SUBSTITUTED AMINOTHIAZOLES FOR THE TREATMENT OF TUBERCULOSIS

Abstract

Disclosed ore substituted aminothiazoles, which can be used for, among other things, the treatment of tuberculosis, pharmaceutical compositions containing the same, and methods of using the same.

Claims

1-46. (canceled)

47. A method of treating non-replicating tuberculosis comprising administering to a subject a compound of Formula I, or a pharmaceutically acceptable salt, ester or prodrug thereof. ##STR00100## wherein: A.sub.1 is NR.sub.18, O, or S, (CH.sub.2).sub.q, or —NH(CH.sub.2), —N(H)—N═C*—(CH).sub.mCH.sub.3 R.sub.1 is optionally substituted alkyl, optionally substituted aryl, optionally substituted pyridyl, optionally substituted pyrimidinyl, optionally substituted pyrazinyl, optionally substituted pyridazinyl, or triazinyl; dd-dd R.sub.2 is, optionally substituted aryl, optionally substituted pyridyl, optionally substituted pyrimidinyl, optionally substituted pyrazinyl, optionally substituted pyridazinyl, or triazinyl; R.sub.3 is H, OR.sub.4, NR.sub.5R.sub.6, NO.sub.2, SO.sub.2NH.sub.2, halo, optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted carbocycle, optionally substituted heterocycle, optionally substituted carboxyl, optionally substituted alkoxycarbonyl, or optionally substituted aryloxycarbonyl; R.sub.4, R.sub.5, and R.sub.6 are each independently H, halo, optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted carbocycle, or optionally substituted heterocycle; R.sub.18 is H, optionally substituted alkyl, optionally substituted aryl optionally substituted aryl alkyl, optionally substituted heteroaryl, optionally substituted carbocycle, or optionally substituted heterocycle; C* is where R.sub.2 bonds with the group —N(H)—N═C*—(CH).sub.mCH.sub.3; m is 0-6; and q is 1-6.

48-49. (canceled)

50. The method of claim 47, wherein the compound is selected from the group consisting of: ##STR00101## ##STR00102## ##STR00103##

51. The method of claim 47 wherein in the compound of Formula I, A.sub.1 is NR.sub.18, O or S.

52. The method of claim 47 wherein in the compound of Formula I, A.sub.1 is (CH.sub.2).sub.q, NH(CH.sub.2).sub.q or —N(H)—N═C*—(CH.sub.2).sub.mCH.sub.3.

53. The method of claim 47 wherein in the compound of Formula I, R.sub.3 is H, halo, or optionally substituted C.sub.1-C.sub.6 alkyl.

54. The method of claim 47 wherein in the compound of Formula I, R.sub.3 is OR.sub.4, NR.sub.5R.sub.6, NO.sub.2, SO.sub.2NH.sub.2.

55. The method of claim 47 wherein in the compound of Formula I, R.sub.1 is: ##STR00104## wherein: R.sub.7 and R.sub.8 are each independently H, OR.sub.11, NR.sub.12R.sub.13, NO.sub.2, SO.sub.2NH.sub.2, halo, optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted carbocycle, or optionally substituted heterocycle, R.sub.11, R.sub.12, and R.sub.13 are each independently H, halo, optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted carbocycle, or optionally substituted heterocycle.

56. The method of claim 47 wherein in the compound of Formula I, R.sub.1 is ##STR00105## and R.sub.17 is C.sub.1-C.sub.6 alkyl.

57. The method of claim 47 wherein in the compound of Formula I, R.sub.2 is: ##STR00106## wherein: R.sub.9 and R.sub.10 are each independently H, OR.sub.14, NR.sub.15R.sub.16, NO.sub.2, SO.sub.2NH.sub.2, halo, optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted carbocycle, or optionally substituted heterocycle, R.sub.14, R.sub.15, and R.sub.16 are each independently H, halo, optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted carbocycle, or optionally substituted heterocycle.

