Inhibitors of heat shock factors and uses thereof

Abstract

The present disclosure relates to a class of mammalian heat shock factor (HSF) inhibitors, to pharmaceutical compositions comprising these inhibitors as well as to methods for using the inhibitors. The inhibitors inhibit stress-induced expression from heat shock gene promoters. Furthermore, the inhibitors are cytotoxic to a variety of human cancer cells types.

Claims

1. An HSF inhibitor, wherein the inhibitor is an acrylamide derivative of (a) formula (II) ##STR00155## or a salt thereof, wherein Z.sub.1 is a 5-8 membered, non-aromatic heterocycle, n is 0-8 (or 0-6 if Z1 is a 5-membered heterocycle); X.sub.1 is N or C(—R.sub.1); X.sub.2 is C(—R.sub.2), N, or S or O (double bond between X.sub.3 and X.sub.4 instead of between X.sub.2 and X.sub.3); X.sub.3 is C(—R) or N; X.sub.4 is S or O, or C(—R.sub.4) or N if X.sub.2 is O or S (double bond between X.sub.3 and X.sub.4 instead of between X.sub.2 and X.sub.3), X.sub.6 is O, S, N, NH, (C.sub.1-C.sub.4)alkylN, CH or C((C.sub.1-C.sub.4)alkyl), CH.sub.2 or CH((C.sub.1-C.sub.4)alkyl); R.sub.1 is H, halo, (C.sub.1-C.sub.4)alkyl or halo(C.sub.1-C.sub.4)alkyl; R.sub.2, R.sub.3, R.sub.5 and R.sub.8 are independently selected, and each can be H or any substituent, provided that its presence is compatible with the small molecule nature of the inhibitor and, statistically, no more than 10% of any ionizable group it may contain is ionized at neutral pH; whereby R.sub.2 and R.sub.3 can form a substituted or unsubstituted 5 to 8 membered aromatic or non-aromatic ring or heterocycle including X.sub.2 and X.sub.3; R.sub.4 is H, halo, (C.sub.1-C.sub.4)alkyl or halo(C.sub.1-C.sub.4)alkyl, (b) formula (V), ##STR00156## or a salt thereof, whereby R.sub.2 and R.sub.5 are independently selected, and each can be any substituent, provided that its presence is compatible with the small molecule nature of the inhibitor and, statistically, no more than 10% of any ionizable group it may contain is ionized at neutral pH; X.sub.5 is O, S or N(—R.sub.6); R.sub.6 is H or is selected from halogen, OH, SH, NH.sub.2 and Z.sub.2 whereby Z.sub.2 is a residue comprising from 1 to 4 carbon atoms; R.sub.7 is —O—Z.sub.2, —S—Z.sub.2, —NH—Z.sub.2 or —N—(Z.sub.2).sub.2; X.sub.7 is CH or N (with 3 or less of the X.sub.7 being N); R.sub.10 is defined as R.sub.5; and s is 0, 1, 2, 3 or 4, with the proviso that the inhibitor in which both R.sub.2 and R.sub.5 are H, s is 0, R.sub.7 is ethoxy, X.sub.5 is O and all X.sub.7 are CH is excluded, (c) formula (IX) ##STR00157## or a salt thereof, whereby R.sub.5 can be any substituent, provided that its presence is compatible with the small molecule nature of the inhibitor and, statistically, no more than 10% of any ionizable group it may contain is ionized at neutral pH; X.sub.5 is O, S or N(—R.sub.6); R.sub.6 is H or is selected from halogen, OH, SH, NH.sub.2 and Z.sub.2 whereby Z.sub.2 is a residue comprising from 1 to 4 carbon atoms; R.sub.7 is —O—Z.sub.2, —S—Z.sub.2, —NH—Z.sub.2 or —N—(Z.sub.2).sub.2; r is 0-4 and R.sub.11 is defined as R.sub.5, with the proviso that, if r is 0, R.sub.5, R.sub.7 and X.sub.5 cannot be simultaneously H, methoxy and O, respectively, or (d) formula (X) ##STR00158## or a salt thereof, whereby Z1 is a 5-8 membered, non-aromatic heterocycle; n is 0-8 (or 0-6 if Z1 is a 5-membered heterocycle); X.sub.6 is O, S, N, NH, (C.sub.1-C.sub.4)alkylN, CH or C((C.sub.1-C.sub.4)alkyl), CH.sub.2 or CH((C.sub.1-C.sub.4)alkyl), R.sub.5 and R.sub.8 are independently selected, and each can be any substituent, provided that its presence is compatible with the small molecule nature of the inhibitor and, statistically, no more than 10% of any ionizable group it may contain is ionized at neutral pH; r is 0-4 and R.sub.11 is defined as R.sub.5.

2. The HSF inhibitor of claim 1, wherein the inhibitor is a compound of formula (IV) or a salt thereof, whereby n, R.sub.2, R.sub.3, R.sub.5, R.sub.8, X.sub.6 and Z.sub.1 are as defined in claim 1 ##STR00159##

3. The HSF inhibitor of claim 1, wherein the inhibitor is a compound of formula (VI) or a salt thereof, whereby n, s, R.sub.2, R.sub.5, R.sub.8, R.sub.10, X.sub.6, X.sub.7 and Z.sub.1 are as defined in claim 1. ##STR00160##

4. The HSF inhibitor of claim 1, wherein the inhibitor is a compound of formula (VIII) or a salt thereof, whereby n, s, R.sub.2, R.sub.5, R.sub.8, R.sub.10, X.sub.6, X.sub.7 and Z.sub.1 are as defined in claim 1. ##STR00161##

5. The HSF inhibitor of claim 1, wherein the inhibitor is (E)-3-(5,6-dihydro-4H-[1,3]oxazin-2-yl)-N-methyl-N-(5-(pyridin-3-yl-thiazol-2-yl)acrylamide or a salt thereof.

6. A pharmaceutical composition comprising an effective amount of an HSF inhibitor or a pharmaceutically acceptable salt or prodrug thereof and one or more pharmaceutically acceptable carriers or excipients, wherein the HSF inhibitor is a compound of (a) formula (I) ##STR00162## or a salt thereof, whereby X.sub.1, X.sub.2, X.sub.3, X.sub.4, X.sub.5, R.sub.5 and R.sub.7 are defined as in claim 1, (b) formula (II) ##STR00163## or a salt thereof wherein Z.sub.1, n, X.sub.1, X.sub.2, X.sub.3, X.sub.4, X.sub.6, R.sub.5 and R.sub.8 are defined as in claim 1, (c) formula (IX) ##STR00164## or a salt thereof, whereby r, R.sub.5, R.sub.7, R.sub.11 and X.sub.5 are defined as in claim 1, or (d) formula (X) ##STR00165## or a salt thereof, whereby Z.sub.1, n, r, R.sub.5, R.sub.8, R.sub.11 and X.sub.6 are defined as in claim 1.

7. The HSF inhibitor of claim 1, wherein the inhibitor is E-3-(5-Pyridin-3-yl-thiazol-2-yl carbamoyl)-acrylic acid ethyl ester or a salt thereof.

Description

BRIEF DESCRIPTION OF FIGURES

(1) FIG. 1. Characterization of Z74 cells. (A) Z74 cells contain an rluc gene linked to a hsp70B gene promoter controlled by endogenous HSF1 (eHSF1), an expressible gene for chimeric transcription factor LexA-HSF1, and a fluc gene driven by a promoter responsive to LexA-HSF1. (B, C) Transient heat stimulates the transcriptional activities of eHSF1 and LexA-HSF1, which results in an enhanced expression of fluc and rluc genes, respectively. Cells were untreated (−), heat-treated at 43° C. (HS) for the indicated time periods (B) or heated at different temperatures for 30 min (C). RLuc (filled bars) and FLuc activities (empty bars) were assayed 6 h after heat treatment. CMV: cytomegalovirus early promoter.

(2) FIG. 2. (A) Inhibition of RLuc expression by I.sub.HSF 001 in Z74 cells. Cultures were mock-exposed (−) or exposed to increasing concentrations of the I.sub.HSF and, 2 hours later, were heated (HS) at 43° C. for 30 min. RLuc (black bars) and FLuc activities were measured 6 hours after HS. Relative activities are shown. (B) Inhibition of RLuc expression by I.sub.HSF 115 in Z74 cells. (C) SPR sensorgrams indicating concentration-dependent binding of I.sub.HSF 058 and I.sub.HSF 115 to recombinant His-tagged human HSF1 (HSF1wt-His) and His-tagged HSF1 DNA-binding domain fragment (HSF1DBD-His). Responses shown are to 500, 250, 125, 62.5, 31.3 and 15.6 μM I.sub.HSF 115, and to 250, 125, 62.5, 31.3, 15.6, 7.81 and 3.91 μM I.sub.HSF 058, respectively.

(3) FIG. 3. (A) HeLa cell cultures were mock-treated (−) or exposed for 2 hours to the indicated concentrations of compounds 058 or 115 and then heat-treated (HS) at 43° C. for 30 min. Top panels: HSF1 DNA-binding activities in cell extracts were determined by EMSA. The panels show the major HSF1-HSE DNA probe complexes. Middle panels: HSF1 levels assayed by western blot (WB). GAPDH served as a loading control. Bottom panels: HSF1 homooligomerization. Aliquots of extracts were EGS-treated and then analyzed by anti-HSF1 WB. The positions of pre-stained molecular weight marker proteins (MW, in thousands) are indicated to the right. Asterisks indicate the positions of monomeric and trimeric HSF1, respectively. (B) Relative occupancy of the hspa1a promoter was assessed by chromatin immunoprecipitation assay of HeLa cell cultures mock-treated (−) or treated with compounds 058 or 115 at 12.5 μM concentrations for 2 hours and then heat-treated (HS) at 43° C. for 30 min. Chromatin fragments were immunoprecipitated using HSF1 antibodies or an F4/80 LR (control) antibody, and associated hspa1a promoter DNA was quantified by real-time PCR.

(4) FIG. 4. (A) rluc, hspa7 and hspa1a mRNA levels in Z74 cells mock-exposed (−) or exposed to the indicated concentrations of compounds 058 or 115 for 2 hours, heat-treated (HS) at 43° C. for 30 min and post-incubated for one hour at 37° C. mRNA levels were quantified by reverse transcription-real-time PCR. (B) HSP72 levels in HeLa cells mock-exposed (−) or exposed to the indicated concentrations of compounds 058 or 115 for 2 hours, heat-treated (HS) at 43° C. for 30 min and post-incubated for 6 hours at 37° C. HSP72 levels were estimated by HSP72 WB. GAPDH levels served as “loading controls”.

(5) FIG. 5. (A) Viability of HeLa cell cultures mock-exposed (−) or exposed to the indicated concentrations of compounds 001, 058 or 115. (B) Viability of MM.1S cells mock-exposed (−) or exposed to the indicated concentrations of compound 115. Viability was assayed after 96 hours of exposure using an Alamar Blue assay.

(6) FIG. 6. (A) HeLa cell cultures were mock-exposed (−) or exposed to the indicated concentrations of compound 115 for 24 (left panel) or 96 (right panel) hours. Cell viability of total cultures, floating cells and attached cells was assayed using an Alamar Blue assay. (B) HeLa cell cultures were mock-exposed (−) or exposed to the indicated concentrations of compound 115 for 15-96 hours. Trypan Blue dye exclusion was employed to determine numbers of live cells (left panel) and relative levels of necrotic cells (right panel). (C) HeLa cell cultures were mock-exposed (−) or exposed to the indicated concentrations of compound 115 for 6 and 24 h. Shown are relative levels of early apoptotic cells (annexin-positive, 7 aminoactinomycin D-negative). (D) Flow cytometric analysis of DNA of HeLa cell cultures mock-exposed (−) or exposed to the indicated concentrations of compound 115 for 6, 15 and 24 hours. Relative levels of apoptotic cells are indicated.

(7) FIG. 7. (A) Upper panel: HSF1 levels in cultures of KD control and KD13 cells as assayed by WB. Lower panel: number of alive KD control and KD13 cells after 1-4 days of culture as determined by Trypan Blue dye exclusion. (B) Viability of KD control and KD13 cells mock-exposed (−) or exposed to the indicated concentrations of compound 115 for 96 h. Cell viability was assayed by Alamar Blue assay.

(8) FIG. 8. General experimental procedures for the synthesis of HSF1 inhibitors

(9) FIG. 9. Inhibition of tumor growth mediated by I.sub.HSF 115. QD: daily.

DETAILED DESCRIPTION

Definitions

(10) Unless otherwise defined, all terms shall have their ordinary meaning in the relevant art. The following terms are defined and shall have the following meanings:

(11) The term “alkenyl,” as used herein, refers to a straight or branched chain hydrocarbon containing from 2 to 10 carbons and containing at least one carbon-carbon double bond formed by the removal of two hydrogens. Representative examples of alkenyl include, but are not limited to, ethenyl, 2-propenyl, 2-methyl-2-propenyl, 3-butenyl, 4-pentenyl, 5-hexenyl, 2-heptenyl, 2-methyl-1-heptenyl, and 3-decenyl.

(12) The term “alkenyloxy,” as used herein, refers to an alkenyl group as defined herein, appended to the parent molecular moiety through an oxy group, as defined herein. Representative examples include, but are not limited to, ethoxy, 2-propoxy, 2-methyl-2-propoxy, 3-butoxy, and the like.

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

(14) The term “alkoxyalkyl,” as used herein, refers to an alkoxy group, as defined herein, appended to the parent molecular moiety through an alkyl group, as defined herein. Representative examples of alkoxyalkyl include, but are not limited to, tert-butoxymethyl, 2-ethoxyethyl, 2-methoxyethyl, and methoxymethyl.

(15) The term “alkoxyalkoxy,” as used herein, refers to an alkoxy group, as defined herein, appended to the parent molecular moiety through an alkoxy group, as defined herein.

(16) The term “alkoxyalkenyl,” as used herein, refers to an alkoxy group, as defined herein, appended to the parent molecular moiety through an alkene group.

(17) The term “alkoxyalkynyl,” as used herein, refers to an alkoxy group, as defined herein, appended to the parent molecular moiety through an alkyne group.

(18) The term “alkoxycarbonyl,” as used herein, refers to 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, ethoxycarbonyl, and tert-butoxycarbonyl.

(19) The term “alkoxycarbonylalkoxy,” as used herein, refers to an alkoxycarbonyl group, as defined herein, appended to the parent molecular moiety through an alkoxy group, as defined herein.

(20) The term “alkyl,” as used herein, refers to a straight or branched chain hydrocarbon containing from 1 to 10 carbon atoms. The term “alkyl,” as related to the compounds of the present disclosure, refers to C.sub.1-alkyl, C.sub.2-alkyl, C.sub.3-alkyl, C.sub.4-alkyl, C.sub.5-alkyl, C.sub.6-alkyl, C.sub.7-alkyl, C.sub.8-alkyl, C.sub.9-alkyl or C.sub.10-alkyl. Representative examples of alkyl include, but are not limited to, methyl, ethyl, n-propyl, iso-propyl, n-butyl, sec-butyl, iso-butyl, tert-butyl, n-pentyl, isopentyl, neopentyl, n-hexyl, 3-methylhexyl, 2,2-dimethylpentyl, 2,3-dimethylpentyl, n-heptyl, n-octyl, n-nonyl, and n-decyl.

(21) The term “alkyl(alkyl)N—,” as used herein, refers to a nitrogen atom, appended to the parent molecular moiety which is substituted with two alkyl group, as defined herein.

(22) The term “alkyl(alkyl)N-alkyl-,” as used herein, refers to an alkyl(alkyl)N—, as defined herein, appended to the parent molecular moiety through an alkyl group, as defined herein.

(23) The term “alkyl(alkyl)N-alkyl-NHC(O)—,” as used herein, refers to an alkyl(alkyl)N-alkyl, as defined herein, appended to the parent molecular moiety through an NHC(O)— group, as defined herein.

(24) The term “alkyl(alkyl)N-alkyl-NHC(O)-alkyl,” as used herein, refers to an alkyl(alkyl)N-alkyl-NHC(O)—, as defined herein, appended to the parent molecular moiety through an alkyl group, as defined herein.

(25) The term “alkylcarbonyl,” as used herein, refers to 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, acetyl, 1-oxopropyl, 2,2-dimethyl-1-oxopropyl, 1-oxobutyl, and 1-oxopentyl.

(26) The term “alkylcarbonyl-NH—,” as used herein, refers to an alkylcarbonyl group, as defined herein, appended to the parent molecular moiety through an unsubstituted nitrogen atom, as defined herein.

(27) The term “alkylcarbonyl-NH-alkyl,” as used herein, refers to an alkylcarbonyl-NH— group, as defined herein, appended to the parent molecular moiety through an alkyl group, as defined herein.

(28) The term “alkylcarbonyl-NH-alkyl-NHC(O)—,” as used herein, refers to an alkylcarbonyl-NH-alkyl group, as defined herein, appended to the parent molecular moiety through a —NHC(O)group, as defined herein.

(29) The term “alkylcarbonyl-NH-alkyl-NHC(O)-alkyl,” as used herein, refers to an alkylcarbonyl-NH-alkyl-NHC(O)— group, as defined herein, appended to the parent molecular moiety through an alkyl group, as defined herein.

(30) The term “alkylsulfonyl,” as used herein, refers to an alkyl group, as defined herein, appended to the parent molecular moiety through a sulfonyl group, as defined herein. Representative examples of alkylsulfonyl include, but are not limited to, methylsulfonyl and ethylsulfonyl.

(31) The term “alkylthio,” as used herein, refers to an alkyl group, as defined herein, appended to the parent molecular moiety through a sulfur atom.

(32) The term “alkylthioalkyl,” as used herein, refers to an alkylthio group, as defined herein, appended to the parent molecular moiety through an alkyl group, as defined herein.

(33) The term “alkynyl,” as used herein, refers to a straight or branched chain hydrocarbon group containing from 2 to 10 carbon atoms and containing at least one carbon-carbon triple bond. Representative examples of alkynyl include, but are not limited, to acetylenyl, 1-propynyl, 2-propynyl, 3-butynyl, 2-pentynyl, and 1-butynyl.

(34) The term “aryl,” as used herein, means a phenyl group or a bicyclic aryl ring or a tricyclic aryl ring. The aryl groups of the present disclosure can be attached to the parent molecular moiety through any carbon atom within the aryl group while maintaining the proper valence. The bicyclic aryl ring consists of a phenyl group fused to a distal cycloalkyl group or a phenyl group fused to a distal cycloalkenyl group, or a phenyl group fused to a distal heteroaryl group, or a phenyl group fused to a distal heterocycle group. Representative examples of the bicyclic aryl ring include, but are not limited to, 2,3-dihydro-1H-indenyl, 1H-indenyl, naphthyl, 7,8-dihydronaphthalenyl, and 5,6,7,8-tetrahydronaphthalenyl. The tricyclic aryl ring consists of the bicyclic aryl ring fused to a cycloalkyl group or the bicyclic aryl ring fused to a cycloalkyl group or the bicyclic aryl ring fused to another phenyl group. Representative examples of tricyclic aryl ring include, but are not limited to, anthracenyl, azulenyl, 9,10-dihydroanthracenyl, fluorenyl, and 4b,8a,9,10-tetrahydrophenanthrenyl.

(35) The aryl groups of the present disclosure can be substituted with 0, 1, 2, 3 or 4 substituents that are independently selected. The independent substituents can be selected from the group consisting of hydroxy, formyl, alkylcarbonyl, alkoxy, alkylthio, alkylthioalkyl, alkoxyalkoxy, alkoxycarbonyl, alkoxyalkenyl, alkoxyalkynyl, alkoxycarbonyl-alkoxy, alkyl, alkenyl, alkenyloxy, alkenylthio, alkynyl, alkynylthio, cycloalkyl, cycloalkylthio, cycloalkylalkoxy, cycloalkylalkylthio, cycloalkenylalkoxy, cycloalkenylalkylthio, alkylSO.sub.2—, acryl, arylalkyl, aryloxy, arylthio, arylalkenyl, arylalkynyl, arylcarbonyl, arylalkoxycarbonyl, heteroaryl, heterocycle, heterocyclealkyl, heterocyclealkenyl, heterocyclealkoxy, heterocyclealkylthio, heterocyclealkynyl, heterocyclecarbonyl, heterocycleoxycarbonyl, cyano, cyanoalkyl, cyanoalkoxy, cyanoalkylthio, cyanoalkynyl, cyanoalkenyl, cyanoalkenylalkoxy, cyanoalkenylalkylthio, halogen, haloalkyl, trihaloalkyl, trihaloalkoxy, hydroxyalkyl, hydroxyalkenyl, hydroxyalkynyl, hydroxyalkoxy, hydroxyalkylthio, dihydoxyalkoxy, dihydoxyalkylthio, nitro, R.sub.f—O—, R.sub.f—S—, HO—N═CH—(CH2).sub.0, 1 or 2-, R.sub.aR.sub.bN—, R.sub.aR.sub.bNalkyl-, R.sub.aR.sub.bNalkenyl-, R.sub.aR.sub.bNalkynyl-, R.sub.aR.sub.bNC(O)—, R.sub.aR.sub.bNC(O)alkynyl-, R.sub.aR.sub.bNC(O)alkoxy-, R.sub.aR.sub.bNSO.sub.2alkoxy-, and R.sub.w—O—N═CH—, wherein alkyl may be optionally substituted with O═ and R.sub.tN═; R.sub.a and R.sub.b are each individually selected from the group consisting of hydrogen, alkyl, alkenyl, alkenylcarbonyl, alkylsulfonyl, alkylcarbonyl, alkoxycarbonyl, alkoxycarbonylalkyl, alkoxycarbonylalkylcarbonyl, alkynyl, aryl, arylalkyl, arylalkenyl, arylcarbonyl, aryloxycarbonyl, arylalkyloxycarbonyl, heteroaryl, heterocycle, heterocyclealkyl, heterocyclealkenyl, heterocyclecarbonyl, heterocyclealkylcarbonyl, haloalkyl, trihaloalkyl, trihaloalkylcarbonyl, haloalkylcarbonyl, heterocycleoxycarbonyl hydroxyalkyl, and hydroxyalkylcarbonyl; R.sub.t is selected from the group consisting of hydrogen, alkyl and HO—; R.sub.f is selected from the group consisting of alkyl, cycloalkyl, cycloalkylalkyl, alkoxyalkyl, alkylthioalkyl, and haloalkyl; R, is selected from the group consisting of hydrogen, alkylcarbonyl-NH-alkyl-NHC(O)-alkyl, alkyl, alkyl(alkyl)N-alkyl-NHC(O)-alkyl, hydroxyalkyl-NHC(O)-alkyl, heterocyclealkyl-NHC(O)-alkyl, heterocycle-NHC(O)-alkyl, and heteroarylalkyl-NHC(O)-alkyl. Alternatively, the substituents can be selected from the group consisting of hydroxy, formyl, alkylcarbonyl, alkoxy, alkylthio, alkylthioalkyl, alkoxyalkoxy, alkoxycarbonyl, alkoxyalkenyl, alkoxyalkynyl, alkoxycarbonylalkoxy, alkyl, alkenyl, alkenyloxy, alkynyl, cycloalkyl, cycloalkylalkoxy, cycloalkenylalkoxy, alkylSO.sub.2—, acryl, arylalkyl, aryloxy, arylalkenyl, arylalkynyl, arylcarbonyl, arylalkoxycarbonyl, heteroaryl, heterocycle, heterocyclealkyl, heterocyclealkenyl, heterocyclealkoxy, heterocyclealkynyl, heterocyclecarbonyl, heterocycleoxycarbonyl, cyano, cyanoalkyl, cyanoalkoxy, cyanoalkynyl, cyanoalkenyl, cyanoalkenylalkoxy, halogen, haloalkyl, trihaloalkyl, trihaloalkoxy, hydroxyalkyl, hydroxyalkenyl, hydroxyalkynyl, hydroxyalkoxy, dihydoxyalkoxy, nitro, R.sub.f—O—, HO—N═CH—(CH2).sub.0, 1 or 2-, R.sub.aR.sub.bN—, R.sub.aR.sub.bNalkyl-, R.sub.aR.sub.bNalkenyl-, R.sub.aR.sub.bNalkynyl-, R.sub.aR.sub.bNC(O)—, R.sub.aR.sub.bNC(O)alkynyl-, R.sub.aR.sub.bNC(O)alkoxy-, R.sub.aR.sub.bNSO.sub.2alkoxy-, and R.sub.w—O—N═CH—, wherein alkyl may be optionally substituted with O═ and R.sub.tN═; R.sub.a and R.sub.b are each individually selected from the group consisting of hydrogen, alkyl, alkenyl, alkenylcarbonyl, alkylsulfonyl, alkylcarbonyl, alkoxycarbonyl, alkoxycarbonylalkyl, alkoxycarbonylalkylcarbonyl, alkynyl, aryl, arylalkyl, arylalkenyl, arylcarbonyl, aryloxycarbonyl, arylalkyloxycarbonyl, heteroaryl, heterocycle, heterocyclealkyl, heterocyclealkenyl, heterocyclecarbonyl, heterocyclealkylcarbonyl, haloalkyl, trihaloalkyl, trihaloalkylcarbonyl, haloalkylcarbonyl, heterocycleoxycarbonyl hydroxyalkyl, and hydroxyalkylcarbonyl; R.sub.t is selected from the group consisting of hydrogen, alkyl and HO—; R.sub.f is selected from the group consisting of alkyl, cycloalkyl, cycloalkylalkyl, alkoxyalkyl, alkylthioalkyl, and haloalkyl; R.sub.w is selected from the group consisting of hydrogen, alkylcarbonyl-NH-alkyl-NHC(O)-alkyl, alkyl, alkyl(alkyl)N-alkyl-NHC(O)-alkyl, hydroxyalkylNHC(O)-alkyl, heterocyclealkyl-NHC(O)-alkyl, heterocycle-NHC(O)-alkyl, and heteroarylalkylNHC(O)-alkyl.

