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

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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

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SLIDE 1

Design and Development of Bioactive Compounds: Targeting HIV Protease, Neuronal PDZ, and Anticancer Analogs of Heterolignans

01/23/2007

Joseck Muchiri Muhuhi

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SLIDE 2

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
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SLIDE 3

O O O O MeO OMe OMe N NH O OH MeO OMe R O O NH R1 R2 O NR O OH MeO OMe

+

HO OH O OMe MeO H2N OH O

+

O O O O O O O O MeO OMe OH

4'

Me H HO OH X NH

  • 1. Etoposide
  • 2. Etopophos = C4'-phosphate
  • 3. NPF (X = F)
  • 4. GL-331 (X = NO2)

O O O O MeO OMe OMe OH Podophyllotoxin H N O O O O MeO OMe OMe 1 IC50 = 50 ng/mL P388 Leukemia Cells R

Figure 1

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SLIDE 4

Scheme 1

  • J. Org. Chem. 2006, 71, 5516-26

O O H O O O N H O O O O N O O O O H2N O O Et3N, NaB(OAc)3H ClCH2CH2Cl, rt quantative + O OH O EDC, HOBt DMF:CH2Cl2 91% O O N OH O O LiAl(OtBu)3H THF, rt, 6h 94% O O N H O O O (COCl)2, DMSO Et3N, CH2Cl2

  • 78 0C, 96%

H N R1 N N O DMB O N N O DMB O Lewis acid MeCN, rt 96% + 7:1 1 2 3 4 6a 7a R2 5a-h R2 R2 O O DMB = HCl.

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SLIDE 5

Figure 2. Mechanism for trans Cycloadduct. Figure 3. X-ray crystal structure

N N O O O O N N O O O O H N N O O O O 6a

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SLIDE 6

Figure 4. Proposed Mechanism for Minor Product. NOE

N N O O O N N O O O H H H H N N O O O O H H H N N H O O O H H O O O N N O O O O H N N O O O O 6

7a

N N O DMB O N N O DMB O H H H H H H 8.4% 5.9% 8.1%

1 2 1 2 3 3

7a 7a

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SLIDE 7

Table 1. Various Catalysts Experimented.

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SLIDE 8

Table 2. Solvents Experimented

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SLIDE 9

Figure 5. Other Alkyl Groups Used

O O N H O O O H N R2 N N R2 O DMB O N N R2 O DMB O TFA, MeCN rt, 30 min + H N H N F H N O H N H N H N H N 7:1 4:1 5:1 8:1 4:1 6:1 R1 H N 2:1 2:1 96% 95% 95% 91% 98% 89% 91% 42% Ratio Yield

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SLIDE 10

N-Ethyl X-ray Crystal Structure

N N O O O O

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SLIDE 11

N-Allyl X-ray Crystal Structure

N N O O O O

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SLIDE 12

Scheme 2. Rigid Alkyl Chain

  • J. Org. Chem. 2006, 71, 5516-26

O O N H O O O N N O DMB O N N O DM B O TFA, M eCN rt, 30 m in 93% + H N 6.2:1 8 9 4

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SLIDE 13

Scheme 3. Facial Selectivity

O O H O O O N H O O O O N O O O O HCl.H2N O O Et3N, NaB(OAc)3H ClCH2CH2Cl, rt 87% + O OH O EDC, HOBt DMF:CH2Cl2 95% O O N OH O O LiAl(OtBu)3H THF, rt, 6h 86% O O N H O O O (COCl)2, DMSO Et3N, CH2Cl2

  • 78 0C, 82%

H N R N N R O O TFA, MeCN reflux 62% 10 11 12 13 14a-d H O O

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Scheme 4. Correct Stereochemistry

  • J. Org. Chem. 2006, 71, 5516-26

O H O O O N O O O LiAl(OtB u) 3H THF 94% H N N N O DM B O N N O DM B O TF A, MeCN 67% 6a 7a ( CF3CH2O)P (O) CH2CO2M e KN(TM S) 2, 18-Crown-6 THF, -78 oC 87% O O O O OH O LiOH aq O 1, E DC, HOBt DM F:CH2Cl2 88% 87% O O N O H O O O O N O O O H S wer n

  • x idation

91% N N O DMB O 20 + + Ins eparable mixtur e, dis tinguis hable by HNM R 53% 15 16 17 18 19 14%

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SLIDE 15

Figure 6. Proposed Mechanism involved in 6a Formation

O H N N O O O N N O O O O H N N O O O O O N N O O O O H H s tepwise addition N N O O O H bond rotation N N O O O O 20 6a

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SLIDE 16

Scheme 5. Changing the Alkene.

