(12) Patent Application Publication (10) Pub. No.: US 2003/0236225A1 Protopopova Et Al

US 20030236225A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2003/0236225A1 Protopopova et al. (43) Pub. Date: Dec. 25, 2003

(54) METHODS OF USE AND COMPOSITIONS (22) Filed: May 17, 2002 FOR THE DAGNOSS AND TREATMENT OF INFECTIOUS DISEASE Publication Classification (76) Inventors: Marina Nikolaevna Protopopova, (51) Int. Cl.' ...... A61 K 31/695; A61K 31/46; Silver Spring, MD (US); Richard A61K 31/445; A61K 31/137; Edward Lee, Cordova, TN (US); A61K 31/13 Richard Allan Slayden, Collins, CO (52) U.S. Cl...... 514/63; 514/304, 514/649; (US); Clifton E. Barry III, Rockville, 514/659; 514/331; 54.6/124; MD (US) 546/246; 564/367; 564/453 Correspondence Address: (57) ABSTRACT JOHN S. PRATT, ESQ Methods and compositions for treating disease caused by KILPATRICK STOCKTON, LLP microorganisms, particularly tuberculosis. In particular, 1100 PEACHTREE STREET methods and compositions comprising Substituted ethylene SUTE 2800 diamines for the treatment of infectious diseases are pro ATLANTA, GA 30309 (US) Vided. In one embodiment, these methods and compositions are used for the treatment of mycobacterial infections, (21) Appl. No.: 10/147,587 including, but not limited to, tuberculosis. Patent Application Publication Dec. 25, 2003. Sheet 1 of 61 US 2003/0236225A1

RNH2/ENG-Pr). MeO ulCICHCHCl, RT MeO Py/THF Me PyBroP X,Y = Cl, Br Rink-acidO resin X= Cl, Br EtN(iso-Pr)2 R=H,4. Me, Et, Bu

-NR O --O Me R4 X NHR2R3, Etn(-Pr). NHR2R3, Etn(-Pr). DMF, 70°C DMF, 70°C

CCl6

10% TFACIC

FIGURE 1 Patent Application Publication Dec. 25, 2003 Sheet 2 of 61 US 2003/0236225A1 Primary Amines

1. NH2

1. 4-Methylbenzylamine H3C

2. Cyclopentylamine 3. N

3. 2-(Aminomethylo)pyridine 2

O NH2 6. Furfurylamine r C H 7. o-3 O HC1 Cu NH O 2 7. 3,4,5-Trimethoxybenzylamine C H3

NH 2 CH3 8. 1-Methyl-3-phenylproplyamine

9. NH2 9. Cyclobutylamine ?

10. H2

10. 1,2,3,4-Trimethoxybenzylamine

FIGURE 2(a) Patent Application Publication Dec. 25, 2003. Sheet 3 of 61 US 2003/0236225A1

NH2 11. c 11. 2,3-Dimethylcyclohexylamine CH

12. Tyramine HO

13. 2.-Fluorobenzylamine CreF (R) H erro 16. (R)-2-Amino-1-butanol NH2

1 CH 3 1 7. O

H 3

17. 3,4-Dimethoxyphenethylamine “Cul NH 2

18 NH2

18. 3,3-Diphenylpropylamine 19 r 19. Propylamine CH3 21. r NH2 N 21. 1-(2-Aminoethyl)piperidine O

FIGURE 20b) Patent Application Publication Dec. 25, 2003. Sheet 4 of 61 US 2003/0236225A1

22.

22. Phenethylamine rNH,

C)O 23. 4-(2-Aminoethyl)morpholine NH2 24. (S)ch 24. (S)-Phenylglycinol

25 H N

25. Tryptamine

27. Cyclohexylamine 28a. ACH NH, H3C 28a. (+)-Isopinocampheylamine H3C

29.

29. Benzylamine

30.

30. 3-Amino-1-propanol H2N 1N1 No

31.

31. 2-Fluorophenethylamine F

FIGURE 20c) Patent Application Publication Dec. 25, 2003 Sheet 5 of 61 US 2003/0236225A1

CH3 33. NH2

33. b-Methylphenethylamine

34.

1. O H3C Olus 34. 4-Methoxyphenethylamine NH2 35. ill 1. N n-1N 35. N,N-Dimethylethylenediamine H3C NH2

36. L - S H3C 1. Y-rro 36. L-Methioninol NH2

37. O

37. Tetrahydrofurfurylamine

38.

38. Amylamine H3C 1N1-N-N-2

NH2 39. Aminomethylcyclopropane 41. rN, N

41. 1-(2-Aminoethyl)piperazine CD

FIGURE 20d) Patent Application Publication Dec. 25, 2003 Sheet 6 of 61 US 2003/0236225A1

42a. CH3 H2N CH3 CH3 42a. (+)-Bornylamine ' CH, CH3

43. tert-Octylamine H3CDuk CH3

44. t > ) 44. 1-Adamantanemethylamine

45. 2-Amino-1-propanol, d.l

46. 3-Phenyl-1-propylamine Cr

NH2

47. 2,2-Diphenylamine NH2 48. r N

48. 1-(3-Aminopropyl)-2-pyrrolidinone ( - (tech)

FIGURE 20e) Patent Application Publication Dec. 25, 2003 Sheet 7 of 61 US 2003/0236225A1

H2N F 49. 4-(Trifluoromethyl)benzylamine F

50. Nre, 50. 1-(2-Aminoehtyl)-pyrrolidine O si. -- HC1 O NH2 51. Veratryl amine

52. 1N1N1 N. 52. 5-Amino-1-pentanol H2N OH

53. 53. 2-(1-Cylcohexenyl)ethylamine O N N 54. CO 54. 5-Aminoquinoline NH2 H2N 55

55. 1-Aminomethyl-1-cylcohexanol, HCl Co 56. H.

56. 1-Aminopiperidine C

FIGURE 20f) Patent Application Publication Dec. 25, 2003 Sheet 8 of 61 US 2003/0236225A1

57. F

57. 3-Fluorobenzylamine 59. (1S,2R)- C r 59. (1S,2R)-cis-1-Amino-2-indanol NH2

61. 1N1,N1OH 61. 4-Amino-1-butanol H2N 63. (S)- her rol 63. (S)-2-Amino-1-butanol NH2

66. NH2 CH,

O 66. 2,4-Dimethoxybenzylamine CH3

68. H3C NH2

68. 1-(1-Naphthyl)ethylamine

69. O 69. 2-(2-Aminoethyoxy)ethanol HN1)-1N1 SoH 70. N s NH2 2N 70. 3-Amino-1,2,4-triazine N1

FIGURE 20g) Patent Application Publication Dec. 25, 2003 Sheet 9 of 61 US 2003/0236225A1

71. NH2

71. 2-Ethoxybenzylamine H3C ~5 72. NH2

72. 4-(3-Aminopropyl)morpholine C)O

73. NH2

O 73. 2-Amino-1-methoxypropane -- YoH 3 74a. CH3

74a. cis-(-)-Myrtanylamine re-r NH2

77a

77a. Cyclooctylamine Ore. H2N 78a

78a. 2-Adamantamine, HCl 79.

79. trans-2-Aminocyclohexanol, HCl OH N 80.

80. 3,Amino-5-phenyl pyrazole

FIGURE 2(h) Patent Application Publication Dec. 25, 2003. Sheet 10 of 61 US 2003/0236225A1

82. 2,3-Dimethoxybenzylamine

83a.

83a. Noradamantamine, HCl () 84. 4-Amino-1-benzylpiperidine NH2

85.

85. 4-Methylcyclohexylamine HN

86. OH

86. (1R,2S)-1-Amino-2-indanol NH2 87 N

87. 3-Aminopyrazole

8 8. F 88. 4-Fluorobenzylamine NH2

90a.

90a, trans-2-Phenylcyclopropylamine, HCI O

FIGURE 20i) Patent Application Publication Dec. 25, 2003. Sheet 11 of 61 US 2003/0236225A1

NH2

91. 1-(3-Aminopropyl)pipecoline O o-rrison 92. 2-Amino-1,3-propanediol NH2

93. OH

OH H3C NH2 93. Thiomicamine S

OH NH2 94. (R)-1-Amino-2-propanol H3C -N-

95. OH NH2 95. (S)-2-Amino-3-cyclohexyl-1-propanol, HCI 97. HO

97. 1-Amino-1-cyclopentane methanol O 98. NH2 OH H3C 98. (S)-Isoleucinol CH3

99.

OH 99. 4-Clorophenylalaniol NH2

FIGURE 20) Patent Application Publication Dec. 25, 2003. Sheet 12 of 61 US 2003/0236225A1

100. CH3 NH2

100. l-Leucinol H3C ul-A-oh OH O1. NH2

101. (1S,2S)-(+)-2-Amino-1-phenyl-1,3-propanediol 102. (S)-(+) luteOH 102. (S)-(+)-1-Amino-2-propanol H3C

OH 103. CH3 H2N

103. 2-Amino-2-methyl-1,3-propanediol OH NH2 104. (d.,) HC1 O N OH 104. d, 1-Serine methyl ester, HCl O

105.a. H3 CH3 H3C 105a. (-)-Isopinocampheylamine NH2

107. H N

107. Histidinol

FIGURE 20k) Patent Application Publication Dec. 25, 2003 Sheet 13 of 61 US 2003/0236225A1

108. N N1\ S-4

108. 2-Amino-5-cyclopropyl-1,3,4-thiadiaZol

109. CH3 H2N OH 109. 2-Amino-2-methyl-1-propanol H3C

111. an 1 NH 2 111. Allylamine

112. to-rrs, 112. 3-Amino-1,2-propanediol OH

115. - -,NH 115. Hexamethyleimine NH2 117. N

S 17. 3-Aminorhodamine CH3 NH2 119. OH

119. (R)-(-)-2-Phenylglycinol

126. Methyl-3-aminocrotonate

FIGURE 201) Patent Application Publication Dec. 25, 2003 Sheet 14 of 61 US 2003/0236225A1 29. cS 129. d, 1-Homocysteine, thiolactone, HCl NH2

137. 2-Amino-5trifluoromethyl-1,3,4-thiadiazol H2N 138. Racemic; "test" well H3C ~so 138. d, 1-2-Amino-1-butanol (test, making EMB) NH2

140. 140. 3-Etoxypropylamine HN1N1\o 1N CH3

141. her CH3 141. sec-Butylamine NH2

142. NH2

142. 2-Aminoheptane e-N-n-re.3

143. CH3 H2N OH 143. 2-Amino-2-methyl-1-propanol H3C

144. NH2

144. (S)-1-Amino-2-(Methoxymethyl)pyrrolidine

FIGURE 20m) Patent Application Publication Dec. 25, 2003. Sheet 15 of 61 US 2003/0236225A1

145a. NH2

145a, trans-l,2-Diaminocyclohexane NH2

147. O NH2

147. 3-Amino-2-methoxydibenzofuran O 148. *R OOH3 $-(N 148. 2-Amino-4-methoxybenzothiazole NH2

149.

149. 1-Aminohomopiperidine C) 150. CC H2 150. 2-Amino-3-hydroxypyridine OH

151. N 151. 1-Aminopyrrolidine, HCl NH2

152. CH3 HO 152. d, 1-2-Amino-1-pentanol NH2

154. OH 154. Ethanolamine H2N 1N1

FIGURE 2(n) Patent Application Publication Dec. 25, 2003 Sheet 16 of 61 US 2003/0236225A1

155. NH2

155. 3-Methylbenzylamine CH3

156. Hc1a1 N1\1\CH, o N- NH2 156. 3-(Dibutylamino)propylamine

157. HO CH3 NH2 157. Norephedrine, HCl

158. H2N O y O 158. Piperonylamine

159. O 159. 2-Methoxyethylamine HN1N1 NoH,

100. e-rrsch, 160. 1-Ethylpropylamine NH2 161. N

S.N NH2 161. 1-(3-Aminopropyl)imidazol n-1N1

162.

162. 1-Aminoadamantamine t C

164. NH2 erro,O O 164. Dimethylaminomalonate, HCl O O

FIGURE 200) Patent Application Publication Dec. 25, 2003 Sheet 17 of 61 US 2003/0236225A1

166.

166. Isopropylamine H3C CH3 167 it. 1. N1-a-NH2 167. 3-(Dimethylamino)propylamine H3C 169. cr" 169. 4-Fluorophenethylamine F

70. NH2

170. 2-(4-Aminophenyl)ethylamine 171. NH2

N 171. 3-Aminoisoxazole O

172. NH2 172. 1,2-Diaminopropane he-S- 2 173. D.L. H N NH2 O SCH 173. d, 1-Tryptophan methyl ester, HCl O 174. D- O

174. d-Aspartic acid, dimethyl ester, HCl NH2O

175. L- O CH3 hero- CH 3 175. l-Leucine ethyl ester, HCl NH2

FIGURE 20p) Patent Application Publication Dec. 25, 2003 Sheet 18 of 61 US 2003/0236225A1

176.Hics L- O O

HN s 176. l-Methionine ethyl ester, HCl CH3 177. D.L O HC3 sore, 177. d.l-a-Amino-n-butyric acid methyl ester, HCI NH2 H3C 178. L- o? O H2N

NO2 178. 3-Mnitro-l-tyrosine ethyl ester, HCI HO

179.9. L O o-3CH NH HO 2 179. 1-3,4-Dihydroxyphenylalanine methyl ester, HCl OH NH2

180. L

NH2 o 180. 1-Lysine methyl ester, HCl 181. (S)- O

srso:NH2 181. (S)-Benzyl-l-cysteine ethyle ester, HCl

FIGURE 20q) Patent Application Publication Dec. 25, 2003 Sheet 19 of 61 US 2003/0236225A1

182. L O CH3 H3C --- CH3 182. l-isoleucine methyl ester, HCI NH2 OH3 183. L- O O HN

HN X=NH 183. 1-Arginine methyl ester, HCl H2N 184. D.L. O. H3C so~ch, 184. d.l-Norleucine methyl ester, HCl NH2

185. NH HC1 O ^- 2 185. b-Alanine ethyle ester, HCl O

186. - O O Hc1 No OH 186. l-Glutamic acid ethyl ester, HCl NH2 187, L- O o1 NCH, NH2 187. 1-Phenylalanine ethyl ester, HCl 188. D,L- O -CH3 NH2 188. d.l-Phenylalanin methyl ester, HCl

FIGURE 20r) Patent Application Publication Dec. 25, 2003 Sheet 20 of 61 US 2003/0236225A1

189, L- O HN CH 189. 1-Histidine methyl ester, HCl N "reNH2 3 90. D.L-res--- O 190, d.l-Alanine ethyl ester, HCl NH2

191. O o1 NCH NH2 191. Tyrosine ethyl ester, HCl HO O CH3 192, L Hc1 No CH3 192. 1-Valine ethyl ester, HCl NH2

193. H.N. CH CH3 193. tert-Amylamine H3C

194. CH

CH 194. tert-Butylamine HC 3

97. L

197. S-Benzyl-L-cysteinol OrrorNH2

198. H

198. N-Phenylethyldiamine

FIGURE 20s) Patent Application Publication Dec. 25, 2003. Sheet 21 of 61 US 2003/0236225A1

NH2 201. CH r N 3 201. N,N,2,2-Tetramethyl-1,3-propanediamine CH, CH3

H N 202.

202. Isonipecotamide O1 NH

203. CH3

203. Isobutylamine H3C -N-N-

204. Hexctidine (mixture of stereosiomers)

NH2 206.

206. exo-Aminonorborname

s N-1 2O7. N

207. Ehtyl 4-amino-1-piperidinecarboxylate NH2

211. D OH NH2 or O 211. D-Glucosamine, HCl OH OH FIGURE 20t) Patent Application Publication Dec. 25, 2003 Sheet 22 of 61 US 2003/0236225A1

2 4. D NH2

214. Aminodiphenylmethane

2 5 H N

215. alpha-Methyltryptamine CH 3

2 6 w NH

216. 9-Aminofluorene, HCl

2 9.

219. 4-Phenylbutylamine cr"

2 CH3 NH2 221. 4-Chloroamphetamine, HCI C O. *-to CHH CH 222. 4-Amino-2,2,6,6-tetramethylpiperidine NH2

223. 4-(Hexacylamino)benzylamine

223. H H3C-1a1a1a1a1n 11a-N lulu,

FIGURE 20u) Patent Application Publication Dec. 25, 2003. Sheet 23 of 61 US 2003/0236225A1

225. -CH3 HO

225. 3-o-Methyldopamine, HCl

226.

226. 3-Fluorophenethylamine F

227. (aH 227. 3-Aminopyrrolidine, dihCl NH2

229.

S NH2

229. 2-Thiopheneethylamine (-)-

230. mix of cis/trans

230. 2-Methylcyclohexylamine NH2

23. NH2

231. 2-Methoxyphenethylamine

232. 2-Fluoroethylamine, HCl 1N1 NH2

FIGURE 20v) Patent Application Publication Dec. 25, 2003 Sheet 24 of 61 US 2003/0236225A1

233.

233. 2-Chlorobenzylamine C 234. c NH2 234. 2-Aminoindan, HCl

235. 2-Amino-4-phenyl-5-tetradecylthiazole 235. H2N ne S f CH3

236. OH

236. 2-Amino-1-phenylethanol or "

238.

238. 2,5-Dimethoxyphenethylamine

240.

240. 2,4-Dichlorophcnethylamine

241.

241. 2,2,2-Trifluoroethylamine F

FIGURE 2(w) Patent Application Publication Dec. 25, 2003. Sheet 25 of 61 US 2003/0236225A1

242.

242. 2-(2-Chlorophenyl)ethylamine cr"C

243.

S OH 243. 2-(2-Aminomethyl)phenylthio)benzyl alcohol OC.

245. Ol 245. 1-Aminoindan NH2

246. r OH ON 246. 1-Amino-4-(2-hydroxyethyl)piperazine

247. NH2 CH3

247. 1,3-Dimethylbutylamine e-S-Sc. 3

249. NH 2 CH3 H3C 249. 1,2-Dimethylbutylamine CH3

FIGURE 20x) Patent Application Publication Dec. 25, 2003 Sheet 26 of 61 US 2003/0236225A1 2 53.4Cs 253. 1-(1-Adamantyl)ethylamine, HCl NH2

H N 254. s NH2 254. (S)-(+)-2-(Aminomethyl)pyrrolidine

2 55. NH2

255. (S)-(-)-2-Cyclohexylethylamine

256.

OH NH2 256. (S)-(-)-2-Amino-3-phenyl-l-propanol

257. NH2

257. (R)-(-)-Cyclohexylethylamine

259. (1S,2S) OH O1 CH3 NH2 259. (1S,2S)-(+)-2-Amino-3-methoxy 1-phenyl-1-propanol

FIGURE 20y) Patent Application Publication Dec. 25, 2003. Sheet 27 of 61 US 2003/0236225A1

260. (1R,2S)

260. (1R,2S)-(–)-2-Amino-1,2-diphenylethanol

261. (-) CH3

HO NH2 261. (-)-3,4-Dihydroxynorephedrine OH

262. (1S,2R) ClubNH2 262. (1S,2R)-(+)-2-Amino-1,2-diphenylethanol O

263.

263. Octadecylamine H3C-(CH2) 17-NH2

264. 3-Aminoquinonuclidine, diFICl

265. (R)-(+) H C 265. (R)-Cycloserine NH2

266. 266. Undecylamine H3C-(CH2) 10 -NH2

FIGURE 20z) Patent Application Publication Dec. 25, 2003 Sheet 28 of 61 US 2003/0236225A1

OH 267. CH3

HO NH2 267. 3,4-Dihydroxynorephedrine OH

268. NH2

HO 268. 3-Hydroxytyramine OH

269. Sk 269. 4-(Trifluoromethoxy)benzylamine

272. CH3 CH3 272. Geranylamine

H 275. N

H 275. 5-Methoxytryptamine 3C NO NH2

276 NH2 CH3

276. 6-Amino-2-methyl-2-heptanol, HCI H3C --~k.CH3

277. 1-N-1-N-1-N-O 277. 6-Amino-1-hexanol HN

FIGURE 2(aa) Patent Application Publication Dec. 25, 2003. Sheet 29 of 61 US 2003/0236225A1

CH3 278. CH3 CH3

HC H 278. Dehydroabietylamine HN

2.79. HC NH2

279. 1-(1-Naphthyl)ethylamine

281 GHs

281. 2-(2-Aminoethyl)-1-methylpyrrolidine 282. orsD.L. CH3 282. d, 1-Valinol NH2

283. D,L- re 283. d, 1-2-Amino-1-hexanol NH

284. C H2 284, trans-2-Aminocyclohexanol, HCl OH

FIGURE 20ab) Patent Application Publication Dec. 25, 2003. Sheet 30 of 61 US 2003/0236225A1

285. (S)- S1\-1 NH2 285. S-Benzylcysteamine, HCl Or

288. NH2 CH3

288. 4-Fluoro-a-methylbenzylamine F

FIGURE 20ac) Patent Application Publication Dec. 25, 2003. Sheet 31 of 61 US 2003/0236225A1

Acyclic Secondary Amines

4. AutN 4. N-Propylcyclopropanemethylamine N-1)-ch,

H 15. 2-(Ethylamino)ethanol o1 N1,N1N CH3

CH3 20. HN1

20. N-Methyl-iso-propylamine H3C CH3 60. HC N-NH N 60. N-Methylpropylamine CH3

62. --- 62. 2-Methylaminomethyl 1,3-dioxolane (r.O

64. HN

64. Dibenzylamine

H 65. N-Butylbenzylamine ( N n1 N1

FIGURE 3(a) Patent Application Publication Dec. 25, 2003. Sheet 32 of 61 US 2003/0236225A1

67.

