Pawlotsky Hcv RéSistance

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Pawlotsky Hcv RéSistance

  1. 1. Hépatite C: Résistance aux Traitements Prof. Jean-Michel Pawlotsky CNR des Hépatites B, C et delta Laboratoire de Virologie & INSERM U635 Hôpital Henri Mondor Université Paris XII Créteil
  2. 2. HCV Resistance α • HCV resistance to IFN-α therapy • HCV resistance to ribavirin ? • HCV resistance to specific antiviral molecules
  3. 3. I HCV resistance to α IFN-α Therapy
  4. 4. α Incidence of Peg-IFNα-Ribavirin Treatment Failures 60 58% 54% PEG-IFN-α2a+ribavirin (Fried et al) PEG- IFN- 2a+ribavirin 48% PEG-IFN-α2a+ribavirin (Hadziyannis et al) PEG- IFN- 2a+ribavirin PEG-IFN-α2b+ribavirin (Manns et al) PEG- IFN- 2b+ribavirin 45 30 24% 16% 18% 15 2% 0 Genotype 1 Genotypes 2/3 (Manns et al, Lancet 2001 ; Fried et al, N Engl J Med 2002 ; Hadziyannis et al, Ann Intern Med 2004)
  5. 5. Treatment Failure
  6. 6. Treatment Schedule Treatment Failure
  7. 7. Treatment Host Schedule Factors Treatment Failure
  8. 8. Treatment Host Schedule Factors Treatment Failure Disease Characteristics
  9. 9. Treatment Host Schedule Factors Treatment Failure Disease Viral Factors Characteristics
  10. 10. Treatment Host Schedule Factors Treatment Failure Disease Viral Factors Characteristics
  11. 11. SNP and SVR in the IDEAL Trial 2 4 6 8 10 12 14 16 18 20 22 Y 1 3 5 7 9 11 13 15 17 19 21 X M 30.0 IL28B -log10(P) rs12979860 15.0 P=1.37×10-28 0.0 Chromosome 19 ideogram 0M 10 M 20 M 30 M 40 M 50 M 60 M 39,623 K 39,666 K 39,708 K 39,750 K 39,793 K 39,835K 30.0 -log10(P) 15.0 0.0 PAK4 NCCRP1 SYCN IL28B AC011445.6 IL28A IL29 LRFN1 GMFG 39,711 K 39,721 K 39,732 K 39,743 K 39,753 K 39,764K 30.0 -log10(P) 15.0 0.0 IL28B AC011445.6 IL28A (Ge et al, Nature, 2009;461:399-401) 2009;461:399-
  12. 12. rs12979860 Allele and SVR (Ge et al, Nature, 2009;461:399-401) 2009;461:399-
  13. 13. rs12979860 Allele Frequency Caucasian African American ancestry ancestry n=871 n=191 12% 16% 39% 37% 49% 47% C/C C/T T/T (Ge et al, Nature, 2009;461:399-401) 2009;461:399-
  14. 14. Geographic Distribution (Thomas et al, Nature, 2009;461:798-801) 2009;461:798-
  15. 15. Effect on HCV Kinetics (Caucasians) 0 ∆ HCV RNA (Log10 IU/mL) -1.0 -2.0 -3.0 TT -4.0 CT -5.0 -6.0 CC 0 2 4 12 Weeks (Thompson et al, AASLD 2009)
  16. 16. Effect on HCV Kinetics (African Americans) 0 ∆ HCV RNA (Log10 IU/mL) -1.0 TT -2.0 CT -3.0 -4.0 -5.0 CC -6.0 0 2 4 12 Weeks (Thompson et al, AASLD 2009)
  17. 17. SVR Predictors Odds Ratio 95% CI p-value rs12979860 CC vs non-CC 5.2 4.1 6.7 <0.0001 HCV RNA ≤ 600,000 IU/mL 3.1 2.3 4.1 <0.0001 Caucasian vs African American 2.8 2.0 4.0 <0.0001 Hispanic vs African American 2.1 1.3 3.6 0.004 METAVIR score ≤F2 2.7 1.8 4.0 <0.0001 Fasting blood sugar < 5.6 mmol/L 1.7 1.3 2.2 <0.0001 (Thompson et al, AASLD 2009)