  • J. Org. Chem. 2006, 71, 5516-26

LiAl(OtBu)3H THF 96% O O N O O O 1 O O OH O EDC, HOBt 97% O O N OH O O O O N O O O H Swern

  • xidation

92% N N O DMB N N O DMB O O H N TFA, MeCN 67% + 15:1 21 22 23 24 25

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Scheme 6. Elongating The Tether.

  • J. Org. Chem. 2006, 71, 5516-26

HCl. O O H O O O N H O O O N O O O H2N O Et3N, NaB(OAc)3H ClCH2CH2Cl, rt 87% + O OH O EDC, HOBt DMF:CH2Cl2 88% LiAl(OtBu)3H THF, rt, 6h 91% H N Catalyst MeCN 62% 26 27 O O O O O N OH O O 28 Swern oxidation 85% O O N O O O 29 H O O NH O O 30

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Scheme 7. Tweaking the Groups.

O O H O O O N H OH O O N OH O O H2N OH Et3N, NaB(OAc)3H ClCH2CH2Cl, rt 93% + O OH O EDC, HOBt DMF:CH2Cl2 95% H N Yb(OTf)3, MeCN reflux, 79% 31 32 O O N O O O 33 Swern oxidation 95% H N N O O DMB N N O O DMB + 1.6:1 34 35

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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
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SLIDE 20

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 P0 and P-2 position
  • Inhibition of binding leads to loss of signal or folding
  • PDZ3 bound to KQTSV

(Cell 1996, 85, 1067-76)

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SLIDE 21

Neuronal PDZ3 Inhibitors

  • Dehydroalanine Peptides – Rigid backbone and Michael acceptors
  • (E)-Alkene Dipeptide Isosteres – Amide bond replaced with nonhydrolyzable E-

Alkene double bond

H2N H N N H H N N H H N O NH2 H2N O O OH O O OH O 4 4 OH O H2N H N N H H N N H H N O NH2 H2N O O OH O O OH O 4 4 OH O H2N H N N H H N N H H N O NH2 H2N O O OH O O OH O 4 4 H2N H N N H H N N H H N O NH2 H2N O O OH O O OH O 4 4 H N N H R1 O R2 H N R1 R2 H N N H R1 R2 O O O O OH OH OH H N R1 R2 O OH H N N H R1 O R2 O OH vs d = 3.8 ang. d = 3.9 ang.

  • J. Org. Chem. 1994, 59, 4875
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Rationale

  • Replace scissile amide bond with an alkene
  • Val and Ser at P0 and P-1 replaced with Val-Ala dipeptide

H N N H O O O H N O O

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Figure 7. Methods Used to Synthesize Dipeptide.

N R1 H H CO2R2 SO2Ar R CO2R2 NHSO2Ar R3 R CO2R2 NHSO2Ar OMs R CO2R2 NHSO2Ar R3 MsOH R3Cu(CN)Li.BF3 R3Cu(CN)MgX.2LiX X = Cl or Br (L-Xaa, L-Xaa)- type EADI (L-Xaa, D-Xaa)-type EADI H N R2 OMe O R1 + Olefin Cross Metathesis Boc H N R2 Boc OMe O R1 CO2R2 R1 NHBoc OMs N O R1 CO2R2 O Boc "R3Cu" anti-SN2' R1 NHBoc OR2 O R3 BocHN N O O O Ph Ph

  • 1. LDA, PhCH2Br
  • 2. LiOH, H2O2

BocHN OH O Ph Ph "R3Cu" anti-SN2' R1 NHBoc OR2 O R3

1. 2. 3. 4.