N-'3CH H 67. N-Benzylethylamine

76. QH N N CH3 76. (Methylaminomethyl)benzyl alcohol

8. H N 81. N-Benzyl-2-phenethylamine or-i-O

HO CH3

HN 89. Pseudoephedrine YOH

110. (1R,2S) HO CH3

110. (1R,2S)-(-)-Ephedrine

113. Diethanolamine

118. N-Benzylethanolamine

FIGURE 3(b) Patent Application Publication Dec. 25, 2003 Sheet 33 of 61 US 2003/0236225A1

120. H N 120. 2-(Propylamine)-ethanol Hc1N1'n 1NoH

HN 121. N-methylbutylamine CH

127. CH3

127.

131. N-Methylphenethylamine

132.

132. N-Ethylcyclohexylamine

136.

1NoH, 136. 4-(Ethylaminomethyl)pyridine N

163.

CH Hc-N1NH 1--O- 3 163. Bis(2-methoxyethyl)amiane

FIGURE 3(c) Patent Application Publication Dec. 25, 2003 Sheet 34 of 61 US 2003/0236225A1

CH3 183. L- O O H2N

HN )=NH 183. l-Arginine methyl ester, HCl H2N

196. OH H

196. Synephrine HO

198. H

198. N-Phenylethyldiamine cre,

199, ov'3CH O N-'3CH 199. N-Methylhomoveratrylamine H

200. H N 200. N-Allylcyclopentylamine Or's-ne,

208. pH N YCH

HO 208. Epinephrine OH

FIGURE 3(d) Patent Application Publication Dec. 25, 2003 Sheet 35 of 61 US 2003/0236225A1

209. mix of (+),(-), and meso CH3

HN --CH

209. Di-sec-butylamine (mix of (+), (-) and meso) H3C l

CH3 210.

210. Diisopropylamine H3C CH3s

212. cis-(1S,2R)-(–)-2-(Benzylamino)cyclohexanemethanol 212. cis-(1S,2R) H CC,N 213. cis-(1R,2S)-(+)-2-(Benzylamino)cyclohexanemethanol

213. cis-(1R,2S) H OC,N

223. 4-(Hexacylamino)benzylamine 223. H3C-1N1--1a1a1a1a1a-N Clue,

258. (3S(3a,4Ab),8A b)-N-t-butyl-D-ecahydro-3-isoquinolinecarboxamide

258.3S

FIGURE 3(e) Patent Application Publication Dec. 25, 2003 Sheet 36 of 61 US 2003/0236225A1

274. H N

274. Allylcyclohexyamine Crs

FIGURE 3(f) Patent Application Publication Dec. 25, 2003. Sheet 37 of 61 US 2003/0236225A1

Cyclic Secondary Amines

5. 4-Benzylpiperidine OO

14. 3-Piperidinemethanol

25. Tryptamine

26.

26. Morpholine

32. 4-Piperidinopiperidine O-O-

O O CH

N 40. Ethyl 1-piperazine carboxylate H

FIGURE 4(a) Patent Application Publication Dec. 25, 2003 Sheet 38 of 61 US 2003/0236225A1

41. rN,

CN D 41. 1-(2-Aminoethyl)piperazine H

58. H

58. Decahydroquinoline OC)

75.

75. 1,2,3,4-Tetrahydropyridoindole NH

80. 3-Amino-5-phenyl pyrazole

87. N N1 N-NH2 87. 3-Aminopyrazole --- 96a. C P F

96a, 1-(2-Fluorophenyl)piperazine r

FIGURE 4(b) Patent Application Publication Dec. 25, 2003 Sheet 39 of 61 US 2003/0236225A1

106. O H N -'3CH 106. 1-Proline methyl ester

107. H N OH

107. Histidinola (N 1. 2

114. 1N O y 114. 1-Piperonylpiperazine HNsu O

115. - -,NH 115. Hexamethyleimine H2N

122. H N

122. 4-Hydroxypiperidine OH

123. H N

123. 2-Piperidinemethanol Cre

FIGURE 4(c) Patent Application Publication Dec. 25, 2003. Sheet 40 of 61 US 2003/0236225A1

CH3 124.

HN CH3

124. 1,3,3-Trimethyl-6-azabicyclo[3.2.1]octane CH3

125. H

(a OH 125. 3-Pyrrolidinol

128. CH3 () N 128. 1-Methylpiperazine H 130. (S)-(+)y^r) 130. (S)-(+)-(2-Pyrolidinylmethyl)pyrrolidine

CH3 133.

133. 1-Methylhomopiperazine C)NH

135. O H N -CH3

135. Methylpipecolinate, HCl

139. CreN 139. 2-Ethylpiperidine FIGURE 4(d) Patent Application Publication Dec. 25, 2003 Sheet 41 of 61 US 2003/0236225A1

153.

153. 1,2,3,4-Tetrahydroisoquinoline

165. H N 165. Piperidine O

168. Nu1)NH 168. 1-(4-Fluorophenyl)piperazine O

173, d.l-Tryptophan methyl ester, HCl O

189, L- O H

189. 1-Histidine methyl ester, HCl N roNH2

195. tert-Butyl (1S,4S)-(–)-2,5-diazabicyclo[2.2.1]heptane-2-carboxylate

195. (1S,4S) HN

4 k.CH3

202.

202. Isonipecotamide

FIGURE 4(e) Patent Application Publication Dec. 25, 2003. Sheet 42 of 61 US 2003/0236225A1

H N

205. Heptamethyleneimine

215. H N

215. alpha-Methyltryptamine CH

217. H

217. 6-Fluoro-1,2,3,4-tetrahydro-2-methylquinoline F

218. 6,7-Dimethoxy 1,2,3,4-tetrahydroisoquinoline, HCl 218, O

HN N- O CH3

222. CHH CH3

rtNH2 222. 4-Amino-2,2,6,6-tetramethylpiperidine

224. NH

OH 224. 4-(-4-Chlorophenyl)-4-hydroxypiperidine Cl

FIGURE 4(f) Patent Application Publication Dec. 25, 2003. Sheet 43 of 61 US 2003/0236225A1

227.

227. 3-Aminopyrrollidine, dihCl NH

228. H

228. 3,5-Dimethylpiperidine (cis-and trans-) H3C JCl CH3 ?hH 237. 2,6-Dimethylmorpholine HC O CH

239. O

239. 1,4-Dioxo-8-azaspiro4.5decane HOXOO

244. 1.-Methyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline, HBr 244. CH3 HO NH

HO 248. 1,3,4,6,7,8-Hexahydro-2H-pyrido(1,2-A)pyrimidinc

H 248. CON N

250. 1,2,3,4-Tetrahydroquinoline C NY

FIGURE 4(g) Patent Application Publication Dec. 25, 2003. Sheet 44 of 61 US 2003/0236225A1

251. H N

251. 1-(2-Methoxyphenyl)piperazine XM

252. ro CD 252. 1-(2-(2-Hydroxyethoxy)ethyl)piperazine H

H 254 N 254. (S)-(+)-2-(Aminomethyl)pyrrolidine

258. (3S(3a,4Ab),8A b)-N-t-butyl-D-ecahydro-3-isoquinolinecarboxamide

258.3S

NH H CH "> 3 6H3C CH3

265. (R)-(+) N O

265. 3-Aminoquinonuclidine, diICI CNH2

270.

270. Homopiperazine CD

FIGURE 4(h) Patent Application Publication Dec. 25, 2003. Sheet 45 of 61 US 2003/0236225A1

27. H

stN 271. 2,6-Dimethylpiperazine H

273. Iminodibenzyl

H 275. C N 275. 5-Methoxytryptamine Hacs NH2

280.

280. 4,4'-Bipiperidine, HCl

286. ro CN 286. 1-(2-Hydroxyethyl)piperazine H

287.

287. 4-Methylpiperidine

FIGURE 4(i)

Patent Application Publication Dec. 25, 2003 Sheet 49 of 61 US 2003/0236225A1

8GTHQ5)I,H. Patent Application Publication Dec. 25, 2003 Sheet 50 of 61 US 2003/0236225A1

O s

wn s

v V

s 5.

O O O O O O L O O O D S. CN CN wn wn Spunoduoso Jo Jequn N Patent Application Publication Dec. 25, 2003 Sheet 51 of 61 US 2003/0236225A1

0ICHIRIQ?IH

Spunoduoso go equnN Patent Application Publication Dec. 25, 2003 Sheet 52 of 61 US 2003/0236225A1

CN N. CN

CO CO CN

O CN

wn is is . . cy) S \ E 2... - . . . gig's. . ; OLO w CD w .S. s w S un 2 2 92 5. -- s N C) E

(s CV) i. i* e3, N H O 9 ; 's : e”.g s OCY) C

9) ... :- 3. : *: sä N CS ...... a ri O O O r o r a. . . . co

D -

is . . ; v s "; w

O O LO O LO O CN CN v- v Senoelou emote eu) u Sueuouou eulue eu) JO epuenooC) Patent Application Publication Dec. 25, 2003. Sheet 53 of 61 US 2003/0236225A1

ZIGIRI[15)IH

Patent Application Publication Dec. 25, 2003. Sheet 54 of 61

Patent Application Publication Dec. 25, 2003 Sheet 55 of 61 US 2003/0236225A1

C fron M 6-0200934 - 2.82e.8 cps Mr lytaine 5e 8 2.5e --- NH2 cis-Myttanylamine2-Adamantantine 20e8 A?nino-1-proparot . -Methionino t Cydooctylamine 1.5e8 0.60 (1S,2S)-2-Amino-1-phenyi s 1.3-propandio se 1-Adamantanemethylamine Oe8 2,2-Diphenylethylamine 5-Amino-1-pentario

S.Oe 43 16S

O3 0.6 09 .2 S 8 Time, nin Spectrum from 0.34 min (4 scans) from MP 6-0200034 168e6 ces

g Cl co 2s s d

all. 8O 16 240 320 400 480 Z, at Spectrum from 0.34 mln (4 scans) from MP 6-0200034, subset 25e 6 cms - 286 --

Oe6

ses 8.095 W >. Goes- 217.0 ?

4-Oe5 24.50d V 325.6 2.Oe5 254.6 307.4 juk.k. W Anax s-V------220 240 260 280 300 320

FIGURE 14

Patent Application Publication Dec. 25, 2003 Sheet 57 of 61 US 2003/0236225A1

I, Lily 'kni: METHODS OF USE AND COMPOSITIONS FOR THE DIAGNOSI AND TREATMENT OF INFECTIOUS DISEASE Marina Nikolaevna Protopopova, Ph.D. May 17, 2002

o d rt rts s C A. E : o O

29S

999 --- 89 99. h L9 9900S800 9. 86.9 9. 68 98. 8. 0.8 SE:L ced 68 96. c 6069 i:866 / 9,202

s 6. "-- V 889 L.

08t. 86 65 8 S02. g-1e

Bid eIf,391 S. Patent Application Publication Dec. 25, 2003 Sheet 58 of 61 US 2003/0236225A1

N R22 Š

NNN 3

N 22 Š

s 3 9 Patent Application Publication Dec. 25, 2003 Sheet 59 of 61 US 2003/0236225A1

NSSSSSSSSSSSSSSSSSx is

S -NNNNNNNNNNN %22% S NNNNNNNN O ZS SS

22 OO SS SS CY) r 2 C P O N 9, 9

t S CN O CAD CY) C 8 SSSSSSSSSSSSSSSS O O K (5 O) N

P O t

s st C/O |-E: N s s s s s Patent Application Publication Dec. 25, 2003 Sheet 60 of 61 US 2003/0236225A1

G -arra 6 N2 a 9 S. d) s S2 g S25 S E stO 9S. S.as do. EC en- 5 to N. N. o a ch ?h in n n is 2 > a. CS Cd O O S. C S2Š2N. N Z, gld

IIII & s asses::& x8ss8:8::::::::::::::: &-3-3 &i. 33asix is 3.. *. * . .: .... :. . . . *.... . YYYYYYYYY R222222222222222222222222222 SNS

HIWWILLIVIIIAAA Nxxx xxyy R22 ENN

-HIWILL *s a- -, ~ x 88s ski - is AAAAAAA 222222222222222222222 SNN

*::::::::::::::::::::::HIWILLIHILHL * : ... , is 3: g: - RYANNA 22 N

IIIII NNNNNNNNNNNNNNNNNNNNNNNN. R2222222222222222222222222

O s 3 v Patent Application Publication Dec. 25, 2003. Sheet 61 of 61 US 2003/0236225A1

IIII

F IIII"SS. N.

& y

SSSQ c eaC

& Yit 2 5 SS 83

&- WWYY W. ?

Od V

- -- D O O D C O C g S. cy) cy CN CN va w Suo SeedSA US 2003/0236225A1 Dec. 25, 2003

METHODS OF USE AND COMPOSITIONS FOR as “Directly Observed Treatment Short-Course” (DOTS). THE DAGNOSIS AND TREATMENT OF For the eradication of TB, diagnosis, treatment, and preven INFECTIOUS DISEASE tion are equally important. Rapid detection of active TB patients will lead to early treatment by which about 90% FIELD OF INVENTION cure is expected. Therefore, early diagnosis is critical for the 0001. The present invention relates to methods and com battle against TB. In addition, therapeutic compliance will positions for treating disease caused by microorganisms, ensure not only elimination of infection, but also reduction particularly tuberculosis. The present invention also relates in the emergence of drug-resistance Strains. to methods and compositions having improved anti-myco 0006 The emergence of drug-resistant M. tuberculosis is bacterial activity, namely compositions comprising novel an extremely disturbing phenomenon. The rate of new TB Substituted ethylene diamine compounds. cases proven resistant to at least one Standard drug increased from 10 percent in the early 1980s to 23 percent in 1991. BACKGROUND OF THE INVENTION Compliance with therapeutic regimens, therefore, is also a 0002 Mycobacterial infections often manifest as diseases crucial component in efforts to eliminate TB and prevent the Such as tuberculosis. Human infections caused by mycobac emergence of drug resistant Strains. Equally important is the teria have been widespread since ancient times, and tuber development of new therapeutic agents that are effective as culosis remains a leading cause of death today. Although the vaccines, and as treatments, for disease caused by drug incidence of the disease declined, in parallel with advancing resistant Strains of mycobacteria. Standards of living, Since the mid-nineteenth century, myco 0007 Although over 37 species of mycobacteria have bacterial diseases still constitute a leading cause of morbid been identified, more than 95% of all human infections are ity and mortality in countries with limited medical caused by Six Species of mycobacteria: M. tuberculosis, M. resources. Additionally, mycobacterial diseases can cause avium intracellulare, M. kansasii, M. fortuitum, M. chelo overwhelming, disseminated disease in immunocompro nae, and M. leprae. The most prevalent mycobacterial mised patients. In Spite of the efforts of numerous health disease in humans is tuberculosis (TB) which is predomi organizations worldwide, the eradication of mycobacterial nantly caused by mycobacterial Species comprising M. diseases has never been achieved, nor is eradication immi tuberculosis, M. bovis, or M. africanum (Merck Manual nent. Nearly one third of the world's population is infected 1992). Infection is typically initiated by the inhalation of with mycobacterium tuberculosis complex, commonly infectious particles which are able to reach the terminal referred to as tuberculosis (TB), with approximately 8 pathways in lungs. Following engulfment by alveolar mac million new cases, and two to three million deaths attribut rophages, the bacilli are able to replicate freely, with even able to TB yearly. Tuberculosis (TB) is the cause of the tual destruction of the phagocytic cells. A cascade effect largest number of human deaths attributable to a single ensues wherein destruction of the phagocytic cells causes etiologic agent (see Dye et al., J. Am. Med. ASSociation, 282, additional macrophages and lymphocytes to migrate to the 677-686, (1999); and 2000 WHO/OMS Press Release). Site of infection, where they too are ultimately eliminated. 0003. After decades of decline, TB is now on the rise. In The disease is further disseminated during the initial Stages the United States, up to 10 million individuals are believed by the infected macrophages which travel to local lymph to be infected. Almost 28,000 new cases were reported in nodes, as well as into the blood Stream and other tissueS Such 1990, constituting a 9.4 percent increase over 1989. A as the bone marrow, Spleen, kidneyS, bone and central Sixteen percent increase in TB cases was observed from nervous system. (See Murray et al. Medical Microbiology, 1985 to 1990. Overcrowded living conditions and shared air The C. V. Mosby Company 219-230 (1990)). Spaces are especially conducive to the spread of TB, con 0008. There is still no clear understanding of the factors tributing to the increase in instances that have been observed which contribute to the virulence of mycobacteria. Many among prison inmates, and among the homeless in larger investigators have implicated lipids of the cell wall and U.S. cities. Approximately half of all patients with bacterial Surface as contributors to colony morphology and “Acquired Immune Deficiency Syndrome' (AIDS) will Virulence. Evidence Suggests that C-mycosides, on the Sur acquire a mycobacterial infection, with TB being an espe face of certain mycobacterial cells, are important in facili cially devastating complication. AIDS patients are at higher tating Survival of the organism within macrophageS. Treha risks of developing clinical TB, and anti-TB treatment seems lose 6,6' dimycolate, a cord factor, has been implicated for to be less effective than in non-AIDS patients. Consequently, other mycobacteria. the infection often progresses to a fatal disseminated disease. 0009. The interrelationship of colony morphology and 0004 Mycobacteria other than M. tuberculosis are Virulence is particularly pronounced in M. avium. M. avium increasingly found in opportunistic infections that plague the bacilli occur in several distinct colony forms. Bacilli which AIDS patient. Organisms from the M. avium-intracellulare grow as transparent, or rough, colonies on conventional complex (MAC), especially Serotypes four and eight, laboratory media are multiplicable within macrophages in account for 68% of the mycobacterial isolates from AIDS tissue culture, are virulent when injected into Susceptible patients. Enormous numbers of MAC are found (up to 10' mice, and are resistant to antibiotics. Rough or transparent acid-fast bacilli per gram of tissue), and consequently, the bacilli, which are maintained on laboratory culture media, prognosis for the infected AIDS patient is poor. often Spontaneously assume an opaque R colony morphol 0005) The World Health Organization (WHO) continues ogy, at which time they are not multiplicable in macroph to encourage the battle against TB, recommending preven ages, are avirulent in mice, and are highly Susceptible to tion initiatives Such as the “Expanded Program on Immu antibiotics. The differences in colony morphology between nization” (EPI), and therapeutic compliance initiatives Such the transparent, rough and opaque Strains of M. avium are US 2003/0236225A1 Dec. 25, 2003