  18. 18. Summary • In patients infected with HCV genotype 1, the rs12979860 genotype: • Is strongly associated with the SVR • Explains 60% of the ethnic influence on SVR • Influences HCV kinetics on therapy • Is probably a marker of patient cell “resistance“ α to the effect of IFN-α through mechanisms that remain to be elucidated
  19. 19. Viral Resistance • Intrinsic properties of viral strains that counteract the antiviral action of antiviral drugs
  20. 20. α Incidence of Peg-IFNα-Ribavirin Treatment Failures 60 58% 54% PEG-IFN-α2a+ribavirin (Fried et al) PEG- IFN- 2a+ribavirin 48% PEG-IFN-α2a+ribavirin (Hadziyannis et al) PEG- IFN- 2a+ribavirin PEG-IFN-α2b+ribavirin (Manns et al) PEG- IFN- 2b+ribavirin 45 30 24% 16% 18% 15 2% 0 Genotype 1 Genotypes 2/3 (Manns et al, Lancet 2001 ; Fried et al, N Engl J Med 2002 ; Hadziyannis et al, Ann Intern Med 2004)
  21. 21. HCV Kinetics by Genotype EC-sponsored DITTO-Trial 7 HCV RNA (log IU/ml) * = significant difference,, 4 and 1 vs 3 difference 6 5 4 * 3 Quantitative assay cutoff * Genotype 4 Genotype 1 * 2 Qualitative assay cutoff * * * Genotype 3 1 * 0 -28 -7 01 4 7 8 15 22 29 (Pawlotsky et al., manuscript in preparation)
  22. 22. HCV Kinetics by Genotype EC-sponsored DITTO-Trial 7 HCV RNA (log IU/ml) * = significant difference,, 4 and 1 vs 3 difference 6 5 4 * 3 Quantitative assay cutoff * Genotype 4 Genotype 1 * 2 Qualitative assay cutoff * * * Genotype 3 1 * 0 -28 -7 01 4 7 8 15 22 29 (Pawlotsky et al., manuscript in preparation)
  23. 23. HCV Kinetics by Genotype EC-sponsored DITTO-Trial 7 HCV RNA (log IU/ml) * = significant difference,, 4 and 1 vs 3 difference 6 5 4 * 3 Quantitative assay cutoff * Genotype 4 Genotype 1 * 2 Qualitative assay cutoff * * * Genotype 3 1 * 0 -28 -7 01 4 7 8 15 22 29 (Pawlotsky et al., manuscript in preparation)
  24. 24. HCV Kinetics by Genotype EC-sponsored DITTO-Trial 7 HCV RNA (log IU/ml) * = significant difference,, 4 and 1 vs 3 difference 6 5 4 * 3 Quantitative assay cutoff * Genotype 4 Genotype 1 * 2 Qualitative assay cutoff * * * Genotype 3 1 * 0 -28 -7 01 4 7 8 15 22 29 (Pawlotsky et al., manuscript in preparation)
  25. 25. Antiviral Delayed resistance clearance Longer half-life of Viral Factors infected cells
  26. 26. Summary α • HCV resistance to IFN-α antiviral effect exists • Its molecular mechanisms are unknown and probably complex • It accounts for only a small part of IFN- α-based treatment failures
  27. 27. II HCV Resistance to Ribavirin ?
  28. 28. Ribavirin’s Antiviral Mechanisms • Direct inhibition of HCV RNA-dependent RNA polymerase ? • Depletion of intracellular GTP pools via IMPDH inhibition ? • RNA mutagenesis leading to "error catastrophe" ? • Enhancement of IFN-induced responses in the liver ?