58. The method of claim 57 wherein in the compound of Formula I, R.sub.2 is ##STR00107## and wherein R.sub.10 is C.sub.1-C.sub.6 alkyl, —O—R.sub.21—C(═O)OR.sub.22, wherein. R.sub.21 is C.sub.1-C.sub.6 alkyl and R.sub.22 is C.sub.1-C.sub.6 alkyl or H.

59. The method of claim 47 wherein in the compound of Formula I, R.sub.3 is H, halo, —C(═O)OR.sub.19, ##STR00108## wherein R.sub.19 is C.sub.1-C.sub.6 alkyl and R.sub.20 is C.sub.1-C.sub.6 alkyl.

60. The method of claim 57 wherein in the compound of Formula I, R.sub.9 and R.sub.10 are independently H or OR.sub.14.

61. The method of claim 57 wherein in the compound of Formula I, R.sub.9 is H and R.sub.10 is C.sub.1-C.sub.6 alkyl.

62. The method of claim 57 wherein in the compound of Formula I, R.sub.9 and Rv.sub.10 are independently H or halo.

63. The method of claim 57 wherein in the compound of Formula I, R.sub.2 is ##STR00109## wherein R.sub.9 and R.sub.10 are independently H, halo, haloalkyl, OR.sub.14, NR.sub.15R.sub.16, or C.sub.1-C.sub.6 alkyl.

64. The method of claim 63 wherein in the compound of Formula I, R.sub.9 and R.sub.10 are independently: H or OR.sub.14; H or NR.sub.15R.sub.16; H or C.sub.1-C.sub.6 alkyl; or H or halo.

65. The method of claim 47 wherein in the compound of Formula I, R.sub.15 and R.sub.16 are independently C.sub.1-C.sub.6 alkyl.

66. The method of claim 47 wherein the compound of Formula I is ##STR00110##

Description

EXAMPLES

[0186] Assays used to measure growth or inhibition of TB.

[0187] Bacterial strains and growth conditions. The strain M. tuberculosis H37Rv was used for all experiments. OFF was expressed using a constitutive episomal plasmid driven by the Rv3583c promoter. An inducible firefly luciferase expression plasmid was constructed using an anhydroietracyeltne inducible system, as described previously (30). Mtb H37Rv was grown at 37° C. in Middlebrook 7H9 broth supplemented with 10% OADC (oleic acid-albumin-dextrose complex), 0.2% glycerol and 0.05% Tween-80 or on Middlebrook 71110 plates supplemented with 10% OADC enrichment.

[0188] Carbon Starvation. Freezer stocks of H3Rv were diluted 1:50 in fresh 7H9 OADC media and cultured until late log phase. OD.sub.600 between 0.6 and 1.0. The bacteria were centrifuged at 2800× g for five minutes and resuspended in 50 mL of starvation media (7H9 and 0.05% Tyloxapol without any supplementation). The cells were then washed an additional two times with starvation media. After the final wash the cells were resuspended in starvation media to an OD.sub.600 of 0.2 and 50 ml of culture aliquoted into a sterile roller bottle. The starvation culture was incubated standing at 3° C. for 5 weeks.

[0189] Compound Testing: An initial assay was developed and utilized using a GFP-expressing M. tuberculosis reporter strain that measured fluorescence as a marker for growth and survival after an outgrowth period in rich 7H9 media. This assay was modified to accommodate the scale of the screen involving the MLPCN library. These modifications include using Alamar blue to measure surviving bacteria rather than the fluorescence, and outgrowth in 7H12 media.

[0190] Carbon starvation assays. For M. tuberculosis starvation screening assays, carbon-starved bacteria expressing GFP were diluted and plated into 384 well plates into which compounds had previously been pinned for a final OD600 of 0.05, a final volumes of 40 μL and a final compound concentration of 30 μM. Plates were incubated for a period of 120 h. at which time 10 μL 5× concentrated media was added to each well of the plate (7H9 media with 50% OADC, 1% glyceroln 0.05 % tyloxapol). Plates were then incubated for an additional 96 hours, at which time fluorescence was read using an M5 Spectramax. Each compound was screened in duplicate, and composite z-scores were calculated using DMSO controls as reference. Compounds were compounds that could inhibit growth or kill M. tuberculosis were defined as compounds with a composite z-score of less than −6. This z-score cutoff was selected as the z score of the concentration of the control antibiotic rifampicin that gave a Z′-factor of 0.

