Selected Phenotypic Assays Used to Evaluate Antiviral Resistance and Viral Fitness of Hepatitis B Virus and Its Variants

Abstract

Currently available antiviral therapies specifically target viral replication by blocking reverse transcription with orally given nucleos(t)ide analogues and are able to specifically suppress viral replication. The unique replication strategy of hepatitis B virus (HBV), however, allows long-term persistence of the viral genome within infected hepatocytes in spite of successful therapy. Thus, antiviral therapy needs to be continued for years. Therapy can result either in the emergence and selection of antiviral-resistant variants or the relapse of viral replication after the termination of antiviral therapy. Resistance is a major problem for 4 of the 5 approved HBV nucleos(t)ide analogues, but it is not the only reason for therapy failure. An accurate phenotypic in vitro assay for resistance allows the identification of a viral variant selected in vivo during antiviral therapy and helps to find therapeutic alternatives. Furthermore, these assays can be used to measure viral fitness and pathogenicity in vitro. With the help of these assays, the prediction of emerging viral variants with drug resistance or increased pathogenic potential can be realized. Phenotypic resistance tests for HBV are not trivial because the virus cannot be readily grown in cell culture. This review focuses on currently available phenotypic assays to evaluate antiviral resistance of HBV and fitness of viral variants in general.

Entecavir Allows an Unexpectedly High Residual Replication of HBV Mutants Resistant to Lamivudine

“…Phenotypic resistance assays for HBV are not well standardized and limited by the problem of obtaining sufficiently accurate and sensitive measures of HBV replication (for review see [19]). A major problem is the distinction between the cloned input HBV DNA used for transfection of the hepatoma cells and the newly replicated HBV DNA.…”

Detection of treatment-resistant infectious HIV after genome-directed antiviral endonuclease therapy

“…Drug resistance influences clinical treatments against viral infections [47–50], so the emergence of resistance to novel endonuclease therapies should also be considered. This is especially important for HIV, which has a high mutation rate that drives virus evolution and treatment resistance.…”

10 Jahre Nationales Referenzzentrum für Hepatitis-B-Viren und Hepatitis-D-Viren in Gießen: Tätigkeiten und Erfahrungen

“…und Validierung von bekannten und neu auftretenden, potenziellen Resistenzmutationen verfügt das NRZ über spezielle Inhousetests, mit denen die Resistenzprofile von HBV-Varianten phänotypisch in der Zellkultur quantitativ bestimmt werden können [17,20,21].…”

“…Bekannte virale Mutationen innerhalb der Reverse-Transkriptase-Domäne der HBV-Polymerase, die mit phänotypischer Resistenz gegenüber antiviralen Therapeutika assoziiert sind [ 17 ], konnten stets bei einem variablen Teil (im Mittel ca. 10 %) der untersuchten Isolate festgestellt werden.…”

“…Genotypische Resistenzmutationen gegenüber dem Wirkstoff Tenofovir sind vergleichbar mit denen gegenüber Adefovir, jedoch führen die bislang gefundenen Mutationen in den zirkulierenden HBV-Genomen nicht zu einer klinisch relevanten Resistenz unter Therapie mit verschiedenen Prodrugs des Wirkstoffs Tenofovir im klinischen Alltag [ 18 , 19 ]. Zur phänotypischen Charakterisierung und Validierung von bekannten und neu auftretenden, potenziellen Resistenzmutationen verfügt das NRZ über spezielle Inhousetests, mit denen die Resistenzprofile von HBV-Varianten phänotypisch in der Zellkultur quantitativ bestimmt werden können [ 17 , 20 , 21 ].…”