almost certainly due to the presence of a glycolipid coating treatment is not continued to completion, however, the on the Surface of transparent and rough organisms which patient may experience a relapse, and the relapse rate for acts as a protective capsule. This capsule, or coating, is patients who do not continue treatment to completion is composed primarily of C-mycosides which apparently high. A variety of forms of patient-centered care are used to Shield the virulent M. avium organisms from lysosomal promote adherence with therapy. The most effective way of enzymes and antibiotics. By contrast, the non-virulent ensuring that patients are taking their medication is to use opaque forms of M. avium have very little C-mycoside on directly observed therapy, which involves having a member their Surface. Both the resistance to antibiotics and the of the health care team observe the patient take each dose of resistance to killing by macrophages have been attributed to each drug. Directly observed therapy can be provided in the the glycolipid barrier on the Surface of M. avium. clinic, the patient's residence, or any mutually agreed upon site. Nearly all patients who have tuberculosis caused by 0.010 Diagnosis of mycobacterial infection is confirmed drug-Sensitive organisms, and who complete therapy will be by the isolation and identification of the pathogen, although cured, and the risk of relapse is very low ("Ending Neglect: conventional diagnosis is based on Sputum Smears, chest The Elimination of Tuberculosis in the United States' ed. L. X-ray examination (CXR), and clinical Symptoms. Isolation Geiter Committee on the Elimination of Tuberculosis in the of mycobacteria on a medium takes as long as four to eight United States Division of Health Promotion and Disease weeks. Species identification takes a further two weeks. There are Several other techniques for detecting mycobac Prevention, Institute of Medicine. Unpublished.) teria Such as the polymerase chain reaction (PCR), myco 0014 What is needed are effective therapeutic regimens bacterium tuberculosis direct test, or amplified mycobacte that include improved vaccination and treatment protocols. rium tuberculosis direct test (MTD), and detection assays Currently available therapeutics are no longer consistently that utilize radioactive labels. effective as a result of the problems with treatment compli ance, and these compliance problems contribute to the 0.011) One diagnostic test that is widely used for detecting development of drug resistant mycobacterial Strains. infections caused by M. tuberculosis is the tuberculin skin test. Although numerous versions of the skin test are avail 0.015 Ethambutol (EMB) is a widely used antibiotic for able, typically one of two preparations of tuberculin antigens the treatment of TB, with over 300 million doses delivered are used: old tuberculin (OT), or purified protein derivative for tuberculosis therapy in 1988. (PPD). The antigen preparation is either injected into the skin intradermally, or is topically applied and is then inva Sively transported into the Skin with the use of a multiprong OH inoculator (Tine test). Several problems exist with the skin test diagnosis method. For example, the Tine test is not HN generally recommended because the amount of antigen n1 NH injected into the intradermal layer cannot be accurately controlled. (See Murray et al. Medical Microbiology, The C. HO V. Mosby Company 219–230 (1990)). Ethambutol 0012 Although the tuberculin skin tests are widely used, they typically require two to three days to generate results, and many times, the results are inaccurate Since false posi 0016 Ethambutol, developed by Lederle Laboratories in tives are Sometimes Seen in Subjects who have been exposed the 1950s, has low toxicity and is a good pharmacokinetic. to mycobacteria, but are healthy. In addition, instances of However, ethambutol has a relatively high Minimum Inhi mis-diagnosis are frequent Since a positive result is observed bition Concentration (MIC) of about 5ug/ml, and can cause not only in active TB patients, but also in perSons vaccinated optic neuritis. Thus, there is an increasing need for new, and with Bacille Calmette-Guerin (BCG), and those who had more effective, therapeutic compositions (See for example, been infected with mycobacteria, but have not developed the U.S. Pat. No. 3,176,040, U.S. Pat. No. 4,262,122; U.S. Pat. disease. It is hard therefore, to distinguish active TB patients No. 4,006,234; U.S. Pat. No. 3,931,157; U.S. Pat. No. from the others, such as household TB contacts, by the 3,931,152; U.S. Re. No. 29.358; and Hausler et al., Bioor tuberculin skin test. Additionally, the tuberculin test often ganic & Medicinal Chemistry Letters 11 (2001) 1679-1681). produces a croSS-reaction in those individuals who were In the decoder years since the discovery of the beneficial infected with mycobacteria other than M. tuberculosis effects of ethambutol, few pharmacological advances in TB (MOTT). Therefore, diagnosis using the skin tests currently treatment have been developed. Moreover, with the com available is frequently Subject to error and inaccuracies. bined emergence of drug resistant Strains, and the more prevalent spread of mycobacterial disease, it is becoming 0013 The standard treatment for tuberculosis caused by drug-Sensitive organisms is a six-month regimen consisting Seriously apparent that new therapeutic compositions are of four drugs given for two months, followed by two drugs crucial in the fight against tuberculosis. given for four months. The two most important drugs, given 0017 Clearly effective therapeutic regimens that include throughout the Six-month course of therapy, are isoniazid improved vaccination and treatment protocols are needed. A and rifampin. Although the regimen is relatively simple, its therapeutic Vaccine that would prevent the onset of tuber administration is quite complicated. Daily ingestion of eight culosis, and therefore eliminate the need for therapy is or nine pills is often required during the first phase of desirable. Although currently available therapeutics Such as therapy; a daunting and confusing prospect. Even Severely ethambutol are effective, the emergence of drug resistant ill patients are often Symptom free within a few weeks, and Strains has necessitated new formulations and compositions nearly all appear to be cured within a few months. If the that are more versatile than ethambutol. Currently available US 2003/0236225A1 Dec. 25, 2003 therapeutics are no longer consistently effective as a result of compositions comprise Substituted ethylene diamines, the problems with treatment compliance, lending to the wherein one amine group is derived from a primary amine, development of drug resistant mycobacterial Strains. What is and wherein the other amine group is derived from a primary needed are new anti-tubercular drugs that provide highly or Secondary amine. In another embodiment of the present effective treatment, and Shortens or simplifies tuberculosis invention, the methods and compositions comprise Substi chemotherapy. tuted ethylene diamines, wherein one amine is derived from cis-(-)myrtanylamine, cyclooctylamine, 2,2-diphenylethy SUMMARY OF THE INVENTION lamine, 3,3-diphenylpropylamine, (+)-bornylamine, 1-ada mantanemethylamine, (+)-isopinocampheylamine; or isopi 0.018. The present invention comprises methods and nocampheylamine. compositions for the treatment of microorganisms, particu larly methods and compositions comprising ethylene 0022. The present invention contemplates various salt diamine compounds effective in the treatment of infectious complexes and other substituted derivatives of the substi organisms. The present invention also provides methods and tuted ethylene diamines. The present invention also contem compositions comprising Substituted ethylene diamines hav plates enantiomers and other Stereoisomers of the Substituted ing improved anti-mycobacterial activity, including Substi ethylene diamines and their substituted derivatives. The tuted ethylene diamines having improved anti-tuberculosis present invention further contemplates treatment for ani activity. mals, including, but not limited to, humans. 0019. The present invention contemplates substituted 0023. Accordingly, it is an object of the present invention ethylene diamines, which can derive from a variety of amine to provide methods and compositions for the treatment and compounds. In the present invention, the Substituted ethyl prevention of diseases caused by microorganisms. ene diamines are based on the following structure. 0024. Accordingly, it is an object of the present invention to provide methods and compositions for the treatment and prevention of infectious diseases. R4 0025. Another object of the present invention is to pro vide methods and compositions for the treatment and pre RNulls NRR vention of mycobacterial disease, including but not limited to, tuberculosis. Substituted Ethylene Diamine 0026. Yet another object of the present invention is to provide methods and compositions for the treatment and 0020. The Substituted ethylene diamine compounds prevention of infectious diseases using compositions com described herein are Synthesized and Screened for activity as prising Substituted ethylene diamines. follows. A chemical library of substituted ethylene diamines 0027. Another object of the present invention is to pro is prepared on a Solid polystyrene Support using Split and vide methods and compositions for the treatment and pre pool technologies. This technique allows for the Synthesis of vention of mycobacterial disease using compositions com a diverse set of Substituted ethylene diamines. These prising Substituted ethylene diamines. diamines are Screened for anti-TB activity using in vitro, biological assays, including a High-Throughput Screening 0028 Still another object of the present invention is to (HTS) assay, based on the recently completed genomic provide methods and compositions for the treatment and sequence of M tuberculosis, and a Minimum Inhibition prevention of tuberculosis using compositions comprising Concentration (MIC) assay. Substituted ethylene diamines. 0021. The methods and compositions described herein 0029. Another object of the present invention is to pro comprise Substituted ethylene diamines that are effective vide methods and compositions for the treatment and pre against disease caused by microorganisms, but not limited to vention of tuberculosis using compositions comprising Sub bacterial infection. One embodiment of the invention pro stituted ethylene diamines, wherein the diamine has an MIC vides methods and compositions comprising Substituted of 50 uM, or less. ethylene diamines that are effective against mycobacterial 0030. Another object of the present invention is to pro disease. Another embodiment of the invention provides vide methods and compositions for the treatment and pre methods and compositions comprising Substituted ethylene vention of tuberculosis using compositions comprising Sub diamines that have MIC of 50 uM or lower for mycobac stituted ethylene diamines, wherein the diamine has an MIC terial disease. Another embodiment of the present invention of 25 uM, or less. comprises substituted ethylene diamines that have an MIC of 25 uM or lower for mycobacterial disease. Yet another 0031) Another object of the present invention is to pro embodiment of the present invention comprises Substituted vide methods and compositions for the treatment and pre ethylene diamines that have an MIC of 12.5 uM or lower for vention of tuberculosis using compositions comprising Sub mycobacterial disease. Another embodiment of the present stituted ethylene diamines, wherein the diamine has an MIC invention comprises Substituted ethylene diamines that have of 12.5 uM, or less. an MIC of 5 uM or lower for mycobacterial disease In 0032. Yet another object of the present invention is to another embodiment of the present invention, the methods provide methods and compositions for the treatment and and compositions comprise Substituted ethylene diamines prevention of tuberculosis using compositions comprising with HTS Luc activity of 10% or greater. In yet another Substituted ethylene diamines, wherein the diamine has an embodiment of the present invention, the methods and MIC of 5 uM, or less. US 2003/0236225A1 Dec. 25, 2003

0.033 Yet another object of the present invention is to 0048 FIG. 10 is a bar graph providing a summary of provide methods and compositions for the treatment and Luciferase activity of discrete Substituted ethylene diamines prevention of tuberculosis using compositions comprising with at least 10% activity in reference to ethambutol at 3.1 Substituted ethylene diamines, wherein the diamine has luM. HTS/Luc activity of 10% or greater. 0049 FIG. 11 is a bar graph showing the frequency of 0034. Another object of the present invention is to pro occurrences of the Selected amine monomers in the Substi vide methods and compositions for the treatment and pre tuted ethylene diamine compounds that were active against vention of tuberculosis using compositions comprising Sub TB. Amine monomers are represented by their numerical Stituted ethylene diamines, wherein one amine group is designations. derived from a primary amine, and the other amine group is 0050 FIG. 12 represents a flow schematic showing a derived from a primary or Secondary amine. synthesis of N-Geranyl-N'-(2-adamanthyl)ethane-1,2-di 0.035 Yet another object of the present invention is to amine (compound 109). provide methods and compositions for the treatment and/or 0051 FIG. 13 is a flow schematic showing a synthesis of prevention of tuberculosis using compositions comprising N-(Cyclooctyl)-N'-(1R,2R,3R,5S)-(–)-isopinocampheyle Substituted ethylene diamines, wherein one amine is derived from cis-(-)myrtanylamine, cyclooctylamine, 2,2-diphenyl thane-1,2-diamine as hydrochloride (compound 59). ethylamine, 3,3-diphenylpropylamine, (+)-bornylamine, 0052 FIG. 14 is a mass spec profile for one representa 1-adamantanemethylamine, (+)-isopinocampheylamine; or tive Sample well containing pooled Substituted ethylene (-)-isopinocampheylamine. diamine compounds. 0.036 Yet another object of the present invention is to 0053 FIG. 15 is a mass spec profile for compound 109, provide composition for the therapeutic formulation for the N-Geranyl-N'-(2-adamanthyl) ethane-1,2-diamine. treatment and prevention of mycobacterial disease. 0054 FIG. 16 is a proton NMR profile for compound 0037 Another object of the present invention is to pro 109, N-Geranyl-N'-(2-adamanthyl) ethane-1,2-diamine. vide compositions for therapeutic formulations for the treat ment and prevention of mycobacterial disease caused by 0055 FIG. 17 is a bar graph of data from a Colony mycobacterial Species comprising M. tuberculosis complex, Forming Units/Lung (CFU/Lung) study showing CFU/Lung M. avium intracellulare, M. kansarii, M. fortuitum, M. growth over time in days for various compounds. chelonoe, M. leprae, M. africanum, M. microti, or M. bovis. 0056 FIG. 18 is a bar graph of data from a CFU/Lung 0.038. These and other objects, features and advantages of study showing CFU/Lung growth over time in days for the present invention will become apparent after a review of various compounds. the following detailed description of the disclosed embodi 0057 FIG. 19 is a bar graph of data from a CFU/Lung ments and the appended claims. study showing CFU/Lung growth over time in days for various compounds. BRIEF DESCRIPTION OF THE FIGURES 0.058 FIG. 20 is a bar graph of data from a lesion study 0.039 FIG. 1 represents a flow schematic showing vari showing visible lesions over time after treatment with vari ous Solid Support Syntheses used to prepare Substituted ous compounds. ethylene diamines. DETAILED DESCRIPTION 0040 FIG. 2 provides chemical structures of a variety of primary amines. 0059. The present invention may be understood more readily by reference to the following detailed description of 0041 FIG. 3 provides chemical structures of a variety of the specific embodiments included herein. However, acyclic Secondary amines. although the present invention has been described with 0.042 FIG. 4 provides chemical structures of a variety of reference to Specific details of certain embodiments thereof, cyclic Secondary amines. it is not intended that Such details should be regarded as limitations upon the Scope of the invention. 0.043 FIG. 5 represents a flow schematic for a represen tative reaction pool of ten Substituted ethylene diamines. 0060 Mycobacterial infections, such as those causing tuberculosis, once thought to be declining in occurrence, 0044 FIG. 6 is a graph of Luminescence Count per have rebounded, and again constitute a Serious health threat. Second (LCPS) versus concentration showing HTS Luc Tuberculosis (TB) is the cause of the largest number of assay results for pooled Substituted ethylene diamine com human deaths attributed to a single etiologic agent with two pounds. to three million people infected with tuberculosis dying each 004.5 FIG. 7 is a graph of LCPS versus concentration year. Areas where humans are crowded together, or living in showing HTS Luc assay results for individual substituted Substandard housing, are increasingly found to have perSons ethylene diamine compounds. affected with mycobacteria. Individuals who are immuno compromised are at great risk of being infected with myco 0046 FIG. 8 is a graph of LCPS versus concentration bacteria and dying from Such infection. In addition, the showing HTS Luc assay results for individual substituted emergence of drug-resistant Strains of mycobacteria has led ethylene diamine compounds. to treatment problems of Such infected perSons. 0047 FIG. 9 is a bar graph providing a summary of MIC 0061 Many people who are infected with mycobacteria activities for discrete Substituted ethylene diamines. are poor, or live in areas with inadequate healthcare facili US 2003/0236225A1 Dec. 25, 2003 ties. As a result of various obstacles (economical, education animals. For example, the present invention may be particu levels, etc.), many of these individuals are unable to comply larly useful for the treatment of cows infected by M. bovis. with the prescribed therapeutic regimens. Ultimately, per Sistent non-compliance by these and other individuals results 0066. As used herein, the term “tuberculosis” comprises in the prevalence of disease. This noncompliance is fre disease States usually associated with infections caused by quently compounded by the emergence of drug-resistant mycobacteria Species comprising M. tuberculosis complex. Strains of mycobacteria. Effective compositions and vac The term “tuberculosis” is also associated with mycobacte cines that target various Strains of mycobacteria are neces rial infections caused by mycobacteria other than M. tuber Sary to bring the increasing number of tuberculosis cases culosis (MOTT). Other mycobacterial species include M. under control. avium-intracellulare, M. kansarii, M. fortuitum, M. chelo nae, M. leprae, M. africanum, or M. microti. 0.062 Chemotherapy is a standard treatment for tubercu losis. Some current chemotherapy treatments require the use 0067. The anti-infective methods and compositions of the of three or four drugs, in combination, administered daily for present invention contain one or more Substituted ethylene two months, or administered biweekly for four to twelve diamine compounds. In particular, these compounds encom months. Table 1 lists several treatment schedules for stan pass a wide range of Substituted ethylene diamine com dard tuberculosis drug regimens. pounds having the following general formula:

TABLE 1. Treatment Schedules for Standard TB Drug Regimens. INDUCTION STANDARD PHASE CONTINUATION DRUG Dosing PHASE REGIMEN Schedule DURATION DRUG Dosing Schedule DURATION Isoniazid Daily, DOT 8 weeks Isoniazid 2/week, DOT 16 weeks Rifampicin Daily, DOT 8 weeks Rifampien 2/week, DOT 16 weeks Pyrazinamide Daily, DOT 8 weeks Ethambutol or Daily, DOT 8 weeks Streptomycin

0.063 Decades of misuse of existing antibiotics and poor compliance with prolong and complex therapeutic regimens has led to mutations of the mycobacterium tuberculosis and R4 has created an epidemic of drug resistance that threatens tuberculosis control world wide. The vast majority of cur RNulls NR2R3 rently prescribed drugs, including the front line drugs, Such as isoniazid, rifampin, pyrazinamide, ethambutol and Strep Substituted Ethylene Diamine tomycin were developed from the 1950s to the 1970s. Thus, this earlier development of tuberculosis chemotherapy did 0068 where “RNH” is typically derived from a primary not have at its disposal the implications of the genome amine, and “RRN” is typically derived from a primary or Sequence of Mycobacterium tuberculosis, the revolution in Secondary amine. The ethylene diamines of the present pharmaceutical drug discovery of the last decades, and the invention are prepared by a modular approach using primary use of national drug testing and combinational chemistry. and Secondary amines as building blocks, and coupling the amine moieties with an ethylene linker building block. 0.064 Consequently, the treatments of drug-resistant M. Representative primary amines, acyclic Secondary amines, tuberculosis Strains, and latent tuberculosis infections, and cyclic secondary amines are shown in FIGS. 2, 3, and require new anti-tuberculosis drugs that provide highly 4, respectively. effective treatments, and Shortened and Simplified tubercu 0069 Generally, chemical moieties R, R2, and R of the losis chemotherapies. Moreover, it is desirable that these ethylene diamine compounds of the present invention are drugs be prepared by a low-cost Synthesis, Since the demo independently Selected from H, alkyl, aryl; alkenyl; alkynyl, graphics of the disease dictate that cost is a Significant factor. aralkyl, aralkenyl, aralkynyl, cycloalkyl, cycloalkenyl; het eroalkyl, heteroaryl; halide, alkoxy, aryloxy; alkylthio; 0065. The present invention provides methods and com arylthio; Sillyl, Siloxy; a disulfide group; a urea group, amino; positions comprising a class of Substituted ethylene diamine and the like, including Straight or branched chain derivatives compounds effective in treatment and prevention of disease thereof, cyclic derivatives thereof, substituted derivatives caused by microorganisms including, but not limited to, thereof, heteroatom derivatives thereof, heterocyclic deriva bacteria. In particular, the methods and compositions of the tives thereof, functionalized derivatives thereof, salts present invention are effective in inhibiting the growth of the thereof, Such Salts including, but not limited to hydrochlo microorganism, M. tuberculosis. The methods and compo rides and acetates, isomers thereof, or combinations thereof. Sitions of the present invention are intended for the treatment For example, nitrogen-containing heterocyclic moieties of mycobacteria infections in human, as well as other include, but are not limited to, groupS. Such as pyridinyl US 2003/0236225A1 Dec. 25, 2003

