The effects of nonnucleoside reverse transcriptase inhibitor resistance mutations on human immunodeficiency virus type 1 replication and reverse transcriptase function

The development of drug resistant Human Immunodeficiency Virus type 1 (HIV-1) variants limits the clinical efficacy of antiretroviral therapies. This work summarizes studies of the effects of nonnucleoside reverse transcriptase inhibitor (NNRTI) -resistance mutations on HIV-1 replication and reverse transcriptase (RT) function.; The replication capacities of HIVNL4-3 derivatives containing NNRTI-resistance mutations were initially evaluated in both a T cell-lymphoma line and in primary human PBMCs. A hierarchy of replication fitness was determined: wt ≅ V179D > K103N ≅ Y181C > V106A ≅ P236L. The least fit variants V106A and P236L are also uncommon in clinical isolates, suggesting that HIV-1 replication fitness can influence the frequency with which viral variants emerge during therapy.; The underlying biochemical mechanisms for these observed reductions in viral fitness were investigated. No significant differences between mutant and wild-type RTs were found in assays of DNA polymerization. NNRTI-resistant mutants did have specific effects on DNA 3′-end and RNA 5′-end-directed RNase H activities using non-viral RNA:DNA hybrids. K103N demonstrated a 1.6-fold reduction in the former activity only, whereas V179D had little effect on RNase H cleavage. V106A and P236L RTs were approximately 4 fold slower in substrate degradation during both modes of cleavage. Y181C catalyzed secondary RNase H cleavages more rapidly during both modes of cleavage.; Analyses of polypurine tract (PPT) primer processing were also conducted. K103N was similar to wild-type in PPT processing. However, V106A and P236L were slowed in PPT formation only. Y181C was enhanced in the primary cleavages associated with PPT formation and removal. These mutants had no effect on the initiation of DNA synthesis from the PPT.; Collectively, these studies demonstrate that NNRTI-resistance mutations have characteristic effects on RNase H cleavage by HIV-1 RT. These RNase H abnormalities affect processing of the PPT primer and likely result in a slowing of the initiation of plus-strand synthesis. Greater reductions in RNase H cleavage correlate with greater reductions in HIV-1 replication fitness and a lesser likelihood of developing during clinical failure of an NNRTI. These studies suggest that the selection for NNRTI-resistant variants will result in characteristic effects on reverse transcription that may influence viral pathogenicity.