(36) The term “arylalkoxy,” as used herein, refers to an aryl group, as defined herein, appended to the parent molecular moiety through an alkoxy group, as defined herein. Representative examples of arylalkoxy include, but are not limited to, 2-phenylethoxy, 3-naphth-2-ylpropoxy, and 5-phenylpentyloxy.

(37) The term “arylalkyl,” as used herein, refers to 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, 3-phenylpropyl, and 2-naphth-2-ylethyl.

(38) The term “arylalkynyl,” as used herein, refers to an aryl group, as defined herein, appended to the parent molecular moiety through an alkyne group, as defined herein. The term “arylalkyloxy,” as used herein, refers to an arylalkyl group, as defined herein, appended to the parent molecular moiety through an oxy group, as defined herein. Representative examples of arylalkyloxy include, but are not limited to, benzyloxy, phenylpropoxy.

(39) The term “arylalkyloxycarbonyl,” as used herein, refers to an arylalkyloxy group, as defined herein, appended to the parent molecular moiety through a carbonyl group, as defined herein. Representative examples of arylalkyloxycarbonyl include, but are not limited to, benzyl carboxylate, phenylpropyl carboxylate.

(40) The term “arylcarbonyl,” as used herein, refers to an aryl group, as defined herein, appended to the parent molecular moiety through a carbonyl group, as defined herein. Representative examples of arylcarbonyl include, but are not limited to, benzoyl and naphthoyl.

(41) The term “aryloxy,” as used herein, refers to an aryl group, as defined herein, appended to the parent molecular moiety through an oxy group, as defined herein. Representative examples of aryloxy groups include, but are not limited to, phenoxy.

(42) The term “aryloxycarbonyl,” as used herein, refers to an aryloxy group, as defined herein appended to the parent molecular moiety through a carbonyl group, as defined herein. Representative examples of aryloxycarbonyl groups include, but are not limited to, phenoxycarbonyl.

(43) The term “biaryl,” as used herein, refers to an aryl group as defined herein, appended to the parent molecular moiety through an aryl group, as defined herein. Representative examples of biaryl include, but are not limited to 4-biphenyl, 3-biphenyl, 2-biphenyl.

(44) The term “carbonyl,” as used herein, refers to a —C(O)— group.

(45) The term “cyano,” as used herein, refers to a —CN group.

(46) The term “cyanoalkyl,” as used herein, refers to a cyano group, as defined herein, appended to the parent molecular moiety through an alkyl group, as defined herein. Representative examples of cyanoalkyl include, but are not limited to, cyanomethyl, 2-cyanoethyl, and 3-cyanopropyl.

(47) The term “cyanoalkoxy,” as used herein, refers to a cyano group, as defined herein, appended to the parent molecular moiety through an alkyoxy group, as defined herein.

(48) The term “cyanoalkynyl,” as used herein, refers to a cyano group, as defined herein, appended to the parent molecular moiety through an alkyne group, as defined herein.

(49) The term “cycloalkyl,” as used herein, refers to a monocyclic, bicyclic, or tricyclic ring system. Monocyclic ring systems are exemplified by a saturated cyclic hydrocarbon group containing from 3 to 8 carbon atoms. Examples of monocyclic ring systems include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl. Bicyclic ring systems are exemplified by a bridged monocyclic ring system in which two non-adjacent carbon atoms of the monocyclic ring are linked by an alkylene bridge of between one and three additional carbon atoms. Representative examples of bicyclic ring systems include, but are not limited to, bicyclo[3.1.1]heptane, bicyclo[2.2.1]heptane, bicyclo[2.2.2]octane, bicyclo[3.2.2]nonane, bicyclo[3.3.1]nonane, and bicyclo[4.2.1]nonane. Tricyclic ring systems are exemplified by a bicyclic ring system in which two non-adjacent carbon atoms of the bicyclic ring are linked by a bond or an alkylene bridge of between one and three carbon atoms. Representative examples of tricyclic-ring systems include, but are not limited to, tricyclo[3.3.1.0.sup.3,7]nonane and tricyclo[3.3.1.1.sup.3,7]decane (adamantane). The term “cycloalkyl,” as related to the compounds of the present disclosure refer to C.sub.3-cycloalkyl, C.sub.4-cycloalkyl, C.sub.5-cycloalkyl, C.sub.6-cycloalkyl, C.sub.7-cycloalkyl or C.sub.8-cycloalkyl.

(50) The cycloalkyl groups of this disclosure can be substituted with 0, 1, 2, or 3 substituents independently selected from alkenyl, alkoxy, alkoxyalkyl, alkoxycarbonyl, alkoxycarbonylalkyl, alkyl, alkylcarbonyl, alkylcarbonylalkyl, alkylcarbonyloxy, alkylsulfonyl, alkylthio, alkynyl, carboxy, carboxyalkyl, cyano, cyanoalkyl, formyl, halogen, haloalkyl, heterocycle, heterocyclealkyl, hydroxy, hydroxyalkyl, mercapto, nitro, oxo, phenyl and R.sub.ssR.sub.ttN— wherein R.sub.ss and R.sub.tt are defined herein.

(51) The term “cycloalkylalkyl,” as used herein, refers to a cycloalkyl group, as defined herein appended to the parent molecular moiety through an alkyl group, as defined herein. Representative examples of cycloalkylalkyl group include, but are not limited to cyclopentylpropyl, cyclohexyl 2-methylbutyl.

(52) The term “cycloalkenyl,” as used herein, refers to a monocyclic, bicyclic, or tricyclic ring system which contains 1 or 2 double bonds but is not aromatic. Monocyclic ring systems are exemplified by an unsaturated cyclic hydrocarbon group containing from 3 to 8 carbon atoms. Examples of monocyclic ring systems include cyclopropenyl, cyclobutenyl, cyclopentenyl, cyclohexenyl, cycloheptenyl, and cyclooctenyl. Bicyclic ring systems are exemplified by a bridged monocyclic ring system in which two non-adjacent carbon atoms of the monocyclic ring are linked by an alkylene bridge of between one and three additional carbon atoms.

(53) The cycloalkenyl groups of this disclosure can be substituted with 1, 2, or 3 substituents independently selected from alkenyl, alkoxy, alkoxyalkyl, alkoxycarbonyl, alkoxycarbonylalkyl, alkyl, alkylcarbonyl, alkylcarbonylalkyl, alkylcarbonyloxy, alkylsulfonyl, alkylthio, alkynyl, cyano, cyanoalkyl, formyl, halogen, haloalkyl, heterocycle, heterocyclealkyl, hydroxy, hydroxyalkyl, mercapto, nitro, phenyl and R.sub.ssR.sub.ttN— wherein R.sub.ss and R.sub.tt are defined herein.

(54) The term “cycloalkenylalkoxy,” as used herein, refers to a cycloalkenyl group as defined herein appended to the parent molecular moiety through an alkoxy group, as defined herein.

(55) The term “dihydroxyalkoxy,” as used herein, refers to two hydroxy groups as defined herein, appended to the parent molecular moiety through an alkoxy group, as defined herein. Representative examples of alkyl include, but are not limited to, 2-dihydroxyethoxy, 2,3-dihydroxypropoxy, 3,4-dihydroxybutoxy and the like.

(56) The term “formyl,” as used herein, refers to a —C(O)H group.

(57) The term “halo” or “halogen,” as used herein, refers to —Cl, —Br, —I or —F.

(58) The term “haloalkyl,” as used herein, refers to at least one halogen, as defined herein, appended to the parent molecular moiety through an alkyl group, as defined herein. Representative examples of haloalkyl include, but are not limited to, chloromethyl, 2-fluoroethyl, trifluoromethyl, pentafluoroethyl, and 2-chloro-3-fluoropentyl.

(59) The term “heteroaryl,” as used herein, means a monocyclic heteroaryl ring or a bicyclic heteroaryl ring. The monocyclic heteroaryl ring is a 5 or 6 membered ring. The 5 membered ring has two double bonds and contains one, two, three or four heteroatoms independently selected from the group consisting of N, O, and S. The 6 membered ring has three double bonds and contains one, two, three or four heteroatoms independently selected from the group consisting of N, O, and S. The bicyclic heteroaryl ring consists of the 5 or 6 membered heteroaryl ring fused to a distal aryl group or the 5 or 6 membered heteroaryl ring fused to a distal cycloalkyl group or the 5 or 6 membered heteroaryl ring fused to a distal cycloalkenyl group or the 5 or 6 membered heteroaryl ring fused to a distal 5 or 6 membered heteroaryl ring, or the 5 or 6 membered heteroaryl ring fused to a distal 5 or 6 membered heterocycle ring. Nitrogen heteroatoms contained within the heteroaryl may be optionally oxidized to the N-oxide or optionally protected with a nitrogen protecting group known to those of skill in the art. The heteroaryl is connected to the parent molecular moiety through any carbon atom or any nitrogen atom contained within the heteroaryl. Representative examples of heteroaryl include, but are not limited to, benzothienyl, benzoxadiazolyl, cinnolinyl, 5,6-dihydroisoquinolinyl, 7,8-dihydroisoquinolinyl, 5,6-dihydroquinolinyl, 7,8-dihydroquinolinyl, furopyridinyl, furyl, imidazolyl, indazolyl, indolyl, isoxazolyl, isoquinolinyl, isothiazolyl, naphthyridinyl, oxadiazolyl, oxazolyl, pyridinyl, pyridazinyl, pyrimidinyl, pyrazinyl, pyrazolyl, pyrrolyl, pyridinium N-oxide, quinolinyl, 5,6,7,8-tetrahydroisoquinolinyl, 5,6,7,8-tetrahydroquinolinyl, tetrazolyl, thiadiazolyl, thiazolyl, thienopyridinyl, thienyl, triazolyl, and triazinyl.

(60) According to the present disclosure, heteroaryls can be substituted with 0, 1, 2, 3 or 4 substituents independently selected. The substituents can be selected from the groups described under “aryl”.

(61) The term “heteroarylalkyl,” as used herein, refers to a heteroaryl, as defined herein, appended to the parent molecular moiety through an alkyl group, as defined herein.

(62) The term “heteroaryl-NHC(O)—,” as used herein, refers to an heteroaryl-, as defined herein, appended to the parent molecular moiety through an NHC(O)— group, as defined herein. The term “heteroaryl-NHC(O)-alkyl,” as used herein, refers to an heteroaryl-NHC(O)—, as defined herein, appended to the parent molecular moiety through an alkyl-group, as defined herein.

(63) The term “heteroarylalkyl-NHC(O)—,” as used herein, refers to an heteroarylalkyl-, as defined herein, appended to the parent molecular moiety through an NHC(O)— group, as defined herein.

(64) The term “heteroarylalkyl-NHC(O)-alkyl,” as used herein, refers to an heteroarylalkyl-NHC(O)—, as defined herein, appended to the parent molecular moiety through an alkyl-group, as defined herein.

(65) The term “heterocycle” or “heterocyclic” as used herein, means a monocyclic heterocyclic ring or a bicyclic heterocyclic ring or a tricyclic heterocyclic ring. The monocyclic heterocyclic ring consists of a 3, 4, 5, 6, 7 or 8 membered ring containing at least one heteroatom independently selected from oxygen, nitrogen and sulfur. The 3 or 4 membered ring contains 1 heteroatom selected from the group consisting of O, N and S. The 5 membered ring contains zero or one double bond and one, two or three heteroatoms selected from the group consisting of 0, N and S. The 6 or 7 membered ring contains zero, one or two double bonds and one, two or three heteroatoms selected from the group consisting of O, N and S. The 8 membered ring contains zero, one, two or three double bonds and one, two or three heteroatoms selected from the group consisting of O, N and S. Representative examples of the monocyclic heterocyclic ring include, but are not limited to, azetidinyl, azepanyl, aziridinyl, diazepanyl, 1,3-dioxanyl, 1,3-dioxolanyl, 1,3-dithiolanyl, 1,3-dithianyl, imidazolinyl, imidazolidinyl, isothiazolinyl, isothiazolidinyl, isoxazolinyl, isoxazolidinyl, morpholinyl, oxadiazolinyl, oxadiazolidinyl, oxazolinyl, oxazolidinyl, piperazinyl, piperidinyl, pyranyl, pyrazolinyl, pyrazolidinyl, pyrrolinyl, pyrrolidinyl, tetrahydrofuranyl, tetrahydrothienyl, thiadiazolinyl, thiadiazolidinyl, thiazolinyl, thiazolidinyl, thiomorpholinyl, 1,1-dioxidothiomorpholinyl (thiomorpholine sulfone), thiopyranyl, and trithianyl. The bicyclic heterocyclic ring consists of the monocyclic heterocyclic ring fused to a distal aryl ring or the monocyclic heterocyclic ring fused to a distal cycloalkyl ring or the monocyclic heterocyclic ring fused to a distal cycloalkenyl ring or the monocyclic heterocyclic ring fused to a distal monocyclic heterocyclic ring, or the monocyclic heterocyclic ring fused to a distal monocyclic heteroaryl ring. Representative examples of the bicyclic heterocyclic ring include, but are not limited to, 1,3-benzodioxolyl, 1,3-benzodithiolyl, 2,3-dihydro-1,4-benzodioxinyl, 2,3-dihydro-1-benzo furanyl, 2,3-dihydro-1-benzothienyl, 2,3-dihydro-1H-indolyl, and 1,2,3,4-tetrahydroquinolinyl. The tricyclic heterocyclic ring consists of the bicyclic heterocyclic ring fused to a phenyl group or the bicyclic heterocyclic ring fused to a cycloalkyl group or the bicyclic heterocyclic ring fused to a cycloalkenyl group or the bicyclic heterocyclic ring fused to another monocyclic heterocyclic ring. Representative examples of tricyclic heterocyclic rings include, but are not limited to, 2,3,4,4a,9,9a-hexahydro-1H-carbazolyl, 5a,6,7,8,9,9a-hexahydrodibenzo[b,d]furanyl, and 5a,6,7,8,9,9a-hexahydrodibenzo[b,d]thienyl. According to the present disclosure, heterocycles of the present disclosure can be substituted with 0-8, more preferably 0-4, i.e, 0, 1, 2, 3 or 4, substituents that are independently selected. Substituents can be selected from the groups described under “aryl”.

(66) The term “heterocyclealkyl,” as used herein, refers to a heterocycle, as defined herein, appended to the parent molecular moiety through an alkyl group, as defined herein. Representative examples of heterocyclealkyl include, but are not limited to, pyridin-3-ylmethyl and 2-pyrimidin-2-ylpropyl and the like.

(67) The term “heterocyclealkyl-NHC(O)—,” as used herein, refers to an heterocyclealkyl-, as defined herein, appended to the parent molecular moiety through an NHC(O— group, as defined herein.

(68) The term “heterocyclealkyl-NHC(O)-alkyl,” as used herein, refers to an heterocyclealkyl-NHC(O)—, as defined herein, appended to the parent molecular moiety through an alkyl-group, as defined herein.

(69) The term “heterocycle-NHC(O)—,” as used herein, refers to an heterocycle, as defined herein, appended to the parent molecular moiety through an NHC(O)— group, as defined herein.

(70) The term “heterocycle-NHC(O)-alkyl,” as used herein, refers to an heterocycle-NHC(O)—, as defined herein, appended to the parent molecular moiety through an alkyl-group, as defined herein.

(71) The term “heterocyclealkenyl,” as used herein, refers to a heterocycle, as defined herein, appended to the parent molecular moiety through an alkenyl group, as defined herein.

(72) The term “heterocyclealkoxy,” as used herein, refers to a heterocycle, as defined herein, appended to the parent molecular moiety through an alkoxy group, as defined herein.

(73) The term “heterocyclecarbonyl,” as used herein, refers to a heterocycle, as defined herein, appended to the parent molecular moiety through a carbonyl group, as defined herein. Representative examples of heterocyclecarbonyl include, but are not limited to, pyridin-4-ylethanone, pyridin-4-ylpropanone.

(74) The term “heterocycleoxy,” as used herein, refers to a heterocycle, as defined herein, appended to the parent molecular moiety through an oxy group, as defined herein. Representative examples of heterocycleoxy include, but are not limited to, pyridin-2-ol, pyridin-4-ol, thiophen-2-ol.

(75) The term “heterocycleoxycarbonyl,” as used herein, refers to a heterocycleoxy, as defined herein, appended to the parent molecular moiety through a carbonyl group, as defined herein. Representative examples of heterocyclecarbonyl include, but are not limited to, pyridin-4-ylcarboxylate, thiophene-2-ylcarboxylate.

(76) The term “heterocycleoxyalkynyl,” as used herein, refers to a heterocycleoxy, as defined herein, appended to the parent molecular moiety through an alkyne group, as defined herein.

(77) The term “hydroxy,” as used herein, refers to an —OH group.

(78) The term “hydroxyalkyl,” as used herein, refers to a hydroxy group, as defined herein, appended to the parent molecular moiety through an alkyl group, as defined herein. Representative examples of hydroxyalkyl include, but are not limited to, 2-hydroxyethyl, 2-hydroxypropyl, 3-hydroxybutyl and the like.

(79) The term “hydroxyalkyl-NHC(O)—,” as used herein, refers to an hydroxyalkyl-, as defined herein, appended to the parent molecular moiety through an NHC(O)— group, as defined herein.

(80) The term “hydroxyalkyl-NHC(O)-alkyl,” as used herein, refers to an hydroxyalkyl-NHC(O)—, as defined herein, appended to the parent molecular moiety through an alkyl-group, as defined herein.

(81) The term “hydroxyalkenyl,” as used herein, refers to a hydroxy group, as defined herein, appended to the parent molecular moiety through an alkenyl group, as defined herein.

(82) The term “hydroxyalkynyl,” as used herein, refers to a hydroxy group, as defined herein, appended to the parent molecular moiety through an alkyne group, as defined herein.

(83) The term “hydroxyalkoxy,” as used herein, refers to a hydroxy group as defined herein, appended to the parent molecular moiety through an alkoxy group, as defined herein. Representative examples include, but are not limited to, 2-hydroxyethoxy, 2-hydroxypropoxy, 3-hydroxybutoxy and the like.

(84) The term “hydroxycycloalkyl,” as used herein, refers to a hydroxy group, as defined herein, appended to the parent molecular moiety through an cycloalkyl group, as defined herein.

(85) The term “—NHC(O)—,” as used herein, refers to an unsubstituted nitrogen atom, as defined herein, appended to the parent molecular moiety through a carbonyl group, as defined herein.

(86) The term “trihaloalkyl,” as used herein, refers to three halogen atoms, appended to the parent molecular moiety through an alkyl group, as defined herein.

(87) The term “trihaloalkoxy,” as used herein, refers to three halogen atoms, appended to the parent molecular moiety through an alkoxy group, as defined herein.

(88) The terms “alkenylthio”, “alkynylthio”, “cycloalkylthio”, “cycloalkylalkylthio”, “cycloalkenylalkylthio”, “arylthio”, “heterocyclealkylthio”, “cyanoalkylthio”, “cyanoalkenylthio”, “hydroxyalkylthio”, and “dihydroxythio”, as used herein, refer to alkenyl, alkynyl, cycloalkyl, cycloalkylalkyl, cycloalkenylalkyl, aryl, heterocyclealkyl, cyanoalkyl, cyanoalkenyl, hydroxyalkyl and dihydroxy moieties as defined herein that are appended to the parent molecular moiety through a sulfur atom.

(89) The term “inhibit” is understood to encompass both partial and complete inhibition.

(90) The terms “HSF inhibitor”, “inhibitor of HSF” or the abbreviation “I.sub.HSF” mean a small molecule compound that, inter alia, is capable of inhibiting the prototypical function of heat shock factor 1, abbreviated as HSF1 (also known as heat shock transcription factor or HSTF), which function is to mediate stress-induced (typically heat-induced) transient accumulation of transcripts (or protein products) of its target genes, which are genes that are functionally linked to stress-inducible promoters, which are typically promoters comprising one or more HSE sequences.

(91) “I.sub.HSF” is also referred to herein as “compound”, “compound of the disclosure” or “active compound”. It will be clear from the context when the latter terms are meant to refer to I.sub.HSF The nucleotide sequence of human hsf1 cDNA and the human HSF1 amino acid sequence were first reported by Rabindran et al. (1991). Proc. Natl. Acad. Sci. USA 88: 6906-6910. For the corresponding first report for human hsf2 cDNA and amino acid sequence, see Schuetz et al. (1991). Proc. Natl. Acad. Sci. USA 88: 6911-6915. For human hsf4, see Nakai et al. (1997) Mol. Cell. Biol. 17: 469-481. See also UniProtKB-Q00613, Q03933 and Q9ULV5.

(92) “Promoter” is a nucleotide sequence that directs the transcriptional expression of a functionally linked gene. A promoter may be constitutively active in a ubiquitous or a cell type-restricted fashion. Alternatively, it may also be inducible, i.e., become active, or more active, in response to some stimulation. Stress-inducible hsp gene promoters are examples of inducible promoters.

(93) The term “heat shock gene (hsp gene)” is understood herein as referring to a gene whose activity is enhanced when the eukaryotic cell containing the gene is exposed to a temperature above its normal growth temperature. Typically, such genes are activated when the temperature to which the cell is normally exposed is raised by 3-10° C. Heat shock genes comprise genes for the “classical” heat shock proteins, i.e., HSP110, HSP90, HSP70, HSP60, HSP40, and HSP20-30. They also include other heat-inducible genes such as genes for MDR1, ubiquitin, FKBP52, hemoxidase and other proteins. The promoters of these genes, the “heat shock promoters (hsp promoters)”, contain characteristic sequence elements referred to as heat shock elements (HSE) that consist of perfect or imperfect sequence modules of the type NGAAN or AGAAN, which modules are arranged in alternating orientations (Amin, J. et al. (1988) Mol. Cell. Biol 8: 3761-3769; Xiao, H. and Lis, J. T. (1988) Science 239: 1139-1142; Fernandes, M. et al. (1994) Nucleic Acids Res. 22: 167-173). These elements are highly conserved in all eukaryotic cells. HSE sequences are binding sites for heat shock transcription factors (HSFs; reviewed in Wu, C. (1995) Annu. Rev. Cell Dev. Biol. 11, 441-469). The factor primarily responsible for activation of hsp genes in mammalian cells exposed to heat or a proteotoxic stress is HSF1 (Baler, R. et al. (1993) Mol. Cell. Biol. 13: 2486-2496; McMillan, D. R. et al. (1998) J. Biol. Chem. 273: 7523-7528). Preferred promoters for use in assessing HSF1 function are those from inducible hsp70 genes. A particularly preferred hsp promoter is the promoter of the human hsp70B gene (Voellmy, R. et al. (1985) Proc. Natl. Acad. Sci. USA 82: 4949-4953).

(94) The term “treatment” refers to any process, action, application, therapy, or the like, wherein a mammal, in particular a human being, is subject to medical aid with the object of improving the mammal's condition, directly or indirectly.

(95) As used herein, the term “pharmaceutically acceptable salt” refers to those salts which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans or mammalian animals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit/risk ratio. Pharmaceutically acceptable salts are well known in the art. For example, Berge and colleagues describe pharmaceutically acceptable salts in detail in J. Pharm. Sci. 66: 1-19 (1977). The salts can be prepared during the final isolation and purification of the compounds of the disclosure, or separately by reacting the free base function with a suitable organic acid or inorganic acid. Examples of pharmaceutically acceptable nontoxic acid addition salts include, but are not limited to, salts of an amino group formed with inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid and perchloric acid or with organic acids such as acetic acid, maleic acid, tartaric acid, citric acid, succinic acid, lactobionic acid or malonic acid or by using other methods used in the art such as ion exchange. Other pharmaceutically acceptable salts include, but are not limited to, adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate, glucoheptonate, glycerophosphate, gluconate, hemisulfate, heptanoate, hexanoate, hydroiodide, 2-hydroxy-ethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, undecanoate, valerate salts, and the like. Representative alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, and the like. Further pharmaceutically acceptable salts include, when appropriate, nontoxic ammonium, quaternary ammonium, and amine cations formed using counterions such as halide, hydroxide, carboxylate, sulfate, phosphate, nitrate, sulfonate and aryl sulfonate.

(96) The term “pharmaceutically acceptable prodrugs” as used herein refers to those prodrugs of the compounds of the present disclosure which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and mammalian animals with undue toxicity, irritation, allergic response, and the like, commensurate with a reasonable benefit/risk ratio, and effective for their intended use. “Prodrug”, as used herein means a compound which is convertible in vivo by metabolic means (e.g. by hydrolysis) to a compound of the disclosure. Various forms of prodrugs are known in the art, for example, as discussed in Bundgaard, H. (ed.), Design of Prodrugs, Elsevier (1985); Widder, K. J. and Green, R. (eds.), Meth. Enzymol., vol. 112, Academic Press (1985); Krogsgaard-Larsen, P. and Bundgaard, H. (eds.). “Design and Application of Prodrugs, Textbook of Drug Design and Development”, Chapter 13, pp. 351-85 (1991); Nielson, N. M. and Bundgaard, H. (1988) J. Pharm. Sci. 77:285-98; Higuchi, T. and Stella, V. (eds.) Pro-drugs as Novel Drug Delivery Systems, American Chemical Society (1975); and Testa, B. and Mayer, J. M. “Hydrolysis In Drug And Prodrug Metabolism: Chemistry, Biochemistry And Enzymology,” John Wiley and Sons, Ltd. (2003).

(97) As used herein, “pharmaceutically acceptable carrier or excipient” is intended to include any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and the like, compatible with pharmaceutical administration, such as sterile pyrogen-free water. Suitable carriers are described in Remington's Pharmaceutical Sciences (Mack Publishing Co., Easton, Pa., 19th ed. 1995), a standard reference text in the field, which is incorporated herein by reference. Some examples of materials which can serve as pharmaceutically acceptable carriers are sugars such as lactose, glucose and sucrose; cyclodextrins such as alpha-, beta- and gamma-cyclodextrins; starches such as corn starch and potato starch; cellulose and its derivatives such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; powdered tragacanth; malt; gelatin; talc; excipients such as 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 as propylene glycol; esters such as ethyl oleate and ethyl laurate; agar; buffering agents such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic saline; Ringer's solution; ethyl alcohol, and phosphate buffer solutions, as well as other non-toxic compatible lubricants such as sodium lauryl sulfate 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 the formulator. Also encompassed are emulsifiers/surfactants such as cremophor EL and solutol HS15, lecithin and phospholipids such as phosphatylcholine. Liposomes may also be used. The use of such media and agents for pharmaceutically active substances is well known in the art. Except insofar as any conventional media or agent is incompatible with the active compound, use thereof in the compositions is contemplated. Supplementary active compounds can also be incorporated into the compositions.