(derived from pyridine, and bonded through a ring carbon), peridinopiperidine, ethyl 1-piperazine carboxylate, 1-(2- piperidinyl (derived from piperidine and bonded through the amino-ethyl)-piperazine; decahydroquinoline; 1,2,3,4- ring nitrogen atom or a ring carbon), and pyrrolidinyl tetrahydro-pyridoindole (reaction at either amine); 3-amino (derived from pyrrolidine and bonded through the ring 5-phenyl pyrazole, 3-aminopyrazole, 1-(2-fluorophenyl) nitrogen atom or a ring carbon). Examples of Substituted, or piperazine, 1-proline methyl ester; histidinol, 1-piperonyl functionalized, derivatives of R, R, and R include, but are piperazine; hexamethyleimine, 4-hydroxypiperidine, 2-pip not limited to, moieties containing Substituents Such as acyl, eridinemethanol; 1,3,3-trimethyl-6-azabicyclo3.2.1 formyl, hydroxy, acyl halide, amide, amino, azido, acid, octane; 3-pyrrolidinol, 1-methylpiperazine, (S)-(+)-(2-pyro alkoxy, aryloxy, halide, carbonyl, ether, ester, thioether, lidinylmethyl) pyrrolidine, 1-methylhomopiperazine; thioester, nitrile, alkylthio, arythio, Sulfonic acid and Salts 2-ethyl-piperidine, 1,2,3,4-tetrahydroisoquinoline, 1-(4- thereof, thiol, alkenyl, alkynyl, nitro, imine, imide, alkyl, fluorophenyl) piperazine; d.l-tryptophan methyl ester; tert aryl, combinations thereof, and the like. Moreover, in the butyl (15, 45)-(-)-2,5-diazabiclyclo[2.2.1 heptane-2-car case of alkylated derivatives of the recited moieties, the boxylate; isonipecotamide, heptamethyleneimine; alpha alkyl Substituent may be pendant to the recited chemical methyltryptamine; 6,7-dimethoxy-1,2,3,4- moiety, or used for bonding to the amine nitrogen through tetrahydroisoquinoline; 3-aminopyrrolidine, 3,5- the alkyl Substituent. dimethylpiperidine, 2,6-dimethylmorpholine, 1,4-dioxo-8- azaspiro4.5decane; 1-methol-6,7-dihydroxy-1,2,3,4- 0070) Examples of chemical moieties R., R., and Ra of tetrahydroisoquinoline; 1,3,4,6,7,8-hexahydro-2H-pyrido the present invention include, but are not limited to: H; methyl, ethyl, propyl; butyl, pentyl; hexyl, heptyl; octyl, (1,2-A) pyrimidine, 1,2,3,4-tetrahydroquinoline, 1-(2-meth ethenyl; propenyl; butenyl, ethynyl, propynyl; butynyl, oxyphenyl) piperazine, 1-(2-(2-hydroxyethoxy)ethyl) pip cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclooc erazine; (S)-(+)-2-(aminomethyl) pyrroli-dine; (3S(3a,4Ab), tyl cyclobutenyl, cyclopentenyl, cyclohexenyl; phenyl; 8Ab)-N-t-butyl-D-ecahydro-3-isoquino-linecarboxamide; tolyl; Xylyl; benzyl, naphthyl, pyridinyl; furanyl, tetrahydro (R)-cycloSerine; homopiperazine; 2,6-dimethylpiperazine 1-napthyl, piperidinyl; indolyl; indolinyl, pyrrolidinyl; (reaction at either amine); iminodibenzyl, 5-methox 2-(methoxymethyl) pyrrolidinyl, piperazinyl, quinolinyl; ytryptamine; 4,4'-bipiperidine, 1-(2-hydroxyethyl) pipera quinolyl; alkylated-1,3-dioxolane; triazinyl; morpholinyl; Zine, 4-methylpiperidine, 1-histidine methyl ester; or methyl phenyl pyrazolyl; indanyl; indonyl, pyrazolyl; thiadiazolyl, pipecoliate. rhodaninyl; thiolactonyl, dibenzofuranyl; benzothiazolyl, 0073. The RHN substituent is derived from a primary homopiperidinyl; thiazolyl; quinonuclidinyl, isoxazolidi amine. The RRN substituent is typically derived from a nonyl; any isomers, derivatives, or Substituted analogs primary or Secondary amine, but may also arise from an thereof, or any Substituted or unsubstituted chemical Species amino acid, or an amino acid precursor. The amino acid can Such as alcohol, ether, thiol, thioether, tertiary amine, Sec transform into an amino alcohol. When an amino acid is ondary amine, primary amine, ester, thioester, carboxylic employed as the Source of the RRN moiety, the precursor acid, diol, diester, acrylic acid, acrylic ester, methionine compound may be Selected from, among others, the follow ethyl ester, benzyl-1-cysteine ethyl ester, imine, aldehyde, ing compounds and their derivatives: d.l-tryptophan methyl ketone, amide, or diene. Further examples of chemical ester, 1-methionine ethyl ester, 1-lysine methyl ester (via moieties R, R2, and R of the present invention include, but reaction at either primary amine), (S)-benzyl-1-cysteine are not limited to, the following Species or Substituted or ethyl ester, 1-arginine methyl ester (via reaction at either alkylated derivatives of the following species, covalently primary amine), 1-glutamic acid ethyl ester, 1-histidine bonded to the amine nitrogen: furan, tetrahydrofuran; methyl ester; or (3S(3a,4Ab),8Ab)-N-t-butyl-D-ecahydro-3- indole; piperazine, pyrrolidine, pyrrolidinone, pyridine, iso-quino linecarboxamide. quinoline, anthracene, tetrahydroquinoline; naphthalene; 0074 The R moiety of the substituted ethylene diamine pyrazole; imidazole; thiophene, pyrrolidine, morpholine; compounds of the present invention is typically Selected and the like. One feature of the recited species or substituted from H, alkyl or aryl, but R can also constitute alkenyl, or alkylated derivatives of these species, is that they may be alkynyl, aralkyl, aralkenyl, aralkynyl, cycloalkyl, cycloalk covalently bonded to the amine nitrogen in any fashion, enyl, and the like. Examples of the R chemical moiety including through the pendant Substituent or alkyl group, include, but are not limited to: H; methyl, ethyl, propyl; through the heteroatom as appropriate, or through a ring butyl, pentyl; hexyl, heptyl; octyl, ethenyl, propenyl; bute atom as appropriate, as understood by one of ordinary skill nyl, ethynyl; propynyl; butynyl, cyclobutyl, cyclopentyl; in the art. cyclohexyl, cyclobutenyl, cyclopentenyl, cyclohexenyl, 0071. The chemical moieties R, R2, and Ra of the phenyl, tolyl, Xylyl; benzyl, naphthyl, Straight or branched present invention also include, but are not limited to, cyclic chain derivatives thereof; cyclic derivatives thereof; substi alkanes and cyclic alkenes, and include bridged and non tuted, functionalized, and heteroatom derivatives thereof; bridged rings. Examples of bridged rings include, but are not and heterocyclic derivatives thereof, and the like. Typically, limited to, the following groups: isopinocamphenyl; bornyl; R is selected from H., methyl, ethyl, butyl or phenyl. norbornyl; adamantanetetyl, cis-(-)myrtanyl; adamantyl; However, when R is “H” the ethylene diamine does not noradamantyl, 6-azabicyclo3.2.1]octane; exo-norbornane; contain ethambutol. and the like. 0075. A majority of the ethylene diamine compounds 0.072 In one embodiment of the present invention, described hrein are preferably prepared using a Solid Support NRRs is derived from a cyclic secondary amine. Examples Synthesis, as Set forth in one of the representative reaction of a cyclic chemical moiety, NRR, of the present invention schemes shown in FIG. 1. However, when R is H, the include, but are not limited to, 4-benzyl-piperidine, 3-pip reaction does not proceed well when Sterically hindered eridinemethanol; piperidine, tryptamine; moropholine, 4-pi amines are used for RNH, or when diamines, Such as US 2003/0236225A1 Dec. 25, 2003 amino alkylenemorpholine, or aminoalkylene-piperidines, Simple amino acids, to allow for greater diversity in the are used for RNH. When R is methyl, or phenyl, sterically building-block monomers. The first three steps of each hindered amines used for RRNH do not work well due to Support Synthesis: the activation of the Rink-acid resin, the Steric hindrance at the reaction Site. In this case, a competing addition of the first amine, and the acylation Step are carried hydrolysis reaction producing the corresponding amino out in 10 ml tubes on a QUESTOR 210 Synthesizer manu alcohols, and incomplete reduction of the amidoethylene factured by ARGONAUTTECHNOLOGIES(R), Inc., Foster amines, interfere with the reaction Scheme. As a result, the City, Calif. The synthesizer handles up to twenty simulta desired diamine products form in low yields. neous reactions in 5 ml or 10 ml reaction vessels to allow for 0.076 The preparation of the ethylene diamines is pref rapid Synthesis of target compounds. The Synthesizer pro erably accomplished in Six Steps, using a rink-acid resin. The vides programmable temperature control and agitation, and first Step of the Synthesis is converting the rink-acid resin to the automated delivery of Solvents into the reaction vessels. rink-chloride by treatment with triphenylphosphine and The addition of the second amine, the reduction with Red hexachloroethane in tetrahydrofuran (THF). This step is Al, and the cleavage from the Solid Support are carried out followed by addition of the primary amine in the presence of in 2 ml wells in a 96-well, chemically resistant plate. Hunig's base (EtN(i-Pr).) in dichloroethane. The third step 0080 Prior to the solid support synthesis, each amine, is the acylation of the resin-attached amine using either one within numbers 1 to 288, as shown in FIGS. 2, 3, and 4, is of the two acylation routes shown in FIG. 1. The acylation dissolved in DMF as a one molar Solution, and organized in Step is preferably accomplished using either chloroacetyl three, 96-well plates (one amine per well), to yield three chloride, C.-bromo-O-methyl acetylbromide, C.-bromo-C.- master plates of these amines. An individual haloacetyl ethylacetyl bromide, C.-bromo-C.-butyl acetylbromide, or amide from each primary amine and a particular R group, C-chloro-O-phenyl-acetylchloride, each in the presence of is formed in the first three Steps of the Support Synthesis. pyridine in THF. Other acylation reagents known to those Individual haloacetyl amides are then pooled into groups of skilled in the art may also be used, however, the C-bro ten or thirty. A Suspension of the pooled resins in a 2:1 moacetyl halides result in low product yields, which may be mixture of DCM/THF is evenly distributed into one, two or attributed to HBr elimination. The acylation may also be three reaction plates to assure 15-20 mg of the Suspension accomplished via a peptide coupling mechanism using per well. The number of reaction plates used is based on the C.-bromo-C.-methylacetic acid, or C-chloro-O-methylacetic amount of Suspension available. Each well of pooled resins acid, in the presence of benzotriazole-1-yl-oxy-tris-pyrroli is reacted with a corresponding amine from the master dino-phosphonium hexafluorophosphate (PyBrop) and N,N- plates. FIG. 5 provides a flow schematic for a representative diisopropylethyl amine (EtN(i-Pr)) in dichloromethane pool. Each reaction occurs in a separate well, in the presence (DCM) and dimethylformamide (DMF). Again, other acy of Hunig's base in DMF at 70-75° C. for 16-20 hours. Each lation reagents known to those skilled in the art may also be resulting amine-amide is reduced using 65+w % Red-Al at used. The acylation Step is preferably performed twice to room temperature. The reduction is followed by cleavage achieve better acylated product yields. with 10% vol. TFA in DCM. The Solvents in each reaction 0.077 Introduction of the second nitrogen moiety is pref well are evaporated, and the TFA salts of the diamines erably achieved in the presence of Hunig's base in dimeth analyzed (mass spec), and Screened against M. tuberculosis. ylformamide (DMF). Reduction of the intermediate amine One plate of pooled diamines are Screened against M. amide is carried out using Red-Al (3.4M solution of sodium Smegmatis. Two randomly Selected rows in each plate; i.e., bis (2-methoxyethoxy) aluminum hydride in toluene). The 24 samples per 96-well plate, or 25% of the library, are final product is cleaved from the resin Support using a 10% examined by mass SpectroScopy. Specific protocols and solution (by volume) of trifluoroacetic acid (TFA) in dichlo detailed methods are provided below in the Examples. romethane (DCM). The solvent is evaporated, and the TFA Salts of the final diamine products are analyzed by mass 0081 Screening Against M. tuberculosis Spec, and Screened against M. tuberculosis for effectiveness. 0082) An entire library of synthesized substituted ethyl Some of the Substituted ethylene diamines, prepared using ene diamines (targeted number of compounds about 100, the above-described Solid-Support Synthesis, are also pre 000), prepared as described above, was Screened, in vitro, pared using a Solution phase Synthesis described below. against M. tuberculosis in ethambutol (EMB) sensitive Luc 0078 Formation of the Substituted Ethylene Diamine assay. The MIC (Minimum Inhibition Concentration) was Library also determined. The MIC is the concentration of a growth inhibitor, here the substituted ethylene diamine, in which 0079 The solid support syntheses, shown in FIG. 1, are there is no multiplication of the microorganism under preferably used to prepare a Substituted ethylene diamine examination. Screening was done using a High-Throughout library. Solid phase Synthesis offers at least three principal Screening (HTS) Luc assay with recombinant mycobacteria advantages: (i) a reduced need for chromatographic proce containing a promoter fusion of a luciferase to the EB dures, (ii) the use of excess reagents to drive a reaction inducible gene (Luc assay). The Luc-assay and MIC assay forward in high yields, and (iii) the use of Split and pool are described in detail below. These assays are well known technologies for the Synthesis of a large number of com to those skilled in the art. Based on this initial Screening, pounds. Solid Support Syntheses of 1,2-diamine libraries 300+ compound mixtures showed anti-TB activity. FIG. 6 have previously been accomplished by the reduction of short represents typical assay data in a luciferase reporter Strain peptides (Cuervo et al., Peptides 1994: Proceedings of the containing an RV0341 EMB-inducible promoter. FIG. 6 European Peptide Symposium; Maia HSL Ed., Esom: represents percent maximum Luminescense Count per Sec Leiden, 1995, 465-466). However, as described herein, an ond (% Max. LCPS) for pooled compound mixtures in one ethylene diamine library is created using amines, rather than row (row D) in one of the 96-well plates. US 2003/0236225A1 Dec. 25, 2003

0083 Deconvolution of the Reactive Wells chemical yields of approximately 20%, based on an assumed 0084. The M. tuberculosis screening revealed approxi 80% yield at each reaction step. In the Luc assay, 32 mately 300 active compounds mixtures that were selected compounds exhibited activity at 1.56 uM, and in the MIC for deconvolution. In particular, wells possessing activity of assay, at least 11 compounds had an MIC of 3.13 uM. approximately <12.5 uM in the HTS Luc assay, and/or an 0089. The total frequency of the top thirteen amines that MIC of approximately <12.5 uM, were selected for a total of contributed to the activity of the substituted ethylene 336 wells. diamines are shown in FIG. 11, with each amine represented 0085 Deconvolutions were performed by discrete re by its numerical designation. These amines include the Synthesis of each Substituted ethylene diamine compound in following: each active compound pool. The pooled compounds in each 0090 #11 2,3-Dimethylcylochexy amines active well were individually Synthesized, and Screened. Syntheses of the targeted diamine compounds in each active 0091) #183,3-Diphenylpropylamine pool were done in the 96-well plates using Stored archived 0092) #44 1-Adamantanemethylamine C-haloacetyl amides (resin attached haloacetyl amides), according to the previously described reaction Steps (the 0093) #47 2,2-Diphenylethylamine addition of the second amine, the reduction with Red-Al, 0094) #63 (S)-2-Amino-1-butanol and the cleavage from the Solid Support). The archived resins were stored as individual compounds at 4 C. The 96-well 0.095 #74.1 (-)-cis-Myrtanyl amine plates were used for the remaining Synthesis Steps as pre 0096) #77.1 Cyclooctylamine viously described. 0.086 The same screening tests, MIC and HTS Lucassay, 0097) #78.1 2-Adamantamine were performed on each deconvoluted compound. Repre 0.098 #105a (1R,2R,3R,5S)-(-)-Isopinocamphey Sentative Luminescence data for deconvoluted compounds lamine are shown in FIGS. 7 and 8. FIGS. 7 and 8 represent the Luminescence Count per Second (LCPS) for individual 0099) #231 2-Methoxyphenethylamine compounds. 0100) #255 (S)-Cylcohexylethylamine 0087 Summary of Screening Results 0101) #266 Undecylamine 0088 Overall, the deconvolution screening results 0102) #272 Geranylamine (revealed about 2,000 ethylene diamine compounds with inhibitory activity against M. tuberculosis. More than 150 of 0103). Other amines that contributed to the activity of the these compounds exhibited MICs equal to or lower than substituted ethylene diamines are shown in Table 2. The approximately 12.5 M. FIG. 9 summarizes the MIC data compounds in Table 2 are sorted by their MIC results. Some for all synthesized discrete compounds with an MIC of 50 compounds, Synthesized in larger quantities (2-60 mg) on luM or less. FIG. 10 Summarizes Luc assay data for all the Quest(R) Synthesizer, and purified by HPLC using semi compounds that exhibit at least 10% activity at each con preparative C18-column, are shown in Table 3. Generally, centration (the results are not cumulative). The MIC and Luc the final purity of each compound in Table 3 was at least activities were obtained for non-purified Samples, with 90%.

TABLE 2 Synthetic Substituted Diethylene Diamines Sorted by Minimum Inhibition Concentration MIC % N1 N2 R4 (uM) Induction 3,3-Diphenylpropylamine exo-Aminonorborname Hydrogen 3.13 53.70 2,2-Diphenylamine (+)-Isopinocampheylamine Hydrogen 3.13 93.94 2,2-Diphenylamine cis-(-)-Myrtanylamine Hydrogen 3.13 64.49 2,2-Diphenylamine Cyclooctylamine Hydrogen 3.13 63.44 2,2-Diphenylamine 3,4-Dihydroxynorephedrine Hydrogen 3.13 428O 5-Aminoquinoline Cyclohexylamine Hydrogen 3.13 18.33 5-Aminoquinoline tert-Octylamine Hydrogen 3.13 20.85 5-Aminoquinoline 4-Methylcyclohexylamine Hydrogen 3.13 26.33 cis-(-)-Myrtanylamine (+)-Bornylamine Hydrogen 3.13 100.00 cis-(-)-Myrtanylamine 1-Adamantanemethylamine Hydrogen 3.13 85.2O cis-(-)-Myrtanylamine (-)-Isopinocampheylamine Hydrogen 3.13 60.94 1-Adamantanemethylamine tert-Octylamine Hydrogen 4.7 9.81 3,4-Dimethoxyphenethylamine 1-Adamantanemethylamine Hydrogen 6.25 11.45 3,4-Dimethoxyphenethylamine Hexetidine (mixture of isomers) Hydrogen 6.25 O 3,4-Dimethoxyphenethylamine Dehydroabietylamine Hydrogen 6.25 O 3,3-Diphenylpropylamine 1-Adamantanemethylamine Hydrogen 6.25 9.53 3,3-Diphenylpropylamine 2-Methylcyclohexylamine Hydrogen 6.25 SO.08 (mix of cis and trans) 3,3-Diphenylpropylamine 1,3-Dimethylbutylamine Hydrogen 6.25 39.40 3,3-Diphenylpropylamine 1-(1-Adamantyl)ethylamine, Hydrogen 6.25 45.14 US 2003/0236225A1 Dec. 25, 2003

TABLE 2-continued Synthetic Substituted Diethylene Diamines Sorted by Minimum Inhibition Concentration MIC % N1 (uM) Induction 3,3-Diphenylpropylamine (S)-(-)-Cyclohexylethylamine Hyd rogen 6.25 43.49 3,3-Diphenylpropylamine (R)-(-)-Cyclohexylethylamine Hyd rogen 6.25 34.54 3,3-Diphenylpropylamine 1-Adamantanemethylamine Methyl 6.25 16.14 Propylamine Hexetidine (mixture of isomers) Hyd rogen 6.25 Phenethylamine Hexetidine (mixture of isomers) Hyd rogen 6.25 b-Methylphenethylamine Hexetidine (mixture of isomers) Hyd rogen 6.25 b-Methylphenethylamine Undecylamine Hyd rogen 6.25 2,2-Diphenylamine (+)-Bornylamine Hyd rogen 6.25 2,2-Diphenylamine (-)-Isopinocampheylamine Hyd rogen 6.25 2,2-Diphenylamine alpha-Methyltryptamine Hyd rogen 6.25 2,2-Diphenylamine alpha-Methyltryptamine Hyd rogen 6.25 2,2-Diphenylamine 4-Phenylbutylamine Hyd rogen 6.25 2,2-Diphenylamine 2,5-Dimethoxyphenethylamine Hyd rogen 6.25 2,2-Diphenylamine 2,4-Dichlorophenethylamine Hyd rogen 6.25 2,2-Diphenylamine 2-(2-Aminomethyl) Hyd rogen 6.25 phenylthio)benzyl alcohol 2,2-Diphenylamine 1-(1-Naphthyl)ethylamine Hyd rogen 6.25 7.20 Veratryl amine 2,5-Dimethoxyphenethylamine Hyd rogen 6.25 Veratryl amine 2-(2-Aminomethyl) Hyd rogen 6.25 phenylthio)benzyl alcohol 5-Aminoquinoline 2-Aminoheptane Hyd rogen 6.25 26.22 5-Aminoquinoline 1-Adamantanamine Hyd rogen 6.25 18.91 1-Aminomethyl-1- Hexetidine (mixture of isomers) Hyd rogen 6.25 cyclohexanol, HCl cis-(-)-Myrtanylamine 2,3-Dimethylcyclohexylamine Hyd rogen 6.25 1OO.OO cis-(-)-Myrtanylamine 3,3-Diphenylpropylamine Hyd rogen 6.25 87.78 cis-(-)-Myrtanylamine (+)-Isopinocampheylamine Hyd rogen 6.25 93.10 cis-(-)-Myrtanylamine 2,2-Diphenylamine Hyd rogen 6.25 81.84 cis-(-)-Myrtanylamine cis-(-)-Myrtanylamine Hyd rogen 6.25 68.24 cis-(-)-Myrtanylamine 1,3,3-Trimethyl-6- Hyd rogen 6.25 68.18 azabicyclo3.2.1]octane cis-(-)-Myrtanylamine 1-Adamantanemethylamine Methyl 6.25 24.22 cis-(-)-Myrtanylamine cis-(-)-Myrtanylamine Methyl 6.25 44.14 Cyclooctylamine 3,3-Diphenylpropylamine Hyd rogen 6.25 1OO.OO Cyclooctylamine (-)-Isopinocampheylamine Hyd rogen 6.25 59.13 sec-Butylamine Hexetidine (mixture of isomers) Hyd rogen 6.25 3-Methylbenzylamine Hexetidine (mixture of isomers) Hyd rogen 6.25 3-Methylbenzylamine Undecylamine Hyd rogen 6.25 2-Methoxyethylamine Hexetidine (mixture of isomers) Hyd rogen 6.25 Geranylamine 2-Adamantanamine, HCl Hyd rogen 6.25 25.66 1-Adamantanemethylamine 4-Benzylpiperidine Hyd rogen 9.4 1-Adamantanemethylamine 2,3-Dimethylcyclohexylamine Hyd rogen 9.4 1-Adamantanemethylamine 3,3-Diphenylpropylamine Hyd rogen 9.4 40.06 1-Adamantanemethylamine 1-Adamantanemethylamine Hyd rogen 9.4 15.25 1-Adamantanemethylamine 2,2-Diphenylamine Hyd rogen 9.4 1-Adamantanemethylamine 1,3,3-Trimethyl-6- Hyd rogen 9.4 azabicyclo3.2.1]octane 1-Adamantanemethylamine 138 Hyd rogen 9.4 3-Phenyl-1-propylamine 138 Hyd rogen 9.4 2,2-Diphenylamine 1-Adamantanemethylamine Hyd rogen 9.4 65.89 2,2-Diphenylamine 138 Hyd rogen 9.4 Furfurylamine Hexetidine (mixture of isomers) Hyd rogen 2.5 3,4,5-Trimethoxybenzylamine Hexetidine (mixture of isomers) Hyd rogen 2.5 1-Methyl-3-phenylpropylamine Dehydroabietylamine Hyd rogen 2.5 Cyclobutylamine Hexetidine (mixture of isomers) Hyd rogen 2.5 2-Fluorobenzylamine Hexetidine (mixture of isomers) Hyd rogen 2.5 2-Fluorobenzylamine Dehydroabietylamine Hyd rogen 2.5 3,4-Dimethoxyphenethylamine Undecylamine Hyd rogen 2.5 O 3,3-Diphenylpropylamine exo-Aminonorborname Hyd rogen 2.5 14.38 3,3-Diphenylpropylamine Decahydroquinoline Hyd rogen 2.5 22.52 3,3-Diphenylpropylamine Hexetidine (mixture of isomers) Hyd rogen 2.5 3,3-Diphenylpropylamine 4-Phenylbutylamine Hyd rogen 2.5 3,3-Diphenylpropylamine 2-Methoxyphenethylamine Hyd rogen 2.5 6.82 3,3-Diphenylpropylamine 2,4-Dichlorophenethylamine Hyd rogen 2.5 3,3-Diphenylpropylamine 1-Aminoindan Hyd rogen 2.5 18.05 3,3-Diphenylpropylamine Undecylamine Hyd rogen 2.5 3,3-Diphenylpropylamine Dehydroabietylamine Hyd rogen 2.5 3,3-Diphenylpropylamine 2-(1-Cyclohexenyl)ethylamine Methyl 2.5 9.5 3,3-Diphenylpropylamine cis-(-)-Myrtanylamine Methyl 2.5 18.41 3,3-Diphenylpropylamine Cyclooctylamine Methyl 2.5 20.84 Propylamine Dehydroabietylamine Hyd rogen 2.5 US 2003/0236225A1 Dec. 25, 2003 10