[0191] Alternative Carbon starvation assay. Carbon starved bacteria were diluted and plated into 384 well plates into which compounds had already been pinned for a final OD600 of 0.005 and final volume of 50 uL. The plates were incubated for 96 hours, at which time 12 uL of concentrated media was added. Plates were incubated for an additional 72 hours. For Alamar blue detection, a solution of 3 parts 18.2% Tween-80 to 4 parts Alamar Blue (3/7th Tween-80 to 4/7th Alamar Blue) is made and 9 ul added to each well in the plate. The plates are incubated (stacked 2-3 high) overnight at 37 ° C. in humidified incubator. The plates are removed from the incubator and sealed with aluminum seals. The fluorescence is read using the Envision plate reader (bottom read) with an excitation wavelength of 531 nm and an emission wavelength of 595 nm (Excitation filter=BODIPY TMR FP 531, barcode 105; Emission filter=Photometric 595, barcode 315; Mirror=BODIPY TMR, barcode 405).

[0192] Replicating, logarithmic assay. For M. tuberculosis screening assays for logarithmically growing, actively replicating activity, bacteria expressing GFP was grown to mid-log phase (OD600=0.6−0.8), diluted, and plated into 384 well plates into which compounds had previously been pinned for a final OD600 of 0.025. Elates were incubated for a period of 72 hr, at which time GFP fluorescence is read. Each compound is screened in duplicate, and composite z-scores were calculated using DMSO controls as reference. Compounds that inhibit replicating M. tuberculosis were defined as compounds with a composite z-score of less than −4. This z-score cutoff was selected using average of the z-scores of the concentrations of the control antibiotics clofazimine and rifampicin that gave a Z′-factor of 0.

[0193] Replicating IC99 determination by OD600. For dose response curves and IC90 determinations by OD600, bacteria w ere grown to mid-log phase and plated in 96 well plates at OD6O0=0.05 in the presence of small molecule inhibitors for 7 days unless otherwise indicated, and growth was assessed by reading OD600. The IC90 was defined as the minimum concentration that inhibited growth by 90% relative to the DMSO control (31).

[0194] Non-replicating IC99 determination by luciferase. For the luciferase assay it test for activity of small molecules directly on non-replicating cells without an outgrowth phase, carbon-starved M. tuberculosis H37Rv containing an inducible firefly luciferase plasmid was dispersed into 96-well plates containing the small molecules and anhydotetracydine 50 nM (to induce luciferase expression). After 5 days the cells were lysed, luciferase reagent added and luminescence measured (Promega Corporation, Madison Wis.) in a Spcctramax M5 (Molecular Devices). The antibiotic rifampicin (at 80X the MIC) was used as a positive control for the assay.

[0195] Replicating IC90 determinations by CFU. To confirm the replicating IC90 values determined using OD600, the activity of selected small molecules were tested by plating fur colony forming units (CFU). M. tuberculosis H3Rv was grown to mid-log phase and plated in 96 well plates at OD600=0.025 in the presence of small molecule inhibitors for specified time periods. The number of surviving bacteria was then determined by plating a dilution series of the culture for colony forming units (CFU). The IC90 was defined the concentration tested that inhibited growth by at least 90% relative to the DMSO control.

[0196] Non-replicating IC90 determinations by CFU. To confirm the non-replicating IC90 values determined using the luciferase reporter, the activity of selected small molecules was tested by plating for CFU. Carbon-starved bacteria were diluted to OD600=0.05 in starvation media and plated in 96 well plates, in the presence of small molecule inhibitors for indicated time periods. The number of surviving bacteria was then determined by plating a dilution series of the culture for colony forming units (CFU). The IC90 was defined as the concentration tested that inhibited survival by at least 90% relative to the DMSO control.

