Selection of a multiple drug-resistant hepatitis B virus strain in a liver-transplanted patient.
OBJECTIVE
Sequential anti-hepatitis B virus (HBV) therapy may lead to the selection of complex mutants. We analyzed the genetic and phenotypic evolution of the viral quasispecies of a patient who received successively lamivudine, add-on adefovir+lamivudine, followed by lamivudine+adefovir+hepatitis B immunoglobulins (HBIg) after orthotopic liver transplantation.


METHODS
For genotypic analysis, a 1310-bp region of the polymerase gene was amplified, cloned, and sequenced. Huh-7 cells were transfected to compare the replication fitness of HBV mutants and their susceptibility to drugs.


RESULTS
At baseline, all HBV genomes carried a wild-type (wt) RT gene but 22% harbored the sP120S and 55% the sC107stop mutations within the surface (S) gene associated with vaccine escape. Following viral breakthrough to lamivudine monotherapy, a complex mixture of lamivudine-resistant HBV strains prevailed. Interestingly, among these mutants emerged a population harboring only the rtL180M+A181V mutations, conferring lamivudine-resistance in vitro. After addition of adefovir to the ongoing treatment, viral load dropped, and the patient underwent an orthotopic liver transplantation and received HBIg. As viral load rose again, a single viral population was progressively selected, harboring the rtV173L+L180M+A181V+N236T and sP120S mutations. In vitro, this last mutant showed a level of replication reduced by only 30% compared to wt HBV and a strong resistance to both lamivudine (>1000-fold) and adefovir (>10-fold). It remained sensitive to tenofovir both in vitro and in vivo.


CONCLUSIONS
We report the selection of a complex HBV mutant that escaped the antiviral pressure of lamivudine, adefovir, and HBIg, and provide insight on the process of selection via genotypic and phenotypic analysis.
Villet
Stéphanie
S
INSERM U271, Laboratoire des Virus Hépatiques et Pathologies Associées, 151 cours Albert Thomas, 69424 Lyon cedex 03, France.
Pichoud
Christian
C
Villeneuve
Jean-Pierre
JP
Trépo
Christian
C
Zoulim
Fabien
F
D000225:Adenine / Q000008:administration & dosage / Q000031:analogs & derivatives; D000328:Adult; D000998:Antiviral Agents / Q000008:administration & dosage; D005075:Biological Evolution; D002478:Cells, Cultured; D018432:Drug Resistance, Multiple / Q000235:genetics*; D004359:Drug Therapy, Combination; D005838:Genotype; D006514:Hepatitis B Surface Antigens / Q000235:genetics / Q000276:immunology; D006515:Hepatitis B virus / Q000145:classification / Q000187:drug effects* / Q000235:genetics*; D019694:Hepatitis B, Chronic / Q000188:drug therapy*; D006801:Humans; D007136:Immunoglobulins / Q000008:administration & dosage; D019259:Lamivudine / Q000008:administration & dosage; D016031:Liver Transplantation; D008297:Male; D063065:Organophosphonates / Q000008:administration & dosage; D010641:Phenotype; D018894:Reverse Transcriptase Inhibitors / Q000008:administration & dosage; D014779:Virus Replication
D000998:Antiviral Agents; D006514:Hepatitis B Surface Antigens; D007136:Immunoglobulins; D063065:Organophosphonates; D018894:Reverse Transcriptase Inhibitors; D019259:Lamivudine; C053001:adefovir; D000225:Adenine; C045213:hepatitis B hyperimmune globulin