TABLE 2-continued Synthetic Substituted Diethylene Diamines Sorted by Minimum Inhibition Concentration MIC % N1 (uM) Induction Phenethylamine Dehydroabietylamine Hyd rogen 2.5 Cyclohexylamine Hexetidine (mixture of isomers) Hyd rogen 2.5 3-Amino-1-propanol Hexetidine (mixture of isomers) Hyd rogen 2.5 b-Methylphenethylamine Dehydroabietylamine Hyd rogen 2.5 4-Methoxyphenethylamine 2-Fluorophenethylamine Hyd rogen 2.5 4-Methoxyphenethylamine 2-(1-Cyclohexenyl)ethylamine Hyd rogen 2.5 4-Methoxyphenethylamine 2,4-Dimethoxybenzylamine Hyd rogen 2.5 4-Methoxyphenethylamine 4-Fluorophenethylamine Hyd rogen 2.5 1. 4-Methoxyphenethylamine Hexetidine (mixture of isomers) Hyd rogen 2.5 Tetrahydrofurfurylamine Hexetidine (mixture of isomers) Hyd rogen 2.5 Amylamine 4-Fluorophenethylamine Hyd rogen 2.5 3-Phenyl-1-propylamine 2-(1-Cyclohexenyl)ethylamine Hyd rogen 2.5 3-Phenyl-1-propylamine 4-Fluorophenethylamine Hyd rogen 2.5 12.94 2,2-Diphenylamine tert-Amylamine Hyd rogen 2.5 9.05 2,2-Diphenylamine Undecylamine Hyd rogen 2.5 2,2-Diphenylamine Dehydroabietylamine Hyd rogen 2.5 2,2-Diphenylamine cis-(-)-Myrtanylamine Methyl 2.5 45.18 1-(3-Aminopropyl)-2- 2,5-Dimethoxyphenethylamine Hyd rogen 2.5 pyrrolidinone (tech) 1-(3-Aminopropyl)-2- 2-(2- Hyd rogen 2.5 pyrrolidinone (tech) Aminomethyl)phenylthio)benzyl alcohol 4-(Trifluoromethyl)benzylamine 2,5-Dimethoxyphenethylamine Hyd rogen 2.5 4-(Trifluoromethyl)benzylamine 1-(1-Naphthyl)ethylamine Hyd rogen 2.5 Veratryl amine 4-Phenylbutylamine Hyd rogen 2.5 5-Amino-1-pentanol 2,5-Dimethoxyphenethylamine Hyd rogen 2.5 5-Amino-1-pentanol 2-(2- Hyd rogen 2.5 Aminomethyl)phenylthio)benzyl alcohol 2-(1-Cyclohexenyl)ethylamine 2-(1-Cyclohexenyl)ethylamine Hyd rogen 2.5 2-(1-Cyclohexenyl)ethylamine 4-Fluorophenethylamine Hyd rogen 2.5 2-(1-Cyclohexenyl)ethylamine 4-Phenylbutylamine Hyd rogen 2.5 2-(1-Cyclohexenyl)ethylamine 2,5-Dimethoxyphenethylamine Hyd rogen 2.5 2-(1-Cyclohexenyl)ethylamine 2-(2-Aminomethyl) Hyd rogen 2.5 phenylthio)benzyl alcohol 1-Aminomethyl-1- 2,5-Dimethoxyphenethylamine Hyd rogen 2.5 cyclohexanol, HCl 3-Fluorobenzylamine 2,5-Dimethoxyphenethylamine Hyd rogen 2.5 4-Amino-1-butanol Hexetidine (mixture of isomers) Hyd rogen 2.5 2-Ethoxybenzylamine Hexetidine (mixture of isomers) Hyd rogen 2.5 cis-(-)-Myrtanylamine Cyclooctylamine Hyd rogen 2.5 67.73 cis-(-)-Myrtanylamine 4-Methylcyclohexylamine Hyd rogen 2.5 18.39 cis-(-)-Myrtanylamine 1-Adamantanamine Hyd rogen 2.5 60.16 cis-(-)-Myrtanylamine 3,3-Diphenylpropylamine Methyl 2.5 22.32 Cyclooctylamine (+)-Isopinocampheylamine Hyd rogen 2.5 57.83 Cyclooctylamine (+)-Bornylamine Hyd rogen 2.5 1OO.OO Cyclooctylamine 1-Adamantanemethylamine Hyd rogen 2.5 52.95 Cyclooctylamine 2,2-Diphenylamine Hyd rogen 2.5 71.43 Cyclooctylamine cis-(-)-Myrtanylamine Hyd rogen 2.5 84.56 Cyclooctylamine Cyclooctylamine Hyd rogen 2.5 59.21 Cyclooctylamine Hexetidine (mixture of isomers) Hyd rogen 2.5 Cyclooctylamine Aminodiphenylmethane Hyd rogen 2.5 Cyclooctylamine Undecylamine Hyd rogen 2.5 5.61 Cyclooctylamine 3,3-Diphenylpropylamine Methyl 2.5 53.92 Cyclooctylamine (+)-Isopinocampheylamine Methyl 2.5 Cyclooctylamine cis-(-)-Myrtanylamine Methyl 2.5 33.89 4-Chlorophenylalaninol Hexetidine (mixture of isomers) Hyd rogen 2.5 (-)-Isopinocampheylamine 3,3-Diphenylpropylamine Hyd rogen 2.5 23.68 (-)-Isopinocampheylamine (+)-Bornylamine Hyd rogen 2.5 44.85 (-)-Isopinocampheylamine 2-Amino-1-propanol, d.1 Hyd rogen 2.5 46.19 (-)-Isopinocampheylamine cis-(-)-Myrtanylamine Hyd rogen 2.5 33.87 (-)-Isopinocampheylamine 2-Adamantanamine, HCl Hyd rogen 2.5 24.29 (-)-Isopinocampheylamine Aminodiphenylmethane Hyd rogen 2.5 48.35 Allylamine Hexetidine (mixture of isomers) Hyd rogen 2.5 3-Ethoxypropylamine Hexetidine (mixture of isomers) Hyd rogen 2.5 sec-Butylamine Dehydroabietylamine Hyd rogen 2.5 2-Aminoheptane Dehydroabietylamine Hyd rogen 2.5 Ethanolamine Hexetidine (mixture of isomers) Hyd rogen 2.5 3-Methylbenzylamine 4-Phenylbutylamine Hyd rogen 2.5 3-Methylbenzylamine 2,4-Dichlorophenethylamine Hyd rogen 2.5 3-Methylbenzylamine Dehydroabietylamine Hyd rogen 2.5 US 2003/0236225A1 Dec. 25, 2003 11

TABLE 2-continued Synthetic Substituted Diethylene Diamines Sorted by Minimum Inhibition Concentration MIC % N1 (uM) Induction Piperonylamine Hexetidine (mixture of isomers) Hyd rogen 2.5 Piperonylamine Dehydroabietylamine Hyd rogen 2.5 2-Methoxyethylamine Dehydroabietylamine Hyd rogen 2.5 4-Fluorophenethylamine Hexetidine (mixture of isomers) Hyd rogen 2.5 3-O-Methyldopamine, HCl Hexetidine (mixture of isomers) Hyd rogen 2.5 3-O-Methyldopamine, HCl Undecylamine Hyd rogen 2.5 3-O-Methyldopamine, HCl Dehydroabietylamine Hyd rogen 2.5 3-Fluorophenethylamine Hexetidine (mixture of isomers) Hyd rogen 2.5 3-Fluorophenethylamine Dehydroabietylamine Hyd rogen 2.5 2-Methoxyphenethylamine Hexetidine (mixture of isomers) Hyd rogen 2.5 2-Methoxyphenethylamine Aminodiphenylmethane Hyd rogen 2.5 34.67 2-Fluoroethylamine, HCl Hexetidine (mixture of isomers) Hyd rogen 2.5 2-Amino-1-phenylethanol Hexetidine (mixture of isomers) Hyd rogen 2.5 2-Amino-1-phenylethanol Dehydroabietylamine Hyd rogen 2.5 2,5-Dimethoxyphenethylamine 2-Adamantanamine, HCl Hyd rogen 2.5 22.18 2-(2-Chlorophenyl)ethylamine N-Allylcyclopentylamine Hyd rogen 2.5 62.31 2-(2-Chlorophenyl)ethylamine Hexetidine (mixture of isomers) Hyd rogen 2.5 3-Hydroxytyramine Hexetidine (mixture of isomers) Hyd rogen 2.5 4 2-Adamantanamine, HCl Hyd rogen 2.5 28.34 (Trifluoromethoxy)benzylamine Geranylamine (+)-Bornylamine Hyd rogen 2.5 Geranylamine 1,3,3-Trimethyl-6- Hyd rogen 2.5 37.42 azabicyclo3.2.1]octane Geranylamine 2-Ethylpiperidine Hyd rogen 2.5 29.81 Geranylamine 1-Adamantanamine Hyd rogen 2.5 16.63 Geranylamine N-Allylcyclopentylamine Hyd rogen 2.5 74.86 Geranylamine Aminodiphenylmethane Hyd rogen 2.5 57.93 Geranylamine Dehydroabietylamine Hyd rogen 2.5 -Adamantanemethylamine Decahydroquinoline Hyd rogen 8.8 -Adamantanemethylamine 1-Adamantanamine Hyd rogen 8.8 2,2-Diphenylamine 2,3-Dimethylcyclohexylamine Hyd rogen 8.8 23.60 2,2-Diphenylamine tert-Octylamine Hyd rogen 8.8 1929 2,2-Diphenylamine Decahydroquinoline Hyd rogen 8.8 8.96 4-Methylbenzylamine Furfurylamine Hyd rogen 25 13:46 4-Methylbenzylamine Benzylamine Hyd rogen 25 17.07 4-Methylbenzylamine Hexetidine (mixture of isomers) Hyd rogen 25 4 Dehydroabietylamine Hyd rogen 25 Hexetidine (mixture of isomers) Hyd rogen 25 Dehydroabietylamine Hyd rogen 25 Furfurylamine Furfurylamine Hyd rogen 25 -Methyl-3-phenylpropylamine Hexetidine (mixture of isomers) Hyd rogen 25 -Methyl-3-phenylpropylamine Undecylamine Hyd rogen 25 2,3,4-Tetrahydro-1- Undecylamine Hyd rogen 25 naphthylamine 2,3,4-Tetrahydro-1- Dehydroabietylamine Hyd rogen 25 naphthylamine 2,3-Dimethylcyclohexylamine Undecylamine Hyd rogen 25 2,3-Dimethylcyclohexylamine Dehydroabietylamine Hyd rogen 25 Tyramine Hexetidine (mixture of isomers) Hyd rogen 25 Tyramine Undecylamine Hyd rogen 25 Tyramine Dehydroabietylamine Hyd rogen 25 Tyramine cis-(-)-Myrtanylamine Methyl 25 2-Fluorobenzylamine Undecylamine Hyd rogen 25 (R)-2-Amino-1-butanol Hexetidine (mixture of isomers) Hyd rogen 25 3,3-Diphenylpropylamine (S)-(+)-1-Amino-2-propanol Hyd rogen 25 O 3,3-Diphenylpropylamine 2-Ethylpiperidine Hyd rogen 25 11.32 3,3-Diphenylpropylamine N-Allylcyclopentylamine Hyd rogen 25 11.63 3,3-Diphenylpropylanime Aminodiphenylmethane Hyd rogen 25 3,3-Diphenylpropylamine 3,5-Dimethylpiperidine (cis Hyd rogen 25 30.28 and trans-) 3,3-Diphenylpropylamine Allylcyclohexylamine Hyd rogen 25 1. O Propylamine Undecylamine Hyd rogen 25 Phenethylamine Undecylamine Hyd rogen 25 Tryptamine (S)-(+)-1-Amino-2-propanol Hyd rogen 25 Tryptamine 2-Amino-2-methyl-1-propanol Hyd rogen 25 Cyclohexylamine Undecylamine Hyd rogen 25 Cyclohexylamine Dehydroabietylamine Hyd rogen 25 (+)-Isopinocampheylamine Dehydroabietylamine Hyd rogen 25 Benzylamine Hexetidine (mixture of isomers) Hyd rogen 25 Benzylamine Undecylamine Hyd rogen 25 3-Amino-1-propanol Dehydroabietylamine Hyd rogen 25 US 2003/0236225A1 Dec. 25, 2003 12

TABLE 2-continued Synthetic Substituted Diethylene Diamines Sorted by Minimum Inhibition Concentration MIC % N1 (uM) Induction 2-Fluorophenethylamine 2-Fluorophenethylamine Hyd rogen 25 2-Fluorophenethylamine Veratryl amine Hyd rogen 25 2-Fluorophenethylamine 2,4-Dimethoxybenzylamine Hyd rogen 25 2-Fluorophenethylamine 2-Amino-2-methyl-1-propanol Hyd rogen 25 2-Fluorophenethylamine 4-Fluorophenethylamine Hyd rogen 25 2-Fluorophenethylamine Hexetidine (mixture of isomers) Hyd rogen 25 2-Fluorophenethylamine 1-(1-Naphthyl)ethylamine Hyd rogen 25 2-Fluorophenethylamine 1-Adamantanemethylamine Methyl 25 2-Fluorophenethylamine cis-(-)-Myrtanylamine Methyl 25 s3 b-Methylphenethylamine 4-Phenylbutylamine Hyd rogen 25 b-Methylphenethylamine 2,4-Dichlorophenethylamine Hyd rogen 25 b-Methylphenethylamine 1-(1-Naphthyl)ethylamine Hyd rogen 25 4-Methoxyphenethylamine 1-Adamantanemethylamine Hyd rogen 25 4-Methoxyphenethylamine 1-(3-Aminopropyl)-2- Hyd rogen 25 pyrrolidinone (tech) 4-Methoxyphenethylamine Veratryl amine Hyd rogen 25 4-Methoxyphenethylamine Undecylamine Hyd rogen 25 4-Methoxyphenethylamine Dehydroabietylamine Hyd rogen 25 Tetrahydrofurfurylamine Dehydroabietylamine Hyd rogen 25 Amylamine 2-Fluorophenethylamine Hyd rogen 25 Amylamine 2-(1-Cyclohexenyl)ethylamine Hyd rogen 25 Amylamine 2,4-Dimethoxybenzylamine Hyd rogen 25 3-Phenyl-1-propylamine 2-Fluorophenethylamine Hyd rogen 25 3-Phenyl-1-propylamine -Adamantanemethylamine Hyd rogen 25 3-Phenyl-1-propylamine 2,4-Dimethoxybenzylamine Hyd rogen 25 3-Phenyl-1-propylamine Hexetidine (mixture of isomers) Hyd rogen 25 3-Phenyl-1-propylamine 4 -Phenylbutylamine Hyd rogen 25 3-Phenyl-1-propylamine 2,4-Dichlorophenethylamine Hyd rogen 25 3-Phenyl-1-propylamine Undecylamine Hyd rogen 25 3-Phenyl-1-propylamine Dehydroabietylamine Hyd rogen 25 2,2-Diphenylamine 4 -(2-Aminoethyl)morpholine Hyd rogen 25 2,2-Diphenylamine -(3-Aminopropyl)-2- Hyd rogen 25 pyrrolidinone (tech) 2,2-Diphenylamine 2-(1-Cyclohexenyl)ethylamine Hyd rogen 25 2,2-Diphenylamine 2,4-Dimethoxybenzylamine Hyd rogen 25 2,2-Diphenylamine 4-(3-Aminopropyl)morpholine Hyd rogen 25 2,2-Diphenylamine 4 -Fluorophenethylamine Hyd rogen 25 2,2-Diphenylamine Hexetidine (mixture of isomers) Hyd rogen 25 2,2-Diphenylamine (S)-(-)-Cyclohexylethylamine Hyd rogen 25 2,2-Diphenylamine -Adamantanemethylamine Methyl 25 5.84 1-(3-Aminopropyl)-2- 4 -Phenylbutylamine Hyd rogen 25 pyrrolidinone (tech) 4-(Trifluoromethyl)benzylamine -Adamantanemethylamine Hyd rogen 25 4-(Trifluoromethyl)benzylamine tert-Amylamine Hyd rogen 25 4-(Trifluoromethyl)benzylamine alpha-Methyltryptamine Hyd rogen 25 6.06 4-(Trifluoromethyl)benzylamine 4 -Phenylbutylamine Hyd rogen 25 4-(Trifluoromethyl)benzylamine 2-(2-Aminomethyl) Hyd rogen 25 5.13 phenylthio)benzyl alcohol 4-(Trifluoromethyl)benzylamine Undecylamine Hyd rogen 25 4-(Trifluoromethyl)benzylamine (-)-3,4-Dihydroxynorephedrine Hyd rogen 25 4-(Trifluoromethyl)benzylamine Dehydroabietylamine Hyd rogen 25 Veratryl amine tert-Amylamine Hyd rogen 25 5-Amino-1-pentanol 4-Phenylbutylamine Hyd rogen 25 2-(1-Cyclohexenyl)ethylamine 2-Fluorophenethylamine Hyd rogen 25 2-(1-Cyclohexenyl)ethylamine 1-Adamantanemethylamine Hyd rogen 25 1-Aminomethyl-1- 4-Phenylbutylamine Hyd rogen 25 cyclohexanol, HC 3-Fluorobenzylamine 4-Phenylbutylamine Hyd rogen 25 3-Fluorobenzylamine 2-(2- Hyd rogen 25 Aminomethyl)phenylthio)benzyl alcohol 2,4-Dimethoxybenzylamine 1-Adamantanamine Hyd rogen 25 2,4-Dimethoxybenzylamine Hexetidine (mixture of isomers) Hyd rogen 25 2,4-Dimethoxybenzylamine Undecylamine Hyd rogen 25 2,4-Dimethoxybenzylamine Dehydroabietylamine Hyd rogen 25 2-Ethoxybenzylamine 1-Adamantanamine Hyd rogen 25 2-Ethoxybenzylamine N-Phenylethyldiamine Hyd rogen 25 2-Ethoxybenzylamine 2,4-Dichlorophenethylamine Hyd rogen 25 2-Ethoxybenzylamine 2-(2-Chlorophenyl)ethylamine Hyd rogen 25 3.89 2-Ethoxybenzylamine Undecylamine Hyd rogen 25 2-Ethoxybenzylamine Dehydroabietylamine Hyd rogen 25 US 2003/0236225A1 Dec. 25, 2003 13