[0197] Generating resistant mutants. The MIC of each compound on solid media was identified by plating 10.sup.7 bacteria on agar containing a dose response in 96 well plate format. The MIC was defined as the lowest concentration resulting in inhibition of bacterial growth. Resistant mutants were generated by plating M. tuberculosis cells onto agar pads containing 2× and 10× the agar MIC of each compound using four independently derived wild-type clones. Colonies that arose on inhibitor containing plates were inoculated into liquid media containing 1× the liquid MIC of the inhibitor. These cultures were grown to mid-log and samples were retested in a liquid MIC assay to confirm that a shift relative to the wild-type MIC was observed.

[0198] Macrophage toxicity assay. To determine macrophage toxicity, J774 macrophages were plated in 96 well plates at a concentration of 6.25×10.sup.4 cells/well and rested overnight. A dilution series of the small molecule being tested was then added to the plates in quadruplicate. The top concentration tested was 50 uM. The plates were incubated for 48 hours, upon which time CellTiter-Glo (Promega Corporation) was used as a readout for macrophage viability.

Example 2

Synthesis of Compounds

[0199] Reagents and solvents were obtained from commercial suppliers and were used without further purification. Analytical LCMS was conducted on an Agilent Poroshell 120 EC-C18 column (30 mm×3.0 mm i.d.), eluting with 0.01% formic acid in water (solvent A) and 0.01% in acetonitrile (solvent B), using the following elution gradient: 0.00 min-0.03 min (5% B), 0.03 min-1.78 min (5% to 95% B, linear gradient), 1.78-2.28 min (95% B). 2.28-2.30 min (95% to 5% B, linear gradient), 2.30-2.50 min (5% B) at a flow rate of 1.75 ml/min and the mass spectra were recorded in elcctrospray positive and/or negative ion modes (ESI+and ESI−) on a Waters ZQ mass spectrometer. Preparative MPLC was performed on Varian ProStar system on a Agilent ZORRAX XD8-C18 column (150 mm×21/2 mm i.d.), eluting with water (solvent A) and acetonitrile (solvent B), using the following elution gradient: 0 in (50% B), 5 min (50% B), 14 min (95% B), 17 min (95% B) at a flow rate of 20 ml/min. .sup.1H NMR spectra (300 MHz or 400 MHz) and .sup.13C-NMR (75 MHz) were recorded on a Bruker spectrometer and chemical shifts are reported in parts per million (ppm, δ) downfield from tetramethylsilane (TMS). Coupling constants (J) are reported in Hz. Spin multiplicities are described as s (singlet), brs (broad singlet), t (triplet), q (quartet), and m (multiplet). Microwave reactions were performed in a Biotage Initiator 60 EXP microwave reactor.

[0200] Synthesis of N-(3-methylpyridin-2-yl)-4-(pyridin-2-yl)thiazol-2-amine (1). The compound was prepared according to the following scheme:

##STR00070##

Specifically, a mixture of 2-(2-bromoacetyl)pyridine hydrobromide (421 mg, 1.50 mmol) and 1-(3-methyl-2-pyridyl)thiourea (251 mg, 1.50 mmol) in ethanol (15 ml) was stirred at 80° C. for 5 h. After cooled to room temperature the mixture was evaporated and the residue was partitioned between AcOEt and 10% aqueous K.sub.2CO.sub.3. Organic phase was washed with brine, dried over Na.sub.2SO.sub.4 and evaporated. The residue was crystallized from ethanol to give compound I (325 mg, 81% yield). .sup.1H NMR (300 MHz, DMSO-d.sub.6) δ 10.52 (s. 1H), 8.63-8.56 (m, 1H), 8.19 (dd, J=4.8, 1.2 Hz, 1H), 8.05 (d, J=7.9 Hz, 1H), 7.87 (td, J=7.7, 1.8 Hz, 1H), 7.66 (s, 1H), 7.57 (dd, J=7.2, 0.6 Hz, 1H), 7.31 (ddd, J=7.5, 4.8, 1.1 Hz, 1H), 6.92 (dd, J=7.3, 5.0 Hz, 1H), 2.37(s, 3H). 13C NMR (75 MHz, DMSO-d.sub.6)δ 160.10, 152.56, 150.00, 149.37, 148.83, 143.29, 138.54, 137.05, 122.44, 119.93, 119.30, 116.37, 109.91, 16.85.