(98) The term “subject” as used herein refers to a mammalian subject. Preferably, the subject is a human subject.

(99) The term “small molecule” means a molecule with a molecular weight of less than about 2,000 daltons, preferably less than about 1,000 daltons and, most preferably, less than 500 daltons. Further to the latter size limitation, to avoid any possible doubt, the term “small molecule” as used herein shall be specifically defined to exclude nucleic acids or nucleic acid aptamers.

(100) By an “effective amount” of an HSF inhibitor of the disclosure is meant an amount of the compound which, when administered once or multiple times over the course of a treatment, confers a therapeutic effect on the treated subject, at a reasonable benefit/risk ratio applicable to any medical treatment. The therapeutic effect may be objective (i.e., measurable by some test or marker) or subjective (i.e., subject gives an indication of or feels an effect). An effective amount of an HSF inhibitor of the disclosure may range from about 0.01 mg/kg body weight to about 50 mg/kg body weight, preferably from about 0.1 to about 30 mg/kg body weight. Effective doses will also vary depending on route of administration, as well as the possibility of co-usage with other agents. It will be understood, however, that the total daily usage of the compounds and compositions of the present disclosure will be decided by the attending physician within the scope of sound medical judgment. The specific effective dose level for any particular patient will depend upon a variety of factors including the disorder being treated and the severity of the disorder; the activity of the specific active 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; drugs used in combination or contemporaneously with the specific compound employed; and like factors well known in the medical arts. It is noted that when used in the context of prophylaxis or prevention, an “effective amount” of a compound of the disclosure is meant to be an amount of the compound which, when administered once or multiple times over the course of a treatment, confers a desired prophylactic effect on the treated subject.

(101) HSF Inhibitors

(102) Mammalian HSF1 is regulated at multiple levels. Zou, J. et al. (1998) Cell 94: 471-480; Guo, Y. et al. (2001) J. Biol. Chem. 276, 45791-45799; Guettouche, T. et al. (2005) BMC Biochemistry 6: 4; Voellmy, R. (2006) Handb. Exp. Pharmacol. (172): 43-68; Boyault et al. (2007) Genes Dev. 21: 2172-2181. In an unstressed mammalian or human cell, HSF1 is present as a heterocomplex with HSP90 and other co-chaperones and co-factors, including CHIP and HDAC6. In this form, HSF1 is incapable of binding to and transactivating an HSE-containing promoter. Inactive HSF1 complex appears to be distributed throughout the cell. When the cell experiences a physical or chemical stress, the HSF1 heterocomplex disassembles partially or completely, and HSF1 homotrimerizes. Homotrimeric HSF1 is capable of binding to HSE DNA sequences. Also, homotrimeric factor concentrates in the nucleus. While activation of DNA-binding ability appears to be a direct consequence of factor trimerization, activation of the transcription-enhancing ability of the factor is controlled by additional factors. Depending on the level of stress experienced by the cell, homotrimeric factor can be sidelined by formation of an HSP90-containing complex. The lower the level of stress, the more likely is inactivation of trimeric HSF1 by formation of such a complex. Transactivation ability is also enhanced/reduced by post-translational modification, primarily phosphorylation/dephosphorylation and deactylation/acetylation. Guettouche, T. et al. (2005); Westerheide, S. D. et al. (2009) Science 323: 1063-1066. HSF1 is also stabilized by acetylation at lysine residues that are not critical to factor activity. Raychaudhuri, S. et al. (2014) Cell 156: 975-985. A number of drug substances used in human therapy cause activation of HSF1, resulting in induced expression of HSPs and a buildup of tolerance (and inhibition of apoptotic mechanisms). Well known examples of drug substances that cause HSF1 activation are inhibitors of HSP90, inhibitors of proteasome function and inhibitors of HDAC6. This induction of HSP expression appears to result in a suboptimal efficacy of the drugs. Bagatell, R. et al. (2000) Clin. Cancer Res. 6: 3312-3318.

(103) Structure-function analyses on mammalian HSF1 located the HSE DNA-binding domain to the N terminus of the HSF1 polypeptide. Further C-terminal are 4-3 hydrophobic repeats (or 4-3 leucine zippers) HR-A and HR-B that play critical roles in trimerization as well as in the regulation of trimerization of the factor. Following these repeats is a region known as “regulatory domain” that modulates the transcriptional activity of the factor. This domain contains a binding site for HSP90 complex as well as at least two sites for activating phosphorylation. C-terminal of this region is another 4-3 hydrophobic repeat, HR-C, that plays a critical role in maintaining (unactivated) HSF1 in a non-homotrimeric state. Towards the C-terminal end of the HSF1 polypeptide are located two transcription activation domains. Voellmy, R. (2006) Handb. Exp. Pharmacol. 172: 43-68. HSF1 has a modular nature. When the DNA-binding domain of human HSF1 was replaced with a DNA-binding domain from bacterial protein LexA, the resulting chimeric factor LexA.sub.87-hHSF1.sub.79 was non-homotrimeric and inactive in the absence of heat or stress, but could be induced by heat or other stress to homotrimerize as well as to transactivate a reporter gene under the control of a LexA-responsive promoter. Zuo et al. (1994) Mol. Cell. Biol. 14: 7557-7568; Zuo et al. (1995) Mol. Cell. Biol. 15: 4319-4330.

(104) A complete structure of an HSF1 molecule was not available when the work disclosed herein was initiated. However, eleven crystal structures of the HSF DNA-binding domain, 9 from yeast and 2 from Drosophila, had been deposited in the Brookhaven Protein Databank at the time. (Structures of the human HSF1 and HSF2 DNA-binding domains became available only very recently.) Based on the former structures, a comparative model of adequately high quality could be generated of the DNA-binding domain of human HSF1. This model predicted four potential binding cavities (cavities A-D). Nine appropriate 3-point pharmacophores were defined. Using these pharmacophores, a proprietary library (Leadbuilder NICE) of about 300,000 commercially available drug-like molecules was searched, and a biased sublibrary of about 2,000 compounds was assembled.

(105) For screening this sublibrary, a cell-based assay was developed. We opted for a cell-based assay, because a cell-free assay would have had to be a DNA-binding assay. Such an assay would have limited discovery to molecules capable of interfering with the DNA-binding function of HSF1. This was thought to be too narrow a focus, considering that three of the four cavities identified in the HSF1 DNA-binding domain are not near the DNA interaction region. A straightforward cell-based assay, in which a HSF1-responsive gene would be assayed, was considered likely to be over-inclusive. Compounds would be scored as hits that inhibit any factor that is involved in HSF1 activation or prevents HSF1 deactivation. These factors include protein kinases, activators of protein phosphatases, factors affecting HSF1 mRNA stability, HSP90, HDAC6, etc. For developing a more discriminating cell-based assay, advantage was taken of the modular nature of HSF1. As discussed above, replacement of the HSF1 DNA-binding domain with an unrelated DNA-binding domain (bacterial LexA) resulted in a chimeric transcription factor whose activity is regulated like that of wild type HSF1 but that binds to a different target promoter. The cell-based assay developed (see below for the particular format employed) compared transactivation by wild type HSF1 and chimeric HSF1 in cells exposed to an HSF1-activating heat treatment in the presence of a sublibrary compound. A compound was scored as a hit, if it inhibited the activity of wild type HSF1 but not of chimeric HSF1.

(106) The cell-based assay made use of HeLa-derived cell line Z74 that stably contains a gene for chimeric transcription factor LexA-(human) HSF1 under the control of a CMV early promoter, a firefly luciferase (flue) gene driven by a promoter responsive to the latter chimeric transcription factor and a Renilla luciferase (rluc) gene functionally linked to an hsp70b promoter and therefore controlled by endogenous HSF1 (FIG. 1A). As is shown in FIGS. 1B & C, the reporter genes responded similarly to heat dose, i.e., their activation depended on both duration of heat exposure and temperature of the exposure. These data demonstrated that transcription factors LexA-HSF1 and (endogenous) wild type HSF1 are similarly stress-regulated in the Z74 cell line. Depletion of HSF1 (and LexA-HSF1) by siRNA supported the notion that this stress regulation is mediated by HSF1 (and LexA-HSF1). Therefore, the Z74 line was well suited for use in a screening assay aimed at discovering molecules that inhibit heat-induced HSF1 activity through interaction with the transcription factor's DNA-binding domain.

(107) The compounds of the afore-mentioned biased sublibrary were screened in cell line Z74 at 12.5 and 25 μM concentrations. Typically, Z74 cultures in 96-well plates were exposed to compound or vehicle for 2 h, heat-treated at 43.0° C. for 30 min, and incubated further for 6 h at 37° C. to allow for accumulation of the luciferases. RLuc and FLuc activities were determined, typically using the Dual-Glo® Luciferase Assay System (Promega Corp., Madison, Wis.), and reading luciferase light counts in a Wallac Microbeta Trilux-1450 Luminometer (PerkinElmer, Waltham, Mass.). I.sub.HSF 001 was one of two library compounds that inhibited RLuc expression but not FLuc expression in heat-treated Z74 cells (Table 1; FIG. 2A). Therefore, I.sub.HSF 001 is an inhibitor of HSF1 function that specifically acts through the HSF1 DNA-binding domain. I.sub.HSF 001 fulfills the pharmacophoric criteria for a cavity A binder. A corroborating experiment utilized a cell line containing an hsp70b promoter-driven fluc gene. I.sub.HSF 001 was found to inhibit FLuc expression in this line (data not shown). I.sub.HSF 001 is (E)-ethyl 4-oxo-4-(thiazol-2-ylamino)but-2-enoate. The compound can be obtained from commercial sources (e.g., AKE-PB-90340538, Akos).

(108) To determine structure-activity relationships as well as to improve upon the inhibitory activity of I.sub.HSF 001, a series of related compounds was synthesized and tested for activity using the above-described cell-based assay. The structure of some of these compounds and their inhibitory activities are presented in Table 2. The compounds were prepared from known starting materials using chemical methods known to the skilled artisan and further described in the example section. The most potent I.sub.HSF was compound 115 that had substantial activity in the high nanomolar range in the Z74-based screening assay (Table 2; FIG. 2B).

(109) TABLE-US-00001 TABLE 1 Inhibition of HSF1 activity in Z74 cells (I.sub.HSF 001 tested at 25 μM) Inhibition of Inhibition of LexA-hHSF HSF1 activity activity I.sub.HSF Structure Experiment No. [RLuc] (%) [FLuc] (%) 001 1 2 3 4 5 6 7 8 9 66 69 69 67 68 63 56 51 45  3  0  0  0  0  0  0  0 15

(110) TABLE-US-00002 TABLE 2 Inhibition of HSF1 activity in Z74 cells Inhibition of Inhibition HSF1 of HSF1 activity at activity at 25 μM I.sub.HSF 12.5 μM I.sub.HSF I.sub.HSF Structure [RLuc] (%) [RLuc] (%) IC.sub.50 (μM) 001 64 +/− 6 37 +/− 9 17.8 +/− 2.6 010 56 56 ND 011 83 69 7.0 012 86 68 11.3 013 50 34 25.0 014 0 89 60 7.1 015 20  0 ND 016 25  0 ND 017 37 32 ND 018 53 20 23.6 020 53 30 23.5 027 42 18 >25 028 41 13 >25 030 18  5 >25 031 24 15 ND 042 0 52  1 ND 043  5 32 ND 045 53 34 ND 046 70 33 ND 048 67 37 ND 049 85 53 ND 050 61 40 ND 051 92 72 9.6 052 71 57 7.4 053 29 15 >25 054 0 69 22 20.2 055 74 27 18.7 056 61 35 19.8 057 65 40 ND 058 97 +/− 2 79 +/− 7 4.8 +/− 0.4 059 72 45 14.7 060 19 16 ND 061 18 12 ND 070 90 +/− 8 72 +/− 7 5.7 +/− 0.8 076 66 56 ND 077 0 67 42 ND 078 54 32 ND 079 79 42 ND 080 71 55 ND 081 65 42 ND 082 91 58 ND 083 34 24 ND 086 78 47 ND 088 31 25 >25 089 61 41 17.9 090 0 98 +/− 2 78 +/− 19 5.4 +/− 0.4 091 27 22 >25 092 99 +/− 1 90 +/− 6 4.7 +/− 0.2 095 44  6 >25 099 95 17 18.0 101 48 −30* >25 105 96 69 10.8 107 92 62 10.6 108 89  −6* 19.6 109 88 53 11.2 112 0 26 13 >25 115 100+/− 99 +/− 1 0.7 +/− 0.1 *Compound enhanced rLuc activity. ND: not determined.

(111) Based on experimental results obtained with the compounds shown in Table 2 as well as a multitude of other related compounds that were made and tested for their inhibitory activity, an HSF inhibitor can be defined as a compound of formulae I or II below,

(112) ##STR00072##
wherein
n is 0-8, preferably 0-4 (0, 1, 2, 3 or 4); o is 0, 1, 2, 3 or 4; p is 0, 1 or 2;
X.sub.1 is N or C(—R.sub.1);
X.sub.2 is C(—R.sub.2), N, or O or S (double bond between X.sub.3 and X.sub.4 instead of between X.sub.2 and X.sub.3);
X.sub.3 is C(—R.sub.3) or N;
X.sub.4 is S or O, or C(—R.sub.4) or N if X.sub.2 is O or S (double bond between X.sub.3 and X.sub.4 instead of between X.sub.2 and X.sub.3);
X.sub.5 is O, S or N(—R.sub.6);
X.sub.6 is O, S, N, NH, (C.sub.1-C.sub.4)alkylN, CH or C((C.sub.1-C.sub.4)alkyl), CH.sub.2 or CH((C.sub.1-C.sub.4)alkyl).sup.or C((C.sub.1-C.sub.4)alkyl).sub.2;
R.sub.1 is H, halo, (C.sub.1-C.sub.4)alkyl; halo(C.sub.1-C.sub.4)alkyl, H being preferred;
R.sub.2, R.sub.3, R.sub.5 and R.sub.8 are independently selected, and each can be H or any substituent, provided that its presence is compatible with the small molecule nature of the inhibitor and, statistically, no more than 10% of any ionizable group it may contain is ionized at neutral pH (R.sub.8 not being a substituent of X.sub.6);

(113) whereby R.sub.2 and R.sub.3 can form a substituted or unsubstituted 5 to 8 membered aromatic or non-aromatic ring or heterocycle as exemplified by formulae I A and I B below:

(114) ##STR00073##
R.sub.4 is H, halo, (C.sub.1-C.sub.4)alkyl; halo(C.sub.1-C.sub.4)alkyl, H being preferred;
R.sub.6 is H or is selected from halogen, OH, SH, NH.sub.2 and Z.sub.2, whereby Z.sub.2 is a residue comprising from 1 to 4 carbon atoms; R.sub.7 is —OH, —SH, —NH.sub.2, —O—Z.sub.2, —S—Z.sub.2, —NH—Z.sub.2 or —N—(Z.sub.2).sub.2;
R.sub.9 is independently selected from (H), alkenyl, alkoxy, alkoxyalkyl, alkoxyalkynyl, alkoxycarbonyl, alkoxycarbonylalkyl, alkoxy-NH.dbd.C(alkyl)-, alkyl, alkylcarbonyl, alkylcarbonylalkyl, alkylcarbonyloxy, alkylsulfonyl, alkylthio, alkynyl, aryl, arylalkoxy, arylalkyl, arylcarbonyl, aryloxy, cyano, cyanoalkyl, cycloalkyl, cycloalkylalkyl, formyl, halogen, haloalkyl, hydroxy, hydroxyalkyl, hydroxycycloalkyl, mercapto, nitro, oxo, phenyl, and R.sub.ssR.sub.ttN—, R.sub.ssR.sub.ttN carbonyl, R.sub.ssR.sub.ttN alkyl, wherein R.sub.ss and R.sub.tt are each independently selected from the group consisting of hydrogen, alkyl, alkylcarbonyl, alkoxycarbonyl, and alkylSO.sub.2—;
and Z.sub.1 is a 5-8 membered heterocycle.

(115) In alternative embodiments, substituents R.sub.2, R.sub.3, and R.sub.5 in formula I or R.sub.2, R.sub.3, R.sub.5 and R.sub.8 in formula II (or in any of formulae III-XII in which one or more of these substituents are indicated) are independently selected from the group consisting of (H), hydroxy, formyl, alkylcarbonyl, alkoxy, alkylthio, alkylthioalkyl, alkoxyalkoxy, alkoxycarbonyl, alkoxyalkenyl, alkoxyalkynyl, alkoxycarbonyl-alkoxy, alkyl, alkenyl, alkenyloxy, alkenylthio, alkynyl, alkynylthio, cycloalkyl, cycloalkylthio, cycloalkylalkoxy, cycloalkylalkylthio, cycloalkenylalkoxy, cycloalkenylalkylthio, alkylSO.sub.2—, acryl, arylalkyl, aryloxy, arylthio, arylalkenyl, arylalkynyl, arylcarbonyl, arylalkoxycarbonyl, heteroaryl, heterocycle, heterocyclealkyl, heterocyclealkenyl, heterocyclealkoxy, heterocyclealkylthio, heterocyclealkynyl, heterocyclecarbonyl, heterocycleoxycarbonyl, cyano, cyanoalkyl, cyanoalkoxy, cyanoalkylthio, cyanoalkynyl, cyanoalkenyl, cyanoalkenylalkoxy, cyanoalkenylalkylthio, halogen, haloalkyl, trihaloalkyl, trihaloalkoxy, hydroxyalkyl, hydroxyalkenyl, hydroxyalkynyl, hydroxyalkoxy, hydroxyalkylthio, dihydoxyalkoxy, dihydoxyalkylthio, nitro, R.sub.f—O—, R.sub.f, —S—, HO—N═CH—(CH2).sub.0, 1 or 2-, R.sub.aR.sub.bN—, R.sub.aR.sub.bNalkyl-, R.sub.aR.sub.bNalkenyl-, R.sub.aR.sub.bNalkynyl-, R.sub.aR.sub.bNC(O)—, R.sub.aR.sub.bNC(O)alkynyl-, R.sub.aR.sub.bNC(O)alkoxy-, R.sub.aR.sub.bNSO.sub.2alkoxy-, and R.sub.w—O—N═CH—, wherein alkyl may be optionally substituted with O═ and R.sub.tN═; R.sub.a and R.sub.b are each individually selected from the group consisting of hydrogen, alkyl, alkenyl, alkenylcarbonyl, alkylsulfonyl, alkylcarbonyl, alkoxycarbonyl, alkoxycarbonylalkyl, alkoxycarbonylalkylcarbonyl, alkynyl, aryl, arylalkyl, arylalkenyl, arylcarbonyl, aryloxycarbonyl, arylalkyloxycarbonyl, heteroaryl, heterocycle, heterocyclealkyl, heterocyclealkenyl, heterocyclecarbonyl, heterocyclealkylcarbonyl, haloalkyl, trihaloalkyl, trihaloalkylcarbonyl, haloalkylcarbonyl, heterocycleoxycarbonyl hydroxyalkyl, and hydroxyalkylcarbonyl; R.sub.t is selected from the group consisting of hydrogen, alkyl and HO—; R.sub.f is selected from the group consisting of alkyl, cycloalkyl, cycloalkylalkyl, alkoxyalkyl, alkylthioalkyl, and haloalkyl; R, is selected from the group consisting of hydrogen, alkylcarbonyl-NH-alkyl-NHC(O)-alkyl, alkyl, alkyl(alkyl)N-alkyl-NHC(O)-alkyl, hydroxyalkyl-NHC(O)-alkyl, heterocyclealkyl-NHC(O)-alkyl, heterocycle-NHC(O)-alkyl, and heteroarylalkyl-NHC(O)-alkyl.

(116) In further alternative embodiments, substituents R.sub.2, R.sub.3, and R.sub.5 in formula I or R.sub.2, R.sub.3, R.sub.5 and R.sub.8 in formula II (or in any of formulae III XII in which one or more of these substituents are indicated) are independently selected from the group consisting of (H), hydroxy, formyl, alkylcarbonyl, alkoxy, alkylthio, alkylthioalkyl, alkoxyalkoxy, alkoxycarbonyl, alkoxyalkenyl, alkoxyalkynyl, alkoxycarbonyl-alkoxy, alkyl, alkenyl, alkenyloxy, alkynyl, cycloalkyl, cycloalkylalkoxy, cycloalkenylalkoxy, alkylSO.sub.2—, acryl, arylalkyl, aryloxy, arylalkenyl, arylalkynyl, arylcarbonyl, arylalkoxycarbonyl, heteroaryl, heterocycle, heterocyclealkyl, heterocyclealkenyl, heterocyclealkoxy, heterocyclealkynyl, heterocyclecarbonyl, heterocycleoxycarbonyl, cyano, cyanoalkyl, cyanoalkoxy, cyanoalkynyl, cyanoalkenyl, cyanoalkenylalkoxy, halogen, haloalkyl, trihaloalkyl, trihaloalkoxy, hydroxyalkyl, hydroxyalkenyl, hydroxyalkynyl, hydroxyalkoxy, dihydoxyalkoxy, nitro, R.sub.f—O—, HO—N═CH(CH2).sub.0, 1 or 2-, R.sub.aR.sub.bN—, R.sub.aR.sub.bNalkyl-, R.sub.aR.sub.bNalkenyl, R.sub.aR.sub.bNalkynyl-, R.sub.aR.sub.bNC(O)—, R.sub.aR.sub.bNC(O)alkynyl-, R.sub.aR.sub.bNC(O)alkoxy-, R.sub.aR.sub.bNSO.sub.2alkoxy-, and R.sub.w—O—N═CH—, wherein alkyl may be optionally substituted with O═ and R.sub.tN═; R.sub.a and R.sub.b are each individually selected from the group consisting of hydrogen, alkyl, carbonyl, alkenyl, alkenylcarbonyl, alkylsulfonyl, alkylcarbonyl, alkoxycarbonyl, alkoxycarbonylalkyl, alkoxycarbonylalkylcarbonyl, alkynyl, aryl, arylalkyl, arylalkenyl, arylcarbonyl, aryloxycarbonyl, arylalkyloxycarbonyl, heteroaryl, heterocycle, heterocyclealkyl, heterocyclealkenyl, heterocyclecarbonyl, heterocyclealkylcarbonyl, haloalkyl, trihaloalkyl, trihaloalkylcarbonyl, haloalkylcarbonyl, heterocycleoxycarbonyl hydroxyalkyl, and hydroxyalkylcarbonyl; R.sub.t is selected from the group consisting of hydrogen, alkyl and HO—; R.sub.f is selected from the group consisting of alkyl, cycloalkyl, cycloalkylalkyl, alkoxyalkyl, alkylthioalkyl, and haloalkyl; R, is selected from the group consisting of hydrogen, alkylcarbonyl-NH-alkyl-NHC(O)-alkyl, alkyl, alkyl(alkyl)N-alkyl-NHC(O)-alkyl, hydroxyalkylNHC(O)-alkyl, heterocyclealkyl-NHC(O)-alkyl, heterocycle-NHC(O)-alkyl, and heteroarylalkylNHC(O)-alkyl.

(117) Preferably (not applicable to R.sub.10-R.sub.13), R.sub.5 is selected from H, (C.sub.1-C.sub.4)alkyl, aromatic heterocycle(C.sub.1-C.sub.4)alkyl and non-aromatic heterocycle(C.sub.1-C.sub.4)alkyl. Among the most preferred R.sub.5 substituents is CH.sub.3. Preferably, R.sub.8 is selected from H, halogen, halo(C.sub.1-C.sub.4)alkyl, (C.sub.1-C.sub.4)alkyl, OCH.sub.3, OH, NH.sub.2, N(CH.sub.3).sub.2, —CN, —(C.sub.2-C.sub.4)alkyl-, and —U(C.sub.1-C.sub.3)alkyl-, wherein U is CH.sub.2, O, S, NH or NCH.sub.3. R.sub.8 can also form a 5- or 6-membered aromatic or non-aromatic cycle or heterocycle, incorporating two adjacent methylene groups of Z1. Most preferably, R.sub.8 is H, halo, (C.sub.1-C.sub.4)alkyl or halo(C.sub.1-C.sub.4)alkyl.

(118) In alternative embodiments, substituents R.sub.6 and Z.sub.2 in formula I or in any of formulae III, V and VII are independently selected from H (R.sub.6 only), (C.sub.1-C.sub.4)alkyl, (C.sub.1-C.sub.4)alkenyl, (C.sub.1-C.sub.4)alkynyl, (C.sub.1-C.sub.4)alkoxy, (C.sub.1-C.sub.4)alkenyloxy, (C.sub.1-C.sub.4)alkynyloxy, mercapto, (C.sub.1-C.sub.4)alkylthio, (C.sub.1-C.sub.4)alkenylthio, (C.sub.1-C.sub.4)alkynylthio, (C.sub.1-C.sub.4)alkylNH—, (C.sub.1-C.sub.4)alkenylNH—, (C.sub.1-C.sub.4)alkynylNH—, (C.sub.1-C.sub.3)alkylN(C.sub.1-C.sub.3)alkyl, (C.sub.1-C.sub.3)alkyl((C.sub.1-C.sub.3)alkyl)N—, (C.sub.1-C.sub.2)alkyl((C.sub.1-C.sub.2)alkyl)N(C.sub.1-C.sub.2)alkyl-, formyl, halo, halo(C.sub.1-C.sub.4)alkyl, halo(C.sub.1-C.sub.4)alkenyl, halo(C.sub.1-C.sub.4)alkynyl, halo(C.sub.1-C.sub.4)alkoxy, halo (C.sub.1-C.sub.4)alkenyloxy, halo (C.sub.1-C.sub.4)alkynyloxy, hydroxy(C.sub.1-C.sub.4)alkyl, hydroxy(C.sub.1-C.sub.4)alkenyl, and hydroxy(C.sub.1-C.sub.4)alkynyl.

