Design and Development of Bioactive Compounds: Targeting HIV - PowerPoint PPT Presentation

Design and Development of Bioactive Compounds: Targeting HIV Protease, Neuronal PDZ, and Anticancer Analogs of Heterolignans 01/23/2007 Joseck Muchiri Muhuhi

Thesis Outline Synthesis of Heterolignans • Methodology Study Using Hetero Diels-Alder/Aza-Mannich Reaction Targeting Podophyllotoxin and Justicidin Analogs PDZ3 Domain Inhibitors • Dehydroalanine and ( E )-Alkene Peptide Isosteres MDR HIV-1 Protease • Synthesis of Reduced Peptides Targeting HIV-1 MDR Protease Today’s Talk Focus Synthesis of Heterolignans • Methodology Study Using Hetero Diels-Alder/Aza-Mannich Reaction Targeting Podophyllotoxin and Justicidin Analogs PDZ3 Domain Inhibitors • ( E )-Alkene Peptide Isosteres

Figure 1 H O O Me O O HO OH OH H O N O O O O O O O O O O O MeO OMe MeO OMe MeO OMe 4' OH OMe OMe 1. Etoposide 2. Etopophos = C 4 '-phosphate 1 Podophyllotoxin X IC 50 = 50 ng/mL P388 Leukemia Cells 3. NPF (X = F) 4. GL-331 (X = NO 2 ) NH R 1 O R R NH + NR N R 2 O O O NH O O O O O MeO OMe OH MeO OMe MeO OMe OMe OH O O MeO + H 2 N OH OH HO OMe

Scheme 1 O O O HCl . Et 3 N, NaB(OAc) 3 H O O N H + H 2 N H ClCH 2 CH 2 Cl, rt O O O O quantative 1 O OH O EDC, HOBt DMF:CH 2 Cl 2 91% O O O (COCl) 2 , DMSO OH O H N N N Et 3 N, CH 2 Cl 2 LiAl(OtBu) 3 H O O O O O O O O -78 0 C, 96% THF, rt, 6h 94% 4 H 2 N R 1 3 O O O R 2 5a-h Lewis acid MeCN, rt 96% N N O N DMB R 2 N DMB R 2 + DMB = O O O 7:1 7a 6a O O J. Org. Chem . 2006 , 71, 5516-26

Figure 2 . Mechanism for trans Cycloadduct. N N O N O N O O N H O O N O O O O 6a O O Figure 3 . X-ray crystal structure

Figure 4 . Proposed Mechanism for Minor Product. O O O O H H N H N N N H O O O O H O H O H H O N H N N O O N H O O O N N N O N O O H O O O O 6 O 7a NOE 8.4% H H 8.1% N N 3 3 N DMB N DMB 2 2 1 1 H H H H O O 5.9% 7a 7a O O

Table 1 . Various Catalysts Experimented.

Table 2 . Solvents Experimented

Figure 5 . Other Alkyl Groups Used R 2 R 2 O H N N N R 2 H N DMB N DMB N + O R 1 O O O O TFA, MeCN rt, 30 min O O O H Ratio Yield N 96% 7:1 H N 95% 4:1 F H N 5:1 95% O H N 91% 8:1 H N 98% 4:1 H N 89% 2:1 H N 6:1 91% H N 2:1 42%

N -Ethyl X-ray Crystal Structure N N O O O O

N -Allyl X-ray Crystal Structure N N O O O O

Scheme 2 . Rigid Alkyl Chain O N H N N H N DMB N DM B N + O O O O O TFA, M eCN rt, 30 m in 6.2:1 93% O O O 4 9 8 J. Org. Chem . 2006 , 71, 5516-26

Scheme 3 . Facial Selectivity O O O Et 3 N, NaB(OAc) 3 H H O O + N HCl.H 2 N ClCH 2 CH 2 Cl, rt H O O O 87% O 10 O EDC, HOBt OH DMF:CH 2 Cl 2 95% O O O OH O LiAl(OtBu) 3 H N N O O O THF, rt, 6h O O 86% 12 11 O O (COCl) 2 , DMSO Et 3 N, CH 2 Cl 2 -78 0 C, 82% H R N R H N O N O H N TFA, MeCN O O O O reflux 62% O 13 14a-d O O