  • Chem. Commun. 1997, 2327 & J. Chem. Soc., Perkin Trans. 1, 1999, 2983
  • J. Am. Chem. Soc. 2005, 127, 15366
  • J. Org. Chem. 2002, 67, 6152
  • J. Org. Chem. 1991, 57, 4370
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Figure 8. PDZ-3 Dipeptide Needed. Figure 9. Retrosynthesis

N H OH Fmoc N H OAc O O Fmoc N H OMs O O Fmoc N H O O Fmoc N H OH O Fmoc H3N O O

H N N H O H N R1 H N N H R1 O O O O OH OH OH H2N H N N H H N N H H N O NH2 H2N O O OH O O OH O 4 4 OH O H H H2N H N N H H N N H O NH2 H2N O O OH O OH O 4 4 OH O H H

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SLIDE 25

Scheme 8

H3N O O AcCl MeOH reflux quantitative H2N O O (Boc)2O NaHCO3 N H O O Boc DIBALH MgBr N H OH Boc 60-65% 2 steps 58% 2 steps 55:45 O H BrMg N Boc H Me Nu 36 37 39 N H H O Boc 38 HCl.

N H OTBS Boc TBSCl Imidazole 90%

  • 1. O3, CH2Cl2
  • 2. Ph3PCH2CO2tBu

N H OTBS O Boc

X

N H OH Boc 40 41 39

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SLIDE 26

Scheme 9

N H OTBS Boc N H OTBS Boc O O

  • 1. O3, CH2Cl2/MeOH
  • 2. Me2S
  • 2. (EtO)2P(O)CH2CO2Me

iPr2NEt; LiCl; MeCN 66 %, 3 steps

  • 1. TFA, CH2Cl2
  • 2. FmocCl, Et3N

> 1% 2 steps N H OTBS O O Fmoc + Minor N H OTBS O O Fmoc Major 73 75 77 76

N H O O Fmoc HF MeCN 87%, N H O O Fmoc CuCN, iPrMgCl BF3.Et2O, THF 88% N H O O Fmoc

X

LiOH or HCl various conditions N H OH O Fmoc OH OTBS 78 77 80 MsCl 92% N H O O Fmoc OMs 79

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SLIDE 27

Scheme 10

N H O O Fmoc DIBAL-H MgBr N H OTBDMS Fmoc N H OH Fmoc

  • 1. O3, CH2Cl2/MEOH
  • 2. Me2S
  • 3. (EtO)2P(O)CH2CO2Me

iPr2NEt; LiCl; MeCN 23% 3 steps 70%, 2 steps Imidazole 76% N H OTBS O O Fmoc N H OTBS O O Fmoc + SOCl2 MeOH crude 100 % TBDMSCl 81 83 84 77 76 N H OH O Fmoc N H H O Fmoc 82 N H OTBS O O Fmoc HF MeCN 87% N H OH O O Fmoc 76 84

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Scheme 11. Stereochemistry Proof

HN O O O O 85 N H OH O O Fmoc 84 86

X

Piperidine, DMF

  • r

piperidine, DMF imidazole, DMAlP H2N O O OH N H OH O O Fmoc 84 86

X

N(CH2CH2NH2)3 CH2Cl2

  • r

morpholine, DMF 87

N H OH O O Fmoc 84

X

N O O Fmoc O (Me)2C(OMe)2 p-TsOH, reflux

  • r

(Me)2C(OMe)2 BF3OEt, reflux N H OH O O Fmoc 84 88

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SLIDE 29

Scheme 12

N H O H O O Fm oc CF3 Cl O Ph OM e N H O O O Fm oc O CF3 P h OM e N H O O O Fm oc O CF3 MeO P h N H OH O O Fmoc CF3 Cl O M eO P h S R 42% DM AP , P y CH2Cl2 81% DMA P, Py CH2Cl2 84 84 89 90 N H Fm oc O O O

  • 0.07

+0.08 +0.11 MTP A

  • 0.02

H H H value change of MTP A es ter s 89 and 90

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SLIDE 30

Figure 13. New Strategy Needed

1. 2.

N H O Fmoc N H N O Fmoc O MgBr THF N H Fmoc Reduction OH N H OMe O Fmoc BuLi CH3P(O)(OMe)2 N H O Fmoc P O (OMe)2 N H O Fmoc CO2R Reduction N H Fmoc CO2R OH HEW

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Scheme 13

N H N O Fmoc O MgCl 1.

