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Prevention, Diagnosis, and Overcoming Resistance to Nucleoside Analogs

Learn about the clinical definitions of resistance, treatment failures, and factors contributing to resistance to nucleoside analogs. Discover methods for diagnosing resistance and managing drug resistance in HBV treatment.

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Prevention, Diagnosis, and Overcoming Resistance to Nucleoside Analogs

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  1. How to prevent, diagnose and overcome resistance to nucleoside analogs ? Fabien Zoulim Liver department, Hôtel Dieu Hospital & Hepatitis research laboratory, INSERM U871 Lyon, France

  2. Clinical definitions

  3. Définition of resistance • Virologic Breakthrough: Rebound in serum HBV DNA levels (e.g. 1 log10 above nadir) • Genotypic Resistance: Detection of mutations known to confer resistance while on therapy • Virologic Breakthrough with Genotypic Resistance: Viral rebound associated with a mutation(s) known to cause resistance. • Primary non response:<1log10 decrease of viral load after 3 months • Partial response: detectable HBV DNA levels during therapy Zoulim & Perrillo, J Hepatol 2008; EASL CPG, J Hepatol 2009

  4. Treatment failure Primary non response Partial response Secondary treatment failure Antiviral drug resistance Host factors Drug metabolism Patient’s compliance Drug factors Antiviral potency Drug factors Barrier to resistance Viral factors Resistant mutants Zoulim & Perrillo J Hepatol 2008; EASL CPG J Hepatol 2009

  5. Incidence of Resistance in Nucleoside Naive Patients % of patients with resistance mutations Lai et al CID 2003; Hadzyiannis et al Gastroenterology 2006; Colonno et al AASLD 2006; Di Bisceglie et al AASLD 2006; Lai et al NEJM 2007; Marcellin et al NEJM 2008

  6. Incidence of Resistance in Lamivudine Refractory Patients % of patients with resistance mutations Lampertico et al Hepatology 2005 & Gastroenterology 2007; Colonno et al AASLD 2007; Lacombe et al AIDS 2006

  7. VirologicBreakthrough BiochemicalBreakthrough Detection of Genotypic Resistance 1 log10 Kinetics of drug resistance emergence BiochemicalBreakthrough Genotypic resistance VirologicalBreakthrough 6 5 4 ALT (IU/L) HBV DNA (log10 IU/mL) Nadir 3 2 ULN 1 Antiviral drug 0 Time Si Ahmed et al. Hepatology. 2000;32:1078-1088; Zoulim Antivir Chem Chemother 2001;12: 131-142; Yuen et al Hepatology 2001; 34:784-791; Locarnini et al Antiviral Therapy 2004;9:679-693

  8. Lamivudine Resistance Accelerates Progression of Liver Disease Placebo 21% YMDDm 13% WT 5% Liaw YF et al. N Engl J Med. 2004;351:1521-1531

  9. Polymerase gene mutations reponsible for drug resistance Pol/RT Spacer RNaseH Terminal protein 349 692 845 a.a. 1 183 (rt1) (rt 344) YMDD GVGLSPFLLA I(G) II(F) A B C D E V173L L180M M204I/V LAM/ FTC A181V/T N236T I233V ? ADV M204I/V I169T T184G S202G/I M250V ETV M204I LdT A194T ? TDF Allen et al. Hepatology 1998; Gish et al. J Hepatol 2005; Qi et al. J Hepatol 2004; Tenney et al. AAC 2004 & 2007; Lai et al. Gastroenterology 2005; Sheldon et al. Antivir Ther 2005; Delaney et al. AAC 2006 ; Schildgen et al NEJM 2006 ; Borroto-Esoda JID 2007; Durantel et al Antiviral Therapy 2008; Villet et al Gastroenterology 2006, J Hepatol 2007 & 2008; Warner et al Hepatology 2008

  10. Tools for the diagnosis of resistance

  11. Dynamic ranges of quantificationof HBV DNA assays 10 102 103 104 105 106 107 108 109 Amplicor HBV Monitor v2.0 (Roche) HBV Hybrid-Capture II (Digene) Ultra-sensitive HBV Hybrid-Capture II Versant HBV DNA 3.0 (bDNA, Siemens) Cobas Taqman HBV (Roche) RealArt HBV LC PCR (Artus Biotech) Abbot Real-time HBV (Abbott) Versant HBV DNA 1.0 (kPCR, Siemens)* *in development

  12. Methods to detect genotypic resistance • Direct PCR sequence analysis • Reverse hybridization assay (INNO-LiPA DR) • Others • Restriction fragment length polymorphism (RFLP) analyses • Matrix-assisted laser desorption/ionization-Time of light Mass Spectrometry (MALDI-TOF MS)

  13. Purple precipitate Chromogen (NBT/BCIP) Alkaline Phosphatase Streptavidin Biotin Amplified target The INNO-LiPA principle Marker line Conj. Cont. Amp. Cont. L528 M528 M552 V552 I552 V555 L555 M555 DNA probe I555 Nitrocellulose strip Stuyver et al. J. Clin. Microbiol. 2000; 38: 702; Sablon E. et al Int. J. Med. Sci. 2005; 2: 8-16

  14. Phenotypic resistance testing • Determines in vitro inhibitory concentrations (IC)/effective concentration (EC) of specific HBV inhibitors relative to a wild type or reference strain • Allows the quantification of the magnitude of resistance to a drug without the need to know the responsible mutation(s) • Confirms the drug susceptibility associated with a given amino-acid change in HBV polymerase (eg. M204V/I) and determines its cross-resistance profile