Scheme 4 . Correct Stereochemistry O ( CF 3 CH 2 O)P (O) CH 2 CO 2 M e OH O LiOH aq H O 16 O 87% KN(TM S) 2 , 18-Crown-6 O O 15 O THF, -78 o C 1 , E DC, HOBt 87% DM F:CH 2 Cl 2 88% O O O O O LiAl(OtB u) 3 H S wer n O H N N N H O THF ox idation O O O O O O 94% 91% 19 17 18 O O O H N TF A, MeCN 67% N N N N DM B N DMB + N DM B + O O O 7a 20 6a O O O 53% Ins eparable mixtur e, dis tinguis hable by HNM R 14% J. Org. Chem . 2006 , 71, 5516-26

Figure 6 . Proposed Mechanism involved in 6a Formation N N O N O N O N N H O O O O O 20 O O O s tepwise O addition N O O N O H N H bond N O N O N H rotation O O O O 6a O H O O

Scheme 5 . Changing the Alkene. O O O O OH LiAl(OtBu) 3 H 1 N N O OH O EDC, HOBt THF O O O O 97% 96% 22 21 O O Swern oxidation 92% O O H N N N N N DMB H N DMB + O O O O 15:1 23 O TFA, MeCN O O 24 25 67% J. Org. Chem . 2006 , 71, 5516-26

Scheme 6 . Elongating The Tether. O O O O O Et 3 N, NaB(OAc) 3 H HCl . N O H + H 2 N O H ClCH 2 CH 2 Cl, rt 26 O 87% O O EDC, HOBt OH DMF:CH 2 Cl 2 88% O O O O N O LiAl(OtBu) 3 H N OH THF, rt, 6h O O O O 91% 28 27 O O Swern oxidation 85% O H O H N NH N O O O O O Catalyst MeCN 30 62% O 29 O J. Org. Chem . 2006 , 71, 5516-26

Scheme 7 . Tweaking the Groups. O O O Et 3 N, NaB(OAc) 3 H N OH H 2 N OH + H H ClCH 2 CH 2 Cl, rt O 31 93% O O OH O EDC, HOBt DMF:CH 2 Cl 2 O 95% O H Swern oxidation N OH N O 95% O O O O 33 32 O O H N Yb(OTf) 3 , MeCN reflux, 79% N N + N N DMB DMB O O 1.6:1 35 34 O O

Conclusion • Aza Diels-Alder/ Mannich reaction proceeds with good yield • Reaction accomplished with variety of anilines & alkenes • Catalyst role is to assist in losing water molecule • Solvent dielectric constant influences product ratios • Aza Diels-Alder reaction is a stepwise and not concerted

Neuronal PDZ3 Inhibitors • Named from proteins (PSD95/SAP90, Disc large, ZO-1) • Mediate protein-protein interactions – signal transduction and assembly • Bind to short C-terminal peptides • Binding dominated by P 0 and P -2 position • Inhibition of binding leads to loss of signal or folding • PDZ3 bound to KQTSV ( Cell 1996 , 85, 1067-76)

Neuronal PDZ3 Inhibitors • Dehydroalanine Peptides – Rigid backbone and Michael acceptors O OH O OH H 2 N H 2 N O O O O O O H H H 4 H H H 4 N N N N N N H 2 N N N OH H 2 N N N OH H H H H O O O O O O 4 NH 2 OH 4 NH 2 OH H 2 N H 2 N O O O O O O H H H H H H 4 4 N N N N N N H 2 N N N OH H 2 N N N OH H H H H O O O O O O 4 4 NH 2 OH NH 2 OH • ( E )-Alkene Dipeptide Isosteres – Amide bond replaced with nonhydrolyzable E - Alkene double bond O R 2 O R 2 H H R 2 N OH N OH O R 2 H N H N N OH H N OH vs H R 1 O R 1 O N H R 1 O R 1 O R 2 H N OH d = 3.8 ang. d = 3.9 ang. R 1 O J. Org. Chem . 1994 , 59, 4875

Rationale • Replace scissile amide bond with an alkene • Val and Ser at P 0 and P -1 replaced with Val-Ala dipeptide O H H N O N O N H O O

