1. Hepatitis B Virus Resistance to Nucleos(t)ide Analogues
Fabien Zoulim, Stephen Locarnini 
Gastroenterology 
Volume 137, Issue 5, Pages e2 (November 2009)
DOI: /j.gastro
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2. Figure 1
Kinetics of drug resistance emergence. Panel A: Evolution of viral load and ALT levels. After an initial drop in viral load following the initiation of antiviral therapy, virologic breakthrough may occur as a consequence of antiviral drug resistance. It corresponds to the rise in serum HBV DNA levels of at least 1-log10 IU/mL compared with the lowest value during therapy (nadir value), in 2 consecutive samples 1 month apart, in patients who have previously responded and have a good treatment compliance. It may be followed by an elevation in serum ALT levels in patients who previously showed transaminases normalization under treatment. It may result in hepatitis flares and in worsening of liver histology. Panel B: Evolution of the viral quasispecies with respect to primary and secondary resistance mutations. At the beginning of therapy, wild-type virus is the major strain circulating in the patient's blood, whereas viral genomes harboring polymorphic mutations may be detected. Because of the spontaneous error rate of the viral polymerase, primary resistance mutations are usually present at levels that are undetectable by conventional diagnostic techniques. At the time of virologic breakthrough, viral genomes harboring primary resistance mutations start to emerge and become the dominant viral strains. The continuation of viral replication under the selective pressure of the drug may lead to the accumulation of additional mutations that increase the resistant mutant replication capacity (ie, secondary resistance mutations).
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3. Figure 2
Mechanisms of selection and emergence of HBV drug-resistant mutants. The main factors involved in the selection of escape mutants are: (i) the long half-life of hepatocytes and viral cccDNA; (ii) the HBV genome variability leading to a complex viral quasispecies and mutant archiving in cccDNA. The composition of the viral quasispecies evolves over time depending on the selective pressure including antiviral therapy and the host immune response. Escape mutants may then spread in the liver and become the dominant species depending on their fitness (ie, their capacity to replicate and dominate wild-type strain in the presence of antiviral pressure) and the replication space available for their dissemination in the liver. Their selection results in treatment failure.
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4. Figure 3
Primary antiviral drug resistance mutations. Polymerase gene mutations conferring resistance to nucleos(t)ide analogs are depicted. Resistance to lamivudine (LMV) and telbivudine (LdT) is conferred by mutations in the YMDD motif within the C domain of the polymerase, ie, rtM204V or rtM204I, often associated with compensatory mutations in the B domain restoring a higher replication capacity, ie, rtL180M and/or rtV173L. Resistance to adefovir (ADV) is conferred by a rtA181V or rtA181T substitution or a rtN236T substitution. The rtA181V/T substitution can also confer decreased susceptibility to LMV and LdT. Resistance to entecavir (ETV) is conferred by a combination of mutations in the B, C, or D domain of the viral polymerase, in addition to a background of substitutions at position rt204. Resistance to tenofovir (TDF) may be conferred by amino acid substitution at position rt194, which needs to be confirmed.
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5. Figure 4
Management flow chart for first virologic breakthrough/partial virologic response.
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6. Figure 5
Impact of drug resistance mutations in the viral polymerase gene on the overlapping surface gene. Panel A: Physical map showing the impact of drug resistance mutations in the viral polymerase gene on the envelope gene. Resistance mutations may therefore result in viral envelope changes leading to altered virion secretion, altered infectivity, and escape to anti-HBs antibodies. Panel B: Antiviral drug-associated HBsAg changes. The main amino acid substitutions in the viral polymerase and their corresponding changes in the envelope proteins are shown.
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7. Supplementary Figure 1
The heptatitis B virus replication cycle and the potential antiviral targets. The Figure shows all the major steps required in the viral replication cycle. HBV is able to infect hepatocytes, successful infection defined by the formation of covalently closed circular (cccDNA). Linear genomes have a high propensity to integrate into host DNA; viral genome integration is not required for viral genome replication. Following formation of cccDNA), viral messenger RNA are transcribed, including pregenomic RNA (pgRNA), the template for reverse transcription of viral DNA. In the cytoplasm, pgRNA can be bound by the viral reverse transcriptase and packaged into viral nucleocapsids, where reverse transcription takes place. Following completion of the minus strand DNA and partial completion of the plus strand DNA, virions are formed via budding into the endoplasmic reticulum. Nucleoside(tide) analogs mainly inhibit the viral minus strand DNA synthesis (reverse transcription) and plus strand DNA synthesis (DNA dependent DNA polymerase activity) within viral nucleocapsids. These drugs do not affect directly viral cccDNA, which is maintained in the nucleus of infected cells, therefore requiring long-term treatment for a sustained control of viral replication. All other steps of the viral life cycle are potential targets for antiviral therapy.
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8. Supplementary Figure 2
Public health significance of antiviral drug associated potential vaccine escape mutants (modified from Clements et al, ). Antiviral drug-associated potential vaccine escape mutants (ADAPVEMs) have the potential to infect naïve but also immunized people and may therefore jeopardize both the antiviral treatment efficacy and the hepatitis B immunization program.
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