  29. 29. Ribavirin’s Antiviral Effect Mean HCV RNA decrease (log IU/ml) 0.5 0.0 Controls Ribavirin monotherapy -0.5 -1.0 0 2 4 6 8 10 12 14 16 18 20 22 24 26 28 Time (days) (Pawlotsky et al., Gastroenterology 2004;126:703-14)
  30. 30. Ribavirin’s Antiviral Effect HCV RNA changes (log IU/ml) HCV RNA changes (log IU/ml) +0.5 +0.5 0.0 0.0 -0.5 -0.5 -1.0 -1.0 -1.5 -1.5 0 2 4 6 8 10 12 14 0 2 4 6 8 10 12 14 Days Days HCV RNA changes (log IU/ml) HCV RNA changes (log IU/ml) +0.5 +0.5 0.0 0.0 -0.5 -0.5 -1.0 -1.0 -1.5 -1.5 0 2 4 6 8 10 12 14 0 2 4 6 8 10 12 14 Days Days (Pawlotsky et al., Gastroenterology 2004;126:703-14)
  31. 31. Summary • Ribavirin mechanisms of action in chronic hepatitis C remain unknown • Ribavirin direct antiviral effect is modest and transient • Long-term ribavirin administration does not select for specific resistance substitutions
  32. 32. III HCV Resistance to Specific Inhibitors
  33. 33. HCV Resistance • Definition Selection of viral variants bearing amino acid substitutions that alter the drug target and thereby confer reduced susceptibility to the drug • Resistant variants are pre-existing at baseline as minor viral populations All single mutants ~10% of double mutants (Pawlotsky JM, Ther Adv Gastroenterol 2009;2:205-219; Perelson AS, unpublished data) 2009;2:205-
  34. 34. Mechanisms of Resistance sensitive resistant
  35. 35. Mechanisms of Resistance Drug sensitive resistant
  36. 36. Mechanisms of Resistance Drug sensitive sensitive resistant resistant
  37. 37. Mechanisms of Resistance Drug Stop drug sensitive sensitive resistant resistant
  38. 38. Mechanisms of Resistance Drug Stop drug sensitive sensitive sensitive resistant resistant resistant
  39. 39. Mechanisms of Resistance Drug Stop drug sensitive sensitive sensitive resistant resistant resistant + fit
  40. 40. Mechanisms of Resistance Drug Stop drug sensitive sensitive sensitive resistant resistant resistant + very fit
  41. 41. DAA Drugs Targets Receptor binding and endocytosis Transport and release Fusion and uncoating (+) RNA Virion assembly RNA replication Translation and polyprotein processing
  42. 42. HCV DAA Drug Development Phase of Development Viral entry inhibitors Preclinical I II III IV Hepatitis C immunoglobulin HCIg) HCIg) HCV-Ab 68 and Ab 65 (monoclonal Ab) HCV- HCV RNA translation inhibitors ISIS 14803 (antisense) * AVI – 4065 (antisense) Heptazyme (ribozyme) * VGX-410C (small molecule IRES inhibitor) VGX- TT 033 (siRNA) (siRNA) Posttranslational processing inhibitors NS3-4A serine proteinase inhibitors NS3- BILN 2061 * ITMN 191 VX-950 VX- SCH 503034 ACH-806/GS-9132 ACH- 806/GS- * HCV replication inhibitors NS5B polymerase inhibitors MK-0608 MK- HCV-796 HCV- * R1626 JTK-003 JTK- * NM-283 NM- * XTL 2125 Cyclophilin B inhibitors DEBIO-025 DEBIO- NIM 811 NS5A inhibitors A-831, A-689 A- Helicase inhibitors QU663 Recombinant Ab fragments Virus assembly and release inhibitors UT-231B (iminosugar-glucosidase inhibitor) UT- (iminosugar- Celgosivir (glucosidase inhibitor) (Pawlotsky JM, Chevaliez S, McHutchison JG, Gastroenterology 2007;132:1979-98)
  43. 43. Targets for New HCV Inhibitors IRES NS3 Protease RNA-Dependent RNA Polymerase
  44. 44. Resistance to NS3/4A Protease Inhibitors