(119) In more specific embodiments, an HSF inhibitor can be defined as a compound according to formulae III or IV presented below,

(120) ##STR00074##
wherein n, R.sub.2, R.sub.3, R.sub.5, R.sub.6, R.sub.7, R.sub.8, X.sub.5, X.sub.6, Z.sub.1 and Z.sub.2 are as defined before for the inhibitors of formulae I and II, respectively.

(121) In yet more specific embodiments, an HSF inhibitor can be defined as a compound according to formulae V or VI,

(122) ##STR00075##
wherein n, R.sub.2, R.sub.5, R.sub.6, R.sub.7, R.sub.8, X.sub.5, X.sub.6, Z.sub.1 and Z.sub.2 are as provided before. X.sub.7 is CH or N (or C(—R.sub.10)). Preferably, 3 or less of the X.sub.7 are N, and most preferably only one of the X.sub.7 is N; R.sub.10 is defined as R.sub.5 ands is 0, 1, 2, 3 or 4.

(123) More particularly, an HSF inhibitor can be defined as a compound according to formulae VII or VIII,

(124) ##STR00076##
wherein n, s, R.sub.2, R.sub.5, R.sub.6, R.sub.7, R.sub.8, R.sub.10, X.sub.5, X.sub.6, X.sub.7, Z.sub.1 and Z.sub.2 are as provided before.

(125) Alternatively, an HSF inhibitor can be defined as a compound according to formulae IX or X below,

(126) ##STR00077##
wherein n, R.sub.5, R.sub.6, R.sub.7, R.sub.8, X.sub.5, X.sub.6, Z.sub.1 and Z.sub.2 are as defined before, r is 0-4 and R.sub.11 is defined as R.sub.5.

(127) More particularly, an HSF inhibitor can be defined as a compound according to formulae XI or XII below, wherein n, R.sub.5, R.sub.6, R.sub.7, R.sub.8, X.sub.5, X.sub.6, Z.sub.1 and Z.sub.2 are as previously defined; o is 0, 1, 2, 3 or 4; R.sub.12 and R.sub.13 are as R.sub.5; and X.sub.8 is CH or N. Preferably, 3 or less of the X.sub.8 are N, and most preferably only one X.sub.8 is N.

(128) ##STR00078##

(129) Specific compounds of formulae I to XII include, but are not limited to: (E)-ethyl 4-oxo-4-(thiazol-2-ylamino)but-2-enoate (E)-ethyl 4-(benzo[d]thiazol-2-ylamino)-4-oxobut-2-enoate (E)-ethyl 4-((4-methylthiazol-2-yl)amino)-4-oxobut-2-enoate (E)-ethyl 4-oxo-4-((4-phenylthiazol-2-yl)amino)but-2-enoate (E)-ethyl 4-((5-methylthiazol-2-yl)amino)-4-oxobut-2-enoate (E)-ethyl 4-oxo-4-((5-phenylthiazol-2-yl)amino)but-2-enoate (E)-ethyl 4-(oxazol-2-ylamino)-4-oxobut-2-enoate (E)-ethyl 4-((1-methyl-1H-pyrazol-3-yl)amino)-4-oxobut-2-enoate (E)-ethyl 4-(isoxazol-3-ylamino)-4-oxobut-2-enoate (E)-ethyl 4-((1,3,4-thiadiazol-2-yl)amino)-4-oxobut-2-enoate (E)-ethyl 4-oxo-4-(pyridine-4-ylamino)but-2-enoate (E)-methyl 4-oxo-4-(thiazol-2-ylamino)but-2-enoate (E)-isopropyl 4-oxo-4-(thiazol-2-ylamino)but-2-enoate N.sup.1-ethyl-N.sup.4-(thiazol-2-yl)fumaramide N.sup.1-isopropyl-N.sup.4-(thiazol-2-yl)fumaramide Ethyl 3-(thiazol-2-ylcarbamoyl)benzoate (E)-butyl 4-oxo-4-(thiazol-2-ylamino)but-2-enoate (E)-ethyl 4-oxo-4-((4-phenylthiophen-2-yl)amino)but-2-enoate (E)-ethyl 4-oxo-4-((2-phenylthiazol-5-yl)amino)but-2-enoate (E)-ethyl 4-((3-methylisoxazol-5-yl)amino)-4-oxobut-2-enoate (E)-ethyl 4-((2-methylpyridin-4-yl)amino)-4-oxobut-2-enoate (E)-ethyl 4-oxo-4-((2-phenylpyridin-4-yl)amino)but-2-enoate (E)-ethyl 4-oxo-4-(quinolin-4-ylamino)but-2-enoate (E)-ethyl 4-((4-methyl-5-phenylthiazol-2-yl)amino)-4-oxobut-2-enoate (E)-ethyl 4-((4,5-dimethylthiazol-2-yl)amino)-4-oxobut-2-enoate (E)-ethyl 4-((4-cyclopropylthiazol-2-yl)amino)-4-oxobut-2-enoate (E)-ethyl 4-oxo-4-((4-(pyridin-2-yl)thiazol-2-yl)amino)but-2-enoate (E)-ethyl 4-oxo-4-((4-(pyridin-3-yl)thiazol-2-yl)amino)but-2-enoate (E)-ethyl 4-oxo-4-((4-(pyridine-4-yl)thiazol-2-yl)amino)but-2-enoate (E)-ethyl 4-((5-ethynylthiazol-2-yl)amino)-4-oxobut-2-enoate (E)-ethyl 4-oxo-4-((5-(pyridine-3-yl)thiazol-2-yl)amino)but-2-enoate (E)-ethyl 4-oxo-4-((5-(o-tolyl)thiazol-2-yl)amino)but-2-enoate N.sup.1, N.sup.1-diethyl-N.sup.4-(4-methylthiazol-2-yl)fumaramide N.sup.1, N.sup.1-dimethyl-N.sup.4-(4-methylthiazol-2-yl)fumaramide (E)-ethyl 4-(methyl(thiazol-2-yl)amino)-4-oxobut-2-enoate Ethyl 2-(thiazol-2-ylamino)thiazol-5-carboxylate (E)-ethyl 4-4([2,4′-bipyridin]-4-ylamino)-4-oxobut-2-enoate (E)-ethyl 4-((2-morpholinopyridin-4-yl)amino)-4-oxobut-2-enoate (E)-ethyl 4-((2-(4-methylpiperazin-1-yl)pyridine-4-yl)amino)-4-oxobut-2-enoate (E)-ethyl 4-oxo-4-((2-(o-tolyl)pyridine-4-yl)amino)but-2-enoate (E)-ethyl 4-oxo-4-((2-(m-tolyl)pyridine-4-yl)amino)but-2-enoate (E)-ethyl 4-oxo-4-((2-(p-tolyl)pyridine-4-yl)amino)but-2-enoate (E)-ethyl 4-((2,6-dimethylpyridin-4-yl)amino)-4-oxobut-2-enoate (E)-ethyl 4-((3-methylpyridin-4-yl)amino)-4-oxobut-2-enoate (E)-ethyl 4-([2,3′-bipyridin]-4-ylamino)-4-oxobut-2-enoate (E)-3-(pyrimidin-2-yl)-N-(thiazol-2-yl)acrylamide (E)-3-(1,2,4-oxadiazol-5-yl)-N-(thiazol-2-yl)acrylamide (E)-3-(5,6-dihydro-4H-1,3-oxazin-2-yl)-N-(thiazol-2-yl)acrylamide (E)-3-(1,3,4-oxadiazol-2-yl)-N-(thiazol-2-yl)acrylamide (E)-3-(4,5-dihydrooxazol-2-yl)-N-(thiazol-2-yl)acrylamide (E)-4-oxo-N-(thiazol-2-yl)hept-2-enamide (E)-3-(oxazol-2-yl)-N-(thiazol-2-yl)acrylamide (E)-4-oxo-N-(thiazol-2-yl)hept-2-enamide (E)-ethyl 4-oxo-4-((2-(pyridine-4-yl)ethyl)(thiazol-2-yl)amino)but-2-enoate (E)-ethyl 4-((morpholinoethyl)(thiazol-2-yl)amino)-4-oxobut-2-enoate (E)-methyl 2-(4-ethoxy-4-oxobut-2-enamido)thiazol-4-carboxylate (E)-ethyl 4-((4-ethyl-5-phenylthiazol-2-yl)amino)-4-oxobut-2-enoate (E)-ethyl 4-((2-(1-methyl-1H-imidazol-4-yl)ethyl)(thiazol-2-yl)amino)-4-oxobut-2-enoate (E)-ethyl 4-((4-carbamoylthiazol-2-yl)amino)-4-oxobut-2-enoate (E)-ethyl 4-oxo-4-((pyridin-2-yl)ethyl)(thiazol-2-yl)amino)but-2-enoate (E)-3-(5,6-dihydro-4H-[1,3]oxazin-2-yl)-N-methyl-N-(5-(pyridin-3-yl-thiazol-2-yl) acrylamide.

(130) The HSF inhibitors of the present disclosure may be prepared by a number of methods well known to those skilled in the art, including, but not limited to those described in the example section, or through modifications of these methods by applying standard techniques known to those skilled in the art of organic synthesis. All processes disclosed in association with the present disclosure are contemplated to be practiced at any scale, including milligram, gram, multigram, kilogram, multikilogram or commercial industrial scale.

(131) The present disclosure is meant to also encompass pharmaceutically acceptable derivatives or prodrugs of the HSF inhibitors described herein. Such pharmaceutically acceptable derivatives or prodrugs may be designed to enhance biological properties such as oral absorption, clearance, metabolism or compartmental distribution. Such derivations are well known in the art. As the skilled practitioner will recognize, the compounds of this disclosure may be modified by appending appropriate functionalities to enhance selective biological properties. Such modifications are known in the art and include those which increase biological penetration into a given biological compartment (e.g., blood, lymphatic system, central nervous system), increase oral availability, increase solubility to allow administration by injection, alter metabolism and alter rate of excretion.

(132) Further Characterization of Selected HSF Inhibitors

(133) I.sub.HSF 001 was included in the sublibrary as a compound that satisfied the parametric values of pharmacophore A4 modeled to fit the largest predicted cavity of the human HSF1 DNA-binding domain (cavity A). Because this cavity is far removed from the presumed DNA interaction region, compound 001 and its analogs were not expected to be capable of directly interfering with the binding of DNA to the HSF1 DNA-binding domain. However, they may affect DNA binding indirectly, if their binding altered the conformation of the HSF1 DNA-binding domain. Alternatively, or in addition, the compounds may interfere with homo-oligomerization of HSF1 or the binding of co-factors. Initial experiments had shown that certain compounds including I.sub.HSF 001 had apparent effects on HSF1 DNA-binding, whereas others compounds did not. We selected I.sub.HSF 058 and 115 for further characterization. I.sub.HSF 058 appeared to affect HSF1 DNA binding (but see the discussion below), whereas I.sub.HSF 115 did not. Direct interactions between the I.sub.HSF and HSF1 were detected by surface plasmon resonance (SPR). Recombinant full-length human HSF1 or a recombinant DNA-binding domain fragment of human HSF1 served as ligands. The SPR sensorgrams in FIG. 2C demonstrate that both I.sub.HSF 058 and I.sub.HSF 115 interacted in a dose-dependent fashion with full-length HSF1 as well as with the HSF1 DNA-binding domain fragment. Interactions were detected at compound concentrations of 15.6 μM and higher. These results demonstrate that the compounds are capable of directly binding the HSF1 DNA-binding domain. However, the compound concentrations at which direct interactions were observed were higher than those that caused inhibition of induced RLuc expression in Z74 cells. There are several factors that may account for this difference in sensitivity. One such factor may be the limited aqueous solubility of I.sub.HSF 058 and 115 that may have negatively affected the interaction experiments. Recombinant HSF1 or its DNA-binding domain fragment may not possess native conformations (or may be limited to conformations that support interactions with the I.sub.HSF less well than other conformations that occur in the cell). They lack posttranslational modifications and are not associated, as HSF1 is in its cellular environment, with multichaperone complexes and/or other co-factors. Moreover, immobilization of recombinant HSF1 and HSF1 DNA-binding fragment on a sensor chip may have further impaired their respective conformations (or limited their conformational flexibility), further jeopardizing their ability to interact with the I.sub.HSF.

(134) Electrophoretic mobility shift assays (EMSA) using a radiolabeled HSE DNA probe were carried out to assess HSF1 DNA-binding activity in extracts of heat-treated (43° C./30 min) HeLa cells that had been exposed to different concentrations of compounds 058 or 115 (FIG. 3A, top panels). Exposure to I.sub.HSF 058 at 25 or 50 μM concentrations resulted in important reductions in heat-induced DNA-binding activity. I.sub.HSF 115 failed to have a similar effect on DNA-binding activity. Anti-HSF1 western blot (WB) of the same extracts revealed reduced levels of HSF1 in the cells that had been exposed to I.sub.HSF 058 (FIG. 3A, middle panels), indicating that the compound had induced degradation of HSF1. I.sub.HSF 115 did not affect HSF1 levels. HSF1 degradation correlated well with the observed decrease in DNA-binding activity. Therefore, HSF1 degradation provides a sufficient explanation for the reduced HSE DNA-binding activity in cells treated with I.sub.HSF 058. We conclude that compound 058 does not impair the DNA-binding ability but the stability of HSF1. EMSA were also carried out using extracts of HeLa cells that had been heat-treated at 43° C. for 30 min. In these assays, no reduction of HSE DNA-binding activity was observed in the presence of I.sub.HSF 001, 011, 049, 050, 051, 052, 057 or 059 at concentrations as high as about 0.5 mM.

(135) We also investigated whether the compounds were capable of interfering with HSF1 oligomerization. Aliquots of the extracts were subjected to cross-linking with EGS (ethylene glycolbis(succinimidylsuccinate)) and then re-analyzed by anti-HSF1 WB. No impairment of oligomerization could be observed (FIG. 3A, bottom panels). I.sub.HSF 058-induced degradation of HSF1 was observed again in this analysis. Chromatin immunoprecipitation experiments were conducted to find out whether the I.sub.HSF could affect HSF1 DNA binding in vivo. In one such experiment, cultures of HeLa cells were exposed for two hours to 12.5 μM I.sub.HSF 058 or 115, respectively, or to vehicle (FIG. 3B). The cultures were subsequently heat-treated and then processed using a routine procedure. DNA fragments were co-precipitated by HSF1 antibodies, and a promoter segment of the hspa1a gene including the gene-proximal HSE sequence and the TATA box sequence was amplified by real-time PCR. Results indicated that HSF1 occupancy of the hspa1a promoter was not reduced substantially in cells that had been exposed to I.sub.HSF 115, indicating that the inhibitor of HSF1 also did not affect in vivo HSF1 DNA binding. The effect seen with I.sub.HSF 058 was ascribed to the aforementioned HSF1 destabilization.

(136) To assess the transcriptional effects of the I.sub.HSF, cultures of Z74 cells were exposed for two hours to I.sub.HSF 058 or I.sub.HSF 115 at different concentrations, or to vehicle. Following heat treatment and a one-hour post-incubation at 37° C., extracts were prepared, and polyadenylated rluc, hsp70b (hspa7) and hspa1a RNA was quantified by reverse transcription and real-time PCR. Exposure to the compounds resulted in a dose-dependent reduction of transcript levels of the hsp70b/hspa7 promoter-driven rluc gene (FIG. 4A, top panel). I.sub.HSF 115 had somewhat more important effects than I.sub.HSF 058. It is noted that rluc mRNA levels were already substantially reduced at 1 μM concentrations of the I.sub.HSF These data appear to reflect the true effects of the I.sub.HSF on heat-induced hsp promoter-driven gene transcription. We tend to discount the possibility that the latter data report effects on transcript stability. After all, the compounds were designed to bind to HSF1 and were found to do so. Nothing suggests that the I.sub.HSF could also interact with transcripts of HSF1 target genes. Dose-dependent inhibitory effects on transcript accumulation were also observed for the hspa1a and hspa7 (hsp70b) genes, although the latter genes appeared to be somewhat less sensitive to the I.sub.HSF than the hsp70b/hspa7-rluc gene, perhaps owing to compensatory mechanism(s) (FIG. 4A, lower panels).

(137) Effects of the I.sub.HSF could also be demonstrated at the level of heat-induced accumulation of inducible Hsp70 (mainly products of the hspa1a and hspa1b genes). WB experiments (FIG. 4B) reported detectable inhibitory effects of I.sub.HSF 115 at 3.125 μM and more substantial effects at 6.25 μM. I.sub.HSF 058 was less effective than I.sub.HSF 115.

(138) Recruitment of HSF1 to target promoters in response to a stress is mediated by ATF1/CREB. Takii, R. et al. (2015) Mol. Cell. Biol. 35: 11-25. ATF1/CREB regulates the stress-induced HSF1 transcription complex that includes BRG1 chromatin-remodeling complex and p300/CBP. The former complex promotes an active chromatin state in the promoters, whereas p300/CBP accelerates the shutdown of HSF1 DNA-binding activity as well as stabilizes HSF1 against proteasomal degradation during recovery from stress. To assess the effects of the I.sub.HSF on the assembly of the transcription complex, use was made of a HeLa-derived cell line that stably expresses a C-terminally FLAG-tagged HSF1. Cultures were heat-treated for 30 min at 43° C. in the presence or absence of I.sub.HSF 115, extracts were prepared, and tagged HSF1 was immunoprecipitated using an anti-FLAG antibody. WB analysis of immunoprecipitates revealed that I.sub.HSF 115 dramatically reduced the HSF1-ATF1 interaction. These findings strongly suggested that I.sub.HSF 115 interferes with the formation of ATF1-based transcription complexes that are instrumental in heat-induced transcription of HSF1 target genes.

(139) HSF Inhibitors Reduce the Viability of a Wide Range of Cancer Cells

(140) Cultures of HeLa cells were exposed for 96 hours to different concentrations of I.sub.HSF 001, 058 and 115, and viability was assessed using an Alamar Blue assay. It is noted that this standard assay examines attached cells only. This subject will be discussed in more detail below. Viability decreased in a dose-dependent fashion (FIG. 5A). Compound 115 was considerably more effective than compound 058, which in turn was more effective than compound 001. The relative cytotoxicity of the compounds appears to parallel their respective activities as inhibitors of HSF1 function (inhibition of RLuc expression in Z74 cells). We next examined the viability of different human cancer cell lines subsequent to a 96-hour exposure to compound 115 (Table 3). HSF1 was reported as being required for optimal p21 expression and for p53-mediated cell cycle arrest in response to genotoxins. Logan, I. R. et al. (2009) Nucleic Acids Res. 37: 2962-2971. These findings suggest that HSF1 may also play a p53-dependent pro-apoptotic function. Cell lines of different p53 status were included in the experiments in order to find out whether sensitivity to I.sub.HSF 115 was related to p53 status. Exposure to I.sub.HSF 115 reduced viability in all cell lines tested, although their sensitivity to the compound varied greatly. A comparison of moderately sensitive HeLa cells and highly sensitive multiple myeloma MM.1S cells is shown in FIGS. 5 A & B. Complete loss of viability of MM.1S cells occurred at 6.25 μM I.sub.HSF 115, in some experiments already at 3.125 μM. No effect of p53 status could be discovered. It is noted that a number of additional HSF inhibitors of the present disclosure were tested and found to effectively reduce the viability of human cancer cells.

(141) Further studies were aimed at finding out whether I.sub.HSF-exposed cancer cells were only growth-arrested or were actually killed and, if they were killed, by what mechanism. In one type of experiment, parallel sets of HeLa cell cultures were exposed for either 24 or 96 hours to different concentrations of I.sub.HSF 115. The first set was assayed for viability by the Alamar Blue assay without prior removal of floating cells. Attached and floating cells were assayed separately in the second set; floating cells were subsequently re-plated and incubated for 72 hours without compound. Cells were found to detach from the plates in a I.sub.HSF concentration-dependent fashion (FIG. 6A). The viability signal (in whole cultures and, more dramatically, when only attached cells were analyzed) decreased as a function of I.sub.HSF concentration. Most floating cells were incapable of re-attaching in the absence of compound (not shown); apparently, these cells had died. More profound effects were observed after 96 hours (right panel) than after 24 hours of exposure (left panel). Hence, the HeLa cells were killed by I.sub.HSF 115 in a concentration- and exposure time-dependent fashion. In a different type of experiment, sets of HeLa cell cultures that had been exposed to I.sub.HSF 115 for 15, 24 and 96 hours were stained with Trypan Blue. Stained (necrotic) and unstained (live) cells were counted (FIG. 6B). I.sub.HSF 115 caused numbers of live cells to decrease and numbers of necrotic cells to increase in a concentration- and time-dependent fashion. An increase in necrotic cells was already apparent after a 15-hour exposure to 3.125 μM I.sub.HSF 115. HeLa cells exposed to I.sub.HSF 115 appeared to die by a non-apoptotic mechanism. A cytometric analysis of HeLa cells exposed to 12.5 or 25 μM I.sub.HSF 115 for up to 24 h revealed only minor sub-G0/G1 fractions (6% or less of cells included) (FIG. 6D). In an annexin V/7 aminoactinomycin D double staining assay on HeLa cells exposed to 25 μM I.sub.HSF 115 for up to 24 hours, no more than about 15% of live cells were annexin V-stained (FIG. 6C). A similar analysis was carried out with multiple myeloma MM.1S cells. In contrast to what was found for HeLa cells, the mechanism of cell death of MM.1S cells included a prominent apoptotic component.

(142) TABLE-US-00003 TABLE 3 Effectiveness of I.sub.HSF 115 in reducing viability of human cancer cell lines - Alamar Blue assay EC.sub.50 Cell line Cancer P53 status (μM) HeLa Cervical Wt 6.2 BT-20 Breast Mut 8.2 BT-474 Breast Mut 5.2 MCF-7 Breast Wt 5.2 MDA-MB-231 Breast Mut 2.2 T47D Breast Mut 4.4 CAMA-1 Breast Mut 3.5 HCC1143 Breast Mut 4.8 PC-3 Prostate Null 4.0 NCI-H460 Lung Wt 9.4 A549 Lung Wt 18.8 NCI-H3122 Lung Wt 11.0 NCI-H1975 Lung Wt 5.1 NCI-H2228 Lung Wt 12.8 THP-1 Acute monocytic Mut 6.9 leukemia Saos-2 Osteosarcoma Null 8.0 MG-63 Osteosarcoma Mut 4.6 U-2 OS Osteosarcoma WT 5.2 HepG2 Hepatocellular Wt 5.2 MM.1S Multiple myeloma Wt 1.7 IM9 Multiple myeloma Wt 2.3 sNF02.2 Malignant peripheral Wt 8.4 nerve sheath sNF96.2 Malignant peripheral Wt 4.2 nerve sheath SK-OV-3 Ovarian carcinoma Mut 15 SK-N-SH Neuroblastoma Wt 4.0 MNNG/HOS Osteosarcoma Mut 9.2 OV56 Ovarian carcinoma Mut 6.8 PEA1 Ovarian carcinoma Mut 4.1 MDA-MB-453 Breast Mut 4.2 U266B1 Multiple myeloma Mut 3.9 RPMI 8266 Multiple myeloma Mut 3.6 A673 Ewing’s sarcoma Mut 1.9 MM.1R Multiple myeloma Wt 1.7

(143) To obtain evidence that the cytotoxic effect of I.sub.HSF 115 is at least in part a consequence of inhibition of HSF1, we transduced HeLa cells with HSF1 siRNA-expressing lentivirus and selected cell lines that exhibited low levels of HSF1. The most severely HSF1-deficient lines grew slowly. FIG. 7 reports on the properties of one such line, KD 13, which was compared to control lentivirus-transduced cells (KD control). The WB in FIG. 7A (upper panel) confirms that the KD 13 line is essentially devoid of HSF1. The lower panel compares growth of the cell line with that of the control cells (the graph showing numbers of Trypan Blue-excluding (live) cells after 1-4 days of culture). We conclude that the KD 13 line has partially overcome its HSF1 dependency. If the cytotoxicity of I.sub.HSF 115 was mediated through inhibition of HSF1, the cell line that had partially lost its HSF1 dependency should have been more resistant to killing by I.sub.HSF 115 than the control lentivirus-transduced cells. The results of Alamar Blue assays performed after 96 hours of exposure to different concentrations of the I.sub.HSF showed this to be the case (FIG. 7B).

(144) Uses of HSF Inhibitors

(145) In general, HSF inhibitors of the present disclosure can be used to inhibit the stress-induced transactivation ability of mammalian HSF1 in a mammalian cell or animal, in a human cell, tissue, organ or organism. Under “stress-induced transactivation ability” is understood the ability of activated HSF1 to cause a transient accumulation of transcripts of its target genes. Because HSF1 is the key regulator of the stress protein response, i.e., is a key regulator of proteostasis, a major application for the HSF inhibitors of the present disclosure will be in the therapy of conditions or diseases that are characterized by an increased demand for HSF1 activity by certain cells, tissues or organs compared to corresponding cells, tissues or organs in healthy subjects. Often, cells, tissues or organs in an afflicted subject display increased levels of HSF1 and/or of HSF1 activity (as detected by any suitable assay, e.g., DNA-binding assays, transactivation assays, nuclear accumulation assays, WB, etc.). Diseases that generally appear to have an increased demand for HSF1 activity include cancers of different types. Furthermore, certain infections also result in activation of HSF1, and progagation of the infectious agent is dependent on this induced HSF1 activity. A recently discovered example involves orthopoxviruses. Filone et al. (2014) PLoS Pathog. 2014, 10, e1003904. See also Wang et al. (2010) J. Transl. Med. 8: 44. In such infections, administration of an inhibitor of HSF1 can be an effective therapeutic measure. Moreover, HSF1 inhibitors may find uses in situations in which pharmacologically induced HSF1 activity may have negative or undesired consequences (e.g., during therapy with HSP90 inhibitors). Experiments with an HSF1−/− mouse strain revealed that HSF1 is a maternal effect gene. No embryos lacking HSF1 activity developed to the blastocyst stage. Christians et al. (2000) Nature 407: 693-4. Hence, HSF1 inhibitors are expected to have contraceptive activity.

