HIV-1 proviral DNA as a potential target for studying antiretroviral drug resistance. Preliminary analyses

Abstract

Background: Selection of mutations associated to resistance (MARS) is one of the factors contributing to virological failure in HIV-1 treatment. Therefore, identification of MARS in circulating virus (plasmatic viral RNA) is critical. For viral loads < 500-1000 copies/ml is very difficult to genotype. Assuming that the HIV-1 reservoirs are a genetic archive of the viral population, analyses of genotypes in proviral DNA would be useful in these patients and those that need switch with undetectable viral loads. Some limitations are the low proportion of circulating infected cells together with the sensitivity of routine tests. Objetives: Comparison of MARS and basal mutations in viral RNA and proviral DNA in HIV-1 patients from our Institution. Evaluate if the proviral DNA genotype could be a predictor of virological failure. Methods & Materials: We studied from 20 HIV-1-patients, plasma (P) and peripheral blood (B) samples, and 10 B samples from patients in failure with low viral load. Nucleic acids were extracted and the pol gene was amplified by nested PCR. Sequencing of protease (PR) and reverse transcriptase (RT) were analyzed in the Stanford database. Results: We observed a good concordance of the mutational profiles in P and B samples in 18 of 20 patients. The 2 remaining ones demonstrated mutations only in B: D30 N (highly resistant to Nelfinavir) and T215NT (low resistant to AZT and d4 T). We also observed mutations in unusual residues and more proportion of mixed bases in proviral DNA. Five of the 10 patients in failure, demonstrated MARS (M41L, K65R, A98G, K101E, K103 N, M184 V/I, G190S) coincidentally with their therapeutic schemas Conclusion: We conclude that P and B are generally concordant for studying HIV-1 genotype in certain patients. Two discordant patients were naïve explaining the lack of mutations in circulation and their presence in archival proviral DNA. The absence of mutations in DNA does not exclude their existence in lymphocytic reservoirs. It appears that genotyping the proviral HIV-1 DNA is useful in patients virologically suppressed in need of switch for treatment simplification or better tolerance/adherence and in those in failure that failed the plasma study. More data is required to evaluate the clinical impact in therapeutic decisions.