DOI: 10.1097/01.aids.0000189566.64729.1f
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PMID: 16227812
Issn Print: 0269-9370
Publication Date: 2005/11/04
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Excerpt
Lacombe et al. [2] reported a biphasic long-term decline in HBV-DNA load in HIV-infected patients receiving tenofovir regardless of resistance to lamivudine. We found their results very interesting, but have some doubts concerning the suitability of the assay used to monitor HBV viral load in patients with high pretreatment HBV-DNA titres above the sensitivity limit of the assay. The Roche Amplicor Monitor test (Roche Diagnostic Systems, Inc., Pleasanton, California, USA) is a very sensitive polymerase chain reaction (PCR)-based quantitative means of detecting low virus levels in patients responding to antiviral treatment if their serum HBV-DNA load decreases to levels that may be undetectable by other commercially available assays. Its linear range of detection without sample dilution is from 2 × 102 to 2 × 105 copies/ml [3], and therefore, as 10-fold serial dilution is required in all cases with a viral titre above the limit of 2 × 105 copies/ml, it is not useful for large-scale analyses of specimens with high viral loads. In our clinical and laboratory experience, the serial dilution of clinical specimens is not supported by the linearity and reproducibility of the curve because we have found that 10-fold serial dilutions of serum samples with viral loads above the dynamic range of the Roche Monitor assay give discordant under and overestimates of HBV titres, thus making it difficult to take clinical decisions concerning disease management and treatment optimization.
In an attempt to clarify the meaning of decreased linearity and reproducibility in clinical specimens with a high viral titre, we compared an in-house real-time PCR quantitation method with the Amplicor Monitor assay after the serial dilution of the HBV plasmid DNA used to construct the standard curve. The real-time PCR results were highly linear (range of linearity 103–1010 HBV-DNA copies/ml), whereas the linearity of the Amplicor Monitor test was less in samples with more than 106 HBV-DNA copies/ml. In this regard, a previous study [4] comparing the Versant HBV 3.0 (branched DNA) assay with the Amplicor Monitor test showed that the differences between the measured HBV-DNA concentrations increased with the quantitative values because of the different dynamic ranges of the two methods.
On the basis of these laboratory data, the effect of the rapid initial slope in HBV-DNA load found by Lacombe et al. [2] in the sera of patients on antiviral treatment may have been clearer if they had given important information such as the linearity of the serial sample dilution and intra-sample reproducibility of the assay, not least because 22 of their 28 patients had HBV-DNA loads greater than 2 × 105. The steeper slope of the decrease in HBV-DNA they found in patients with high HBV-DNA levels before tenofovir treatment may have been caused by a loss of linearity between two timepoints leading to an over or underestimate of HBV-DNA levels, rather than the effect of an intrahepatic immune response, particularly as lamivudine and tenofovir inhibit HBV replication and have no direct effect on the immune response.
Furthermore, the rapid initial decline in viral load was associated with the detection of YMDD mutants, thus suggesting that the YMDD motif (which induces the emergence of lamivudine-resistant strains) or other unrecognized mutations within the polymerase gene may influence viral fitness in the presence of other HBV replication inhibitors.
