Genotypic and phenotypic resistance: longitudinal and sequential analysis of hepatitis B virus polymerase mutations in patients with lamivudine resistance after liver transplantation.
OBJECTIVE
Lamivudine-resistant strains appear in 27-62.5% of liver transplant recipients treated with lamivudine for hepatitis B virus (HBV) recurrence, and may lead to failure of antiviral therapy. In an extension of our previous study, we investigated the molecular events associated with the emergence of lamivudine-resistant mutants in this population.


METHODS
Sequential serum samples from 10 consecutive patients with lamivudine resistance after liver transplantation were analyzed for viral genotype, precore mutants, and viral polymerase gene mutants (L528M, M552V, M552I) using restriction fragment length polymorphism. Quantitative analysis of HBV DNA was performed using hybridization assay and polymerase chain reaction.


RESULTS
Eight patients (80%) were infected with genotype D and two (20%) with genotype C. Polymerase mutants (genotypic resistance) were identified in all the patients. Phenotypic resistance (rise in serum HBV DNA titers above the detection limit of the hybridization assay) developed in five patients (50%); of the remainder, three (30%) did not have phenotypic resistance, and two were primary nonresponders. Genotypic resistance was detected earlier than phenotypic resistance (median 285 days [range 42-510] vs median 387 days [range 320-420], p = 0.055). In five patients (50%), the emergence of the YMDD mutants took over the wild type; in three (30%), the YMDD mutant took over the wild type, but the wild type re-emerged during lamivudine therapy; and in two (20%), the YMDD mutants were detected in a mixture with the wild type (in different percentages). The mean pretreatment serum ALT level was significantly lower in the patients who did not develop phenotypic resistance (p = 0.0002). The M552I pure viral population was found mainly in these patients, and all retained stable graft function (median follow-up 33 months). A high pretreatment HBV DNA level (>50 x 10(6) copies/ml) was highly statistically significantly correlated with the rapid occurrence of phenotypic resistance (r = -0.90, p = 0.04).


CONCLUSIONS
We reached the following conclusions: 1) In our area, liver transplant recipients who develop resistance to lamivudine given for recurrent HBV infection seem to be mainly infected with genotype D. 2) Re-emergence of the wild type can occur during lamivudine therapy. 3) Genotypic resistance precedes phenotypic resistance, although phenotypic resistance does not always follow genotypic resistance. 4) Quantitative determination of viremia and analysis of polymerase gene mutants are recommended for monitoring antiviral therapy of liver transplant patients with HBV reinfection in the graft.
Ben-Ari
Ziv
Z
The Liver Institute, Department of Medicine D, Rabin Medical Center, Beilinson Campus, Petah Tiqva, Israel.
Daudi
Nili
N
Klein
Athalia
A
Sulkes
Jaqueline
J
Papo
Orit
O
Mor
Eytan
E
Samra
Zmira
Z
Gadba
Rachamim
R
Shouval
Daniel
D
Tur-Kaspa
Ran
R
D000328:Adult; D002648:Child; D004259:DNA-Directed DNA Polymerase / Q000235:genetics*; D004351:Drug Resistance / Q000235:genetics*; D005260:Female; D005838:Genotype; D006515:Hepatitis B virus / Q000201:enzymology* / Q000235:genetics*; D006801:Humans; D019259:Lamivudine / Q000627:therapeutic use*; D016031:Liver Transplantation; D008137:Longitudinal Studies; D008297:Male; D008875:Middle Aged; D009154:Mutation; D010641:Phenotype; D018894:Reverse Transcriptase Inhibitors / Q000627:therapeutic use*
D018894:Reverse Transcriptase Inhibitors; D019259:Lamivudine; D004259:DNA-Directed DNA Polymerase