  15. Results of phenotypic assays 100% Wild-type virus Patient’s virus IC50 = Drug susceptibility Inhibition of viral replication 50% IC50 patient = Fold change IC50 wild type 0% IC50 IC50 Concentration of drug

  16. Choice of drug for treatment adaptation

  17. Cross-resistance data for the most frequent resistant mutants *: (+ L180M + M204I/V). Durantel et al Hepatology 2004; Brunelle et al Hepatology 2005; Yang et al Antiviral Therapy 2005; Villet et al Gastroenterology 2006 & 2008; Delaney et al AAC 2006; Villet et al J Hepatol 2007 & 2008; Brunelle et al AAC 2007; Qi et al Antiviral therapy 2007; Tenney et al AAC 2004 & 2007

  18. Management of HBV Drug Resistance Lamivudine / Telbivudine resistance Not valid LAM FTC LdT Switch to ETV Add ADV Add TDF Switch to TVD: TDF+FTC Zoulim and Perrillo, J Hepatol, 2008; EASL CPG J Hepatol 2009

  19. Management of HBV Drug Resistance Adefovir non response Adefovir resistance Switch to TDF TVD ETV LdT Add Lamivudine Add ETV Add Telbivudine Switch to TVD: TDF+FTC Zoulim and Perrillo, J Hepatol, 2008; EASL CPG J Hepatol 2009

  20. Management of HBV Drug Resistance Entecavir resistance Not valid LAM LdT Add ADV Add TDF Switch to TVD: TDF+FTC Zoulim and Perrillo, J Hepatol, 2008; EASL CPG J Hepatol 2009

  21. Add-on or switch ?

  22. Wild type LAM LAM LAM-R ADV ADV ADV-R + Add-on therapy with drugs having a complementary cross-resistance profile Zoulim Antivir Res 2004;Villeneuve et al J Hepatol 2003; Lampertico et al Gastroenterology 2007

  23. The problem of sequential therapy and switching strategy LAM LAM 300 250 200 150 100 50 10 ADV 9 8 L180M+M204V 7 N236T Serum HBV DNA (Log10 copies/mL) 6 Reverted to wild type ALT (IU/L) 5 4 3 2 janv-98 janv-99 janv-00 janv-01 janv-02 janv-03 janv-04 janv-05 HBV DNA ALT Villeneuve et al, J Hepatol 2003

  24. Patients with lamivudine resistance: adefovir add-on strategy 3-yr cumulative probability Virologic breakthrough* and ADV resistance** Virologic breakthrough* 100 100 80 80 ADV mono ADV mono ADV+LAM Patients with virological breakthrough ADV+LAM Patients with ADV-R 60 60 40 40 30% P<0.001 P<0.001 20 20 16% 6% 0% 0 0 Months 0 3 6 9 12 15 18 21 24 27 30 33 36 0 3 6 9 12 15 18 21 24 27 30 33 36 Patients still at risk 273 268 256 225 201 158 61 229 225 217 194 179 146 57 255 238 223 213 200 177 103 242 227 214 205 200 174 92 * > 1 log rebound of HBV DNA compared to on-treatment nadir ** N236T or A181T-V in patients with a virological breakthrough Lampertico et al Gastroenterology 2007

  25. When changing treatment ?

  26. Rescue therapy in patients with clinical breakthrough Drug A Drug B Serum HBV DNA (Log10 copies/mL) and ALT (x ULN) Month of therapy

  27. Rescue therapy in patients at the time of virologic breakthrough Drug A Drug B Serum HBV DNA (Log10 copies/mL) and ALT (x ULN) Month of therapy Zoulim and Perrillo, J Hepatol, 2008; EASL CPG J Hepatol 2009

  28. Telbivudine Lamivudine Viral Load at Week 24 is a Predictor of Resistance at Week 104 of Therapy (Telbivudine vs. Lamivudine trial) HBeAg Positive, n=921 HBeAg Negative, n=446 Lai et al , NEJM, 2007

  29. Early add-on therapy to prevent drug resistance Drug A Drug B Serum HBV DNA (Log10 copies/mL) and ALT (x ULN) Month of therapy Zoulim and Perrillo, J Hepatol, 2008; EASL CPG J Hepatol 2009

  30. Secondary Treatment Preferences Based on Virologic Monitoring Nucleoside analog treatment Monitor at 12-24 weeks Monitor every 12 weeks Partial virologic response Virologic breakthrough Early non reponse Add a more potent agent* or switch to a more potent combination (emtricitabine/tenofovir*) Switch to more potent agent* * Choice based on cross-resistance data Zoulim and Perrillo, J Hepatol, 2008; EASL CPG J Hepatol 2009

  31. Very Early Add-on Therapy to Keep Viral Load as Low as Possible 1. Start with a drug having high potency and low rate of resistance 2. Add a drug with a different cross-resistance profile Drug A Drug A + Drug B Long-term viral load suppression, or risk of selection of MDR mutants ? Month of therapy outgrowth of drug resistant mutant?

  32. Prevention of drug resistance • First line therapy • Use of antivirals with high antiviral potency and high barrier to resistance • Combination therapy with complementary drugs • Second line treatment • Add-on strategies with complementary drugs preferred to sequential monotherapies • Early treatment adaptation to prevent accumulation of mutations • Choice always based on cross-resistance data • It’s prime time for the next treatment objective • Clearance of HBsAg !

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