[0201] Synthesis of N-(6-methylpyridin-2-yl)4-(pyridin-2-yl)thiazol-2-amine (7). The compound was prepared according to the following scheme:

##STR00071##

Specifically, a mixture of 4-(pyridine-2-yl)thiazole-2-amine (53 mg, 0.30 mmol), 2-chloro-6-methylpyridine (27.7 ul. 0.25 mmol), powdered Na.sub.2CO.sub.3 (39 mg, 0.37 mmol), Xantphos (25 mg, 0.04 mmol) and Pd.sub.2 (20 mg. 0.02 mmol) in toluene (1 ml) was heated at 150° C. for 30 min under microwave irradiation. After cooled to room temperature the mixture was diluted with AcOEt and filtered to remove insoluble. The filtrate was evaporated. The residue was purified by preparative reversed phase HPLC (C18, 50˜95% acetonitrile in water) to give compound 7(17 mg, 25% yield). .sup.1H NMR (300 MHz, DMSO-d.sub.6) δ 11.39 (s, 1H), 8.62-8.53 (m, 1H), 7.96 (dd. J=7.7, 0.8, 1H), 7.87 (td, J=7.9, 1.8, 1H), 7.65 (s, 1H), 7.60 (t, J=7.8. 1H), 7.35-7.25 (m, 1H), 6.89 (d, J=8.2, 1H), 6.80 (d, J=7.3, 1H), 2.47 (s, 3H).

[0202] Synthesis of 4-(pyridin-2-yl)-N-(o-tolyl)thiazol-2-amine dihydrohormide (8). The compound was prepared according to the following scheme:

##STR00072##

Specifically, a mixture of 2-(2-bromoacetyl)pyridine hydrobromide (56 mg. 10.20 mmol) and 1-(o-tolyl)thiourea (35 mg, 0.21 mmol) in ethanol (1 ml) was stirred at 70° C. for 6 h. After cooled to room temperature the mixture was evaporated and the residue was triturated with AcOEt. The solid was collected by filtration to give compound 8 (80 mg, 93% yield as dihydrobormide salt). .sup.1H NMR (300 MHz. DMSO-d.sub.6)δ 9.70(s. 1H), 8.73(d, J=5.7, 1H). 8.52 (t, J=7.7, 1H), 8.45 (d, J=7.8, 1H), 8.21-8.11 (m, 1H), 8.03-7.94 (m, 1H), 7.91-7.80 (m, 1H), 7.26 (t, J=6.8, 2H), 7.14-7.03 (m, 1H), 2.30 (s, 3H).

[0203] Synthesis of 5-chloro-N-(3-methylpyridin-2-yl)-4-(pyridin-2-yl)thiazol-2-amine (16). The compound was prepared according to the following scheme:

##STR00073##

Specifically, a mixture of N-(3-methylpyridin-2-yl)-(pyridin-2-yl)thiazil-2-amine 1 (54 mg, 0.20 mmol) and N-chlorusuccinimide (32 mg, 0.24 mmol) in dichloromethane (2 ml) was stirred at room temperature for 64 h. After diluted with dichloromethane, the mixture was washed with saturated aqueous sodium bicarbonate and saturated aqueous sodium chloride, dried over anhydrous sodium sulfate and evaporated. The residue was chromatographed on a silica gel (ethyl acetate:chloroform=1:4−1:2) to give compound 16 (57 mg, 93% yield). 1H NMR (400 MHz, DMSO-d6)δ 10.86(s, 1H), 8.70-8.65 (m, 1H), 8.23-8.18 (m, 1H), 8.01 (d, J=7.6 Hz, 1H), 7.92 (t, J=7.5 Hz, 1H), 7.60 (d, J=6.7 Hz, 1H), 7.41-7.35 (m, 1H), 6.99-6.93 (m, 1H), 2.36 (s, 3H), Rt 1.34 min, MS +3.03/304.9.