(146) In certain embodiments, one or more of the HSF inhibitors of the present disclosure are used either alone or in combination with other active anti-cancer therapeutic agents or regimens such as radiation therapy to prevent or treat cancer or neoplastic disease. Cancers to be treated include, but are not limited to, B cell lymphoma, T cell lymphoma, myeloma, leukemia (AML, ALL, CML, CLL), hematopoietic neoplasias, thymoma, lymphoma, sarcoma, lung cancer, liver cancer, non-Hodgkins lymphoma, Hodgkins lymphoma, Burkitt's lymphoma, breast cancer, pancreatic cancer, colon cancer, lung cancer, renal cancer, bladder cancer, liver cancer, prostate cancer, ovarian cancer, primary or metastatic melanoma, squamous cell carcinoma, basal cell carcinoma, sebaceous cell carcinoma, brain cancer (astrocytoma, glioma, glioblastoma, ependymoma, medulloblastoma, meningioma, oligodendroglioma, oligoastrocytoma), angiosarcoma, hemangiosarcoma, adenocarcinoma, liposarcoma, head and neck carcinoma, thyroid carcinoma, soft tissue sarcoma, osteosarcoma, testicular cancer, uterine cancer, cervical cancer, gastrointestinal cancer, colorectal cancer, oral cancer, nasopharyngeal cancer, oropharyngeal cancer, esophageal cancer, stomach cancer, multiple myeloma, bile duct cancer, cervical cancer, laryngeal cancer, penile cancer, urethral cancer, anal cancer, vulvar cancer, vaginal cancer, gall bladder cancer, thymoma, salivary gland cancer, lip and oral cavity cancer, adenocortical cancer, non-melanoma skin cancer, pleura mesothelioma, joint cancer, hypopharyngeal cancer, ureter cancer, peritoneum cancer, omentum cancer, mesentery cancer, Ewing's sarcoma, rhabdomyosarcoma, spinal cord cancer, endometrial cancer, neuroblastoma, pituitary cancer, retinoblastoma, eye cancer, and islet cell cancer, and any other cancer now known or later identified to be dependent or to benefit from HSF activity.

(147) Examples of types of anti-cancer therapeutic agents with which the HSF inhibitors of the present disclosure may be combined include, but are not limited to, e.g., chemotherapeutic agents, growth inhibitory agents, agents used in radiation therapy (radioactive isotopes (e.g., I.sub.131, I.sub.125, Y.sub.90 and Re.sub.186), toxins such as enzymatically active toxins of bacterial, fungal, plant or animal origin, or fragments thereof, anti-angiogenesis agents, apoptotic agents, anti-tubulin agents, and other agents to treat cancer, such as inhibitors of HSP90, the proteasome or histone deacetylases (HDAC), in particular HDAC6, anti-HER-2 antibodies (e.g., Herceptin (Trastuzumab)), anti-CD20 antibodies, epidermal growth factor receptor (EGFR) antagonist (e.g., a tyrosine kinase inhibitor), HER1/EGFR inhibitor (e.g., erlotinib (Tarceva™)), platelet derived growth factor inhibitors (e.g., Gleevec™ (Imatinib Mesylate)), a COX-2 inhibitor (e.g., celecoxib), interferons, cytokines, antagonists (e.g., neutralizing antibodies) that bind to one or more of the following targets: ErbB2, ErbB3, ErbB4, PDGFR-beta, BlyS, APRIL, BCMA or VEGF receptor(s), TRAIL/Apo2, and other bioactive and organic chemical agents, etc. Combinations thereof are also included in the disclosure.

(148) Example chemotherapeutic agents include one or more chemical compounds useful in the treatment of cancer. Non-limiting examples of chemotherapeutic agents include alkylating agents such as thiotepa and CYTOXAN, cyclosphosphamide; alkyl sulfonates such as busulfan, improsulfan and piposulfan; aziridines such as benzodopa, carboquone, meturedopa, and uredopa; ethylenimines and methylamelamines including altretamine, triethylenemelamine, triethylenephosphoramide, triethylenethiophosphoramide and trimethylol melamine; acetogenins (such as bullatacin and bullatacinone); a camptothecin (including the synthetic analogue topotecan); bryostatin; callystatin; CC-1065 (including its adozelesin, carzelesin and bizelesin synthetic analogues); cryptophycins (particularly cryptophycin 1 and cryptophycin 8); dolastatin; duocarmycin (including the synthetic analogues, KW-2189 and CB1-TM1); eleutherobin; pancratistatin; a sarcodictyin; spongistatin; nitrogen mustards such as chlorambucil, chlornaphazine, cholophosphamide, estramustine, ifosfamide, mechlorethamine, mechlorethamine oxide hydrochloride, melphalan, novembichin, phenesterine, prednimustine, trofosfamide, uracil mustard; nitrosureas such as carmustine, chlorozotocin, fotemustine, lomustine, nimustine, and ranimustine; antibiotics such as the enediyne antibiotics (e.g., calicheamicin, especially calicheamicin gamma1); dynemicin, including dynemicin A; bisphosphonates, such as clodronate; an esperamicin; as well as neocarzinostatin chromophore and related enediyne chromoprotein antibiotics, aclacinomysins, actinomycin, authramycin, azaserine, bleomycins, cactinomycin, carabicin, caminomycin, carzinophilin, chromomycins, dactinomycin, daunorubicin, detorubicin, 6-diazo-5-oxo-L-norleucine, ADRIAMYCIN™, doxorubicin (including morpholino-doxorubicin, cyanomorpholino-doxorubicin, 2-pyrrolino-doxorubicin and deoxydoxorubicin), epirubicin, esorubicin, idarubicin, marcellomycin, mitomycins such as mitomycin C, mycophenolic acid, nogalamycin, olivomycins, peplomycin, potfiromycin, puromycin, quelamycin, rodorubicin, streptonigrin, streptozocin, tubercidin, ubenimex, zinostatin, zorubicin; anti-metabolites such as methotrexate and 5-fluorouracil (5-FU); folic acid analogues such as denopterin, methotrexate, pteropterin, trimetrexate; purine analogs such as fludarabine, 6-mercaptopurine, thiamiprine, thioguanine; pyrimidine analogs such as ancitabine, azacitidine, 6-azauridine, carmofur, cytarabine, dideoxyuridine, doxifluridine, enocitabine, floxuridine; androgens such as calusterone, dromostanolone propionate, epitiostanol, mepitiostane, testolactone; anti-adrenals such as aminoglutethimide, mitotane, trilostane; folic acid replenisher such as frolinic acid; aceglatone; aldophosphamide glycoside; aminolevulinic acid; eniluracil; amsacrine; bestrabucil; bisantrene; edatraxate; defofamine; demecolcine; diaziquone; elformithine; elliptinium acetate; an epothilone; etoglucid; gallium nitrate; hydroxyurea; lentinan; lonidamine; maytansinoids such as maytansine and ansamitocins; mitoguazone; mitoxantrone; mopidanmol; nitraerine; pentostatin; phenamet; pirarubicin; losoxantrone; podophyllinic acid; 2-ethylhydrazide; procarbazine; PSK, polysaccharide complex (JHS Natural Products, Eugene, Oreg.); razoxane; rhizoxin; sizofuran; spirogermanium; tenuazonic acid; triaziquone; 2,2′,2″-trichlorotriethylamine; trichothecenes (such as T-2 toxin, verracurin A, roridin A and anguidine); urethan; vindesine; dacarbazine; mannomustine; mitobronitol; mitolactol; pipobroman; gacytosine; arabinoside (“Ara-C”); cyclophosphamide; thiotepa; taxoids, e.g., TAXOL™, paclitaxel (Bristol-Myers Squibb Oncology, Princeton, N.J.), ABRAXANE™, Cremophor-free, albumin-engineered nanoparticle formulation of paclitaxel (American Pharmaceutical Partners, Schaumberg, Ill.), and TAXOTERE™ doxetaxel (Rhone-Poulenc Rorer, Antony, France); chloranbucil; GEMZAR™ gemcitabine; 6-thioguanine; mercaptopurine; platinum analogs such as cisplatin and carboplatin; vinblastine; platinum; etoposide (VP-16); ifosfamide; mitoxantrone; vincristine; NAVELBINE vinorelbine; novantrone; teniposide; edatrexate; daunomycin; aminopterin; xeloda; ibandronate; irinotecan (Camptosar, CPT-11) (including the treatment regimen of irinotecan with 5-FU and leucovorin); topoisomerase inhibitor RFS 2000; difluoromethylornithine (DMFO); retinoids such as retinoic acid; capecitabine; combretastatin; leucovorin (LV); oxaliplatin, including the oxaliplatin treatment regimen (FOLFOX); inhibitors of PKC-alpha, Raf, H-Ras, EGFR (e.g., erlotinib (Tarceva™)) and VEGF-A that reduce cell proliferation and pharmaceutically acceptable salts, acids or derivatives of any of the above. Also encompassed are anti-hormonal agents that act to regulate or inhibit hormone action on tumors such as anti-estrogens and selective estrogen receptor modulators (SERMs), including, for example, tamoxifen (including NOLVADEX™ tamoxifen), raloxifene, droloxifene, 4-hydroxytamoxifen, trioxifene, keoxifene, LY117018, onapristone, and FARESTON toremifene; aromatase inhibitors that inhibit the enzyme aromatase, which regulates estrogen production in the adrenal glands, such as, for example, 4(5)-imidazoles, aminoglutethimide, MEGASE™ megestrol acetate, AROMASIN™ exemestane, formestane, fadrozole, RIVISOR™ vorozole, FEMARA™ letrozole, and ARIMIDEX™ anastrozole; and antiandrogens such as flutamide, nilutamide, bicalutamide, leuprolide, and goserelin; as well as troxacitabine (a 1,3-dioxolane nucleoside cytosine analog); antisense oligonucleotides, particularly those which inhibit expression of genes in signaling pathways implicated in aberrant cell proliferation, such as, for example, PKC-alpha, Raf and H-Ras; ribozymes such as a VEGF expression inhibitor (e.g., ANGIOZYME™ ribozyme) and a HER2 expression inhibitor; vaccines such as gene therapy vaccines, for example, ALLOVECTIN™ vaccine, LEUVECTIN™ vaccine, and VAXID™ vaccine; PROLEUKIN™, rIL-2; LURTOTECAN topoisomerase 1 inhibitor; ABARELIX™ rmRH; vinorelbine and esperamicins, and pharmaceutically acceptable salts, acids or derivatives of any of the above.

(149) “Radiation therapy” refers to the use of directed gamma rays or beta rays to induce sufficient damage to a cell to limit its ability to function normally or to destroy the cell altogether. It will be appreciated that there will be many ways known in the art to determine the dosage and duration of treatment. Typical treatments are given as a one-time administration and typical dosages range from 10 to 200 units (Grays) per day.

(150) Because the HSF inhibitors of the present disclosure are directed to hitherto unexploited tumor cell targets (i.e., HSF), they can enhance, additively or, possibly, synergistically, the activity of the aforementioned anti-cancer therapeutic agents or of radiation therapy. Preferred combinations will comprise an HSF inhibitor of the present disclosure and an anti-cancer therapeutic agent that induces a stress protein response, i.e., that enhances expression of heat shock proteins. Elevated levels of heat shock proteins have cytoprotective effects and thereby may, or are known to, negatively affect the anticancer efficacy of such agents. Agents that induce heat shock protein expression include HSP90 inhibitors, HDAC6 inhibitors and proteasome inhibitors.

(151) Examples of HSP90 inhibitors that can be used in combination with an HSF inhibitor of the present disclosure include, but are not limited to, quinone ansamycin antibiotics, such as the macbecins, geldanamycin, including derivatives of geldanamycin, such as 17-AAG, herbimycin A, radicicol, and synthetic compounds that can bind into the N-terminal ATP-binding site of HSP90. HSP90 inhibitors that bind the C-terminal ATP-binding site have also been identified such as novobiocin, cisplatin, epigallocatechin-3-gallate (EGCG) and taxol (Donelly and Blagg (2008) Curr. Med. Chem. 15: 2702-2717). Another HSP90 inhibitor that appears to interact with the C-terminal ATP-binding site is celastrol and its derivatives (Zhang et al. (2009) J. Biol. Chem. 284: 33381-33389). HSP90 inhibitors are also described in U.S. Pat. Publ. Nos. 2008/0269218, 2008/0306054, 2010/0249231 and 2011/0112099.

(152) Inhibitors of HDAC6 activity that can be used in combination with an HSF inhibitor of the present disclosure include but are not limited to hydroxamic acid-based HDAC inhibitors, suberoylanilide hydroxamic acid (SAHA) and its derivatives, NVP-LAQ824, trichostatins including trichostatin A, scriptaid, m-carboxycinnamic acid bishydroxamic acid (CBHA), ABHA, pyroxamide, propenamides, oxamflatin, 6-(3-chlorophenylureido)caproic hydroxamic acid (3-C1-UCHA), A-161906, JNJ-16241199, tubacin and tubacin analogs, siRNA (see, e.g., U.S. Pat. Publ. No. 2007/0207950), butyrate, phenylbutyrate, sodium butyrate, valproate, (−)-depudecin, sirtinol, hydroxamic acid, epoxyketone-containing cyclic tetrapeptides, trapoxins, HC-toxin, chlamydocin, diheteropeptide, WF-3161, Cy1-1, Cy1-2, non-epoxyketone-containing cyclic tetrapeptides, PXD101, dimeric HDAC inhibitors, certain depsipeptides, FR901228 (FK228), apicidin, APHA compound 8, cyclic-hydroxamic-acid-containing peptides (CHAPS), benzamides and benzamide analogs, MS-275 (MS-27-275), CI-994, LBH589, deprudecin, organosulfur compounds and any combination thereof. An inhibitor of HDAC6 activity can be an inhibitor that acts at the level of transcription and/or translation of the HDAC6 protein, whereby such an inhibitor alters HDAC6 activity by decreasing the amount of functional HDAC6 protein produced. An inhibitor of HDAC6 activity can be, but is not limited to, an antisense nucleic acid, a ribozyme (e.g., as described in U.S. Pat. No. 5,877,022), RNAs that effect spliceosome-mediated trans-splicing (Puttaraju et al. (1999) Nature Biotech. 17:246; U.S. Pat. Nos. 6,013,487; 6,083,702), RNAs that trigger RNA interference mechanisms (RNAi), including small interfering RNAs (siRNA) that mediate gene silencing (Kawaguchi et al., (2003) Cell 115:727-738; Sharp et al. (2000) Science 287:2431) and/or other non-translated RNAs, such as “guide” RNAs (Gorman et al., (1998) Proc. Nat. Acad. Sci. USA 95:4929; U.S. Pat. No. 5,869,248) and the like, as are known in the art.

(153) Examples of a proteasome inhibitor (European Journal of Cancer, 42, 1623, (2006)) that can be used in combination with an HSF inhibitor of the present disclosure include, for example, bortezomib, MG-132, carfilzomib (PR-171; US2005/0245435), NPI-0052 (Br. J. Cancer, 95, 961, (2006)), SC-68896 (WO2007/017284), tyropeptin A, TP-110, TP-104 (WO2005/105826), belactosins (Biochem. Pharmacol., 67, 227, (2004)) and the like.

(154) HSF inhibitors of the disclosure may also be used to mitigate resistance to anti-cancer therapeutic agents that target signaling through an HSF. For example, ErbB2 over-expression results in enhanced aerobic glycolysis as a consequence of over-expression of HSF1 and its target LDH A. Trastuzumab inhibits this activity. Resistance to trastuzumab can be reversed by inhibiting aerobic glycolysis, which can be achieved by inhibition of HSF1. Zhao et al. (2011) Cancer Res. 71: 4585-97; Zhao et al. (2009) Oncogene 28: 3689-3701.

(155) The anti-cancer therapeutic agent and/or radiation therapy can be administered according to therapeutic protocols well known in the art. It will be apparent to those skilled in the art that the administration of the anti-cancer therapeutic agent and/or radiation therapy can be varied depending on the disease being treated and the known effects of the chemotherapeutic agent and/or radiation therapy on that disease. Also, in accordance with the knowledge of the skilled clinician, the therapeutic protocols (e.g., dosage amounts and times of administration) can be varied in view of the observed effects of the administered therapeutic agents (i.e., anti-cancer therapeutic agent or radiation) on the patient, and in view of the observed responses of the disease to the administered therapeutic agents, and observed adverse affects.

(156) Also, in general, an HSF inhibitor of the present disclosure and an anti-cancer therapeutic agent do not have to be administered in the same pharmaceutical composition, and, due to differences in physical and chemical characteristics, may be administered by different routes. For example, compounds of the present disclosure may be administered intravenously, whereas the chemotherapeutic agent may be administered orally. The determination of the mode of administration and the advisability of administration, where possible, in the same pharmaceutical composition, is well within the knowledge of the skilled clinician. The initial administration can be made according to established protocols known in the art, and then, based upon the observed effects, the dosage, modes of administration and times of administration can be modified by the skilled clinician.

(157) The particular choice of chemotherapeutic agent or radiation will depend upon the diagnosis of the physicians and their judgment of the condition of the patient and the appropriate treatment protocol.

(158) An HSF inhibitor of the present disclosure and an anti-cancer therapeutic agent and/or radiation may be administered concurrently (e.g., simultaneously, essentially simultaneously or within the same treatment protocol) or sequentially, depending upon the nature of the proliferative disease, the condition of the patient, and the actual choice of anti-cancer therapeutic agent and/or radiation to be administered in conjunction with the HSF inhibitor.

(159) If an HSF inhibitor of the present disclosure and the anti-cancer therapeutic agent and/or radiation is not administered simultaneously or essentially simultaneously, then the optimum order of administration of the compound of the present disclosure, and the anti-cancer therapeutic agent and/or radiation, may be different for different tumors. Thus, in certain situations the HSF inhibitor of the present disclosure may be administered first followed by the administration of the anti-cancer therapeutic agent and/or radiation, whereas in other situations the anti-cancer therapeutic agent and/or radiation may be administered first followed by the administration of an HSF inhibitor of the present disclosure. Such administration may be repeated during a single treatment protocol. The determination of the order of administration, and the number of repetitions of administration of each therapeutic agent during a treatment protocol, is well within the knowledge of the skilled physician after evaluation of the disease being treated and the condition of the patient. For example, the anti-cancer therapeutic agent and/or radiation may be administered first, especially if it has a cytotoxic effect, and then the treatment may be continued with the administration of a compound of the present disclosure followed, where determined advantageous, by the re-administration of the anti-cancer therapeutic agent and/or radiation, and so on until the treatment protocol is completed.

(160) Thus, in accordance with experience and knowledge, the practicing physician can modify each protocol for the administration of a component (therapeutic agent, i.e., compound of the present disclosure, anti-cancer therapeutic agent or radiation) of the treatment according to the individual patient's needs, as the treatment proceeds.

(161) Pharmaceutical Compositions

(162) The pharmaceutical compositions of the present disclosure comprise an effective amount of an HSF inhibitor of the present disclosure (preferably a compound of formulae (II), (V), (IX) or (X)) formulated together with one or more pharmaceutically acceptable carriers or excipients.

(163) The pharmaceutical compositions of this disclosure may be administered orally, parenterally, by inhalation spray, topically, rectally, nasally, buccally, vaginally or via an implanted reservoir, preferably by oral administration or administration by injection (or infusion).

(164) The pharmaceutical compositions of this disclosure may contain any conventional non-toxic pharmaceutically acceptable carriers, adjuvants or vehicles. In some cases, the pH of the formulation may be adjusted with pharmaceutically acceptable acids, bases or buffers to enhance the stability of the formulated HSF inhibitor or its delivery form. The term parenteral as used herein includes subcutaneous, intracutaneous, intravenous, intramuscular, intraarticular, intraarterial, intrasynovial, intrasternal, intrathecal, intralesional and intracranial injection or infusion techniques.

(165) Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups and elixirs. In addition to an active compound (i.e., an HSF inhibitor of the disclosure), the liquid dosage forms may contain inert diluents commonly used in the art such as, for example, water or other solvents, solubilizing agents and emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide, oils (in particular, cottonseed, groundnut, corn, germ, olive, castor, and sesame oils), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols and fatty acid esters of sorbitan, and mixtures thereof. Besides inert diluents, the oral compositions can also include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents.

(166) Injectable preparations, for example, sterile injectable aqueous or oleaginous 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 injectable solution, suspension or emulsion in a nontoxic parenterally acceptable diluent or solvent. Among the acceptable vehicles and solvents that may be employed are water, Ringer's solution, U.S.P. and isotonic sodium chloride solution. Solubilizing excipients include water-soluble organic solvents such as polyethylene glycol 300, polyethylene glycol 400, ethanol, propylene glycol, glycerin, N-methyl-2-pyrrolidone, dimethylacetamide and dimethylsulfoxide; non-ionic surfactants such as Cremophor EL, Cremophor RH40, Cremophor RH60, Solutol HS15, d-α-tocopherol polyethylene glycol 1000 succinate, polysorbate 20, polysorbate 80, sorbitan monooleate, poloxamer 407, Labrafil M-1944CS, Labrafil M-2125CS, Labrasol, Gellucire 44/14, Softigen 767, and mono- and di-fatty acid esters of PEG 300, 400 and 1750; water-insoluble lipids such as castor oil, corn oil, cottonseed oil, olive oil, peanut oil, peppermint oil, safflower oil, sesame oil, soybean oil, hydrogenated vegetable oils, hydrogenated soybean oil, and medium-chain triglycerides of coconut oil and palm seed oil, various cyclodextrins such as α-cyclodextrin, β-cyclodextrin, hydroxypropyl-β-cyclodextrin (e.g., Kleptose), and sulfobutylether-β-cyclodextrin (e.g., Captisol); and phospholipids such as lecithin, hydrogenated soy phosphatidylcholine, distearoylphosphatidylglycerol, L-α-dimyristoylphosphatidylcholine and L-α-dimyristoylphosphatidylglycerol. Strickley (2004) Pharm. Res. 21: 201-30.

(167) The injectable formulations can be sterilized, for example, by filtration through a bacteria-retaining filter, or by incorporating sterilizing agents in a sterile solid composition (or sterilize the solid composition by irradiation) which subsequently can be dissolved or dispersed in sterile water or other sterile injectable medium prior to use.

(168) 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 accomplished 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 upon 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 accomplished by dissolving or suspending the drug in an oil vehicle. Injectable depot forms are made by microencapsulating the drug in biodegradable polymers such as polylactide-polyglycolide. Depending upon the ratio of drug to polymer and the nature of the particular polymer employed, the rate of drug release can be controlled. Examples of other biodegradable polymers include poly(orthoesters) and poly(anhydrides). Depot injectable formulations are also prepared by entrapping the drug in liposomes or microemulsions that are compatible with body tissues.

(169) Compositions for rectal or vaginal administration are preferably suppositories which can be prepared by mixing the compounds of this disclosure with suitable non-irritating excipients or carriers such as cocoa butter, polyethylene glycol or a suppository wax which are solid at ambient temperature but liquid at body temperature and therefore melt in the rectum or vaginal cavity and release the active compound.

(170) Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In such solid dosage forms, the active compound is mixed with at least one inert, pharmaceutically acceptable excipient or carrier such as sodium citrate or dicalcium phosphate and/or: a) fillers or extenders such as starches, lactose, sucrose, glucose, mannitol, and silicic acid, b) binders such as, for example, carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidone, sucrose, 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, f) absorption accelerators such as quaternary ammonium compounds, g) wetting agents such as, for example, 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 mixtures thereof. In the case of capsules, tablets and pills, the dosage form may also comprise buffering agents.

(171) Solid compositions of a similar type may also be employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugar as well as high molecular weight polyethylene glycols and the like.

(172) 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 composition that they release the active ingredient(s) only, or preferentially, in a certain part of the intestinal tract, optionally, in a delayed manner. Examples of embedding compositions that can be used include polymeric substances and waxes.

(173) Dosage forms for topical or transdermal administration of a compound of this disclosure include ointments, pastes, creams, lotions, gels, powders, solutions, sprays, inhalants or patches. The active component is admixed under sterile conditions with a pharmaceutically acceptable carrier and any preservatives or buffers as may be required. Ophthalmic formulations, ear drops, eye ointments, powders and solutions are also contemplated as being within the scope of this disclosure.

(174) The ointments, pastes, creams and gels may contain, in addition to a compound of this disclosure, excipients such as animal and vegetable fats, oils, waxes, paraffins, starch, tragacanth, cellulose derivatives, polyethylene glycols, silicones, bentonites, silicic acid, talc and zinc oxide, or mixtures thereof.

(175) Powders and sprays can contain, in addition to compounds of this disclosure, excipients such as lactose, talc, silicic acid, aluminum hydroxide, calcium silicates and polyamide powder, or mixtures of these substances. Sprays can additionally contain customary propellants.

(176) Transdermal patches can be made by dissolving or dispensing the active in the proper medium. Absorption enhancers can also be used to increase the flux of the compound across the skin. The rate can be controlled by either providing a rate controlling membrane or by dispersing the compound in a polymer matrix or gel.

(177) For pulmonary delivery, a therapeutic composition of the disclosure is formulated and administered to the patient in solid or liquid particulate form by direct administration e.g., inhalation into the respiratory system. Solid or liquid particulate forms of the active compound prepared for practicing the present disclosure include particles of respirable size: that is, particles of a size sufficiently small to pass through the mouth and larynx upon inhalation and into the bronchi and alveoli of the lungs. Delivery of aerosolized therapeutics, particularly aerosolized antibiotics, is known in the art (see, for example U.S. Pat. Nos. 5,767,068, 5,508,269 and WO 98/43650). A discussion of pulmonary delivery of antibiotics is also found in U.S. Pat. No. 6,014,969.