  45. 45. Antiviral Efficacy of NS3/4A Protease Inhibitors Median/mean log Drug Phase Dose Duration HCV RNA reduction Telaprevir III 750 mg q8h 14 days -4.4 Boceprevir III 400 mg tid 7 days -1.6 TMC435 II 200 mg qd 7 days -4.1 ITMN-191/R7227 II 200 mg q8h 14 days -3.8 MK-7009 II 700 mg bid 8 days -4.7 BI201335 II 240 mg qd 14 days -4.0 Narlaprevir II 400 mg bid 7 days -4.2 BMS-650032 I 300 mg bid 3 days -3.3
  46. 46. Amino Acid Substitutions Associated with Resistance Macrocyclic Linear BILN 2061 R7227 VX-950 SCH503034 (Ciluprevir) (ITMN-191) (Telaprevir) (Boceprevir) A156S/V, In vitro A156V/T, D168A, Q41R, A156S/T, D168A/V, A156S/T/V F43S, S138T, T54A, V170A R155Q S489L In vivo A156S/T/V, V36M/A, No data No data R155K/T, T54A/S, R155K T54A, V36A/M (Kwong et al., Curr Opin Pharmacol, 2008;8:522-531; Susser et al., J Hepatol 2009;50(Suppl.1):S7) Pharmacol, 2008;8:522- 2009;50(Suppl.1):S7)
  47. 47. Amino Acid Substitutions Associated with Resistance Arg155 Asp168 Ala156 Thr54 Val36 (Pawlotsky J-M, Ther Adv Gastroenterol 2009;2: 205-219) J- 205-
  48. 48. Telaprevir (Vertex & Tibotec)
  49. 49. Paramètres Associés à la Résistance • Niveau de résistance au médicament conféré par la ou les substitutions amino acidiques • In vivo “fitness“ des variants viraux sélectionnés
  50. 50. PROVE 2 Trial (Phase II) Naïve, Genotype 1, Europe PR48 α Placebo + Peg-IFNα2a + Ribavirin (RBV) Follow-up Follow- Telaprevir 750 mg q8h T12/PR24 α + Peg-IFNα2a α Peg-IFNα2a + RBV Follow-up Follow- + RBV Telaprevir 750 mg q8h T12/PR12 α + Peg-IFNα2a Follow-up Follow- + RBV Telaprevir 750 mg q8h T12/P12 Follow-up Follow- α + Peg-IFNα2a 0 12 24 48 72 Weeks on therapy (Hézode et al., N Engl J Med 2009;360:1839-50) 2009;360:1839-
  51. 51. PROVE 2 Trial (Phase II) Breakthoughs through week 12 100 80 12-week SVR rates (%) 60 40 24% 20 3% 0 T12/P12 (no RBV) T12/PR12 + T12/PR24 (Hézode et al., N Engl J Med 2009;360:1839-50) 2009;360:1839-
  52. 52. PROVE 2 Trial (Phase II) Relapses after treatment 100 80 SVR rates (%) 60 48% 40 30% 22% 20 14% 0 PR48 T12/P12 (no RBV) T12/PR12 T12/PR24 (n=82) (n=78) (n=82) (n=81) (Hézode et al., N Engl J Med 2009;360:1839-50) 2009;360:1839-
  53. 53. Boceprevir (SP/Merck)
  54. 54. Patterns of Response to Boceprevir N=22 non-responders to previous standard treatment who were dosed for 14 days with BOC (Phase 1b) (Susser et al., Hepatology 2009;50:1709-18) 2009;50:1709-
  55. 55. In vitro Resistance to In Boceprevir (replicon) (replicon) (Susser et al., Hepatology 2009;50:1709-18) 2009;50:1709-
  56. 56. Relative in vivo Fitness of Boceprevir-Resistant Variants Boceprevir-Resistant (Susser et al., Hepatology 2009;50:1709-18) 2009;50:1709-
  57. 57. Dynamics of NS3 Protease Variants (Susser et al., Hepatology 2009;50:1709-18) 2009;50:1709-
  58. 58. Summary • During telaprevir or boceprevir monotherapy, pre- existing resistant HCV variants are rapidly selected in virtually all treated patients • Both single mutants (36, 54, 155 and 156) and double mutants (36/155, 26/156) can be selected • The corresponding viral variants generally have reduced fitness compared to wild-type HCV • Most of these amino acid substitutions confer cross- resistance to all of the NS3/4A protease inhibitors in development
  59. 59. Summary (cont’d) • Failure of the triple combination of pegylated IFN-α, ribavirin and telaprevir to clear HCV is associated with the selection of telaprevir-resistant variants • These variants may persist after therapy or be progressively replaced by wild-type virus • Telaprevir resistant variants remain sensitive to pegylated interferon and ribavirin
