the compounds of present invention are potently antibacterial t a variety of antibiotic—resistant strains, and in many cases (particularly with respect to VRE), more potent than vancomycin.

Example 12 Measurement of minimum tory concentrations against Streptococcus pneumoniae Group A W0 2012/116452 Generally following the ure of Example 9, but using Todd Hewitt broth (beef heart digest) rather than Mueller-Hinton broth, MICs were initially measured against Streptococcuspneumoniae Group A for compounds of the present invention as follows: 8 <0.25 uM DNM0576 0.5 uM DNM0620 <0.25 pM DNM0629 <0.25 uM DNM0631 <0.25 uM DNM0636 0.8 uM DNM0640 <0.25 pM Repeated measurement ofMIC ofthe above seven compounds in Todd Hewitt broth or in CASO broth (casein, soybean) indicates the MIC range for these compounds is between 125 nM and 8 nM, the average MIC being 1.6 14M. These results te that the compounds ofthe present invention have potent antibacterial s against Streptococcus.

Claims (27)

The invention claimed is:

1. A compound having a -containing ring, or prodrugs or pharmaceutically acceptable salts thereof, of one of Formulas I: R1 R3 N NH N S N R5 R10 (Ia) R1 R3 N NH N S N R5 N R6 R10 (Ib) in which E is –CH2– or is absent whereby the sulfur-containing ring is ly connected to R1, R2, R9, and R10, are each independently selected from the group ting of hydrogen, methyl, ethyl, propyl, cyclopropyl, butyl, cyclobutyl, pentyl, hexyl, isopropyl, isobutyl, neopentyl, y, and ethoxy; when the compound is a compound of Formula Ia, R11 is independently selected from the group consisting of hydrogen, methyl, ethyl, propyl, cyclopropyl, butyl, cyclobutyl, pentyl, hexyl, isopropyl, isobutyl, neopentyl, methoxy, and ethoxy; onally, R1 and R2 may connect to form a phenyl or benzofuran ring; additionally, R9 and R10 may connect to form a phenyl or benzofuran ring; R3 and R8 are each independently selected from the group consisting of hydrogen, methyl, ethyl, propyl, pyl, chlorine, fluorine, tert-butyl, methoxy, and ethoxy; R4 and R7 are each independently selected from the group consisting of hydrogen, chlorine, methyl ester, ethyl ester, methyl, ethyl, propyl, cyclopropyl, butyl, cyclobutyl, isopropyl, isobutyl, methoxy, and ethoxy; and R5 and R6 are each independently selected from the group consisting of hydrogen, cyclopentyl, cyclopropyl, furan, thiophene, trifluoromethyl, trifluoromethyl ether, methylthiol, formaldehyde, chlorine, ne, bromine, phenyl, methyl, ethyl, isopropyl, propyl, butyl, cyclobutyl, isobutyl, neopentyl, pentyl, methoxy, and ethoxy.

2. The compound of claim 1 in which the compound is according to Formula Ia; R1 and R2 are either independently selected from the group consisting of hydrogen and methyl, or form a phenyl ring whereby the ring system is naphthyl; R9 and R10 are either ndently selected from the group consisting of hydrogen and methyl, or form a phenyl ring whereby the ring system is naphthyl; R11 is hydrogen; R3 and R8 are each independently selected from the group consisting of hydrogen, methyl, chlorine, fluorine, isopropyl, utyl, and methoxy; R4 and R7 are each independently selected from the group consisting of hydrogen, , chlorine, and ethyl ester; and R5 and R6 are each independently selected from the group consisting of methyl, ethyl, , hydrogen, ne, isopropyl, cyclopentyl, bromine, cyclopropyl, trifluoromethyl, trifluoromethyl ether, methylthiol, formaldehyde, furan, and thiophene.

3. The compound of claim 2 in which E is absent.

4. The compound of claim 3 in which R1, R2, R4, R7, R9, and R10 are each

5. The compound of claim 4 in which R3 and R8 are each chlorine; and R5 and R6 are selected from the group consisting of hydrogen, methyl, and isopropyl.

6. 5-(4,5-bis(4-chloromethylphenyl)thiophenyl)-1H-tetrazole or a prodrug or pharmaceutically acceptable salt thereof.

7. 5-(4,5-bis(4-chloroisopropylphenyl)thiophenyl)-1H-tetrazole or a g or pharmaceutically acceptable salt thereof.

8. 5-(4-(3-butylphenyl)(4-chlorobenzyl)thiophenyl)-1H-tetrazole or a prodrug or pharmaceutically acceptable salt thereof.

9. 5-(4,5-bis(4-chloroisopropylphenyl)methylthiophenyl)-1H-tetrazole or a prodrug or pharmaceutically able salt thereof.

10. The compound of claim 1 in which the compound is selected from the group consisting of those compounds listed in or prodrugs or ceutically acceptable salts thereof.

11. A method of treatment of a microbial infection in a non-human mammal, comprising administering an effective amount of an antimicrobial compound of any one of the preceding claims to a non-human mammal in need thereof.

12. The method of treatment of claim 11 in which the microbial infection is ntially caused by Gram-positive bacteria.

13. The method of ent of claim 12 in which the microbial infection comprises lococcal infection.

14. A method of treatment of a microbial infection in a non-human mammal, comprising administering an effective amount of an antimicrobial adjuvant compound of any one of claims 1 to 10 and an effective amount of an antimicrobial compound to a non-human mammal in need thereof.

15. The method of treatment of claim 14 in which the microbial infection is substantially caused by Gram-negative ia.

16. The method of ent of claim 14 in which the microbial infection is substantially caused by ositive bacteria.

17. The method of treatment of claim 14 in which the microbial infection is polymicrobial.

18. The method of treatment of claim 14 in which the microbial infection is substantially caused by Pseudomonas aeruginosa.

19. Use of an antimicrobial compound of any one of claims 1 to 10 in the preparation of a medicament for the treatment of a microbial infection in a human patient.

20. The use of claim 19 in which the microbial infection is substantially caused by Gram-positive bacteria.

21. The use of claim 20 in which the microbial infection comprises Staphylococcal infection.

22. Use of an antimicrobial adjuvant compound of any one of claims 1 to 10 and an antimicrobial compound in the preparation of a medicament for the treatment of a microbial ion in a human patient.

23. The use of claim 22 in which the microbial infection is substantially caused by egative bacteria.

24. The use of claim 23 in which the microbial infection is substantially caused by Gram-positive bacteria.

25. The use of claim 23 in which the microbial infection is polymicrobial.

26. The use of claim 23 in which the ial infection is substantially caused by Pseudomonas aeruginosa.

27. A pharmaceutical composition comprising a compound of any one of claims 1- 10 and a pharmaceutically acceptable excipient.