TABLE 2-continued Synthetic Substituted Diethylene Diamines Sorted by Minimum Inhibition Concentration MIC % N1 (uM) Induction cis-(-)-Myrtanylamine 2-(1-Cyclohexenyl)ethylamine Hyd rogen 25 cis-(-)-Myrtanylamine Hexetidine (mixture of isomers) Hyd rogen 25 cis-(-)-Myrtanylamine Aminodiphenylmethane Hyd rogen 25 cis-(-)-Myrtanylamine 2,4-Dichlorophenethylamine Hyd rogen 25 cis-(-)-Myrtanylamine (S)-(-)-Cyclohexylethylamine Hyd rogen 25 28.94 cis-(-)-Myrtanylamine Undecylamine Hyd rogen 25 cis-(-)-Myrtanylamine (+)-Isopinocampheylamine Methyl 25 cis-(-)-Myrtanylamine Cyclooctylamine Methyl 25 24.92 Cyclooctylamine 2,3-Dimethylcyclohexylamine Hyd rogen 25 50.55 Cyclooctylamine (S)-2-Amino-1-butanol Hyd rogen 25 1OO.OO Cyclooctylamine 2-Adamantanamine, HCl Hyd rogen 25 29.61 Cyclooctylamine 4-Phenylbutylamine Hyd rogen 25 Cyclooctylamine 2-Chlorobenzylamine Hyd rogen 25 Cyclooctylamine 2-Aminoindan, HCl Hyd rogen 25 Cyclooctylamine Dehydroabietylamine Hyd rogen 25 Cyclooctylamine 1-(1-Naphthyl)ethylamine Hyd rogen 25 4.62 Cyclooctylamine 1-Adamantanemethylamine Methyl 25 14.2O 2,3-Dimethoxybenzylamine Hexetidine (mixture of isomers) Hyd rogen 25 2,3-Dimethoxybenzylamine Undecylamine Hyd rogen 25 2,3-Dimethoxybenzylamine Dehydroabietylamine Hyd rogen 25 4-Methylcyclohexylamine Hexetidine (mixture of isomers) Hyd rogen 25 4-Methylcyclohexylamine Undecylamine Hyd rogen 25 4-Methylcyclohexylamine Dehydroabietylamine Hyd rogen 25 4-Fluorobenzylamine Dibenzylamine Hyd rogen 25 27.98 trans-2- Cyclooctylamine Hyd rogen 25 32.8O Phenylcyclopropylamine, HCI trans-2- 2-Adamantanamine, HCl rogen 25 18.99 Phenylcyclopropylamine, HCI trans-2- 1-Adamantanamine rogen 25 18.84 Phenylcyclopropylamine, HCI Thiomicamine Hexetidine (mixture of isomers) rogen 25 (R)-1-Amino-2-propanol Hexetidine (mixture of isomers) rogen 25 4-Chlorophenylalaninol 2,4-Dichlorophenethylamine rogen 25 4-Chlorophenylalaninol Undecylamine rogen 25 4-Chlorophenylalaninol Dehydroabietylamine rogen 25 I-Leucinol Hexetidine (mixture of isomers) rogen 25 I-Leucinol 2,4-Dichlorophenethylamine rogen 25 I-Leucinol Dehydroabietylamine rogen 25 (-)-Isopinocampheylamine 2-Methoxyphenethylamine rogen 25 29.59 (-)-Isopinocampheylamine Undecylamine rogen 25 Allylamine Dehydroabietylamine rogen 25 3-Amino-1,2-propanediol Hexetidine (mixture of isomers) rogen 25 3-Ethoxypropylamine 3,3-Diphenylpropylamine rogen 25 3-Ethoxypropylamine Undecylamine rogen 25 3-Ethoxypropylamine Dehydroabietylamine rogen 25 sec-Butylamine 2,4-Dichlorophenethylamine rogen 25 sec-Butylamine Undecylamine rogen 25 2-Aminoheptane Hexetidine (mixture of isomers) rogen 25 2-Aminoheptane 4-Phenylbutylamine rogen 25 2-Aminoheptane 2,4-Dichlorophenethylamine rogen 25 1-Naphthalenemethylamine Hexetidine (mixture of isomers) rogen 25 1-Naphthalenemethylamine 4-Phenylbutylamine rogen 25 1-Naphthalenemethylamine 2,4-Dichlorophenethylamine rogen 25 1-Naphthalenemethylamine Undecylamine rogen 25 Ethanolamine Dehydroabietylamine rogen 25 Piperonylamine 4-Phenylbutylamine rogen 25 1-Ethylpropylamine Hexetidine (mixture of isomers) rogen 25 1-Ethylpropylamine Dehydroabietylamine rogen 25 Isopropylamine Hexetidine (mixture of isomers) rogen 25 4-Fluorophenethylamine 4-Phenylbutylamine rogen 25 4-Fluorophenethylamine 2,4-Dichlorophenethylamine rogen 25 4-Fluorophenethylamine Dehydroabietylamine rogen 25 3-Fluorophenethylamine Undecylamine rogen 25 2-Thiopheneethylamine 2-Adamantanamine, HCl rogen 25 19.09 2-Methylcyclohexylamine (mix Hexetidine (mixture of isomers) rogen 25 of cis and trans) 2-Methylcyclohexylamine (mix Dehydroabietylamine rogen 25 of cis and trans) 2-Methoxyphenethylamine 2-Adamantanamine, HCl rogen 25 26.77 2-Methoxyphenethylamine (-)-Isopinocampheylamine rogen 25 31.95 2-Methoxyphenethylamine 1-Adamantanamine rogen 25 24.38 US 2003/0236225A1 Dec. 25, 2003 14

TABLE 2-continued Synthetic Substituted Diethylene Diamines Sorted by Minimum Inhibition Concentration MIC % N1 (uM) Induction 2-Methoxyphenethylamine N-Allylcyclopentylamine Hyd rogen 25 14.56 2-Methoxyphenethylamine 4-Phenylbutylamine Hyd rogen 25 2-Methoxyphenethylamine Undecylamine Hyd rogen 25 2-Methoxyphenethylamine Dehydroabietylamine Hyd rogen 25 2-Fluoroethylamine, HCl Undecylamine Hyd rogen 25 2-Fluoroethylamine, HCl Dehydroabietylamine Hyd rogen 25 2-Aminoindan, HCl 2-Adamantanamine, HCl Hyd rogen 25 17.72 2-Amino-1-phenylethanol Undecylamine Hyd rogen 25 2,5-Dimethoxyphenethylamine (+)-Bornylamine Hyd rogen 25 25.78 2,5-Dimethoxyphenethylamine Noradamantamine, HCl Hyd rogen 25 11.73 2,5-Dimethoxyphenethylamine 1-Adamantanamine Hyd rogen 25 12.57 2-(2-Chlorophenyl)ethylamine 4-Phenylbutylamine Hyd rogen 25 2-(2-Chlorophenyl)ethylamine Undecylamine Hyd rogen 25 2-(2-Chlorophenyl)ethylamine 1-(1-Naphthyl)ethylamine Hyd rogen 25 2-(2- Hexetidine (mixture of isomers) Hyd rogen 25 Aminomethyl)phenylthio)benzyl alcohol 2-(2- 4-Phenylbutylamine Hyd rogen 25 Aminomethyl)phenylthio)benzyl alcohol 2-(2- Undecylamine rogen 25 Aminomethyl)phenylthio)benzyl alcohol 1-Aminoindan Hexetidine (mixture of isomers) rogen 25 1-Aminoindan Undecylamine rogen 25 1-Aminoindan Dehydroabietylamine rogen 25 1,3-Dimethylbutylamine Hexetidine (mixture of isomers) rogen 25 1,3-Dimethylbutylamine Undecylamine rogen 25 5.92 1,3-Dimethylbutylamine Dehydroabietylamine rogen 25 (S)-(-)-Cyclohexylethylamine (-)-Isopinocampheylamine rogen 25 19.31 (S)-(-)-Cyclohexylethylamine Hexetidine (mixture of isomers) rogen 25 (S)-(-)-Cyclohexylethylamine Undecylamine rogen 25 1088 (S)-(-)-Cyclohexylethylamine Dehydroabietylamine rogen 25 (S)-(–)-2-Amino-3-phenyl-1- Hexetidine (mixture of isomers) rogen 25 propanol (S)-(–)-2-Amino-3-phenyl-1- Undecylamine rogen 25 propanol (S)-(–)-2-Amino-3-phenyl-1- Dehydroabietylamine rogen 25 propanol (1S,2S)-(+)-2-Amino-3- Hexetidine (mixture of isomers) rogen 25 methoxy-1-phenyl-1-propanol Octadecylamine (+)-Bornylamine rogen 25 Octadecylamine 1-Adamantanamine rogen 25 Geranylamine 2,3-Dimethylcyclohexylamine rogen 25 14.53 Geranylamine tert-Octylamine rogen 25 15.22 Geranylamine 1-Adamantanemethylamine rogen 25 4.37 Geranylamine Decahydroquinoline rogen 25 31.79 Geranylamine Dibenzylamine rogen 25 6.48 Geranylamine N-Butylbenzylamine rogen 25 16.44 Geranylamine Cyclooctylamine rogen 25 12.37 Geranylamine (-)-Isopinocampheylamine rogen 25 8.95 Geranylamine -(1-Adamantyl)ethylamine, rogen 25 32.95 HCI Geranylamine Undecylamine rogen 25 Geranylamine -(1-Naphthyl)ethylamine rogen 25 Amylamine -Adamantanamine rogen 37.5 3-Phenyl-1-propylamine 3,3-Diphenylpropylamine rogen 37.5 3-Phenyl-1-propylamine 2,2-Diphenylamine rogen 37.5 3-Phenyl-1-propylamine -Adamantanamine rogen 37.5 18.65 2,2-Diphenylamine 3,3-Diphenylpropylamine rogen 37.5 2,2-Diphenylamine 2,2-Diphenylamine rogen 37.5 5.56 2,2-Diphenylamine ,3,3-Trimethyl-6- rogen 37.5 8.67 azabicyclo3.2.1]octane 2,2-Diphenylamine -Adamantanamine rogen 37.5 58.10 4-(Trifluoromethyl)benzylamine ert-Octylamine rogen 37.5 7.47 4-(Trifluoromethyl)benzylamine 38 rogen 37.5 4-Methylbenzylamine 2-Fluorobenzylamine rogen 50 22.10 4-Methylbenzylamine 4 -Fluorobenzylamine rogen 50 14.62 4-Methylbenzylamine alpha-Methyltryptamine rogen 50 4-Methylbenzylamine Undecylamine rogen 50 Cyclopentylamine Undecylamine rogen 50 US 2003/0236225A1 Dec. 25, 2003 15

TABLE 2-continued Synthetic Substituted Diethylene Diamines Sorted by Minimum Inhibition Concentration MIC % N1 (uM) Induction Furfurylamine 2-Fluorobenzylamine Hyd rogen 50 Furfurylamine Benzylamine Hyd rogen 50 Furfurylamine 4-Fluorobenzylamine Hyd rogen 50 Furfurylamine alpha-Methyltryptamine Hyd rogen 50 Furfurylamine Undecylamine Hyd rogen 50 Furfurylamine Dehydroabietylamine Hyd rogen 50 Furfurylamine Furfurylamine Hyd rogen 50 3,4,5-Trimethoxybenzylamine 2-Fluorobenzylamine Hyd rogen 50 3,4,5-Trimethoxybenzylamine Benzylamine Hyd rogen 50 3,4,5-Trimethoxybenzylamine alpha-Methyltryptamine Hyd rogen 50 3,4,5-Trimethoxybenzylamine Undecylamine Hyd rogen 50 3,4,5-Trimethoxybenzylamine Dehydroabietylamine Hyd rogen 50 1-Methyl-3-phenylpropylamine alpha-Methyltryptamine Hyd rogen 50 1-Methyl-3-phenylpropylamine Octadecylamine Hyd rogen 50 Cyclobutylamine Octadecylamine Hyd rogen 50 Cyclobutylamine Undecylamine Hyd rogen 50 Cyclobutylamine Dehydroabietylamine Hyd rogen 50 1,2,3,4-Tetrahydro-1- Hexetidine (mixture of isomers) Hyd rogen 50 naphthylamine 1,2,3,4-Tetrahydro-1- Aminodiphenylmethane rogen 50 4.31 naphthylamine 1,2,3,4-Tetrahydro-1- alpha-Methyltryptamine rogen 50 O naphthylamine 1,2,3,4-Tetrahydro-1- 2-Methoxyphenethylamine rogen 50 O naphthylamine 2,3-Dimethylcyclohexylamine Hexetidine (mixture of isomers) rogen 50 2,3-Dimethylcyclohexylamine Aminodiphenylmethane rogen 50 6 4 2,3-Dimethylcyclohexylamine alpha-Methyltryptamine rogen 50 Tyramine Furfurylamine rogen 50 Tyramine 2-Fluorobenzylamine rogen 50 O 7 Tyramine Benzylamine rogen 50 Tyramine 2,4-Dichlorophenethylamine rogen 50 2-Fluorobenzylamine Aminodiphenylmethane rogen 50 2-Fluorobenzylamine 4-Phenylbutylamine rogen 50 2-Fluorobenzylamine 2-Methoxyphenethylamine rogen 50 2-Fluorobenzylamine 2,4-Dichlorophenethylamine rogen 50 2-Fluorobenzylamine 1,3-Dimethylbutylamine rogen 50 2-Fluorobenzylamine 1-(1-Adamantyl)ethylamine, rogen 50 HCI (R)-2-Amino-1-butanol Dehydroabietylamine rogen 50 3,4-Dimethoxyphenethylamine Aminodiphenylmethane rogen 50 3,4-Dimethoxyphenethylamine 4-Phenylbutylamine rogen 50 3,4-Dimethoxyphenethylamine 2-Methoxyphenethylamine rogen 50 3,4-Dimethoxyphenethylamine 2,4-Dichlorophenethylamine rogen 50 3,4-Dimethoxyphenethylamine 1,3-Dimethylbutylamine rogen 50 3,3-Diphenylpropylamine Piperidine rogen 50 3,3-Diphenylpropylamine 2,3-Dimethylcyclohexylamine Methyl 50 7 8 1. 3,3-Diphenylpropylamine (-)-Isopinocamphenylamine Methyl 50 13.06 Propylamine (S)-(+)-1-Amino-2-propanol Hyd rogen 50 O Phenethylamine (S)-(+)-1-Amino-2-propanol Hyd rogen 50 Phenethylamine 4-Phenylbutylamine Hyd rogen 50 Phenethylamine 2,4-Dichlorophenethylamine Hyd rogen 50 Phenethylamine 1,3-Dimethylbutylamine Hyd rogen 50 Phenethylamine 1-(1-Adamantyl)ethylamine Hyd rogen 50 HCI Phenethylamine 1-(1-Naphthyl)ethylamine Hyd rogen 50 4-(2-Aminoethyl)morpholine 2-Amino-2-methyl-1-propanol Hyd rogen 50 Cyclohexylamine 2,4-Dichlorophenethylamine Hyd rogen 50 exo-Aminonorborname Benzylamine Hyd rogen 50 (+)-Isopinocampheylamine Hexetidine (mixture of isomers) Hyd rogen 50 (+)-Isopinocampheylamine Aminodiphenylmethane Hyd rogen 50 (+)-Isopinocampheylamine 4-Phenylbutylamine Hyd rogen 50 (+)-Isopinocampheylamine 2,4-Dichlorophenethylamine Hyd rogen 50 (+)-Isopinocampheylamine Undecylamine Hyd rogen 50 Benzylamine 3,3-Diphenylpropylamine Hyd rogen 50 Benzylamine 2-Amino-2-methyl-1-propanol Hyd rogen 50 Benzylamine 1-(1-Naphthyl)ethylamine Hyd rogen 50 Benzylamine 2,4-Dichlorophenethylamine Hyd rogen 50 3-Amino-1-propanol Undecylamine Hyd rogen 50 2-Fluorophenethylamine 3,3-Diphenylpropylamine Hyd rogen 50 g 2-Fluorophenethylamine 1-Adamantanemethylamine Hyd rogen 50 US 2003/0236225A1 Dec. 25, 2003 16

TABLE 2-continued Synthetic Substituted Diethylene Diamines Sorted by Minimum Inhibition Concentration MIC % N1 R4 (uM) Induction 2-Fluorophenethylamine 1-(3-Aminopropyl)-2- Hyd rogen 50 O pyrrolidinone (tech) 2-Fluorophenethylamine Decahydroquinoline Hyd rogen 50 2-Fluorophenethylamine 1-Adamantanamine Hyd rogen 50 34 2-Fluorophenethylamine 2,4-Dichlorophenethylamine Hyd rogen 50 2-Fluorophenethylamine Undecylamine Hyd rogen 50 2-Fluorophenethylamine Dehydroabietylamine Hyd rogen 50 2-Fluorophenethylamine 2-(1-Cyclohexenyl)ethylamine Methyl 50 2-Fluorophenethylamine Cyclooctylamine Methyl 50 8 1. b-Methylphenethylamine 3,3-Diphenylpropylamine Hyd rogen 50 b-Methylphenethylamine tert-Octylamine Hyd rogen 50 b-Methylphenethylamine 2-(1-Cyclohexenyl)ethylamine Hyd rogen 50 b-Methylphenethylamine 2-Amino-2-methyl-1-propanol Hyd rogen 50 b-Methylphenethylamine 4-Fluorophenethylamine Hyd rogen 50 b-Methylphenethylamine Geranylamine Hyd rogen 50 b-Methylphenethylamine 5-Methoxytryptamine Hyd rogen 50 4-Methoxyphenethylamine 3,3-Diphenylpropylamine Hyd rogen 50 4-Methoxyphenethylamine 2-Amino-2-methyl-1-propanol Hyd rogen 50 4-Methoxyphenethylamine 2,4-Dichlorophenethylamine Hyd rogen 50 4-Methoxyphenethylamine 1-(1-Naphthyl)ethylamine Hyd rogen 50 L-Methioninol Hexetidine (mixture of isomers) Hyd rogen 50 Tetrahydrofurfurylamine 1-Adamantanemethylamine Hyd rogen 50 Tetrahydrofurfurylamine 2-(1-Cyclohexenyl)ethylamine Hyd rogen 50 Tetrahydrofurfurylamine 4-Fluorophenethylamine Hyd rogen 50 Tetrahydrofurfurylamine Undecylamine Hyd rogen 50 Amylamine 1-Adamantanemethylamine Hyd rogen 50 Amylamine Hexetidine (mixture of isomers) Hyd rogen 50 Amylamine Undecylamine Hyd rogen 50 Amylamine Dehydroabietylamine Hyd rogen 50 1-Adamantanemethylamine cis-(-)-Myrtanylamine Methyl 50 3-Phenyl-1-propylamine 4-(2-Aminoethyl)morpholine Hyd rogen 50 3-Phenyl-1-propylamine 1-(3-Aminopropyl)-2- Hyd rogen 50 pyrrolidinone (tech) 3-Phenyl-1-propylamine Veratryl amine Hyd rogen 50 3-Phenyl-1-propylamine Aminodiphenylmethane Hyd rogen 50 3-Phenyl-1-propylamine 2-(2- Hyd rogen 50 Aminomethyl)phenylthio)benzyl alcohol 2,2-Diphenylamine 2-Fluorophenethylamine Hyd rogen 50 2,2-Diphenylamine 3,3-Diphenylpropylamine Methyl 50 2,2-Diphenylamine (+)-Isopinocampheylamine Methyl 50 2,2-Diphenylamine (+)-Bornylamine Methyl 50 2,2-Diphenylamine Cyclooctylamine Methyl 50 2,2-Diphenylamine (-)-Isopinocampheylamine Methyl 50 3.81 4-(Trifluoromethyl)benzylamine 4-(2-Aminoethyl)morpholine rogen 50 4-(Trifluoromethyl)benzylamine 2-(1-Cyclohexenyl)ethylamine rogen 50 4-(Trifluoromethyl)benzylamine Hexetidine (mixture of isomers) rogen 50 4-(Trifluoromethyl)benzylamine 2,4-Dichlorophenethylamine rogen 50 4-(Trifluoromethyl)benzylamine (S)-(-)-Cyclohexylethylamine rogen 50 Veratryl amine 1-Adamantanemethylamine rogen 50 Veratryl amine 2-(1-Cyclohexenyl)ethylamine rogen 50 Veratryl amine 4-Fluorophenethylamine rogen 50 Veratryl amine Hexetidine (mixture of isomers) rogen 50 Veratryl amine 2,4-Dichlorophenethylamine rogen 50 Veratryl amine (S)-(-)-Cyclohexylethylamine rogen 50 Veratryl amine Undecylamine rogen 50 Veratryl amine Dehydroabietylamine rogen 50 Veratryl amine 1-(1-Naphthyl)ethylamine rogen 50 5-Amino-1-pentanol 1-Adamantanemethylamine rogen 50 5-Amino-1-pentanol Dibenzylamine rogen 50 5-Amino-1-pentanol cis-(-)-Myrtanylamine rogen 50 12.97 2-(1-Cyclohexenyl)ethylamine 2,4-Dimethoxybenzylamine rogen 50 1-Aminomethyl-1- tert-Amylamine rogen 50 cyclohexanol, HCl 1-Aminomethyl-1- 2-(2- rogen 50 cyclohexanol, HCl Aminomethyl)phenylthio)benzyl alcohol 1-Aminomethyl-1- Undecylamine rogen 50 cyclohexanol, HCl 1-Aminomethyl-1- 1-(1-Naphthyl)ethylamine rogen 50 cyclohexanol, HCl US 2003/0236225A1 Dec. 25, 2003 17