  60. 60. Resistance to Inhibitors of HCV Replication
  61. 61. Inhibitors of HCV Replication • RNA-dependent RNA polymerase (RdRp) inhibitors • Nucleoside analogues • Non-nucleoside inhibitors (NNIs) • NS5A inhibitors • Cyclophilin inhibitors • miR122 antagonists
  62. 62. In vitro Resistance 4’ azido-cytidine azido- Ser 96 2’-methyl nucleosides Ser 282 Copyright © 2006 Merck Co., Inc., Whitehouse Station, New Jersey, USA All rights reserved
  63. 63. HCV Resistance to 2’-C-Methyl Nucleoside Inhibitors 2’C-Me-ATP in the catalytic site (Migliaccio et al., J Biol Chem 2003;278:49164-70)
  64. 64. 7-Deaza-2’-C-Methyl-Adenosine (MK-0608) in Chimpanzees wt S282R/T/I wt S282 MK-0608 at 1mg.kg-1 orally 7 6 HCV RNA (Log IU/mL) 5 4 3 2 - 4,6 - 4,1 LOQ 1 (20 IU/mL) TMA + + + + - - - - - - - - + + 0 -15 -10 -5 0 5 10 15 20 25 30 35 40 45 50 55 60 65 70 Days (Carroll et al., Antimicrob Agents Chemother 2009;53:926-934) 2009;53:926-
  65. 65. Valopicitabine (NM283) Phase IIb, IFN Nonresponders Week 24 0 n=7 -0.27 -0.5 HCV RNA reduction -1 NM283 800 mg monotherapy -1.5 -2 -2.5 -3 -3.5 0 4 8 12 16 20 24 28 32 36 40 44 48 Weeks (Afdhal et al, EASL 2006)
  66. 66. Antiviral Efficacy of NNIs Median/mean log Drug Phase Dose Duration HCV RNA reduction GS-9190 II 40 mg bid 8 days -1.4 Filibuvir II 300 mg bid 8 days -2.1 ANA598 II 800 mg bid 3 days -2.9 BI207127 II 800 mg q8h 3 days -3.1 VCH-759 II 400 mg tid 10 days -1.7 ABT-333 II 600 mg bid 2 days -1.5 VX-222 Ib 750 mg bid 3 days -3.7 MK-3281 Ib 800 mg bid 7 days -1.3 (1a), -3.8 (1b)
  67. 67. In vitro Resistance NNI site C Benzothiadiazine NNI site A Asn 411, Met 414, Tyr 448 Indoles, Benzimidazoles Indoles, Pro 495, Pro 496, Val 499 A Allosteric GTP B E C D C NNI site B Thiophene-COOH Thiophene- Met 423 NNI site D Benzofurans NNI site E (hypothetical) Cys 316, Val 201 Cys 445, Tyr 448, Tyr 452 Copyright © 2006 Merck Co., Inc., Whitehouse Station, New Jersey, USA All rights reserved
  68. 68. HCV 796 (ViroPharma Wyeth) Antiviral efficacy 1 Treatment Follow-up Log HCV RNA change 0 -1 Placebo 50 mg 100 mg 250 mg -2 500 mg 1000 mg 1500 mg -3 -1 2 5 8 11 14 17 20 23 26 29 (Chandra et al, DDW 2006) Study Day
  69. 69. Prevention of Resistance
  70. 70. Prevention of Resistance to HCV Inhibitors • Exclude use as a monotherapy • Combine with one or several other antivirals with: • At least additive antiviral effect • No cross-resistance
  71. 71. R7128/R7227 Combination INFORM Trial 1 Mean log10 HCV RNA change from Placebo 0 -1 R7128 500b D1-7/R7227 100t D4-7-Na -2 R7227 100t D1-7/R7128 500b D4-7-Na -3 -4 500b/100t 1000b/200t -5 500b/200t 1000b/100t -6 -7 0 2 4 6 8 10 12 14 Day (Gane et al., EASL 2009)
  72. 72. R7128/R7227 Combination INFORM Trial 1 Mean log10 HCV RNA change from Placebo 0 -1 -2 -3 -4 1000b/600b-TE -5 1000b/900b-TE 1000b/900b-Na -6 -7 0 2 4 6 8 10 12 14 Day (Gane et al., AASLD 2009)
  73. 73. Cure with IFN-Free Regimens ?
  74. 74. Combination of MK-7009 and MK-608 (Merck) in vitro 10 5-10 5 Synergy in the genotype 1b 0-5 replicon at low MK-0608, high MK-7009 doses 0 -5-0 -5 0.3 0.02 0.15 MK-7009 NS3 12 1.20 inhibitor (nM) MK-608 pol inhibitor (mM) (Olsen et al., APASL 2008)
  75. 75. Combination of MK-7009 and MK-608 (Merck) in vivo 6 MK 7009 MK 7009 + MK-608 monotherapy Chimp A 5 Chimp B Chimp C log IU/mL 4 3 SVR 2 LOQ 1 0 0 50 90 130 170 210 250 Day (Olsen et al., APASL 2008)

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