(178) The total daily dose of an HSF inhibitor of this disclosure administered to a human subject or patient in single or in divided doses can be, for example, from 0.01 to 50 mg/kg body weight or more usually from 0.1 to 30 mg/kg body weight. Single dose compositions may contain such amounts or submultiples thereof to make up the daily dose. In general, treatment regimens according to the present disclosure comprise administration to a human subject in need of such treatment from about 1 mg to about 5000 mg of the compound(s) of this disclosure per day in single or divided doses. Doses for mammalian animals can be estimated based on the latter human doses.

(179) An HSF inhibitor of this disclosure can be administered, for example, by injection, intravenously, intraarterially, subdermally, intraperitoneally, intramuscularly, or subcutaneously; or orally, buccally, nasally, transmucosally, topically, in an ophthalmic preparation, or by inhalation, as a daily dose of about 0.01 to about 50 mg/kg of body weight. Alternatively, dosages (based on a daily dose of between about 1 mg and 5000 mg) may be administered every 4 to 120 hours, or according to the requirements of the particular inhibitor drug. The methods herein contemplate administration of an effective amount of HSF inhibitor (in a pharmaceutical composition) to achieve the desired or stated effect. Typically, the pharmaceutical compositions of this disclosure will be administered from about 1 to about 6 times per day or alternatively, as a continuous infusion. Such administration can be used as a chronic or acute therapy. The amount of active compound that may be combined with pharmaceutically acceptable excipients or carriers to produce a single dosage form will vary depending upon the host treated and the particular mode of administration. A typical composition will contain from about 5% to about 95% active compound (w/w). Alternatively, such preparations may contain from about 20% to about 80% active compound. Specific dosage and treatment regimens for any particular patient will depend upon a variety of factors, including the activity of the specific HSF inhibitor employed, the age, body weight, general health status, sex, diet, time of administration, rate of excretion, drug combination, the severity and course of the disease, condition or symptoms, the patient's disposition to the disease, condition or symptoms, and the judgment of the treating physician.

(180) All references cited in this application, including publications, patents and patent applications, shall be considered as having been incorporated in their entirety.

(181) Recitation of ranges of values herein are merely intended to serve as a shorthand method of referring individually to each separate value falling within the range, unless otherwise indicated herein, and each separate value is incorporated into the specification as if it were individually recited herein. Unless otherwise stated, all exact values provided herein are representative of corresponding approximate values (e. g., all exact exemplary values provided with respect to a particular factor or measurement can be considered to also provide a corresponding approximate measurement, modified by “about,” where appropriate).

(182) The description herein of any aspect or embodiment of the invention using terms such as reference to an element or elements is intended to provide support for a similar aspect or embodiment of the disclosure that “consists of,” “consists essentially of” or “substantially comprises” that particular element or elements, unless otherwise stated or clearly contradicted by context (e.g., a composition described herein as comprising a particular element should be understood as also describing a composition consisting of that element, unless otherwise stated or clearly contradicted by context).

(183) This invention includes all modifications and equivalents of the subject matter recited in the aspects or claims presented herein to the maximum extent permitted by applicable law.

(184) The present disclosure, thus generally described, will be understood more readily by reference to the following examples, which are provided by way of illustration and are not intended to be limiting of the present invention.

EXAMPLES

Example 1: Synthesis of HSF Inhibitors

(185) Abbreviations

(186) APCI, atmospheric pressure chemical ionization; Aq., aqueous; Boc, tert-butoxycarbonyl; Boc.sub.2O, Di-tert-butyl dicarbonate; Conc., concentrated; DCE, 1,2-dichloroethane; DCM, dichloromethane; DDQ, 2,3-dichloro-5,6-dicyano-1,4-benzoquinone; DIPEA, N,N-diisopropylethylamine; DMF-DMA, N—N Dimethyl formamide dimethyl acetal; DME, 1,2-dimethoxyethane; DMF, dimethylformamide; EDC, 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide; EL S D, evaporative light scattering detection; equiv., equivalents; Et.sub.2O, diethyl ether; EtOAc, ethyl acetate; EtOH, ethanol; h, hours; HATU, 2-(7-aza-1H-benzotriazole-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate; HOBt, N-hydroxybenzotriazole; HPLC, high performance liquid chromatography; LC-MS, liquid chromatography mass spectrometry; IBCF, isobutyl chloroformate; IPA, 2-propanol; MeI, iodomethane; MeOH, methanol; min, minute; MP-Carbonate, macroporous triethylammonium methylpolystyrene carbonate; NBS, N-bromosuccinimide; N,N,N′,N′-methylmorpholine; NMR, nuclear magnetic resonance; pet. ether, petroleum ether; PTSA, p-Toluenesulfonic acid; Rt, retention time; Sat., saturated; Si—NH.sub.2, aminopropyl bonded silica; Si-Thiol, silica 1-propanethiol; TBTU, O-(benzotriazol-1-yl)N,N,N′,N′-tetramethyluronium tetrafluoroborate; TFA, trifluoroacetic acid; THF, tetrahydrofuran; TLC, thin layer chromatography; TMOF, trimethylorthoformate; TMS, trimethylsilane; % v/v, percentage volume to volume; % w/v, percentage weight to volume.

(187) Analytical Methods

(188) Reverse Phase Preparative LC-MS: Mass-directed purification preparative LC-MS using a preparative C-18 column (Phenomenex Luna C18 (2), 100×21.2 mm, 5 μm).

(189) Analysis of products and intermediates has been carried out using reverse-phase analytical HPLC-MS using the parameters set out below.

(190) HPLC Analytical Methods:

(191) AnalpH2_MeOH_4 min: Phenomenex Luna C18 (2) 3 μm, 50×4.6 mm; A=water+0.1% formic acid; B=MeOH; 45° C.; % B: 0 min 5%, 1 min 37.5%, 3 min 95%, 3.51 min 5%, 4.5 min 5%; 2.25 mL/min.

(192) AnalpH9_MeOH_4 min: Phenomenex Luna C18 (2) 3 μm, 50×4 6 mm; A=water pH9 (Ammonium Bicarbonate 10 mmol); B=MeOH; 45° C.; % B: 0 min 5%, 1 min 37.5%, 3 min 95%, 3.51 min 5%, 4.5 min 5%; 2.25 mL/min.

(193) AnalpH2_MeOH_QC: Phenomenex Luna C18 (2) 3 μm, 50×4.6 mm; A=water+0.1% formic acid; B=MeOH; 35° C.; % B: 0 min 5%, 7.5 min 95%, 10 min 95%, 10.10 min 5%,13.0 min 5%; 1.5 mL/min.

(194) AnalpH2_MeCN_QC: Phenomenex Luna C18 (2) 3 μm, 50×4.6 mm; A=water+0.1% formic acid; B=MeCN+0.1% TFA; 40° C.; % B: 0 min 5%, 0.1 min 5%, 8 min 95%, 10.5 min 95%, 10.55 min 5%, 13.5 min 5%; 1.5 mL/min.

(195) AnalpH9_MeOH_QC: Phenomenex Luna C18 (2) 3 μm, 50×4.6 mm; A=water+0.1% formic acid; B=MeOH; 45° C.; % B: 0 min 5%, 7.5 min 95%, 10 min 95%, 10.10 min 5%, 13.0 min 5%; 1.5 mL/min.

(196) Method_2_TFA_UPLC_2: Acquity UPLC BEH C18 1.7 μm, 100×2.1 mm; 25° C.; A=water+0.025% TFA; B=acetonitrile+0.025% TFA; % B: 0 min 30%, 4 min 80%, 6 min 80%, 6.1 min 30%; 0.4 mL/min.

(197) Method_4_TFA_UPLC_2: Acquity UPLC BEH C18 1.7 μm, 100×2.1 mm; 25° C.; A=water+0.025% TFA; B=acetonitrile+0.025% TFA; % B: 0 min 10%, 4 min 80%, 6 min 80%, 6.1 min 10%; 0.3 mL/min.

(198) AK4: Acquity UPLC BEH C18 1.7 μm, 50×2.1 mm; A=water+0.025% TFA; B=acetonitrile+0.025% TFA; % B: 0 min 15%, 3 min 95%, 4 min 95%, 4.1 min 15%; 0.4 mL/min.

(199) AK4P: Acquity UPLC BEH C18 1.7 μm, 50×2.1 mm; A=water+0.025% TFA; B=acetonitrile+0.025% TFA; % B: 0 min 10%, 3 min 80%, 4 min 80%, 4.1 min 10%; 0.3 mL/min.

(200) XTERRA: Xterra MS C18 3 μm, 50×4.6 mm; A=water+0.1% HCOOH; B=acetonitrile; % B: 0 min 10%, 4 min 90%, 8 min 90%, 8.01 min 10%; 1 mL/min.

(201) LCMS-2: Acquity UPLC BEH C18 1.7 μm, 50×2.1 mm; A=water+0.025% TFA; B=acetonitrile+0.025% TFA; % B: 0 min 20%, 2 min 90%, 3 min 90%, 3.1 min 20%; 0.4 mL/min.

(202) 1.1 Synthesis of Intermediates

(203) General experimental schemes for the synthesis of HSF inhibitors of the disclosure are provided in FIG. 8.

(204) 1.1.1 Synthesis of Amino Heterocycles (AH)

(205) 1.1.1.1 Amino Heterocycle Intermediates AH1, AH2:

(206) ##STR00079##
Step 1—Suzuki Coupling

(207) To a microwave vial was added N-Boc-2-amino-5-bromo-thiazole (0.09 mmol, 1 equiv.) followed by the boronic acid (0.27 mmol, 3 equiv.), Pd(PPh.sub.3).sub.2Cl.sub.2 (0.005 mmol, 0.05 equiv.), aq. Cs.sub.2CO.sub.3 (4 M, 0.36 mmol, 4 equiv.) and DME/EtOH/water 7:2:3 (1 mL). The reaction mixture was de-gassed with nitrogen for 2 min and irradiated in a microwave at 130° C. for 8 min. LC-MS analysis showed that the Boc group had been removed during the reaction. The reaction mixture was passed through 1 g Si-Thiol cartridges, eluting with EtOH. The filtrate was evaporated to dryness, purified by preparative LC-MS and isolated directly as the amine for use in Routes A, D (FIG. 9).

(208) TABLE-US-00004 TABLE 4 Amino Heterocycle Intermediates (AH) Prepared by Step 1 Compound ID Analytical Data Yield 0 5-o-Tolyl-thiazol-2-ylamine formic acid salt AH1 AnalpH2_MeOH_4MIN; Rt 1.65 min; m/z 191 (MH.sup.+); pale yellow solid 22 mg, 34% 5-Pyridin-3-yl-thiazol-2-ylamine AH2 AnalpH2_MeOH_4MIN; Rt 0.62 min; m/z 178 (MH.sup.+); pale brown solid 13 mg, 21%
1.1.1.2 Synthesis of Amino Heterocycle Intermediate AH3:

(209) Step 1: tert-Butyl thiazol-2-ylcarbamate: to a solution of 2-amino thiazole (30 g, 300 mmol) in THF (300 mL) was added Boc.sub.2O (79 mL, 360 mmol) and the reaction mixture was stirred at ambient temperature for 16 h. The reaction mixture was concentrated under reduced pressure, and the precipitated solid was washed with pet.-ether to afford tert-butyl thiazol-2-ylcarbamate (50 g, 83%). MS (APCI); m/z 201 (MH.sup.+).

(210) Step 2: tert-Butyl methyl(thiazol-2-yl)carbamate: to a suspension of NaH (60% in mineral oil suspension, 13.3 g, 330 mmol) in DMF, cooled to 5° C., was added tert-butyl thiazol-2-yl carbamate (55 g, 275 mmol) followed by dropwise addition of iodomethane (58.16 g, 412 mmol). The reaction mixture was allowed to warm to ambient temperature over 2 h. The reaction mixture was poured onto crushed ice, stirred, and the precipitated solid was filtered and dissolved in pet. ether, dried over anhydrous Na.sub.2SO.sub.4 and concentrated under reduced pressure to afford tert-butyl methyl(thiazol-2-yl)carbamate as an off-white solid (48 g, 82%). MS (APCI); m/z 215 (MH.sup.+).

(211) ##STR00083##

(212) Step 3: tert-Butyl 5-bromothiazol-2-yl(methyl)carbamate: to a solution of tert-butyl methyl(thiazol-2-yl)carbamate (52 g, 243 mmol) in THF (520 mL), cooled to 5° C. was added NBS (47 g, 267 mmol) in ten portions. The reaction mixture was allowed to stir at ambient temperature for 2 h. The reaction mixture was concentrated, and triturated with pet.-ether. The precipitated solids were filtered and dried to afford tert-butyl 5-bromothiazol-2-yl(methyl)carbamate as a pale yellow solid (62 g, 87%). .sup.1H NMR (CDCl.sub.3) −δ1.58 (s, 9H), δ3.49 (s, 3H), δ7.33 (s, 1H).

(213) Step 4: tert-Butyl methyl (5-(pyridin-3-yl)thiazol-2-yl)carbamate: to a solution of tertbutyl 5-bromothiazol-2-yl(methyl)carbamate (25 g, 85.32 mmol) and 3-pyridine boronic acid (15.7 g, 127.98 mmol) in DME/water (320 mL/80 mL) under an argon atmosphere was added K.sub.2CO.sub.3 (37.13 g, 273.03 mm) followed by Pd(PPh.sub.3).sub.4 (9.8 g, 8.53 mmol), and the reaction mixture was heated to 100° C. for 16 h. The reaction mixture was cooled to ambient temperature, filtered through a celite pad, concentrated under vacuum and partitioned between water and EtOAc. The organic layer was dried over anhydrous Na.sub.2SO.sub.4 and concentrated. The crude compound was purified by column chromatography (silica gel 100-200 mesh, 30-40% EtOAc/pet. ether) to afford tert-butyl methyl (5-(pyridin-3-yl)thiazol-2-yl)carbamate as an off-white solid (15 g, 52%). AK4; Rt 1.68 min (98%); m/z 292 (MH.sup.+).

(214) Step 5: N-Methyl-5-(pyridin-3-yl)thiazol-2-amine (Amino Heterocycle Intermediate AH3): a solution of tert-butyl methyl (5-(pyridin-3-yl)thiazol-2-yl)carbamate (21 g, 72.4 mmol) in TFA/DCM (1:1, 157 mL) was stirred at ambient temperature for 2 h. The reaction mixture was concentrated and basified with sat. aq. NaHCO.sub.3 solution and extracted with EtOAc. The organic layer was dried over Na.sub.2SO.sub.4 and concentrated in vacuo to afford N-methyl-5-(pyridin-3-yl)thiazol-2-amine as an off-white solid (10.7 g, 78%). AK4P; Rt 0.86 min (100%); m/z 192 (MH.sup.+); .sup.1H NMR (d6-DMSO) −δ2.86-2.87 (d, 3H), δ7.34-7.37 (d of d, 1H), −δ7.64 (s, 1H), δ7.77-7.86 (m, 2H), δ8.36-8.37 (d, 1H), δ8.68 (s, 1H)

(215) 1.1.1.3 Synthesis of Amino Heterocycle Intermediates (AH4-AH6)

(216) ##STR00084##

(217) Step 1: To a solution of 2-bromothiazole (0.6 mmol, 1 equiv.) in 1,4-dioxane (0.5 mL) was added the amine (10 equiv., 6 mmol) and the reaction subjected to microwave irradiation at 180° C. for 30 min after which time the solvent was removed and the crude residue purified by preparative LC-MS to afford the desired product.

(218) TABLE-US-00005 TABLE 5 Amino Heterocycle Intermediates AH4-6 Compound ID Analytical Data Yield (2-Pyridin-4-yl-ethyl)thiazol-2-yl-amine AH4 AnalpH9_MeOH_4min; Rt 2.09 min (90%); m/z 206 (MH.sup.+); Green glass 65 mg (52%) (2-morpholin-4-yl-ethyl)thiazol-2-yl-amine AH5 AnalpH9_MeOH_4min; Rt 2.14 min (>95%); m/z 214 (MH.sup.+); Gold glass 42 mg (32%) (2-Pyridin-2-yl-ethyl)thiazol-2-yl-amine AH6 AnalpH9_MeOH_4min; Rt 2.08 min (90%); m/z 206 (MH.sup.+); Orange glass 38 mg (30%)
1.1.1.4 Synthesis of Amino Heterocycle Intermediate AH7

(219) ##STR00088##

(220) Step 1: [2-(1-Methyl-1H-imidazol-4-yl)-ethyl]thiazol-2-yl-amine—AH7. To a solution of 2-bromothiazole (400 mg, 1 equiv., 2.43 mmol) in IPA (5 mL) was added 1-methylhistamine (610 mg, 2 equiv., 4.87 mmol) and the reaction was heated at 90° C. for 96 h after which time the solvent was removed under vacuum and the crude residue purified by preparative LC-MS to afford the title compound as a golden colored glass (120 mg, 23%). AnalpH2_MeOH_4 min; Rt 0.27 min (>95%); m/z 209 (MH.sup.+).

(221) 1.1.1.5 Synthesis of Amino Heterocycle Intermediates AH8, AH10

(222) ##STR00089##

(223) Step 1: 2-tert-Butoxycarbonylamino-5-phenyl-thiazole-4-carboxylic acid methyl ester. To a solution of 5-bromo-2-tert-butoxycarbonylamino-thiazole-4-carboxylic acid methyl ester (250 mg, 1 equiv., 0.74 mmol) in DME (2.5 mL) was added phenyl boronic acid (135 mg, 1.5 equiv., 1.11 mmol) and Pd(PPh.sub.3).sub.4 (42 mg, 0.05 equiv., 0.037 mmol) and the reaction mixture degassed for 2 min under nitrogen. Potassium phosphate tribasic (aq. 2 M, 1.48 mL, 4 equiv., 2.96 mmol) was added and the reaction mixture degassed for 2 min under nitrogen after which time the reaction was subjected to microwave irradiation at 90° C. for 10 min. The crude reaction mixture was diluted with DCM (5 mL) and water (5 mL) and the organics obtained by means of filtration through a phase separation cartridge after which time the solvent was removed under vacuum and the crude residue purified by silica column chromatography with 0-30% EtOAc/isohexane as the eluent to afford the title compound as a yellow solid (236 mg, 47%). AnalpH2_MeOH_4 min; Rt 3.15 min (>95%); m/z 335 (MH.sup.+).

(224) Step 2: 2-Amino-5-phenyl-thiazole-4-carboxylic acid methyl ester (AH8). To a solution of 2-tert-butoxycarbonylamino-5-phenyl-thiazole-4-carboxylic acid methyl ester (50 mg, 1 equiv., 0.15 mmol) in DCM (0.5 mL) at 0° C. was added 4M HCl/dioxane (0.5 mL), and the reaction was stirred at ambient temperature for 1 h after which time the solvent was removed under vacuum and the crude residue azeotroped with toluene (2×10 mL) to afford the title compound as a gold colored glass (38 mg, 94%). AnalpH2_MeOH_4 min; Rt 2.22 min (>95%); m/z 235 (MH.sup.+).

(225) Step 2a: (4-Carbamoyl-5-phenyl-thiazol-2-yl)-carbamic acid tert-butyl ester. A solution of 2-tert-butoxycarbonylamino-5-phenyl-thiazole-4-carboxylic acid methyl ester (480 mg, 1 equiv., 1.43 mmol) in ammonia (7 M in MeOH) (30 mL) was heated at 110° C. for 16 h in a sealed tube after which time the solvent was removed and the crude residue purified by silica column chromatography with 0-50% EtOAc/isohexane as the eluent to afford the title compound as a white solid (260 mg, 56%). AnalpH2_MeOH_4 min; Rt 2.95 min (>95%); m/z 320 (MH.sup.+).

(226) Step 3a: 2-Amino-5-phenyl-thiazole-4-carboxylic acid amide (AH10). The procedure followed was the same as step 2 (above). This afforded the title compound as a white solid (60 mg, 95%). AnalpH2_MeOH_4 min; Rt 1.75 min (>95%); m/z 220 (MH.sup.+).

(227) 1.1.1.6 Synthesis of Amino Heterocycle Intermediate AH9

(228) ##STR00090##

(229) Step 1: 4-Ethyl-5-phenyl-thiazol-2ylamine (AH9). To a solution of 4-phenyl-2-butanone (500 mg, 1 equiv., 3.37 mmol) in DCE (20 mL) was added NBS (719 mg, 1.2 equiv., 4.04 mmol) and benzoyl peroxide (11 mg, 0.013 equiv., 0.045 mmol), and the reaction was heated at 80° C. for 2 h after which time the reaction was cooled to ambient temperature, diluted with DCM, washed with aq. sodium bisulfate (10% w/v, 20 mL) and sat. aq. sodium bicarbonate (20 mL), dried over MgSO.sub.4, filtered and the solvent removed under vacuum. The resulting residue was dissolved in EtOH (20 mL) followed by the addition of thiourea (739 mg, 2.8 equiv., 9.71 mmol) and the reaction heated at 80° C. for 2 h, after which time sat. aq. sodium bicarbonate (10 mL) was added and the precipitate collected by filtration and dried under vacuum to afford the title compound as a cream solid (688 mg, 100%). AnalpH2_MeOH_4 min; Rt 1.63 min (>95%); m/z 205 (MH.sup.+).

(230) 1.1.2 Synthesis of Acid Intermediates (A)

(231) ##STR00091##
Typical Procedure:

(232) Step 1: Amide Coupling. As route A except the compound was used crude in the next step.

(233) Step 2: Ester Hydrolysis: to a solution of the ester in THF (˜0.1 M) was added 1 M aq. LiOH (5 equiv.). The reaction mixture was stirred at ambient temperature, overnight. The reaction mixture was evaporated to dryness, re-dissolved in water and acidified to pH 2 with 0.1 M or 0.5 M HCl. Product was extracted into EtOAc. The organic layer was washed with brine, dried with MgSO.sub.4 and evaporated to dryness to give the acid intermediate (A1-A2) which was used directly in Route E.

(234) TABLE-US-00006 TABLE 6 Acid Intermediates (A) Compound ID Analytical Data Yield (E)-3-(Thiazol-2-ylcarbamoyl)-acrylic acid A1 AnalpH2_MeOH_4MIN; Rt 1.86 min; m/z 199 (MH.sup.+); yellow solid  68 mg, 82% (E)-3-(4-Methyl-thiazol-2-ylcarbamoyl)- acrylic acid A2 AnalpH2_MeOH_4MIN; Rt 2.18 min; m/z 213 (MH.sup.+); pale yellow solid 127 mg, 82%
1.1.3 Synthesis of Ester Intermediates (E)

(235) Step 1: Amide Formation. To a solution of ethyl hydrogen fumarate (1 equiv.) in THF (1.16 M), cooled to −40° C. was added isobutyl chloroformate (1.5 equiv.) dropwise followed by NMM (2 equiv.). The reaction mixture was stirred for 30 min. Amine (2 equiv.) was added to the above reaction mixture and stirred for 1 h at −40° C. After completion of the reaction (monitored by TLC), the reaction mixture was diluted with water and extracted with EtOAc. The organic layer was dried over anhydrous Na.sub.2SO.sub.4 and concentrated under reduced pressure to obtain crude compound. The crude compound was purified by silica gel column chromatography (1:1 pet. ether/EtOAc) to afford the corresponding amide.

(236) ##STR00094##

(237) Step 2: Cyclisation to the Ester Intermediate (E). To a stirred solution of triphenyl phosphine (1.5 equiv.) in DCM (0.15 M) at ambient temperature was added DDQ (1.5 equiv.). The reaction mixture was stirred for 10 min. A pre-mixed solution of the amide intermediate (1 equiv.) in DCM (2 M) was added slowly at ambient temperature over a period of 15 min and stirred for 20 min. After completion of the reaction (monitored by TLC), the reaction mixture was diluted with 5% w/v aq. NaOH and extracted with DCM. The organic layer was dried with anhydrous Na.sub.2SO.sub.4 and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (1:1 pet. ether/EtOAc) to afford the ester intermediates (E).

(238) TABLE-US-00007 TABLE 7 Ester Intermediates (E) Compound ID Analytical Data Yield (E)-3-(5,6-Dihydro-4H- [1,3] oxazin-2-yl)-acrylic acid ethyl ester E2 MS (APCI); m/z 184 (MH.sup.+); .sup.1H NMR (d.sup.6 DMSO) - δ 1.21-1.24 (t, 3H), δ 1.79-1.84 (m, 2H), δ 3.44-3.47 (t, 2H), δ 4.14-4.19 (q, 2H), δ 4.22-4.24 (t, 2H), δ 6.33-6.37 (d, 1H, J = 16.4 Hz), δ 6.64-6.68 (d, 1H, J = 16.4 Hz). 1-5 g, 34% (E)-3-(4,5-Dihydro-oxazol-2-yl]- acrylic acid ethyl ester E2 MS (APCI); m/z 170 (MH.sup.+); .sup.1H NMR (CDCl.sub.3) - δ 1.30-1.34 (t, 3H), δ 3.99-4.05 (t, 2H), δ 4.23-4.28 (q, 2H), δ 4.33-4.38 (t, 2H), δ 6.56-6.60 (d, 1H, J = 16.4 Hz), δ 7.09- 7.13 (d, 1H, J = 16.4 Hz). 750 mg, 34%
1.2 General Procedures: Route A F
1.2.1 Route A—General Procedure for Amide Synthesis

(239) To the acid (1-1.2 equiv.) and TBTU (1-1.2 equiv.) in anhydrous DCM/DMF (2:1, 0.3M) was added DIPEA/anhydrous DCM, (1-1.2 equiv., 0.3 M). The reaction mixture was stirred at ambient temperature for 50 min. After this time, the amine/anhydrous DCM, (1 equiv., 0.14 M) was added and the reaction mixture stirred at ambient temperature overnight. To the reaction vessel was added tetrafluorophthalic anhydride (0.8-1 equiv.) in anhydrous DMF (0.48 M) and allowed to stir for 2-3 h to scavenge unreacted acid and TBTU. After this time, 2-3 equiv. MP-Carbonate was added and the reaction mixture stirred 6-12 h. The reaction mixture was applied to a preconditioned Si—NH.sub.2 cartridge (1 or 2 g) and eluted with DMF (1×column volume) and MeOH (1×column volume). Evaporation of the eluents yielded the desired amide. In some instances, preparative LC-MS was required for purification.