[0204] Synthesis of 5-bromo-N-(3-methylpyridin-2-yl)-4-(pyridin-2-yl)thiazol-2-amine (17). The compound was prepared according to the following scheme:

##STR00074##

Specifically, a mixture of N-(3-methylpyridin-2-yl)-4-(pyridin-2-yl)thiazol-2-amine 1 (54 mg, 0.20 mmol) and N-bromosuccinimide (42 mg, 0.24 mmol) in dichloromethane (10 ml) was stirred at room temperature for 70 min. After diluted with dichloromethane, the mixture was washed with saturated aqueous sodium bicarbonate, dried over anhydrous sodium sulfate and evaporated. The residue was chromatographed on a silica gel (ethyl acetate:chloroform=1:4−1:2) to give compound 17 (60 mg, 86% yield). .sup.1H NMR (400 MHz, DMSO-d.sub.6)δ 10.89 (s, 1H), 8.67 (d, J=3.4Hz, 1H), 8.21 (d, J=3.8 Hz, 1H), 8.02 (d, J=7.6 Hz, 1H), 7.91 (t, J=7.5 Hz, 1H), 7.60(d, J=6.6 Hz, 1H), 7.42-7.35 (m, 1H), 6.99-6.93 (m, 1H), 2.36 (s, 3H), Rt 1.38 min, MS +346.9/348.8.

[0205] Synthesis of ethyl 2-((3-methylpyridin-2-yl)amino)-4-(pyridin-2-yl)thiazole-5-carboxylate (18). The compound was prepared according to the following scheme:

##STR00075##

Specifically, a mixture of ethyl 2-bromo-3-oxo-3-(pyridio-2-yl)propanoate (142 mg, 0.52 mmol) and 1-(3-mecthylpyridin-2-yl)thiourea (88 mg, 0.53 mmol) in ethanol (5 ml) was stirred at 70° C. for 2 h. After cooled to room temperature, the reaction was quenched with saturated aqueous sodium bicarbonate (5 ml) and the mixture was evaporated to about a half of its volume. Water (5 ml) was added and the mixture was extracted with chloroform. Organic phase was washed with saturated aqueous sodium chloride, dried over anhydrous sodium sulfate and evaporated. The residue was crystallized from ethanol to give compound 18 (100 mg) and the mother liquor was chromatographed on a silica gel (ethyl acetate:hexanes=1:2−1:1) to give additional compound 18 (67 mg, total 167 mg, 94% yield). .sup.1H NMR (300 MHz, DMSO-d.sub.6)δ 11.18 (s, 1H), 8.62 (d, J=4.8 Hz, 1H), 8.30 (d, J=4.7 Hz, 1H), 7.87 (td, J=7.9, 1.6 Hz, 1K), 7.71-7.66 (m, 1H), 7.64 (d, J=7.4 Hz, 1H), 7.46-739 (m, 1H), 7.01 (dd, J=7.2, 5.1 Hz, 1H), 4.14 (q, J=7.0 Hz, 2H), 2.36(s, 3H), 1.14 (t, J=7.1 Hz, 3H), Rt 1.24 min, MS +341.0.

[0206] Synthesis of N-(3-methylpyridin-2-yl)-5-(morpholinomethyl)-4-(pyridin-2-yl)thiazol-2-amine (19). The compound was prepared according to the following scheme:

##STR00076##

Specifically, a mixture of N-(3-methylpyridin-2-yl)-4-(pyridin-2-yl)thiazol-2-amine 1 (113 mg, 0.42 mmol), morpholine (367 μl, 4.21 mmol), aqueous formaldehyde solution (36.5%, 341 μl, 4.21 mmol) in ethanol (2 ml) was stirred at 80° C. for 2 h. After cooled to room temperature, the mixture was diluted with ethyl acetate and washed with saturated aqueous sodium chloride, dried over anhydrous sodium sulfate and evaporated. The residue was chromatographed on a silica gel (chloroform:methanol=39:1 with 0.1% aqueous ammonia) followed by chromatographed on a silica gel (ethyl acetate:chloroform=1:4−1:1 with 0.1% triethylamine) to give compound 19 (144 mg, 93% yield). .sup.1H NMR (400 MHz, DMSO-d.sub.6)δ 10.25 (s, 1H), 8.65-8.59 (m, 1H), 8.22-8.17 (m, 2H), 8.04 (d, J=7.7 Hz, 1H), 7.87 (t, J=7.0 Hz, 1H), 7.55 (d, J=6.6 Hz, 1H), 7.33-7.25 (m, 1H), 6.94-6.87 (m, 1H), 4.21 (s, 2H), 3.59 (s, 8H), 2.35 (s, 3H), Rt 1.00 min. MS +368.1.