  • 2. Al2O3, ether

N H O Fmoc N H OH Fmoc NaBH4 CeCl3.7H20 91% 88:12 N H O Fmoc N O N DMAP benzene reflux HN O O HN O O + HN O O H H 5.0% 4.5% CH3 HN O CH3 O H H 11.4% 23.7% DMAP 96 97 98 99 100 94 89% 84% 2 steps N O N 2

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SLIDE 32

Scheme 14

N H OAc Fmoc N H OH Fmoc

  • 1. O3, CH2Cl2: MeOH
  • 2. (EtO)P(O)CH2CO2CH3

iPr2NEt; LiCl; MeCN N H OAc O O Fmoc N H OH O O Fmoc 61% 2 steps Na2CO3 87% TBSCl Imidazole 75% N H OTBS O O Fmoc N H OTBS O O Fmoc + Ac2O py HF, MeCN 87% N H OH O O Fmoc

  • 1. MsCl, py
  • 2. CuCN, iPrMgCl

BF3.OEt, THF

  • 78 oC, 88%

N H O O Fmoc 102 103 73 74 82 76 79

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SLIDE 33

Scheme 15. Trying to Demethylate

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SLIDE 34

Scheme 16

N H OH O HN(OMe)Me carbonylimidazole CH2Cl2 92% N H N O O Boc N H O Boc MgCl THF 1.

  • 2. Al2O3, ether

94%, 2 steps LiAl(OtBu)3H EtOH, -78 oC 92%, 95:5 N H Boc OH HN O O NaH THF 59% 104 103 104 100 Boc 16 M NaOH THF/MeOH 64% HN O O H H 4.9% 6.0% HCl.

N H B oc O Ac OAllyl O N H Boc OH OA llyl O N H Boc O H OMe O + 1:1 106 108 107

  • 1. O3, MeOH/CH2Cl2
  • 2. M e2S
  • 3. (E tO)2P( O)CH2CO 2Allyl

71%, 3 s teps N H Boc OH Ac 2O DMA P, py 99% 104 N H Boc OA c 105 Na2CO3 M eOH 74%

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SLIDE 35

Scheme 17

N H B oc OH OAlly l O N H Fmoc OH OA llyl O

X

Ms Cl TFA CH2Cl2 N H Boc OA c OA llyl O Na2CO 3 77% OH/H2O N H Fmoc OA llyl O N H Fmoc OH O Pd(P Ph3)4 morpholine M sCl, py 90% 106 108 110 112 Fmoc-O NS u TEA M eCN:H2O 75%, 2 s teps H3N OH OAlly l O 109 N H Boc OM s OA llyl O N H Fmoc OM s OA llyl O 111 CuCN iPrM gCl BF3O Et2 89%

X

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SLIDE 36

Scheme 18

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Scheme 19. Summary

N H OH O HCl.HN(OMe)Me carbonyldiimidazole 92% N H N O O Boc N H O Boc MgCl 1.

  • 2. Al2O3

94%, 2 steps LiAl(OtBu)3H 92%, 95:5 N H Boc OH

  • 1. Ac2O, DMAP, Py
  • 2. O3, MeOH/CH2Cl2
  • 3. Me2S
  • 4. (EtO)2P(O)CH2CO2Allyl

70%, 4 steps N H Boc OAc O O N H Boc OH O O N H Fmoc OH O O Na2CO3 77% TFA CH2Cl2 N H Fmoc O O N H Fmoc OMs O O MsCl 90% CuCN, iPrMgCl BF3.OEt2 89% Pd(PPh3)4 NaBH4 cr 100% N H Fmoc OH O 100 111 107 101 102 103 109 110 105 Boc H3N OH O O Fmoc-ONSu 75%, 2 steps 108

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SLIDE 38

Scheme 20

100 113 H2N H N N H H N N H H N H2N O NH2 O O OH O OH O OH O H2N H N N H H N N H H N H2N O NH2 O O OH O OH O OH O O H2N H N N H H N N H H2N O NH2 O O OH O OH O OH O N H Fmoc O O OH 2,4,6-trichlorobenzoyl chloride py, CH2Cl2

  • r

HOBt, EDC, DMAP N H Fmoc OH O 4 eq 1.0 eq 114 115 116 4 4 4 4 4 4

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Conclusion

  • Synthesis of a dipeptide isostere in gram quantities accomplished
  • Route amenable to other amino acids

H N N H O H N H N N H O H N H N CF3 F Dipole moments and bond length close to amide bond H N CH3

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Acknowledgement

Advisor

  • Dr. Mark R. Spaller

Lab Members Funding

W.S.U NIH (GM 63021) IMSD Fellowship (NIH GM 58905-08) Professors I was lucky to interact with Family

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SLIDE 41

Questions?