TABLE 2-continued Synthetic Substituted Diethylene Diamines Sorted by Minimum Inhibition Concentration MIC % N1 (uM) Induction 3-Fluorobenzylamine tert-Amylamine Hyd rogen 50 3-Fluorobenzylamine Hexetidine (mixture of isomers) Hyd rogen 50 3-Fluorobenzylamine Undecylamine Hyd rogen 50 4-Amino-1-butanol Undecylamine Hyd rogen 50 4-Amino-1-butanol Dehydroabietylamine Hyd rogen 50 2,4-Dimethoxybenzylamine N-Phenylethyldiamine Hyd rogen 50 2,4-Dimethoxybenzylamine Aminodiphenylmethane Hyd rogen 50 2,4-Dimethoxybenzylamine 4 -Phenylbutylamine Hyd rogen 50 2,4-Dimethoxybenzylamine 2-Chlorobenzylamine Hyd rogen 50 2,4-Dimethoxybenzylamine 2,4-Dichlorophenethylamine Hyd rogen 50 2,4-Dimethoxybenzylamine 2-(2-Chlorophenyl)ethylamine Hyd rogen 50 2,4-Dimethoxybenzylamine 4 Hyd rogen 50 (Trifluoromethoxy)benzylamine 2-Ethoxybenzylamine Aminodiphenylmethane Hyd rogen 50 2-Ethoxybenzylamine 4 -Phenylbutylamine Hyd rogen 50 2-Ethoxybenzylamine 2-Chlorobenzylamine Hyd rogen 50 2-Ethoxybenzylamine 2-Aminoindan, HCl Hyd rogen 50 2-Ethoxybenzylamine 2,5-Dimethoxyphenethylamine Hyd rogen 50 2-Ethoxybenzylamine 4 Hyd rogen 50 (Trifluoromethoxy)benzylamine 2-Ethoxybenzylamine -(1-Naphthyl)ethylamine Hyd rogen 50 cis-(-)-Myrtanylamine 4 -(2-Aminoethyl)morpholine Hyd rogen 50 cis-(-)-Myrtanylamine 2-Fluorophenethylamine Hyd rogen 50 cis-(-)-Myrtanylamine -(3-Aminopropyl)-2- Hyd rogen 50 pyrrolidinone (tech) cis-(-)-Myrtanylamine Veratryl amine Hyd rogen 50 cis-(-)-Myrtanylamine N-Butylbenzylamine Hyd rogen 50 cis-(-)-Myrtanylamine 2,4-Dimethoxybenzylamine Hyd rogen 50 cis-(-)-Myrtanylamine 2,3,4-Tetrahydropyridoindole Hyd rogen 50 cis-(-)-Myrtanylamine 4 -Phenylbutylamine Hyd rogen 50 cis-(-)-Myrtanylamine 2-(2-Chlorophenyl)ethylamine Hyd rogen 50 3.91 cis-(-)-Myrtanylamine -(1-Adamantyl)ethylamine, Hyd rogen 50 10.85 HCI cis-(-)-Myrtanylamine (R)-(-)-Cyclohexylethylamine Hyd rogen 50 5.89 cis-(-)-Myrtanylamine Dehydroabietylamine Hyd rogen 50 cis-(-)-Myrtanylamine -(1-Naphthyl)ethylamine Hyd rogen 50 cis-(-)-Myrtanylamine +)-Bornylamine Methyl 50 4.04 Cyclooctylamine 4 -Methylcyclohexylamine Hyd rogen 50 4.55 Cyclooctylamine N-Phenylethyldiamine Hyd rogen 50 Cyclooctylamine 4 -(Hexacylamino)benzylamine Hyd rogen 50 Cyclooctylamine 2,5-Dimethoxyphenethylamine Hyd rogen 50 Cyclooctylamine 2,4-Dichlorophenethylamine Hyd rogen 50 3.36 Cyclooctylamine 2-(2-Chlorophenyl)ethylamine Hyd rogen 50 9.15 Cyclooctylamine -(1-Adamantyl)ethylamine, Hyd rogen 50 10.62 HCI Cyclooctylamine (S)-(-)-Cyclohexylethylamine Hyd rogen 50 5.85 Cyclooctylamine (R)-(-)-Cyclohexylethylamine Hyd rogen 50 Cyclooctylamine 4 Hyd rogen 50 4.54 (Trifluoromethoxy)benzylamine 2-Adamantanamine, HCl cis-(-)-Myrtanylamine Hyd rogen 50 49.73 4-Methylcyclohexylamine N-Phenylethyldiamine Hyd rogen 50 4-Methylcyclohexylamine 4-Phenylbutylamine Hyd rogen 50 4-Fluorobenzylamine N-Benzyl-2-phenethylamine Hyd rogen 50 4-Fluorobenzylamine Hexetidine (mixture of isomers) Hyd rogen 50 4-Fluorobenzylamine Undecylamine Hyd rogen 50 4-Fluorobenzylamine Dehydroabietylamine Hyd rogen 50 trans-2- Hexetidine (mixture of isomers) Hyd rogen 50 Phenylcyclopropylamine, HCI trans-2- Undecylamine rogen 50 Phenylcyclopropylamine, HCI trans-2- Dehydroabietylamine rogen 50 Phenylcyclopropylamine, HCI (R)-1-Amino-2-propanol 4-(Hexacylamino)benzylamine rogen 50 (R)-1-Amino-2-propanol Undecylamine rogen 50 (R)-1-Amino-2-propanol Dehydroabietylamine rogen 50 I-Leucinol Undecylamine rogen 50 (-)-Isopinocampheylamine 2-Ethoxybenzylamine rogen 50 27.27 (-)-Isopinocampheylamine Hexetidine (mixture of isomers) rogen 50 (-)-Isopinocampheylamine 4-Phenylbutylamine rogen 50 (-)-Isopinocampheylamine Dehydroabietylamine rogen 50 (-)-Isopinocampheylamine 1-(1-Naphthyl)ethylamine rogen 50 US 2003/0236225A1 Dec. 25, 2003 18

TABLE 2-continued Synthetic Substituted Diethylene Diamines Sorted by Minimum Inhibition Concentration MIC % N1 (uM) Induction Allylamine 3,3-Diphenylpropylamine Hyd rogen 50 Allylamine 2-Amino-1-propanol, d.1 Hyd rogen 50 Allylamine Undecylamine Hyd rogen 50 3-Amino-1,2-propanediol Dehydroabietylamine Hyd rogen 50 3-Ethoxypropylamine 2,2-Diphenylamine Hyd rogen 50 95.81 3-Ethoxypropylamine cis-(-)-Myrtanylamine Hyd rogen 50 2-Aminoheptane 2-(2- Hyd rogen 50 Aminomethyl)phenylthio)benzyl alcohol 1-Naphthalenemethylamine Geranylamine Hyd rogen 50 1-Naphthalenemethylamine Dehydroabietylamine Hyd rogen 50 1-Aminopyrrolidine, HCl Hexetidine (mixture of isomers) Hyd rogen 50 1-Aminopyrrolidine, HCl Undecylamine Hyd rogen 50 1-Aminopyrrolidine, HCl Dehydroabietylamine Hyd rogen 50 Ethanolamine 3,3-Diphenylpropylamine Hyd rogen 50 3-Methylbenzylamine Geranylamine Hyd rogen 50 3-Methylbenzylamine 5-Methoxytryptamine Hyd rogen 50 Piperonylamine Aminodiphenylmethane Hyd rogen 50 Piperonylamine 2,4-Dichlorophenethylamine Hyd rogen 50 Piperonylamine 2-(2- Hyd rogen 50 Aminomethyl)phenylthio)benzyl alcohol Isopropylamine Dehydroabietylamine Hyd rogen 50 4-Fluorophenethylamine 2,4-Dimethoxybenzylamine Hyd rogen 50 4-Fluorophenethylamine Aminodiphenylmethane Hyd rogen 50 4-Fluorophenethylamine 2-(2- Hyd rogen 50 Aminomethyl)phenylthio)benzyl alcohol 4-Chloroamphetamine, HCI N-Allylcyclopentylamine Hyd rogen 50 10.25 4-Chloroamphetamine, HCl Hexetidine (mixture of isomers) Hyd rogen 50 4-Chloroamphetamine, HCI 4-Phenylbutylamine Hyd rogen 50 4-Chloroamphetamine, HCI 2-Methoxyphenethylamine Hyd rogen 50 4-Chloroamphetamine, HCI Undecylamine Hyd rogen 50 4-Chloroamphetamine, HCI Dehydroabietylamine Hyd rogen 50 3-Fluorophenethylamine (-)-Isopinocampheylamine Hyd rogen 50 3-Fluorophenethylamine 1-Adamantamine Hyd rogen 50 8.59 3-Fluorophenethylamine 4-Phenylbutylamine Hyd rogen 50 2-Methylcyclohexylamine (mix Undecylamine Hyd rogen 50 of cis and trans) 2-Methoxyphenethylamine 3,3-Diphenylpropylamine Hyd rogen 50 2-Methoxyphenethylamine (+)-Bornylamine Hyd rogen 50 2-Methoxyphenethylamine tert-Octylamine Hyd rogen 50 20.46 2-Methoxyphenethylamine 1-Adamantanemethylamine Hyd rogen 50 2-Methoxyphenethylamine Dibenzylamine Hyd rogen 50 2-Methoxyphenethylamine N-Butylbenzylamine Hyd rogen 50 5.20 2-Methoxyphenethylamine 1,3,3-Trimethyl-6- Hyd rogen 50 8.59 azabicyclo3.2.1]octane 2-Methoxyphenethylamine N-Phenylethyldiamine Hyd rogen 50 2-Methoxyphenethylamine 2,4-Dichlorophenethylamine Hyd rogen 50 2-Methoxyphenethylamine 2-(2-Chlorophenyl)ethylamine Hyd rogen 50 2-Methoxyphenethylamine 1-(1-Adamantyl)ethylamine, Hyd rogen 50 3.61 HCI 2-Aminoindan, HCl (+)-Bornylamine Hyd rogen 50 2-Aminoindan, HCl Noradamantamine, HCl Hyd rogen 50 7.43 2-(2-Chlorophenyl)ethylamine N-Phenylethyldiamine Hyd rogen 50 2-(2-Chlorophenyl)ethylamine Aminodiphenylmethane Hyd rogen 50 2-(2-Chlorophenyl)ethylamine 2,4-Dichlorophenethylamine Hyd rogen 50 2-(2-Chlorophenyl)ethylamine 1-(1-Adamantyl)ethylamine, Hyd rogen 50 HCI 2-(2-Chlorophenyl)ethylamine Dehydroabietylamine Hyd rogen 50 2-(2- 2-Methoxyphenethylamine Hyd rogen 50 Aminomethyl)phenylthio)benzyl alcoho 2-(2- 2,5-Dimethoxyphenethylamine rogen 50 Aminomethyl)phenylthio)benzyl alcoho 2-(2- 2-(2-Chlorophenyl)ethylamine rogen 50 Aminomethyl)phenylthio)benzyl alcoho 2-(2- 1-(1-Adamantyl)ethylamine, rogen 50 Aminomethyl)phenylthio)benzyl HCI alcoho US 2003/0236225A1 Dec. 25, 2003 19

TABLE 2-continued Synthetic Substituted Diethylene Diamines Sorted by Minimum Inhibition Concentration MIC % N1 R4 (uM) Induction 2-(2- Dehydroabietylamine Hydrogen 50 Aminomethyl)phenylthio)benzyl alcohol 1-Aminoindan 4-Phenylbutylamine Hydrogen 50 1-Aminoindan 2,4-Dichlorophenethylamine Hydrogen 50 1,3-Dimethylbutylamine 4-Phenylbutylamine Hydrogen 50 (S)-(-)-Cyclohexylethylamine Aminodiphenylmethane Hydrogen 50 (S)-(-)-Cyclohexylethylamine 4-Phenylbutylamine Hydrogen 50 (S)-(-)-Cyclohexylethylamine 2,4-Dichlorophenethylamine Hydrogen 50 (S)-(-)-Cyclohexylethylamine 1-(1-Adamantyl)ethylamine, Hydrogen 50 HCI (1S,2S)-(+)-2-Amino-3- Dehydroabietylamine Hydrogen 50 methoxy-1-phenyl-1-propanol Octadecylamine 2-Adamantanamine, HCl Hydrogen 50 3-Hydroxytyramine (1R,2S)-(–)-2-Amino-1,2- Hydrogen 50 diphenylethanol 3-Hydroxytyramine Dehydroabietylamine Hydrogen 50 Geranylamine 3,3-Diphenylpropylamine Hydrogen 50 Geranylamine N-Phenylethyldiamine Hydrogen 50 Geranylamine Hexetidine (mixture of isomers) Hydrogen 50 Geranylamine 2-Thiopheneethylamine Hydrogen 50 Geranylamine 2-Methoxyphenethylamine Hydrogen 50 Geranylamine 2,5-Dimethoxyphenethylamine Hydrogen 50 Geranylamine 2,4-Dichlorophenethylamine Hydrogen 50 Geranylamine 2-(2-Chlorophenyl)ethylamine Hydrogen 50 2-Fluorophenethylamine 2,3-Dimethylcyclohexylamine Methyl >50 2.07 4-(Trifluoromethyl)benzylamine 2,3-Dimethylcyclohexylamine Hydrogen >50 8.2O 4-(Trifluoromethyl)benzylamine 1-Adamantanamine Hydrogen >50 32O2 5-Aminoquinoline exo-Aminonorborname Hydrogen >50 17.87

01.04]

TABLE 3

Compounds Synthesized in Larger Quantities for Further in vitro Evaluations

Amount, Yields, Cmpd # Name Structure mg %

1 N-(4-Methylphenyl)-N'- 23 25 (furfuryl)ethane-1,2-diamine Cy/- c. (Z)

2 N-(4-Methylphenyl)-N'- 27 29 (benzyl)ethane-1,2-diamine Me -C/-NH 3 N-1-(1,2,3,4-Tetrahydro 11 naphthalene)-N'-(undecenyl)- ethane-1,2-diamine ( NH NH / Yulia

US 2003/0236225A1 Dec. 25, 2003 21

TABLE 3-continued

Compounds Synthesized in Larger Quantities for Further in vitro Evaluations

Amount, Yields, Cmpd # Name Structure mg %

11 N-Cyclooctyl-N'-(3,3- Me 22 18 diphenylpropyl)ethane-1,2- diamine NH

13 N-Allyl-N-cyclopentyl-N-(3,3- 33 27 diphenylpropyl)ethane-1,2- diamine /

14 N-(3,3-Diphenylpropyl)-N'- 17 16 exo-(2-norborny)ethane-1,2- diamine HN

15 1-2-N-(3,3-Diphenylpropyl)- Me 6.2 5 aminoethyl-3,5-dimethyl piperidine /- ()Me

17 N-2-(2-Methoxyphenyl)ethyl- 50 40 N'-(3,3-diphenylethyl)ethane- / NH 1,2-diamine NH OMe

21 N-(3,3-Diphenylpropyl)-N'- 5 4 (1S)-(1-ethylcyclohexane)- / NH ethane-1,2-diamine NH (S) Me

US 2003/0236225A1 Dec. 25, 2003

0105 Formulations Substituted ethylene diamine compounds, and the in Vitro 0106 Therapeutics, including compositions containing and in Vivo Suppression of the growth of colonies of M. the Substituted ethylene diamine compounds of the present tuberculosis. It will be appreciated that other examples, invention, can be prepared in physiologically acceptable including minor variations in chemical procedures, will be formulations, Such as in pharmaceutically acceptable carri apparent to those skilled in the art, and that the invention is ers, using known techniques. For example, a Substituted not limited to these specific illustrated examples. ethylene diamine compound is combined with a pharma ceutically acceptable excipient to form a therapeutic com EXAMPLE I position. Generating the Ethylene Diamine Library 0107 The compositions of the present invention may be administered in the form of a Solid, liquid or aeroSol. 0112 The Rink-acid resin was obtained from NOVA Examples of Solid compositions include pills, creams, Soaps BIOCHEME) Inc., San Diego, Calif. Solvents: acetonitrile, and implantable dosage units. Pills may be administered dichloromethane, dimethylformamide, ethylenedichloride, orally. Therapeutic creams and anti-mycobacteria Soaps may methanol and tetrahydrofuran were purchased from ALD be administered topically. Implantable dosage units may be RICHE), Milwaukee, Wis., and used as received. All other administered locally, for example, in the lungs, or may be reagents were purchased from SIGMA-ALDRICHCR), West implanted for Systematic release of the therapeutic compo Monroe Highland, Ill. Solid phase syntheses were performed Sition, for example, Subcutaneously. Examples of liquid on a QUEST(R 210 Synthesizer, from ARGONAUTTECH compositions include formulations adapted for injection NOLOGIES(R), Foster City, Calif., with the aid of combina intramuscularly, Subcutaneously, intravenously, intraarteri torial chemistry equipment, from WHATMANGE POLYFIL ally, and formulations for topical and intraocular adminis TRONICS(R) (Kent, England; Rockland, Mass.) and tration. Examples of aerosol formulations include inhaler ROBBINS SCIENTIFIC(R), Sunnyvale, Calif. Evaporation formulations for administration to the lungs. of solvents was done using SPEEDVAC(R) AES, from 0108 A Sustained release matrix, as used herein, is a SAVANT(R), Holbrook, N.Y. All necessary chromatographic matrix made of materials, usually polymers, which are separations were performed on a WATERS ALLIANCE HT degradable by enzymatic or acid/base hydrolysis, or by SYSTEMCR, Milford, Mass. Analytical thin-layer chroma dissolution. Once inserted into the body, the matrix is acted tography was performed on MERCKCR silica gel 60Fs, upon by enzymes and body fluids. The Sustained release plates, purchased from SIGMA-ALDRICH(R), West Monroe matrix is chosen desirably from biocompatible materials, Highland, Ill. including, but not limited to, liposomes, polylactides, polyg 0113. The activation of the Rink-acid resin, the addition lycolide (polymer of glycolic acid), polylactide co-glycolide of the first amine, and the acylation Step were carried out in (coplymers of lactic acid and glycolic acid), polyanhydrides, 10 ml tubes using the QUEST(R 210 Synthesizer. The poly(ortho)esters, polypeptides, hyaluronic acid, collagen, addition of the Second amine, the reduction with Red-AL, chondroitin Sulfate, carboxylic acids, fatty acids, phospho and the cleavage from the Solid Support were carried out in lipds, polysaccharides, nucleic acids, polyamino acids, 96-deep (2 ml) well, chemically resistant plates. amino acids Such as phenylalanine, tyrosine, isoleucine, polynucleotides, polyvinyl propylene, polyvinylpyrrollidone 0114 A. Activation of the Rink-Acid Resin and Silicone. A preferred biodegradable matrix is a matrix of 0115 The Rink-acid resin had a coverage of 0.43-0.63 one of either polylactide, polyglycolide, or polylactide co mmol of linker per gram resin. Four to five grams of this glycolide. resin were suspended in 80 ml of a 2:1 mixture of dichlo 0109 The dosage of the composition will depend on the romethane and tetrahydrofuran (THF), and distributed into condition being treated, the particular composition used, and ten, 10 ml tubes, with 8 ml of resin Suspension per tube. other clinical factors, Such as weight and condition of the Each suspension was filtered and washed twice with THF. A patient, and the route of administration. A Suitable dosage solution of triphenylphosphine (3.80 g, 14.5 mmol) in 30 ml may range from 100 to 0.1 mg/kg. A more preferred dosage of THF was prepared, and 3 ml of this solution was added may range from 50 to 0.2 mg/kg. A more preferred dosage to each tube, followed by the addition of 3 ml of a solution may range from 25 to 0.5 mg/kg. Tablets or other forms of of hexachloroethane in THF (3.39 g/14.3 mmol hexachlo media may contain from 1 to 1000 mg of the substituted roethane in 30 ml THF). After agitation for six hours at room ethylene diamine. Dosage ranges and Schedules of admin temperature, each activated resin was washed twice with istration Similar to ethambutol or other anti-tuberculosis THF and twice with dichloromethane. drugs may be used. 0116 B. Addition of the First Amine 0110. The composition may be administered in combi 0.117) Each tube, containing the activated rink resin, was nation with other compositions and procedures for the charged with 3 ml of dichloroethane, 0.3 ml (1.74 mmol) treatment of other disorders occurring in combination with N,N-diisopropylethylamine (EtN(iPr)) and the correspond mycobacterial disease. For example, tuberculosis frequently ing amine (around 1 mmol). If the Selected amine was a Solid occurs as a Secondary complication associated with acquired at room temperature, it was added as a Solution, or a immunodeficiency Syndrome (AIDS). Patients undergoing suspension in DMF. Enough dichloroethane was added to AIDS treatment, which includes procedures Such as Surgery, each tube for a final volume of 8 ml. The reaction mixture radiation or chemotherapy, may benefit from the therapeutic was heated at 45 C. for 6-8 hours. The resins were filtered, methods and compositions described herein. washed with a 2:1 mixture of dichloromethane and methanol 0111. The following specific examples will illustrate the (1x8 ml), then with methanol (2x8 ml), and then dried under invention as it applies to the particular Synthesis of the argon for 10 minutes. US 2003/0236225A1 Dec. 25, 2003 32