(240) 1.2.2 Route B—General Procedure for Amide Synthesis

(241) To a solution of the acid (1.2-1.5 equiv.) in DCM (0.28 M) was added TBTU (1.2-1.5 equiv.) and DIPEA (1.2-1.5 equiv.) The reaction was stirred at ambient temperature for 10 min followed by the addition of the amine (1 equiv.), and the reaction was stirred for 16 h at ambient temperature after which time the reaction was diluted with water (2 mL) and DCM (2 mL) and the organic phase collected by means of a phase separation cartridge. The solvent was removed under vacuum and the crude residue purified by preparative LC-MS to afford the desired compound.

(242) 1.2.3 Route C—General Procedure for Amide Synthesis

(243) To a solution of the amine (1 equiv.) in toluene or DCE (0.4 M) was added trimethylaluminium (2 M in hexanes, 1 equiv.), and the reaction was stirred at ambient temperature for 30 min. This was followed by the addition of a solution of the ester (1 equiv.) in toluene or DCE (0.4 M), and the reaction was heated at 90° C. for 4 h after which time the reaction mixture was cooled to ambient temperature, diluted with water (20 mL) and extracted with EtOAc (3×20 mL). The combined organics were washed with brine (30 mL), dried over MgSO.sub.4, filtered and the solvent removed under vacuum. The crude residue was triturated with DCM/hexane and dried. In some instances, the crude compound required purification by preparative LC-MS to afford the desired compound.

(244) 1.2.4 Route D—General Procedure for Amide Synthesis Via Acid Chloride

(245) Step 1: To a solution of the acid (1 equiv.) in anhydrous DCM (0.46 M) and a catalytic amount of anhydrous DMF under a nitrogen atmosphere was added a 2 M solution of oxalyl chloride (1.1 equiv.) in anhydrous DCM. The reaction mixture was stirred at ambient temperature for 2 h. After this time, the reaction mixture was evaporated to dryness and re-dissolved in anhydrous DCM for direct use in step 2 below.

(246) Step 2: To the amine (0.5-0.67 equiv.) was added a solution of pyridine in anhydrous DCM (3.7-5.15 M, 2.8 equiv.) followed by addition of a solution of the acid chloride in anhydrous DCM (0.3-0.37 M, 1 equiv.), synthesized in step 1, and the reaction mixture was purged with nitrogen and stirred at ambient temperature overnight.* To the reaction mixture was added DCM (3 mL) followed by water (4 mL). The mixture was agitated and passed through a phase-separation cartridge. DCM layer was further washed with water (4 mL). Organic layers were combined, evaporated to dryness, and compound was purified by preparative LC-MS. * In some instances no work-up was performed, the reaction mixture was evaporated to dryness and purified by preparative LC-MS.

(247) 1.2.5 Route E—Amide Coupling

(248) As Route A except amine was used in excess (1.5-2 equiv.). and 1 equiv. was used of all other reagents. In instances where the amine was a salt, amine was free-based with the addition of DIPEA prior to addition to the reaction mixture. In some instances, compounds required further purification via preparative LC-MS.

(249) 1.2.6 Route F—Ester Synthesis

(250) To a microwave vial was added a solution of the acid in the alcohol (0.26 M, 1 equiv.) and conc. sulfuric acid (0.94 equiv.). The vial was sealed and heated in a microwave at 100° C. for 2 min. Reaction mixture was evaporated to dryness and purified by preparative LC-MS.

(251) 1.3 Final Compounds—Routes A—F

(252) 1.3.1 Final Compounds Synthesized by Route A

(253) TABLE-US-00008 TABLE 8 Final Compounds Synthesized by Route A Compound I.sub.HSF Int.* Analytical Data Yield (E)-3-(Benzothiazol-2-ylcarbamoyl)- acrylic acid ethyl ester 010 AnalpH2_MeOH_QC; Rt 8.30 min (98%); m/z 277 (MH.sup.+); .sup.1H NMR (d.sup.6-DMSO) - δ 1.25-1.29 (t, 3H), δ 4.21-4.26 (q, 2H), δ 6.86-6.89 (d, 1H, J = 15 Hz), δ 7.28-7.32 (d, 1H, J = 15 Hz), δ 7.32-7.36 (t of d, 1H), δ 7.44-7.49 (t of d, 1H), δ 7.78- 7.80 (d of t, 1H), δ 8.00-8.03 (d of t, 1H) δ 12.92 (s, br, 1H); pale yellow solid 18 mg, 88% (E)-3-(5-Phenyl-thiazol-2-ylcarbamoyl)- acrylic acid ethyl ester 014 AnalpH2_MeOH_QC; Rt 8.60 min (89%); m/z 303 (MH.sup.+); yellow solid 35.5 mg, 99% (E)-3-(Thiazol-2-ylcarbamoyl)-acrylic acid methyl ester 027 AnalpH2_MeOH_QC; Rt 6.45 min (99%); m/z 241 (MH.sup.+); white solid 9 mg, 7% 00 (E)-3-(Thiazol-2-ylcarbamoyl)-acrylic acid isopropyl ester 028 AnalpH2_MeOH_QC; Rt 7.54 min (97%); m/z 213 (MH.sup.+); orange solid 24 mg 56% 01 (E)-3-(4-Phenyl-thiophen-2-ylcarbamoyl)-acrylic acid ethyl ester 043 AnalpH2_MeCN_QC; Rt 7.38 min (99%); m/z 302 (MH.sup.+); dark yellow solid 4.1 mg 8% 02 (E)-3-(2-Phenyl-thiazol-5-ylcarbamoyl)-acrylic acid ethyl ester 045 AnalpH2_MeOH_QC; Rt 8.34 min (99%); m/z 303 (MH.sup.+); .sup.1H NMR (d.sup.6-DMSO) - δ 1.26-1.29 (t, 3H), δ 4.2-4.26 (q, 2H), δ 6.76-6.80 (d, 1H, J = 15 Hz), δ 7.14-7.18 (d, 1H, J = 15 Hz), δ 7.42-7.50 (m, 3H), δ 7.71 (s, 1H), δ 7.89-7.91 (d of t, 2H), δ 12.21 (s, br, 1H); yellow solid 11.6 mg 21% 03 (E)-3-(4-Methyl-5-phenyl-thiazol-2- ylcarbamoyl)-acrylic acid ethyl ester 051 AnalpH2_MeOH_QC; Rt 8.75 min (87%); m/z 317 (MH.sup.+); yellow solid 45 mg 79% 04 (E)-3-(4,5-Dimethyl-thiazol-2-ylcarbamoyl)- acrylic acid ethyl ester 052 AnalpH2_MeOH_QC; Rt 7.90 min (93%); m/z 255 (MH.sup.+); yellow solid 36 mg 78% 05 (E)-3-(4-Cyclopropyl-thiazol-2-ylcarbamoyl)- acrylic acid ethyl ester 053 AnalpH2_MeOH_QC; Rt 8.05 min (89%); m/z 267 (MH.sup.+); yellow/brown solid 38 mg 79% 06 (E)-3-(4-Pyridin-2-yl-thiazol-2-yl carbamoyl)-acrylic acid ethyl ester 054 AnalpH2_MeOH_QC; Rt 7.18 min (96%); m/z 304 (MH.sup.+); .sup.1H NMR (d.sup.6-DMSO) - δ 1.26-1.29 (t, 3H), δ 4.21-4.26 (q, 2H), δ 6.84-6.88 (d, 1H, J = 15.4 Hz), δ 7.28-7.31 (d, 1H, J = 15.4 Hz), δ 7.34-7.38 (dqd, 1H), δ 7.89-7.97 (m, 3H), δ 8.61-8.63 (d of q, 1H), δ 12.95 (s, br, 1H); brown/yellow solid 40 mg 74% 07 (E)-3-(4-Pyridin-3-yl-thiazol-2-yl carbamoyl)-acrylic acid ethyl ester 055 AnalpH2_MeOH_QC; Rt 7.02 min (86%); m/z 304 (MH.sup.+); yellow solid 15 mg 27% 08 (E)-3-(5-o-Tolyl-thiazol-2-ylcarbamoyl)- acrylic acid ethyl ester 059 AH1 AnalpH2_MeCN_QC; Rt 7.54 min (89%); m/z 317 (MH.sup.+); .sup.1H NMR (d.sup.6-DMSO) δ 1.32-1.35 (t, 3H), δ 2.47 (s, 3H), δ 4.27-4.32 (q, 2H), δ 6.88-6.92 (d, 1H, J = 15.6 Hz), δ 7.33-7.37 (d, 1H, J = 15.6 Hz), δ 7.34-7.38 (m, 2H), δ 7.40-7.42 (d of m, 1H), δ 7.46-7.48 (d of d, 1H), δ 7.71 (s, 1H), δ 12.88 (s, br, 1H); yellow solid 19 mg 66% 09 (E)-3-(Methyl-thiazol-2-yl- carbamoyl)-acrylic acid ethyl ester 070 AnalpH2_MeOH_QC; Rt 7.32 min (92%); m/z 241 (MH.sup.+); .sup.1H NMR (d.sup.6-DMSO) δ 1.26-1.3 (t, 3H), δ 3.78 (s, 3H), δ 4.22-4.27 (q, 2H), δ 6.78-6.82 (d, 1H, J = 15.4 Hz), δ 7.38-7.39 (d, 1H), δ 7.60-7.61 (d, 1H), δ 7.62- 7.66 (d, 1H, J = 15.4 Hz); yellow solid 36 mg 75% * Where the intermediate column is left blank, the starting material(s) was/were commercially available.
1.3.2 Final Compounds Synthesized by Route B

(254) TABLE-US-00009 TABLE 9 Final Compounds Synthesized by Route B Compound I.sub.HSF Int. Analytical Data Yield 0 (E)-3-[(2-Pyridin-4-yl-ethyl)- thiazol-2-yl-carbamoyl]- acrylic acid ethyl ester 099 AH4 AnalpH2_MeOH_QC; Rt 5.75 min (90%); m/z 332 (MH.sup.+); .sup.1H NMR (d.sup.6-DMSO) - δ 1.26 (t, 3H), δ 3.03-3.07 (t, 2H), δ 4.19-4.24 (q, 2H), δ 4.57-4.60 (t, 2H), δ 6.54- 6.58 (d, 1H, J = 15.2 Hz), δ 7.17-7.18 (d, 2H), δ 7.34- 7.42 (d, 1H, J = 15.2 Hz), δ 7.43-7.44 (d, 1H), δ 7.66- 7.67 (d, 1H), δ 8.43-8.44 (d of d, 2H); beige solid 5.2 mg 5% (E)-3-[(2-Morpholin-4-yl- ethyl)-thiazol-2-yl-carbamoyl]- acrylic acid ethyl ester 101 AH5 AnalpH2_MeOH_QC; Rt 4.6 min (92%); m/z 340 (MH.sup.+); .sup.1H NMR (d.sup.6-DMSO) - δ 1.06-1.09 (t, 3H), δ 2.19- 2.22 (t, 4H), δ 2.38-2.41 (t, 2H), δ 3.28-3.30 (t, 4H), δ 4.02-4.07 (q, 2H), δ 4.21- 4.23 (t, 2H), δ 6.61-6.65 (d, 1H, J = 15 Hz), δ 7.2-7.21 (d, 1H), δ 7.41-7.42 (d, 1H), δ 7.51-7.55 (d, 1H, J = 15 Hz); cream solid 30.6 mg 45% (E)-3-[(2-Pyridin-2-yl-ethyl)- thiazol-2-yl-carbamoyl]-acrylic acid ethyl ester 112 AH6 AnalpH2_MeOH_QC; Rt 6.16 min (89%); m/z 332 (MH.sup.+); orange solid 2.4 mg 4% 2-((E)-3 -Ethoxycarbonyl- acryloylamino)-5-phenyl- thiazole-4-carboxylic acid methyl ester 105 AH8 AnalpH2_MeOH_QC; Rt 8.43 min (>95%); m/z 361 (MH.sup.+); pale gold solid 3.5 mg 6% (E)-3-(4-Ethyl-5-phenyl-thiazol-2- ylcarbamoyl)-acrylic acid ethyl ester 107 AH9 AnalpH2_MeOH_QC; Rt 8.79 min (>95%); m/z 331 (MH.sup.+); pale lemon solid 11.2 mg 3.5% (E)-3-(4-Carbamoyl-5-phenyl- thiazol-2-ylcarbamoyl)-acrylic acid ethyl ester 109 AH10 AnalpH2_MeOH_QC; Rt 7.9 min (>95%); m/z 346 (MH.sup.+); .sup.1H NMR (d.sup.6-DMSO) - δ 1.39-1.43 (t, 3H), δ 4.35- 4.40 (q, 2H), δ 6.97-7.01 (d, 1H, J = 15.5 Hz), δ 7.40- 7.44 (d, 1H, J = 15.5 Hz), δ 7.52-7.59 (m, 3H), δ 7.65 (s, br, 1H), δ 7.69-7.71 (d of t, 2H), δ 13.07 (s, br, 2H); yellow solid 6.3 mg 10%
1.3.3 Final Compounds Synthesised by Route C

(255) TABLE-US-00010 TABLE 10 Compounds Synthesised by Route C Compound I.sub.HSF Int. Analytical Data Yield (E)-3-(5,6-Dihydro-4H-[1,3] oxazin-2-yl)-N-methyl-N-(5- pyridin-3-yl-thiazol-2-yl)- acrylamide 115 AH3, E1 AnalpH9_MeOH _QC; Rt 7.19 min (99.2); m/z 329 (MH.sup.+);.sup.1 H NMR (d.sup.6-DMSO) - δ 1.83-1.89 (m, 2H), δ 3.48- 3.51 (t, 2H), δ 3.79 (s, 3H), δ 4.28-4.30 (t, 2H), δ 6.77- 6.80 (d, 1H, J = 15.4 Hz), δ 7.23-7.27 (d, 1H, J = 15.4 Hz), δ 7.44-7.48 (q of d, 1H), δ 8.04-8.07 (d of t, 1H), δ 8.13 (s, 1H), δ 8.51-8.52 (d of d, 1H), δ 8.89-8.90 (d, 1H); cream solid 259 mg 21.5% (E)-3-(5,6-Dihydro-4H-[1,3]oxazin- 2-yl)-N-thiazol-2-yl-acrylamide 090 E1 Method_4_TFA_UPLC_2; Rt 1.94 min (96%); m/z 238 (MH.sup.+); .sup.1H NMR (CD.sub.3CO.sub.2D) - δ 2.29-2.31 (t, 2H), δ 3.79- 3.82 (t, 2H), δ 4.78-4.80 (t, 2H), 87.17-7.21 (d, 1H, J = 15.6 Hz), δ 7.22-7.23 (d, 1H), δ 7.36-7.40 (d, 1H, J = 15.6 Hz), δ 7.53-7.54 (d, 1H); mp - 205-208° C; pale brown solid 38 mg, 9% (E)-3-(4,5-Dihydro-oxazol-2-yl)- N-thiazol-2-yl-acrylamide 092 E2 Method_4_TFA_UPLC_2; Rt 2.30 min (88%); m/z 224 (MH.sup.+); .sup.1H NMR (CD.sub.3CO.sub.2D) - δ 4.06-4.11 (t, 2H), δ 4.53- 4.58 (t, 2H), δ 6.94-6.98 (d, 1H, J = 16 Hz), 87.19-7.20 (d, 1H), 87.32-7.36 (d, 1H, J = 16 Hz), 87.50-7.51 (d, 1H); mp - 221-226° C.; yellow solid 38 mg, 9%
1.3.4 Final Compounds Synthesised by Route D

(256) TABLE-US-00011 TABLE 11 Final Compounds Synthesised by Route D Compound ID Int. Analytical Data Yield (E)-3-(4-Methyl-thiazol-2- ylcarbamoyl)-acrylic acid ethyl ester 011 AnalpH2_MeOH_QC; Rt 7.51 min (95%); m/z 241 (MH.sup.+); .sup.1H NMR (d.sup.6-DMSO) - δ 1.25-1.29 (t, 3H), δ 2.29 (d, 3H) δ 4.20-4.25 (q, 2H), δ 6.79-6.83 (d, 1H, J = 15 Hz), 86.87 (d, 1H) δ 7.21- 7.25 (d, 1H, J = 15 Hz), δ 12.61 (s, br, 1H); yellow solid 31 mg, 64% 0 (E)-3-(4-Phenyl-thiazol-2- ylcarbamoyl)-acrylic acid ethyl ester 012 AnalpH2_MeOH_QC; Rt 8.48 min (100%); m/z 303 (MH.sup.+); tan solid 24 mg, 40% (E)-3-(5-Methyl-thiazol-2- ylcarbamoyl)-acrylic acid ethyl ester 013 AnalpH2_MeOH_QC; Rt 7.58 min (100%); m/z 241 (MH.sup.+); yellow solid 21 mg, 44% (E)-3-(Oxazol-2-ylcarbamoyl)- acrylic acid ethyl ester 015 AnalpH9_MeOH_QC; Rt 5.27 min (88%); m/z 211 (MH.sup.+); cream solid 21 mg 51% (E)-3-(1-Methyl-1H-pyrazol-3- ylcarbamoyl)-acrylic acid ethyl ester 016 AnalpH2_MeOH_QC; Rt 6.30 min (100%); m/z 224 (MH.sup.+); cream solid 32 mg 73% (E)-3-(Isoxazol-3-ylcarbamoyl)- acrylic acid ethyl ester 017 AnalpH2_MeOH_QC; Rt 6.48 min (100%); m/z 211 (MH.sup.+); cream solid .sup.1H NMR (d.sup.6-DMSO) - δ 1.25-1.28 (t, 3H), δ 4.2- 4.25 (q, 2H), δ 6.76-6.80 (d, 1H, J = 15.7 Hz), δ 7.0-7.01 (d, 1H) δ 7.20-7.24 (d, 1H, J = 15.7 Hz), δ 8.85-8.86 (d, 1H), δ 11.63 (s, br, 1H) 33 mg 79% (E)-3-([1,3,4]Thiadiazol-2-yl carbamoyl)-acrylic acid ethyl ester 018 AnalpH2_MeOH_QC; Rt 6.50 min (100%); m/z 228 (MH.sup.+); tan solid 13 mg 29% (E)-3-(Pyridin-4-ylcarbamoyl)- acrylic acid ethyl ester 020 AnalpH2_MeOH_QC; Rt 4.16 min (98%); m/z 221 (MH.sup.+); pale yellow solid  7 mg 15% (E)-3-(3-Methyl-isoxazol-5- ylcarbamoyl)-acrylic acid ethyl ester 046 AnalpH2_MeCN_QC; Rt 5.52 min (100%); m/z 225 (MH.sup.+); .sup.1H NMR (d.sup.6-DMSO) δ 1.24-1.28 (t, 3H), δ 2.20 (s, 3H), δ 4.19-4.24 (q, 2H), δ 6.27 (s, 1H), δ 6.76-6.80 (d, 1H, J = 15.4 Hz), δ 7.13-7.17 (d, 1H, J = 15.4 Hz), δ 12.15 (s, br, 1H); pale brown solid 28 mg 62% (E)-3-(2-Methyl-pyridin-4- ylcarbamoyl)-acrylic acid ethyl ester 048 AnalpH2_MeCN_QC; Rt 5.30 min (100%); m/z 235 (MH.sup.+); pale brown solid 14 mg 30% (E)-3-(2-Phenyl-pyridin-4- ylcarbamoyl)-acrylic acid ethyl ester 049 AnalpH2_MeCN_QC; Rt 6.82 min (97%); m/z 297 (MH.sup.+); pale brown solid 37 mg 63% 0 (E)-3-(Quinolin-4-ylcarbamoyl)- acrylic acid ethyl ester 050 AnalpH2_MeCN_QC; Rt 5.91 min (86%); m/z 271 (MH.sup.+); pale brown solid 25 mg 46% (E)-3-(4-Pyridin-4-yl-thiazol-2- lcarbamoyl)-acrylic acid ethyl ester 056 AnalpH2_MeCN_QC; Rt 6.02 min (87%); m/z 304 (MH.sup.+); pale orange solid 10 mg 16% (E)-3-(5-Cyano-thiazol-2-yl carbamoyl)-acrylic acid ethyl ester 057 AnalpH2_MeCN_QC; Rt 4.27 min (99%); m/z 252 (MH.sup.+); .sup.1H NMR (d.sup.6-DMSO) δ 1.33-1.37 (t, 3H), δ 4.29- 4.34 (q, 2H), δ 6.95-6.99 (d, 1H, J = 15.4 Hz), δ 7.30-7.34 (d, 1H, J = 15.4 Hz), δ 8.54 (s, 1H), δ 13.57 (s, br, 1H); pale brown solid 21 mg 42% (E)-3-(5-Pyridm-3-yl-thiazol-2- ylcarbamoyl)-acrylic acid ethyl ester 058 AH2 AnalpH2_MeCN_QC; Rt 4.28 min (99%); m/z 304 (MH.sup.+); .sup.1H NMR (DMSO- d6): δ 12.91 (brs, 1H), 8.89 (dd, J = 2.3, 0.8 Hz, 1H), 8.51 (dd, J = 4.8, 1.5 Hz, 1H), 8.11 (s, 1H), 8.07-8.04 (m, 1H), 7.48-7.44 (m, 1H), 7.28 (d, J = 15.5 Hz, 1H), 6.86 (d, J = 15.5 Hz, 1H), 4.23 (q, J = 7.2 Hz, 2H), 1.27 (t, J = 7.2 Hz, 3H); pale brown solid  9 mg 43% (E)-3-([2,4′]Bipyridinyl-4- ylcarbamoyl)-acrylic acid ethyl ester 076 AnalpH2_MeOH_QC; Rt 6.56 min (100%); m/z 298 (MH.sup.+); .sup.1H NMR (d.sup.6-DMSO) δ 1.26-1.30 (t, 3H), δ 4.21- 4.26 (q, 2H), δ 6.78-6.81 (d, 1H, J = 15.4 Hz), δ 7.20-7.24 (d, 1H, J = 15.4 Hz), δ 7.67- 7.69 (d of d, 1H), δ 7.93-7.95 (d of d, 2H), 8.31 (d, 1H), δ 8.65-8.67 (d, 1H), δ 8.72- 8.73 (d of d, 2H), δ 11.14 (s, br, 1H); beige solid 21 mg 36% (E)-3-(2-Morpholin-4-yl-pyridin-4- ylcarbamoyl)-acrylic acid ethyl ester formic acid salt 077 AnalpH2_MeOH_QC; Rt 5.08 min (100%); m/z 306 (MH.sup.+); .sup.1H NMR (d.sup.6-DMSO) δ 1.25-1.29 (1, 3H), δ 3.37-3.39 (m, 4H), δ 3.69- 3.71 (m, 4H) δ 4.20-4.25 (q, 2H), δ 6.70-6.74 (d, 1H, J = 15.4 Hz), δ 6.87-6.89 (d of d, 1H) δ 7.16-7.20 (d, 2H, J = 15.4 Hz), δ 8.05-8.06 (d, 1H), δ 8.16 (s, 1H), δ 10.71 (s, br, 1H), δ 12.83 (s, br, 1H); pale yellow solid 42 mg 60% (E)-3-[2-(4-Methyl-piperazin-1-yl)- pyridin-4-ylcarbamoyl]-acrylic acid ethyl ester formic acid salt 078 AnalpH2_MeOH_QC; Rt 3.88 min (95%); m/z 319 (MH.sup.+); yellow solid 29 mg 40% (E)-3-(2-o-Tolyl-pyridin-4- ylcarbamoyl)-acrylic acid ethyl ester 079 AnalpH2_MeOH_QC; Rt 6.78 min (97%); m/z 311 (MH.sup.+); tan solid 40 mg 65% 0 (E)-3-(2-m-Tolyl-pyridin-4- ylcarbamoyl)-acrylic acid ethyl ester 080 AnalpH2_MeOH_QC; Rt 7.43 min (100%); m/z 311 (MH.sup.+); pale brown solid 46 mg 74% (E)-3-(2-p-Tolyl-pyridin-4-ylcarbamoyl)- acrylic acid ethyl ester formic acid salt 081 AnalpH2_MeOH_QC; Rt 7.23 min (99%); m/z 311 (MH.sup.+); tan solid .sup.1H NMR (d.sup.6-DMSO) δ 1.26-1.30 (t, 3H), δ 2.37 (s, 3H), δ 4.21-4.26 (q, 2H), δ 6.76-6.80 (d, 1H, J = 15.6 Hz), δ 7.20-7.24 (d, 1H, J = 15.4 Hz), δ 7.31-7.33 (d, 2H), δ 7.53-7.55 (d of d, 1H), δ 7.88-7.9 (d, 2H), δ 8.15 (s, 1H), δ 8.16-8.17 (d, 1H), δ 8.55-8.56 (d, 1H), δ 10.99 (s, 1H), δ 12.95 (s, br, 1H) 39 mg 55% (E)-3-(2,6-Dimethyl-pyridin-4- ylcarbamoyl)-acrylic acid ethyl ester 082 AnalpH2_MeOH_QC; Rt 4.53 min (88%); m/z 249 (MH.sup.+); yellow solid  9 mg 18% (E)-3-(3-Methyl-pyridin-4- ylcarbamoyl)-acrylic acid ethyl ester 083 AnalpH2_MeOH_QC; Rt 4.17 min (88%); m/z 235 (MH.sup.+); off-white solid 15 mg 32% (E)-3-([2,3′]Bipyridmyl-4- ylcarbamoyl)-acrylic acid ethyl ester 086 AnalpH2_MeOH_QC; Rt 6.92 min (100%); m/z 298 (MH.sup.+); tan solid 28 mg 47% *Where the intermediate column is left blank, the starting material(s) was/were commercially available.