[0207] Synthesis of tert-butyl 2-((2-((4-(pyridin-2-yl)thiazol-2-yl)amino)pyridin-3-yl)oxy)acetate (20). The compound was prepared according to the following scheme:

##STR00077##

Specifically, a mixture of 2-aminopyridin-3-ol (551 mg, 5.00 mmol) and powdered sodium hydroxide (500 mg, 5.00 mg) in methanol (5 ml) was stirred at room temperature for 40 min. The mixture was evaporated to dryness. To the residue was added dimethyl sulfoxide (5 ml) and tert-butyl bromoacetate (807 μl, 5.50 mmol) and the mixture was stirred at room temperature for 20 h. The mixture was diluted with water and extracted with chloroform. Organic phase was washed with water and saturated aqueous sodium chloride, dried over anhydrous sodium sulfate and evaporated. The residue was. chromatographed on a silica gel (ethyl acetate:hexanes=1:2−2:1) to give tert-butyl 2-((2-aminopyridin-3-yl)oxy)acetate (622 mg, 55% yield). .sup.1H NMR (300 MHz, DMSO-d.sub.6)δ 7.53 (d, J=5.0 Hz, 1H), 6.91 (d, J=7.8 Hz, 1H), 6.47 (dd, J=7.8, 5.0 Hz, 1H), 5.64 (s, 1H), 4.65 (s, 2H), 1.43 (s, 9H), Rt 0.69 min, MS+225.2.

[0208] To a solution of tert-butyl 2-((2-aminopyridin-3-yl)oxy)acotate (604 mg, 2.69 mmol) in tetrahydrofuran (5 ml) was added benzoyl isothiocyanate (410 μl, 3.01 mmol) and the mixture was stirred at 70° C. for 3.5 h. After cooled to room temperature, hexanes were added and the mixture was stirred at room temperature overnight. Resulting precipitates were collected by filtration, washed with a mixture of ethyl acetate and hexanes (1:5) and dried under vacuum to give tert-butyl 2-((2-(3-benzoylthioureido)pyridin-3-yl)oxy)acetate (902 mg, 86% yield). .sup.1H NMR (300 MHz, DMSO-d.sub.6)δ 8.09 (d, J=4.7 Hz, 1H), 8.00 (d, J=7.6 Hz, 2H), 7.69 (t, J=7.3 Hz, 1H), 7.61-7.50 (m, 3H), 7.33 (dd, J=8.2, 4.8 Hz, 1H), 4.85 (s, 2H), 1.41 (s, 9H), Rt 1.42 min. MS+388.2.

[0209] A mixture of (m-butyl 2-((2-(3-benzoylthioureido)pyridin-3-yl)oxy)acetate (895 mg, 2.31 mmol) and potassium carbonate (3.87 mg, 2.80 mmol) in ethanol (8 ml) was stirred at 70° C. for 9 h. After cooled to room temperature, water was added and the mixture was extracted with ethyl acetate. Combined extracts was washed with saturated aqueous sodium chloride, dried over anhydrous sodium sulfate and evaporated. The residue was chromatographed on a silica gel (ethyl acctate:chloroform=1:4−1:1) to give tert-butyl 2-((2-thioureidopyridin-3-yl)oxy)acetate (125 mg, 17% yield). .sup.1H NMR (300 MHz, CDCl.sub.3)δ 10.93 (s, 1H), 8.77 (s, 1H), 7.86-7.83 (m, 1H), 7.06-7.02 (m, 1H), 6.94 (dd, J=8.1, 5.0 Hz, 1H), 6.84 (s, 1H), 4.61 (s, 2H), 1.48 (s, 9H), Rt 1.24 min, MS +284.8.