Figure 7 . Methods Used to Synthesize Dipeptide. OMs R 1 CO 2 R 2 1. R MsOH H H N CO 2 R 2 SO 2 Ar NHSO 2 Ar R 3 Cu(CN)MgX.2LiX R 3 Cu(CN)Li.BF 3 X = Cl or Br R 3 R 3 R R CO 2 R 2 CO 2 R 2 NHSO 2 Ar NHSO 2 Ar ( L -Xaa, L -Xaa)- type EADI ( L -Xaa, D -Xaa)-type EADI Chem. Commun . 1997 , 2327 & J. Chem. Soc., Perkin Trans. 1 , 1999 , 2983 R 1 O H H Olefin Cross N OMe N 2. Boc Boc OMe + Metathesis R 2 O R 2 R 1 J. Org. Chem . 2002 , 67, 6152 Ph Ph O O O 1. LDA, PhCH 2 Br 3. BocHN OH BocHN N O 2. LiOH, H 2 O 2 Ph Ph J. Am. Chem. Soc . 2005 , 127, 15366 CO 2 R 2 R 3 R 1 "R 3 Cu" R 1 OR 2 N O anti -S N 2' NHBoc O Boc 4. O R 3 OMs "R 3 Cu" R 1 OR 2 R 1 CO 2 R 2 anti -S N 2' NHBoc O NHBoc J. Org. Chem . 1991 , 57, 4370

Figure 8 . PDZ-3 Dipeptide Needed. O OH O OH H 2 N H 2 N O O O O O H O H H H 4 H H 4 N N N N N H 2 N N N OH H 2 N N N OH H H H H O O O O O H 4 NH 2 OH 4 NH 2 OH O O H H N OH N OH N N H H O R 1 O H H N OH R 1 H O Figure 9 . Retrosynthesis O O O Fmoc Fmoc Fmoc N OH N O N O H H H OMs O O Fmoc Fmoc H 3 N N N O H H O OH OAc

Scheme 8 AcCl (Boc) 2 O MeOH Boc O HCl . O O N H 2 N H 3 N H NaHCO 3 reflux O O O quantitative 60-65% 37 36 2 steps Me DIBALH H Boc Boc MgBr N N Boc H H N BrMg OH 58% 2 steps H O H O 55:45 38 Nu 39 1. O 3 , CH 2 Cl 2 TBSCl Boc Boc X Boc N N N O H H Imidazole H OH 2. Ph 3 PCH 2 CO 2 t Bu OTBS OTBS 90% 39 40 41

Scheme 9 1. O 3 , CH 2 Cl 2 /MeOH O 2. Me 2 S Boc Boc N N O H 2. (EtO) 2 P(O)CH 2 CO 2 Me H OTBS OTBS 75 i Pr 2 NEt; LiCl; MeCN 73 66 %, 3 steps 1. TFA, CH 2 Cl 2 2. FmocCl, Et 3 N > 1% 2 steps O O Fmoc + Fmoc N O N O H H OTBS OTBS 76 77 Major Minor O O HF Fmoc Fmoc N O N O MeCN H H OTBS OH 87%, 77 78 MsCl 92% O O CuCN, i PrMgCl Fmoc Fmoc N O N O BF 3. Et 2 O, THF H H OMs 88% 79 80 O LiOH or HCl Fmoc X N OH H various conditions

Scheme 10 SOCl 2 DIBAL-H Fmoc O Fmoc H Fmoc OH N N N MeOH H H H O O O crude 100 % 82 81 MgBr Fmoc TBDMSCl Fmoc N N H Imidazole H 70%, 2 steps OTBDMS OH 76% 84 83 1. O 3 , CH 2 Cl 2 /MEOH O O 2. Me 2 S Fmoc Fmoc + N O N O 3. (EtO) 2 P(O)CH 2 CO 2 Me H H OTBS OTBS iPr 2 NEt; LiCl; MeCN 23% 3 steps 77 76 O O HF Fmoc Fmoc N O N O MeCN H H OTBS OH 87% 76 84

Scheme 11 . Stereochemistry Proof O X HN O O O 85 Piperidine, DMF O or Fmoc N O H piperidine, DMF OH 84 imidazole, DMAlP 86 O X H 2 N O OH N(CH 2 CH 2 NH 2 ) 3 87 CH 2 Cl 2 O or Fmoc N O H morpholine, DMF OH 84 86 O X Fmoc N O (Me) 2 C(OMe) 2 p-TsOH, reflux O O or 88 Fmoc N O H (Me) 2 C(OMe) 2 OH O BF 3 OEt, reflux 84 Fmoc N O H OH 84