0118 C. Acylation with the Halo-Acylchloride modate 96-well reaction plates during Synthesis, allowing 0119 a. Acylation with Chloroacetyl Chloride. Each for the addition of reagents into the plates, and a proper resin was prewashed with THF (2x8 ml), and then charged Sealing that maintains reagents and Solvents for hours at with THF (8 ml), pyridine (0.3 ml, 3.67 mmole) and elevated temperatures. These clamps consist of a top and chloroacetyl chloride (0.25 ml, 2.5 mmole). The reaction bottom cover provided with changeable, chemically resis mixture was stirred for 8 hours at 45 C., and then for 6-8 tant Sealing gaskets. They are designed to accommodate hours at room temperature. Each resin was filtered, washed 96-well reaction plates between the top and bottom covers. with a 2:1 mixture of dichloromethane/methanol (1x8 ml), The reaction plates were Sealed and kept in an oven at methanol (2x8 ml) and THF (2x8 ml). The acylation was 70-75 C. for 16 hours. After cooling to room temperature, repeated using the same loading of reagents, but a shorter the resins were filtered, washed with a 1:1 mixture of reaction time of 4 hours at 45 C., and 2 hours at room DCM/methanol (1x1 ml), methanol (2x1 ml), and then dried temperature. Each resin was then filtered, washed with a 2:1 in a desiccator under vacuum for 2 hours. mixture of dichloromethane and methanol (1x8 ml), and 0125 E. Reduction with Red-Al then with methanol (3x8 ml). Each resin was dried under argon for 10 minutes. Each resin was then transferred into a 0.126 The reaction plates were placed into COM Vial and dried in a desiccator under Vacuum for 1 hour. BICLAMPS(R). A 1:6 mixture of Red-Al (65+w % in tolu ene) and THF was added, at 0.6 ml per well (0.28 mmole of 0120 b. Acylation with C.-Phenyl-C.-Chloroacetyl Chlo Red-Alper well), and allowed to react for 4 hours. Each ride. The same procedure Set out for the acylation with resin was then filtered, washed with THF (2x1 ml), and chloroacetyl chloride was used. A 2.5 mmol excess of methanol (3x1 ml). The addition of methanol should pro C-phenyl-C-chloroacetyl chloride, relative to mmol amount ceed with caution. Each resin was then dried under vacuum. of linker in the rink-acid resin, was used. 0121 c. Acylation with C.-Halo-O-Methyl, C.-Halo-C.- 0127 F. Cleavage of Final Ethylene Diamine Compound Ethyl and C.-Halo-O-Butylacetyl Bromide. A 1:1:1 mixture 0128. This step was carried out using a cleavage mani (by volume) of the C-bromoproponyl bromide (R=Me), fold, a Teflon coated aluminum, filter/collection vacuum C.-bromobutyryl bromide (R=Et), and C.-bromohexanoyl manifold, designed for recovering cleavage products from bromide (R=Bu) was used to give a molar ratio of the reaction plates into collection plates. The manifold is 0.52:0.56:0.42 (in nunols). This resulted in a molar excess of designed to ensure that the filtrate from each well is directed 1.65, 1.75 and 1.31, respectively, if the original coverage of to a corresponding well in a receiving 96-well collection the resin was 0.63 mmol/g (0.5g resin per tube), and 2.4, 2.6 plate. The reaction plates (placed on the top of the collection and 1.9 if the original coverage of the resins was 0.43 plates in this manifold) were charged with a 10:85:5 mixture mmol/g (0.5g resin per tube). of TFA, dichloromethane, and methanol (0.5 ml of mixture 0.122 d. Acylation with C.-Chloro-O-Methyl Acetic acid. per well). After fifteen minutes, the solutions were filtered Each resin was prewashed with dichloromethane. Each tube and collected into proper wells on the collection plates. The was charged with 3 ml of a solution of PyBrop (0.29 g, 0.62 procedure was repeated. Solvents were evaporated on a mmole) in dichloromethane, a Solution of the C-chloro-O- SPEED VAC(R), Holbrook, N.Y., and the residual samples methylacetic acid (0.095 g, 0.62 mmole) in 3 ml of DMF, (TFA salts) were tested without further purification. and EtN(iPr). (0.2 ml, 1.2 mmole). Each reaction mixture was allowed to react for 16-18 hours at room temperature. EXAMPLE II Each resin was then filtered, washed with dichcloromethane (2x8 ml) and methanol (2x8 ml), and the acylation was Deconvolution Example repeated. Each resin was then filtered, washed with dichlo 0.129 Deconvolution of the active wells was performed romethane (2x8 ml), methanol (3x8 ml), and dried under by re-synthesis of discrete compounds, from the archived argon for about 10 minutes. Each resin was transferred into C-haloacetyl amide resins (10 resins, 0.05-0.10 g each), a Vial, and dried in a desiccator under vacuum for one hour. which were Set aside at the end of the acylation Step before 0123 D. Addition of the Second Amine the pooling. Each resin was assigned a discrete column (1, or 2, or 3, etc., see the template) in a 96 well filterplate, and 0.124 Ten, or thirty prepared C-haloacetyl amide resins was divided between X rows (A, B, C, etc), where X is the from the first three Steps were pooled together, leaving number of hits discovered in the original Screening plate. To 0.05-0.10 gram of each individual resin for necessary decon each well, in a row, a selected N2 (RR-NH) hit amine (0.1 volutions. A suspension of the pooled resin mixture in 100 mmol), DMF (180 ml) and EtNiPra (20 ml) were added: the ml of a 2:1 mixture of dichloromethane and THF was first selected amine was added to the resins in the row “A”, distributed into one, two or three, 96-well reaction plates. the second amine-to the resins in the row “B”, the third For one reaction plate, 1.7 to 2.0 grams of resin were used. amine-to the resins in the row “C”, etc. A lay-out of a For two reaction plates, 3.0 to 3.3 grams of resin were used, representative 96-well filter plate is shown in Table 4. and for three reaction plates, 4.7 to 5.0 grams of resin were used. The distributed Suspension was then filtered using a 0.130. The reaction plates were sealed and kept in an oven filtration manifold, a Small lightweight manifold that is at 70-75 C. for 16 hours. After cooling to room temperature, generally used for drawing Solvents and reagents from the the resins were filtered, washed with a 1:1 mixture of DCM chambers of the 96-well reaction plates. The reaction plates and methanol (1x1 ml), methanol (2x1 ml), and dried in were transferred into COMBICLAMPS(E) (Huntington, desiccator under vacuum for 2 h. Reduction and cleavage W.Va.), and 10% EtN(iPr) in DMF was added at 0.2 ml per were performed according to StepS 5 and 6 in the original well (0.21 mmole of EtN(iPr) per well), followed by the Synthetic protocol. The product wells from the cleavage addition of a 11.OM solution of the appropriate amine from were analyzed by ESI-MS (Electro Spray Ionization Mass the corresponding master plate, 0.1 ml per well (0.1 mmole Spectroscopy) to ensure the identity of the actives, and were amine per well). The COMBICLAMPS(R) are used to accom tested in the same Luc and MIC assayS. US 2003/0236225A1 Dec. 25, 2003 33

TABLE 4 Lay-Out of Representative 96-Well Filter Plate A. A2 A3 A4 A5 A6 A7 A8 A9 A10 Selected

G1 G2 G3 G4 G5 G6 G7 G8 G9 G10 Selected

H1 H2 H3 H4 HS H6 H7 HS H9 H1O Selected

*X* selected

to be added on the step 4 Resin Resin Resin Resin Resin Resin Resin Resin Resin Resin Individu al #1 #2 #3 #4 #5 #6 #7 #8 #9 #10 Resins #1-10, preloaded with proper amine N1.

EXAMPLE III

Solid-Phase Synthesis of Selected Substituted Compound 109 Ethylenediamine Compounds. Using the QUESTOR 210 Synthesizer 0131 The Solid-phase protocol described above in Example I was applied to the Scaled-up Synthesis of the Selected Substituted ethylene diamine compounds. Here, all reaction Steps, from the activation of the Rink-acid resin to the cleavage of the final product, were carried out using the QUESTOR instrument only, which allowed for the simulta neous Syntheses of twenty parallel reactions. Purification of all crude samples was done by HPLC to yield desirable products in purity greater than 90%. Table 3 lists the Scale-ups of Substituted ethylene diamines. Here, the Syn 0.133 1. Activation of the Rink-acid resin. Synthesis of thesis of one of the active compounds, N-Geranyl-N'-(2- Rink-Cl resin. Rink-acid resin, coverage (linker) of 0.43 to adamanthyl)ethane-1,2-diamine is described below as an 0.63 mmol/g (0.8 g., 0.5 mmol), was placed into one of the example. 10 ml tubes of QUESTOR 210 Synthesizer, and washed twice with THF. A solution of triphenylphosphine (0.380 g, 1.45 0132) The Preparation of N-Geranyl-N'-(2-adamanthyl mmol) in THF (3 ml) was added, followed by the addition )ethane-1,2-diamine (compound 109) is set forth in FIG. 12. of a solution of hexachloroethane (0.4g, 1.43 mmol) in THF US 2003/0236225A1 Dec. 25, 2003 34

(3 ml). THF was added up to the volume of the tube (100%) finishing up with acetonitrile (100%). Obtained: 27 (approximately 2 ml). After 6 hours, the resin was filtered, mg of N-Geranyl-N'-(2-adamanthyl)ethane-1,2-diamine washed with THF (2x8 ml) and dichloromethane (2x8 ml). diacetate, 24% yield, 98% purity by NMR. 0134 2. Addition of the first amine. Synthesis of resin attached geranylamine. The tube with activated resin was EXAMPLE IV charged with 3 ml of dichloroethane, EtN(iPr), (0.3 ml, 1.74 mmol), and geranylamine (0.230 g, 1.5 mmol). Dichlo Representative Solution Phase Synthesis of the roethane was added to a volume of 8 ml. The reaction was Active Compounds carried for 8 hours at 45 C., and for 6-8 hours at room temperature. Geranylamine loaded resin was filtered, 0139 Preparation of N-(Cyclooctyl)-N'-(1R,2R,3R,5S)- washed with a 2:1 mixture of dichloromethane and methanol (-)-isopinocampheylethane-1,2-diamine as hydrochloride (1x8 ml), then with methanol (2x8 ml), and Suck dried for (compound 59) is set forth in FIG. 13. 10 minutes under argon. 0135 3. Acylation with chloroacetyl chloride. Synthesis of resin attached N-Geranyl-C-chloroacetamide. The resin Compound 59 was prewashed with THF (2x8 ml). The tube was charged (S) Me with 8 ml of THF, pyridine (0.3 ml, 3.67 mmol), and Me chloroacetylchloride (0.2 ml, 2.5 mmol), and allowed to stir for 8 h at 45 C., and 6-8 hat room temperature (RT). After n1n NY (R) (R) the reaction was complete, the resin was filtered, washed H with a 2:1 mixture of dichloromethane and methanol (1x8 Me ml), methanol (2x8 ml), and THF, and the acylation was repeated using the same loads of the reagents, but shorter reaction time: 4 hours at 45 C. and 2 hours at room temperature. At the end, the C-chloroacetamide loaded resin 0140 Bromocyclooctylacetylamide. To a mixture of was filtered, washed with a 2:1 mixture of dichloromethane cyclooctylamine (3.3 g, 0.026 mol) and pyridine (2.42 g, and methanol (1x8 ml), methanol (3x8 ml), and suck dried 0.031 mmol) in anhydrous THF (80 ml) at 0°C. was added for 15 min under argon. dropwise, via syringe, bromoacetylbromide (5.78 g., 0.029 0136 4. Addition of the second amine. Synthesis of resin mol). The reaction temperature was maintained by an ice attached N-Geranyl-N'-(2-adamantyl)acetamide. The tube bath. The reaction mixture was allowed gradually to warm with the resin was charged with DMF (3 ml) and EtN(iPr). up to room temperature, and was Stirred at room temperature (0.6 ml, 4.4 mmol), followed by the addition of a suspension for 1 hour. The precipitate was removed by filtration, washed of 2-adamantamine hydrochloride (2.0 g, 1.1 mmol) in DMF with ethyl ether (1x30 ml), and the filtrate was concentrated (4 ml), and was allowed to stir at 70-75° C. for 16 hours. to dryneSS on a rotory evaporator. Bromocyclooctylacetyla After cooling down to the room temperature, the resin was mide was forwarded to the second step without additional filtered, washed with a 1:1 mixture of DCM and methanol purification. (1x8 ml), methanol (2x8 ml), and Suck dried for 15 minutes under argon. 0141 N-(Cyclooctyl)-N'-(1R,2R,3R,5S)-(–)-isopi 0137) 5. Reduction with Red-Al. Synthesis of resin nocampheyl-1-carbonylethane-1,2-diamine. To a Solution of attached N-Geranyl-N'-(2-adamantyl)ethane-1,2-diamine. the bromocyclooctylacetylamide in DMF (60 ml) were The resultant resin was suspended in anhydrous THF (3 ml) added Hunig's base (4.64 g., 0.036 mol) and (1R,2R,3R,5S)- in a tube, and stirred for 15 min. Commercially available (-)-isopinocampheylamine (4.5 g., 0.029 mol), and the reac Red-Al, 65+w % in toluene, was added (2.0 ml, 6.4 mmol), tion mixture was stirred at 80° C. for 16 hours. After cooling followed by addition of 2-3 ml of anhydrous THF (to fill up off to the room temperature, the reaction mixture was diluted the volume of the tube). The mixture was allowed to react with 150 ml of ethyl ether, and washed with 1M NaOH for 4 hours. After the reaction, the resin was filtered, washed solution (2x50ml). The organic layer was washed with brine with THF (1x8 ml), a 1:1 mixture of THF and methanol (1x8 (1x50 ml), dried over MgSO, and concentrated to dryness ml) (addition of MeOH should proceed with caution), on the rotory evaporator. The residue (11.04 g) as brown oil methanol (3x8 ml), and then dried. was purified on COMBIFLASKCR (Isco, Lincoln, Nebr., 0138 6. Cleavage from the resin and purification. Syn USA), using Silicagel catridges commercially available thesis of N-Geranyl-N'-(2-adamanthyl)ethane-1,2-diamine from BIOTAGE(R) (Biotage, Inc. of Dyax Corp, Va, USA), acetate. For this last step of the synthesis, the tube with the and the following mobile phase gradient: 30 min run, resin was charged with a 10:90 mixture of TFA and dichlo starting with DCM, 100%, and finishing up with a mixture romethane, and the formed bright red Suspension was DCM:MeOH:NHOH (600:400:10). The final product (7.29 allowed to stir for 30 min. After addition of MeOH (0.5 ml), g) was obtained as a brown oil; 76% yield, purity 90%. the colorleSS Suspension was filtered, and the filtrate was collected into a proper tube. The procedure was repeated, and solvents were evaporated on a SPEEDVAC(R). Half of nocampheylethane-1,2-diamine. To a Solution of the amide, the amount of crude N-Geranyl-N'-(2-adamanthyl)ethane-1, from previous step, in anhydrous THF (160 ml), was added 2-diamine (in a form of trifluoroacetate Salt) was purified by dropwise via syringe commercially available (SIGMA-AL HPLC using following conditions: column C18, flow 4 DRICH(R) Red-Al, as 65 wt % solution in THF (28 ml, 0.09 ml/min, 30 min run, gradient starting with 5%AcOH/MeOH mol). The reaction mixture was stirred at reflux for 20 hours. US 2003/0236225A1 Dec. 25, 2003 35

After cooling down to the room temperature, the reaction 0144 A. Library of Substituted Ethylenediamines mixture was poured into 1.5M NaOH (200 ml), and extracted with ethyl ether (2x100 ml). The organic layer was 0145 Mass spectroscopy served as a means for monitor washed with brine (1x100 ml), dried over MgSO, and ing the reaction results of the library of ethylenediamines. evaporated to dryneSS on the rotory evaporator to yield 7.2 Mass spectroScopy was done on two randomly Selected rows g of a crude product, as a brown oil. Chromatographic (24 Samples) per reaction plate, for roughly 28,000 com purification of the crude using the Same equipment and pounds in pool of 10 or 30 compounds per well. Thus, if ten conditions as for the previous Step, gave 3.5 g of the compounds per well were Synthesized, the mass Spectra for diamine. The diamine was treated with 2.0M Solution of HCl each well should contain ten Signals, correlating with the in ethyl ether (25 ml), and kept in a refrigerator overnight. proper molecular ions for each compound. The presence or A dark yellow solid (4.2 g) formed, and was filtered off, and absence of a particular signal indicated the feasibility of the recrystallized from MeOH and ethyl ether to yield 1.5 g of particular Synthesis. Based on the mass spectral data, and on the diamine as an HCl salt (of purity greater than 98%, NMR a general analysis of the reactivity of the various amines, it and MS are available), 19% overall yield. is estimated that 67,000 compounds were formed out of EXAMPLE V 112,000 compounds. 0146 FIG. 14 is a representative mass spec profile for Mass SpectroScopy Analysis one sample well. Mass spectra for a representative ethylene 0143 Mass spectra data were obtained by Elecrospray diamine compound is shown in FIG. 15. Tables 5 to 8, Ionization technique on a PERKIN ELMER(R)/SCIEX(R), below, list illustrative examples of mass Spec data for API-300, TOMS with an autosampler, manufactured by representative reaction wells, with each well containing ten SCIEXCE), Toronto, Canada. Substituted ethylene diamines.

TABLE 5

ILLUSTRATIVE EXAMPLES OF MASS SPEC DATA FOR REPRESENTATIVE ETHYLENEDAMINES TEN COMPOUNDS PER WELL). RRNH in the 2" position RNH in the 1 position (pool of (from the master plate of M + 1 of the product 10 resins) the amines) RNHCHCHNRR, Plate #4-034-2, well D10 1-(2-Aminoethyl)piperidine 2-Aminoheptane 27O absent Phenethylamine 263 4-(2-Aminoethyl)morpholifle 272 absent Tryptamine 3O2 Cyclohexylamine 241 Exo-2-Aminonorbomane 253 Benzylamine 249 2-Fluorophenethylamine 281 ? -Methylphenethylamine 277 4-Methoxyphenethylamine 293 Plate #4-56-1, well C4 4-Methylbenzylamine exo-2-Aminonorborname 259 Cyclopentylamine 223 2-(Aminomethyl)piperidifle 246 low intensity Furfurylamine 235 3,4,5-Trimethoxybenzylamine 335 1-Methy1-3-phenylpropylamine 287 Cylcobutylamine 209 1,2,3,4-Tetrahydro-1-naphthylamine 258 2,3-Dimethy1cyclohexylamime 265 2-Amino-1-butanol 227 low intensity Plate #4-44-2, well G1

Veratrylamine 4-Fluorophenethylamine 333 2-(1-Cyclohexenyl)ethylamine 291 5-Aminoquinolone 310 absent 1-(1-Naphthyl)ethylamine 337 absent 1-Aminopiperidine 266 3-Fluorobenzylamine 291 2,4-Dimethoxybenzylamine 333 3-Amino-1,2,4-triazine 262 absent 2-Ethoxybenzylamine 317 4-(3-Aminopropyl)morpholine 310 absent US 2003/0236225A1 Dec. 25, 2003 36

0147)

TABLE 6

Mass Spec Data for Synthesized Ethylenediamines

N --- YR-10 r~~ H O IM + 1 of the IM + 1 of the products, R = Ph

RNH in the 1 position products, R = H Diamines, 1 Amino alcohols, 13

Tyramine 3O8 384 258 formed 2-Adamantamine 321 absent 398 absent 272 formed cis-Myrtanylamine 324 400 274 formed 3-Amino-1-propanol 246 322 196 absent L-Methioninol 305 absent 382 absent 256 absent Cyclooctylamine 298 374 248 formed (1S,2S)-2-Amino-1-phenyl 337 absent 414 absent 288 absent 1,3-propandiol 1-Adamantanemethylamine 336 412 absent 286 formed 2,2-Diphenylethylamine 368 444 318 formed 5-Amino-1-pentanol 274 350 224 formed

0148

TABLE 7

Mass Spec Data for Synthesized Ethylenediamines, R = H and Me

N YR-10 H IM + 1" of the IM + 1" of the products, R = Me

RNH in the 1 position products, R = H Diamines, 1 Amino alcohols, 13

Tyramine 278 293 196 absent 2-Adamantamine 293 absent 307 absent 210 low intensity cis-Myrtanylamine 293 309 212 formed 3-Amino-1-propanol 217 231 134 absent L-Methioninol 277 absent 291 absent 194 formed Cyclooctylamine 269 269 absent 186 absent (1S,2S)-2-Amino-1-phenyl 309 low intensity 323 absent 226 formed 1,3-propandiol 1-Adamantanemethylamine 307 321 224 formed 2,2-Diphenylethylamine 339 353 256 formed 5-Amino-1-pentanol 245 259 162 absent