The Body Covers: The 39th Annual Meeting of the Interscience Conference on Antimicrobial Agents and Chemotherapy
Poster Session 35.I: Antiretroviral Resistance and Resistance Testing
September 26, 1999
A broad range of topics related to HIV drug resistance was covered during this session. This included the prevalence of drug resistance in certain patient populations, performance characteristics and comparisons of different resistance assays, mutations associated with certain drugs and combinations of drugs.
Briefly, genotypic resistance assays use molecular biology techniques to determine differences (codon or amino acid changes) in the HIV genetic sequence that are known to confer drug resistance. This analysis is performed primarily in the reverse transcriptase and protease genes of HIV. If there are no resistance mutations found, the virus is said to be wild type. There are several competing assays (VircoGENTM, Virco, TrueGene/Open GeneTM, Visible Genetics Inc.) all using the polymerase chain reaction (PCR) as the platform. Phenotypic resistance assays measure the growth of a patient's virus in the presence of HIV drugs. Phenotypic assays can take one of two forms. The standard assay (AIDS Clinical Trials Group/Department of Defense, ACTG/DOD) takes cells or plasma from the patient and grows that virus in the presence of HIV negative lymphocytes. That virus is then grown again in those cells in the presence of different concentrations of HIV drugs. If the virus grows in the presence of drugs, it is said to be resistant. A new type of phenotypic assay is called the recombinant virus assay (RVA). Two versions of this assay exist (AntivirogramTM, Virco and PhenoSenseTM, ViroLogic Inc). This assay takes the reverse transcriptase and protease genes of a patient's plasma virus and inserts them into another well-characterized strain of HIV. This recombinant virus is then grown in a well-characterized cell line in the presence of different HIV drugs. The result is expressed as the inhibitory concentration (IC) and is stated as a fold change (i.e. 2 fold greater concentration when compared to wild type). This type of assay takes some of the variability out of the traditional phenotypic assay. At this stage, both genotypic and phenotypic assays require at least 500 copies of plasma viral load for detection of resistance.
Four abstracts (423, 443, 444, 445) looked at the presence of HIV drug resistance in treatment naïve patients in Luxembourg, US, Spain and France respectively during the period between 1996 and early 1999. Although many natural changes were found in the sequences of these patients (particularly in the protease gene), the prevalence of drug resistance ranged from about 2-15%, depending on the drug and class. AZT and 3TC resistance appeared to be the most common. Some protease inhibitor (PI) resistance was found in the US and Spanish studies, but not in this French study. This probably reflected the fact that the French analysis was done earlier when there was less PI exposure. Of concern was the approximately 7% resistance to the nonnucleoside reverse transcriptase inhibitors (NNRTI) in the US treatment naïve study.
Abstracts 423, 424 (US), 425, and 442 (France) looked at the prevalence of resistance in a cross section of treated patients. In abstract 423, although the NRTI treated group was PI naive, mutations that conveyed resistance to saquinavir, ritonavir and indinavir were found in 10-20% of the samples. The frequency of PI mutations in a PI treated cohort in abstract 424 was higher, but in both groups the frequency of nelfinavir resistance was low. This may reflect the later introduction of nelfinavir into these populations, and not a resistance advantage of nelfinavir. Both abstracts reported high (>25 %) genotypic resistance to AZT and 3TC in these heavily NRTI treated populations. In abstract 442, phenotypic cross-resistance to PIs was described in a French cohort of patients who had been previously treated with multiple PIs. Well over two thirds of viral isolates from these patients were cross-resistant to indinavir, ritonavir, saquinavir and nelfinavir. The authors also studied whether these patient's isolates were cross-resistant to amprenavir. A significant number of patient isolates (60%) were still sensitive to amprenavir. The authors suggest that amprenavir may still retain activity in some patients who are heavily pretreated with other PIs. Abstract 429 reported on the presence of AZT related mutations in patients who had only been exposed to d4T and not AZT. There was only one patient who had genotypic evidence of d4T resistance, but over half had AZT associated mutations. Interestingly in this study, phenotypic analysis showed that despite the presence of many AZT mutation-containing isolates, all the isolates were completely sensitive to AZT and d4T.
Three abstracts (433, 434, 435) looked at the impact of reverse transcriptase mutations on the activity of adefovir (Preveon). In the presence of AZT related mutations (i.e. 215Y/F), adefovir has markedly less activity in a phenotypic assay. To the contrary, in the presence of the 3TC mutation 184V, adefovir has hypersensitivity (more activity) when compared to wild type virus and patients with this mutation had better viral load responses. In patient strains or laboratory made viruses that contained both the 215Y and 184V mutations, phenotypic activity of adefovir was restored. In addition, analysis of the reverse transcriptase enzyme containing these two mutations found that the enzyme may not function efficiently leading to a decrease in viral replication.
Abstracts 418 and 419 describe some performance characteristics for the PhenoSenseTM phenotypic assay. In abstract 418 two samples from the same patient was drawn one month apart. Since these were treated patients with detectable viral loads, a significant number of isolates were resistant to several of the 15 drugs tested. A two fold difference (considered significant) between time points was seen in about 5% of samples. Furthermore, repeated testing of the same samples on different days and by different operators, demonstrated less than a two-fold difference between samples. It was also shown that different subtypes of HIV could be assayed with equal efficiency. Finally viral mixtures containing about 20% mutant virus in the background of 80% wild type virus could be detected by this phenotyping method.
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