[0210] A mixture of 2-(2-bromoacetyl)pyridine hydrobromide (126 mg, 0.45 mmol) and tert-butyl 2-((2-thioureidopyridin-3-yl)oxy)acetate 1-(3-methyl-2-pyridyl)thiourea (122 mg, 0.43 mmol) in ethanol (4 ml) was stirred at 70° C. for 3.5 h. After cooled to room temperature, the reaction was quenched with saturated aqueous sodium bicarbonate (7 ml) and the mixture was evaporated to about a half of its volume and extracted with ethyl acetate. Combined extracts was washed with water and saturated aqueous sodium chloride, dried over anhydrous sodium sulfate and evaporated. The residue was chromatographed on a silica gel (ethyl acetate:hexanes=1:2−1:0) to give compound 20 (146 mg, 88% yield). .sup.1H NMR (300 MHz, DMSO d.sub.6)δ 10.04 (s, 1H), 8.60 (d, J=4.7 Hz, 1H), 8.03 (d, J=7.9 Hz, 1H), 7.96 (d, J=5.0 Hz, 1H), 7.88 (td, J=7.7, 1.6 Hz, 1H), 7.70 (s, 1H), 7.39-7.28 (m, 2H), 6.96 (dd, J=7.9, 5.0 Hz, 1H), 4.85 (s, 2H), 1.43 (s, 9H), Rt 1.31, MS+385.9.

[0211] Synthesis of 2-((2-((4-(pyridin-2-yl)thiazol-2-yl)amtno)pyridin-3-yl)oxy)acetic acid (21). The compound was prepared according to the following scheme:

##STR00078##

Specifically, a mixture of compound 20 (124 mg, 0.32 mmol), trifluroacetix acid (2 ml) and dichloromethane (3 ml) was stirred at room temperature for 2.5 h. The mixture was diluted with toluene and evaporated to dryness. The residue was triturated with ethyl acetate and the resulting precipitates were collected by filtration, washed with ethyl acetated and dried under vacuum to give compound 21 as mono trifluoroacetic acid salt (114 mg, 80% yield). .sup.1H NMR (400 MHz, DMSO-d.sub.6)δ 13.19 (s, 2H), 10.27 (s, 1H), 8.64 (d, J=3.7 Hz, 1H), 8.17 (d, J=7.7 Hz, 1H), 8.06 (t, J=7.0 Hz, 1H), 7.98 (d, J=4.6 Hz, 1H), 7.89 (s, 1H), 7.50-7.46 (m, 1H), 7.43 (d, J=7.8 Hz, 1H), 7.00 (dd, J=7.5, 5.0 Hz, 1H), 4.88 (s, 2H), Rt 0.84 min, MS+329.0.

[0212] Compounds 2-6 and 9-15 (see table below) disclosed herein could be made by similar methods, or by the schemes provided herein, or can be purchased commercially, for example, from Chembridge (San Diego, Calif.).

Example 3

[0213] The compounds were tested against replicating and non-replicating TB according to the assays described in Example 1. As can be seen some compounds can inhibit the growth of both replicating and non-replicating TB, whereas others are more selective or completely selective. The results are shown in the following table. Although Compound 16 has not yet been tested it is expected to have activity in the assays.

TABLE-US-00003 Non- replicating Mtb Replicating Carbon- Mtb starved Log MIC MIC IC99 IC90 (uM, Compound # Structure (uM, GFP) Luciferase) 1 [00079] 0.5 1 2 [00080] 2 2 3 [00081] 8 4 [00082] 62 5 [00083] 62 8 6 [00084] 125 8 7 [00085] 62 16 8 [00086] 31 2 9 [00087] 62 8 10 [00088] 125 11 [00089] None detected 12 [00090] 125 13 [00091] 8 14 [00092] 16 15 [00093] 62 16 16 [00094] 17 [00095] <1 μM <16 μM 18 [00096] 4 μM 8 μM 19 [00097] 16 μM 32 μM 20 [00098] N/A 4 μM 21 [00099] 125 μM >125 μM