Synthesis of Final Compound 108 via Amino Heterocycle Intermediate AH7: (E)-3-{[2-1-Methyl-1H-imidazol-4-yl)-ethyl]-thiazol-2-yl-carbamoyl}-acrylic Acid Ethyl Ester

(257) ##STR00146##

(258) To a solution of [2-(1-Methyl-1H-imidazol-4-yl)-ethyl]-2-yl-amine (thiazole intermediate AH7), (120 mg, 1 equiv., 0.57 mmol) in DCM (1 mL) was added ethyl fumaroyl chloride (70 μL, 1.1 equiv., 0.63 mmol), dropwise, and triethylamine (204 μL, 3 equiv., 1.72 mmol), and the reaction was stirred at ambient temperature for 1 h after which time the solvent was removed and the crude residue purified by preparative LC-MS to afford the title compound, I.sub.HSF 108 as a golden colored glass (3.4 mg, 1.78%). AnalpH2_MeOH_QC; Rt 4.51 min (>95%); m/z 335 (MH.sup.+).

(259) 1.3.5 Final Compounds Synthesised by Route E

(260) TABLE-US-00012 TABLE 12 Final Compounds Synthesised by Route E Compound I.sub.HSF Int. Analytical Data Yield (E)-But-2-enedioic acid ethylamide thiazol-2-ylamide 030 A1 AnalpH2_MeOH_QC; Rt 5.90 min (85%); m/z 226 (MH.sup.+); beige solid 34 mg, 89% (E)-But-2-enedioic acid isopropylamide thiazol-2-ylamide 031 A1 AnalpH2_MeOH_QC; Rt 6.40 min (98%); m/z 240 (MH.sup.+); beige solid 36 mg, 90% (E)-But-2-enedioic acid diethyl- amide (4-methyl-thiazol-2-yl)-amide 060 A2 AnalpH2_MeOH_QC; Rt 7.20 min (95%); m/z 268 (MH.sup.+); pale brown solid 26 mg, 58% 0 (E)-But-2-enedioic acid dimethyl- amide (4-methyl-thiazol-2-yl)-amide 061 A2 AnalpH2_MeOH_QC; Rt 6.38 min (98%); m/z 240 (MH.sup.+); .sup.1H NMR (d.sup.6-DMSO) - δ 2.37 (s, 3H), δ 3.02 (s, 3H), δ 3.20 (s, 3H), δ 6.93 (s, 1H), δ 7.09-7.12 (d, 1H, J = 15.1 Hz), δ 7.57- 7.61 (d, 1H, J = 15.1 Hz), δ 12.61 (s, br, 1H); off-white solid  5 mg, 13%
1.3.6 Final Compounds Synthesised by Route F

(261) TABLE-US-00013 TABLE 13 Final Compounds Synthesised by Route F Compound I.sub.HSF Int.* Analytical Data Yield (E)-3-(Thiazol-2-ylcarbamoyl)- acrylic acid butyl ester 042 AnalpH2_MeCN_QC; Rt 5.98 min (100%); m/z 255 (MH.sup.+); .sup.1H NMR (CDCl.sub.3) - δ 0.85-0.89 (t, 3H), δ 1.28-1.37 (m, 2H), δ 1.57-1.64 (m, 2H), δ 4.18-4.22 (t, 2H) δ 6.27- 6.30 (d, 1H, J = 13.4 Hz), δ 6.44-6.47 (d, 1H, J = 13.4 Hz), δ 6.93-6.94 (d, 1H), δ 7.44-7.45 (d, 1H) δ 12.90 (s, br, 1H); white solid 15 mg, 46% *Where the intermediate column is left blank, the starting material(s) was/were commercially available.
1.4 Final Compounds Route G
1.4.1 Synthesis of Final Compounds Via Route G
Synthesis of Acid Intermediate (A1):

(262) Step 1: (E)-Ethyl-4-oxo-4-(thiazol-2-ylamino)-but-2-enoate. To a stirred solution of fumaric acid monoethyl ester (1 g, 6.94 mmol) in DCM (100 mL) at 0° C. was added EDC (1.6 g, 8.33 mmol), HOBt (1.14 g, 8.33 mmol) and 2-aminothiazole (694 mg, 6.94 mmol) sequentially and stirred at ambient temperature for 16 h. The reaction mixture was diluted with water, and extracted with DCM (2×200 mL). The combined organics were washed with water (2×100 mL), brine (2×100 mL), dried over Na.sub.2SO.sub.4 and concentrated in vacuo. The crude compound was purified by column chromatography (silica gel, 100-200 mesh, EtOAc/pet. ether) to afford (E)-ethyl 4-oxo-4-(thiazol-2-yl)amino-2-enoate as a pale yellow solid (500 mg, 32%). R.sub.f: 0.4 (20% EtOAc/pet. ether). .sup.1H NMR (d.sup.6 DMSO) −δ1.23-1.28 (t, 3H), δ4.19-4.25 (q, 2H), δ6.8-6.84 (d, 1H), δ7.25-7.29 (d, 1H), δ7.33-7.34 (d, 1H), δ7.55-7.56 (d, 1H), δ12.72 (s, br, 1H).

(263) Step 2: (E)-4-Oxo-4-(thiazol-2-ylamino)-but-2-enoic acid (Acid Intermediate A1). To a solution of (E)-ethyl 4-oxo-4-(thiazol-2-ylamino)-but-2-enoate (500 mg, 2.37 mmol) in THF/water (1:1) was added LiOH (199 mg, 4.74 mmol) and stirred at ambient temperature for 2 h. The reaction mixture was concentrated in vacuo, and the residue was acidified with sat. aq. KHSO.sub.4, diluted with water, and the precipitated solid was collected by filtration to afford (E)-4-oxo-4-(thiazol-2-ylamino)-but-2-enoic acid (acid intermediate A1) as a pale yellow solid (350 mg, 75%). R.sub.f: 0.1 (10% MeOH/CHCl.sub.3). .sup.1H NMR (d.sup.6 DMSO) −δ6.75-6.79 (d, 1H), δ7.17-7.21 (d, 1H), δ7.32-7.33 (d, 1H), δ7.55-7.56 (d, 1H), δ12.72 (s, br, 1H), δ13.16 (s, br, 1H).

(264) Synthesis of Final Compound I.sub.HSF 089 Via Acid Intermediate A1:

(265) Step 3: N.sup.1-(Thiazol-2-yl)fumaramide. To a stirred suspension of (E)-4-oxo-4-(thiazol-2-ylamino)but-2-enoic acid (A1) (200 mg, 1.01 mmol) in dry THF at −40° C. was added NMM (122 mg, 1.21 mmol) and isobutyl chloroformate (166 mg, 1.21 mmol), and the reaction was stirred for 20 min. Ammonium carbonate (484 mg, 5.05 mmol) was added, and the reaction mixture was allowed to warm to ambient temperature over 2 h. The solvent was evaporated in vacuo, and the residue was diluted with water. The precipitated solid was collected by filtration, and washed with Et.sub.2O to obtain N.sup.1-(thiazol-2-yl)fumaramide as a light brown solid (180 mg, 90%). R.sub.f: 0.2 (10% MeOH/CHCl.sub.3). .sup.1H NMR (d.sup.6 DMSO) −δ7.07 (s, br, 2H), 67.3-7.31 (d, 1H), δ7.50 (s, br, 1H), δ7.54-7.55 (d, 1H), δ7.95 (s, br, 1H), δ12.62 (s, br, 1H)

(266) ##STR00152##

(267) Step 4: (E)-N.sup.1-(Dimethylamino)methylene-N.sup.4-(thiazol-2-yl)fumaramide. To a stirred solution of N.sup.1-(thiazol-2-yl)fumaramide (300 mg, 1.52 mmol) in dry 1,4-dioxane (20 mL) was added DMF-DMA and heated at 80° C. for 2 h. The organic volatiles were evaporated in vacuo, and the crude solid was washed with Et.sub.2O (2×20 mL) to afford (E)-N.sup.1-(Dimethylamino)methylene-N.sup.4-(thiazol-2-yl)fumaramide as a brown solid (150 mg, 99%). R.sub.f: 0.3 (10% MeOH/CHCl.sub.3). .sup.1H NMR (d.sup.6 DMSO) −δ3.10 (s, 3H), δ 3.19 (s, 3H), δ6.92-6.96 (d, 1H), δ7.26-7.30 (d, 1H), δ7.30-7.31 (d, 1H), δ7.53-7.54 (d, 1H), δ8.54 (s, br, 1H), δ12.59 (s, br, 1H)

(268) Step 5: (E)-3-(1,2,4-Oxadiazol-5-yl)-N-(thiazol-2-yl)acrylamide (I.sub.HSF 089, identified as DX 089 in the above scheme). To a stirred solution of hydroxylamine hydrochloride (50 mg, 0.72 mmol) in 5 M aq. NaOH (0.15 mL, 0.72 mmol) and acetic acid (1.5 mL) in 1,4 dioxane was added N.sup.1-(Dimethylamino)methylene-N.sup.4-(thiazol-2-yl)fumaramide (150 mg, 0.60 mmol). The reaction mixture was heated to 90° C. for 16 h. The reaction mixture was cooled to ambient temperature and suspended in water (50 mL) and extracted with EtOAc (2×50 mL). The organic layer was washed with brine (50 mL), dried over Na.sub.2SO.sub.4 and concentrated in vacuo. The crude compound was purified by column chromatography (silica gel, 100-200 mesh, 40% EtOAc/pet. ether) to obtain I.sub.HSF 089 as a yellow solid (25 mg, 19%). R.sub.f: 0.7 (10% MeOH/CHCl.sub.3). Method_2 TFA_UPLC_2; Rt 1.74 min (94%); m/z 223 (MH.sup.+); .sup.1H NMR (d.sup.6 DMSO) δ7.35-7.36 (d, 1H), δ7.47-7.51 (d, 1H, J=15.8 Hz), δ7.57-7.58 (d, 1H), δ7.58-7.62 (d, 1H, J=15.8 Hz), δ9.16 (s, 1H), δ12.81 (s, br, 1H).

(269) Synthesis of Final Compound I.sub.HSF 091 Via Acid Intermediate A1:

(270) Step 3a: (E)-tert-Butyl-2-(4-oxo-4-(thiazol-2-ylamino)but-2-enoyl)hydrazinecarboxylate. To a stirred suspension of (E)-4-oxo-4-(thiazol-2-ylamino)but-2-enoic acid (A1) (500 mg, 2.52 mmol) in DCM (50 mL) at 0° C. was added sequentially, DIPEA (0.62 mL, 3.53 mmol), EDC (540 mg, 2.83 mmol), HOBt (390 mg, 2.83 mmol) and Boc-hydrazine (340 mg, 2.59 mmol), and the reaction mixture was stirred at ambient temperature for 16 h. The reaction mixture was diluted with water (50 mL), extracted with DCM (2×200 mL), the organic layer was washed with brine (50 mL), dried over Na.sub.2SO.sub.4 and concentrated in vacuo. The crude compound was purified by column chromatography (silica gel, 100-200 mesh, 60% EtOAc/pet ether) to obtain (E)-tertbutyl-2-(4-oxo-4-(thiazol-2-ylamino)but-2-enoyl)hydrazinecarboxylate as a white solid (200 mg, 26%). R.sub.f: 0.6 (10% MeOH/CHCl.sub.3). .sup.1H NMR (d.sup.6 DMSO) δ1.42 (s, 9H), δ7.02-7.06 (d, 1H), δ7.17-7.21 (d, 1H), δ7.31-7.32 (d, 1H), δ7.54-7.56 (d, 1H), δ9.04 (s, br, 1H), δ10.23 (s, br, 1H), δ12.69 (s, br, 1H); MS (APCI) m/z 313 (MH.sup.+).

(271) Step 4a: (E)-4-Hydrazinyl-oxo-N-(thiazol-2-yl)but-2-enamide hydrochloride. To a stirred solution of (E)-tert-butyl-2-(4-oxo-4-(thiazol-2-ylamino)but-2-enoyl)hydrazinecarboxylate (200 mg, 0.64 mmol) in dry 1, 4-dioxane (20 mL), cooled to 0° C. was added slowly HCl in dioxane (5 mL), and the reaction was allowed to stir at ambient temperature for 16 h. The organic volatiles were evaporated in vacuo, and the precipitated solid was washed with Et.sub.2O (2×20 mL) to give (E)-4-hydrazinyl-oxo-N-(thiazol-2-yl)but-2-enamide hydrochloride as a white solid (150 mg, 99% crude yield). R.sub.f: 0.2 (10% MeOH/CHCl.sub.3). .sup.1H NMR (d.sup.6 DMSO) δ7.10-7.13 (d, 1H), δ7.28-7.31 (d, 1H), δ7.34-7.35 (d, 1H), δ7.56-7.57 (d, 1H), δ11.74 (s, br, 1H), δ12.75 (s, br, 1H) some exchangeable protons not visible; MS (APCI) m/z 213 (MH.sup.+).

(272) Step 5a: (E)-3-(1,3,4-Oxadiazol-2-yl)-N-(thiazol-2-yl)acrylamide (I.sub.HSF 091, identified as DX 091 in the above scheme). To a solution of (E)-4-hydrazinyl-oxo-N-(thiazol-2-yl)but-2-enamide hydrochloride (200 mg, 0.81 mmol) in triethyl orthoformate (6 mL) was added PTSA (catalytic) and heated to 100° C. in a sealed tube for 16 h. The reaction mixture was cooled to ambient temperature, poured onto water (50 mL) and extracted with EtOAc (2×50 mL). The organic layer was washed with brine (50 mL), dried over Na.sub.2SO.sub.4 and concentrated in vacuo. The crude compound was purified by column chromatography (silica gel, 100-200 mesh, 75% EtOAc/pet. ether) to obtain compound (E)-3-(1,3,4-oxadiazol-2-yl)-N-(thiazol-2-yl)acrylamide, I.sub.HSF 091 as a yellow solid (50 mg, 28%). R.sub.f: 0.5 (10% MeOH/CHCl.sub.3). AK4; Rt 1.09 min (96%); m/z 223 (MH.sup.+); .sup.1H NMR (d.sup.6 DMSO) δ7.28-7.32 (d, 1H, J=15.6 Hz), δ7.34-7.35 (d, 1H), δ7.56-7.60 (d, 1H, J=15.6 Hz), δ7.56-7.57 (d, 1H), δ9.38 (s, 1H), δ12.74 (s, br, 1H).

(273) 1.5 Final Compounds—Route H

(274) 1.5.1 Synthesis of Final Compound I.sub.HSF 088 Via Route H

(275) ##STR00153##

(276) Step 1: (E)-Ethyl 3-(pyrimidin-2-yl)acrylate. A pre-mixed solution of triethylphosphonoacetate (270 mg, 1.2 mmol) in THF (5 mL) was added to a suspension of NaH (40 mg, 1.01 mmol) in THF at −15° C. over a period of 10 min. The reaction was stirred at −15° C. for 30 min. To the reaction mixture was added a pre-mixed solution of pyrimidine-2-carbaldehyde (100 mg, 0.92 mmol) in THF (15 mL) at −20° C. over a period of 10 min. The reaction mixture was quenched with ice-cold water and extracted with EtOAc (2×20 mL). The combined organics were washed with water (2×10 mL), brine solution (2×10 mL), dried over Na.sub.2SO.sub.4 and concentrated in vacuo. The crude compound was purified by column chromatography (silica gel, 100-200 mesh, eluting with 10-20% EtOAc/pet. ether) to afford (E)-ethyl 3-(pyrimidin-2-yl)acrylate as a pale yellow liquid (60 mg, 36%). R.sub.f: 0.6 (50% EtOAc/pet. ether). XTERRA; Rt 2.92 min (99%); m/z 179 (MH.sup.+); .sup.1H NMR (CDCl.sub.3) δ1.33-1.37 (t, 3H), δ4.27-4.32 (q, 2H), δ7.17-7.27 (m, 2H), δ7.68-7.72 (d, 1H), δ8.77-8.79 (d, 2H).

(277) Step 2: (E)-3-(Pyrimidin-2-yl)acrylic acid. To a stirred solution of (E)-ethyl 3-(pyrimidin-2-yl)acrylate (80 mg, 0.44 mmol) in THF (10 mL) was added an aqueous solution of LiOH.H.sub.2O (75 mg, 1.79 mmol) at ambient temperature. After 3 h the reaction mixture was concentrated in vacuo, and the residue was extracted with EtOAc (10 mL). The combined organic layers were washed with brine (2×10 mL), dried over anhydrous Na.sub.2SO.sub.4 and concentrated in vacuo to afford (E)-3-(pyrimidin-2-yl)acrylic acid as an off-white solid (40 mg, 59%). R.sub.f: 0.2 (70% EtOAc/pet. ether). .sup.1H NMR (d.sup.6-DMSO) δ6.98-7.02 (d, 1H), δ7.43-7.47 (d, 1H), δ7.49-7.51 (t, 1H), δ8.88-8.90 (d, 2H), δ12.88 (s, br, 1H).

(278) Step 3: (E)-3-(Pyrimidin-2-yl)-N-(thiazol-2-yl)acrylamide (I.sub.HSF 088, identified as DX 088 in the above scheme). To a stirred suspension of (E)-3-(pyrimidin-2-yl)acrylic acid (50 mg, 0.32 mmol) in DMF (3 mL) was added DIPEA (0.1 mL, 0.64 mmol), HATU (187 mg, 0.49 mmol), and 2-aminothiazole (35 mg, 0.35 mmol) sequentially at 0° C. The reaction was slowly warmed to ambient temperature and stirred for 30 min. The reaction mixture was diluted with EtOAc and washed with water then brine, dried over anhydrous Na.sub.2SO.sub.4 and concentrated under reduced pressure to obtain crude product. The crude product was triturated with MeOH, filtered and washed with MeOH to afford (E)-3-(pyrimidin-2-yl)-N-(thiazol-2-yl)acrylamide, I.sub.HSF 088, as a yellow solid (18 mg, 24%). R.sub.f: 0.3 (70% EtOAc/pet. ether). Method_2_TFA_UPLC_2; Rt 1.49 min (99%); m/z 233 (MH.sup.+); .sup.1H NMR (CD.sub.3CO.sub.2D) δ7.19-7.20 (d, 1H), δ7.49-7.53 (d, 1H,), δ7.5-7.52 (d, 2H), δ7.83-7.87 (d, 1H, J=14.95 Hz), δ9.00 (d, 1H), δ11.67 (s, br, 1H).

(279) 1.6 Final Compounds Route I

(280) Synthesis of Final Compound I.sub.HSF 095 Via Route I

(281) Step 1: N.sup.1,N.sup.1-Dimethyl-N.sup.4-(thiazol-2-yl)fumaramide. To a stirred solution of 3-(thiazol-2-ylcarbamoyl)-acrylic acid (500 mg, 2.52 mmol) in dry THF at −40° C. was added NMM (306 mg, 3.03 mmol) and isobutyl chloroformate (413 mg, 3.03 mmol), and the solution was stirred for 20 min. N,O-dimethylhydroxylamine hydrochloride (154 mg, 2.52 mmol) was added to the reaction mixture which was allowed to warm to ambient temperature and was stirred for 2 h. The solvent was evaporated in vacuo, and the residue was diluted with water, extracted with EtOAc (2×100 mL) and the combined organics washed with water (2×100 mL), brine (2×100 mL), dried over Na.sub.2SO.sub.4 and concentrated in vacuo. The crude compound was purified by column chromatography (silica gel, 100-200 mesh, 60% EtOAc/pet. ether as eluent) to afford N.sup.1,N.sup.1dimethyl-N.sup.4-(thiazol-2-yl)fumaramide as an off-white solid (250 mg, 41%). R.sub.f: 0.2 (40% EtOH/pet. ether). .sup.1H NMR (d.sup.6-DMSO) δ3.2 (s, 3H), δ3.73 (s, 3H), δ7.15-7.19 (d, 1H), δ7.3-7.31 (d of d, 1H), δ7.41-7.44 (d, 1H), δ7.53-7.54 (d of d, 1H), δ12.68 (s, br, 1H).

(282) ##STR00154##

(283) Step 2: (E)-4-Oxo-N-(thiazol-2-yl)hept-2-enamide (I.sub.HSF 095, identified as DX 095 in the above scheme). To a stirred solution of N.sup.1,N.sup.1-dimethyl-N.sup.4-(thiazol-2-yl)fumaramide (150 mg, 0.63 mmol) in dry THF, cooled to 0° C., was added slowly a freshly prepared solution of n-propylmagnesium bromide (37 mg, 1.55 mmol). The reaction was stirred at 0° C. for 2 h. The reaction mixture was quenched with sat. aq. NH.sub.4Cl solution, the solvent was evaporated in vacuo, and the residue was diluted with water and extracted with EtOAc (2×100 mL). The combined organics were washed with water (2×100 mL), brine (2×100 mL), dried over Na.sub.2SO.sub.4 and concentrated in vacuo. The crude compound was purified by column chromatography (silica gel, 100-200 mesh, 18% EtOAc/pet. ether) to afford (E)-4-oxo-N-(thiazol-2-yl)hept-2-enamide, I.sub.HSF 095, as a white solid (25 mg, 18%). R.sub.f: 0.6 (30% EtOH/pet. ether). LCMS-2; Rt 1.32 min (97%); m/z 225 (MH.sup.+); .sup.1H NMR (CDCl.sub.3) δ0.96-1.00 (t, 3H), δ1.68-1.77 (m, 2H), δ2.65-2.68 (t, 2H), δ7.09-7.13 (d, 1H, J=15.8 Hz), δ7.12-7.13 (d, 1H), δ7.33-7.37 (d, 1H, J=15.82 Hz), δ7.67-7.68 (d, 1H), δ12.24 (s, br, 1H).

Example 2: Pharmaceutical Compositions for Parenteral Administration

(284) 2.1 Formulation of I.sub.HSF 115 in Kleptose HPB

(285) TABLE-US-00014 TABLE 14 IHSF 115 in aqueous Kleptose HPB Component Amount I.sub.HSF 115 85 mg Kleptose HPB 3.035 g Water 8.5 g
Preparation of a Parenteral Formulation for Use in the Experiments Described Under Example 3:

(286) 1. Prepare 15 ml of a 357 mg/ml stock solution of Kleptose HPB (Roquette Pharma) in water (of which the amount not used under 2 is reserved for vehicle controls).

(287) 2. Dissolve 85 mg of compound 0115 in 8.5 ml of Kleptose stock solution by incubating the solution overnight with slow agitation (at room temperature). Over 90% of the drug substance will be dissolved after this incubation.

(288) 3. Vortex briefly and sterile-filter the solution of step 2 through MillexR-GV, Syringe Driven Filter Unit (cat. No. SLGV 004 SL, Milipore).

(289) 4. For 2 mg and 1 mg doses, dose undiluted at 200 μL, and 100 μL, respectively.

(290) 2.2 Suspension formulations of I.sub.HSF 115

(291) 2.2.1 Formulation with Solutol HS15 and Lecithin

(292) TABLE-US-00015 TABLE 15 Formulation with Solutol HS15 and Lecithin Component Amount/volume I.sub.HSF 115 25 mg Solutol HS15 (1 g/ml in water)* 37.5 μl Phosphatidylcholine 400 μl (0.25 g/ml water)* Water 537.5 μl *Chemicals obtained from Sigma-Aldrich
Preparation of a Parenteral Formulation for Use in the Experiments Described Under Example 4:

(293) 1. Mix solvents (Solutol HS15, phosphatidylcholine and water).

(294) 2. Autoclave and add water to compensate for losses, if any.

(295) 2. Use the autoclaved solvent mixture to dissolve I.sub.HSF 115 (sterilized by β-irradiation (25 kGy)) by stirring the complete mixture at 60° C. for approximately 10 min.

(296) 2.2.2 Formulation with Cremophor EL and Lecithin

(297) TABLE-US-00016 TABLE 16 Formulation with Cremophor EL and Lecithin Component Amount/volume I.sub.HSF 115 25 mg Cremophor EL (1 g/ml in water)* 25 μl Phosphatidylcholine 400 μl (0.25 g/ml water)* Water 270 μl Glucose 5% 280 μl *Chemicals obtained from Sigma-Aldrich
Preparation of a Parenteral Formulation:

(298) 1. Mix solvents (Cremophor EL, phosphatidylcholine, glucose and water).

(299) 2. Autoclave and add water to compensate for losses, if any.

(300) 2. Use the autoclaved solvent mixture to dissolve I.sub.HSF 115 (sterilized by β-irradiation (25 kGy)) by stirring the complete mixture at 60° C. for approximately 10 min.

Example 3: Anti-Tumor Activity of I.SUB.HSF .115 as a Single Agent in a Human Prostate Cancer Xenograft Model

(301) Prostate cancer cell line PC-3 was obtained from ATTC and grown in tissue culture under standard conditions. Groups (8 animals) of athymic mice (6-7 weeks old, male, obtained from Harlan) were inoculated subcutaneously on the right flank with 5×10.sup.6 PC-3 cells. Cells were administered as a 1:1 mixture of cells in medium and Matrigel. Treatment was initiated when tumor size reached about 200 mg, at which time the mice were randomized. Of the various groups in the experiment, one group received a daily intravenous dose (via the tail vein) of 2 mg I.sub.HSF 115 in an aqueous solution of Kleptose HPB (see Example 2.1) for 7 days, followed by additional dosing with 2 mg I.sub.HSF 115 every other day. Another group received a daily dose of 0.8 mg I.sub.HSF 115 in an aqueous solution of Kleptose HPB (see Example 2.1) for 7 days, followed by additional dosing with 0.8 mg I.sub.HSF 115 every other day. A control group was subjected to same regimen, administering vehicle (Kleptose HPB solution) only. Tumor size and weight were assessed periodically and recorded. Tumor size was estimated using calipers. Calculation: tumor weight (mg)=(a×b.sup.2/2) where ‘b’ is the smallest diameter and ‘a’ is the largest diameter. The results for the period of daily administration are shown in FIG. 9. Daily administration of 2 mg I.sub.HSF 115 resulted in a substantial (and statistically significant) reduction in tumor growth. The 0.8 mg dose had a smaller effect. Loss of weight for the period was less than 4%. No toxicity was observed, except for some local swelling in the inoculation region.