title: The P236L delavirdine-resistant human immunodeficiency virus type 1 mutant is replication defective and demonstrates alterations in both RNA 5'-end- and DNA 3'-end-directed RNase H activities.
abstract: The nonnucleoside reverse transcriptase (RT) inhibitor (NNRTI) delavirdine (DLV) selects in vitro for the human immunodeficiency virus type 1 (HIV-1) RT mutation P236L, which confers high-level resistance to DLV but not other NNRTIs. Unexpectedly, P236L has developed infrequently in HIV-1 isolates obtained from patients receiving DLV; K103N is the predominant resistance mutation observed in that setting. We characterized the replication fitness of viruses derived from pNL4-3 containing P236L or K103N in both H9 and primary human peripheral blood mononuclear cell cultures infected in parallel with the two mutants. In the absence of DLV, p24 production by wild-type virus occurred more rapidly and to higher levels than with either mutant; P236L consistently demonstrated a two- to threefold decrease in p24 relative to K103N. At low levels of DLV, growth of wild-type virus was severely inhibited, and K103N replicated two- to threefold more efficiently than P236L. At high concentrations of DLV, P236L replication and K103N replication were both inhibited. Recombinant RTs containing K103N or P236L were analyzed for DNA polymerization on heteropolymeric RNA templates and RNase H degradation of RNA-DNA hybrids. Neither mutant demonstrated defects in polymerization. K103N demonstrated normal RNA 5'-end-directed RNase H cleavage and slowed DNA 3'-end-directed RNase H cleavage compared to wild-type RT. P236L demonstrated slowing of both DNA 3'-end- and RNA 5'-end-directed RNase H cleavage, consistent with its reduced replication efficiency relative to K103N. These data suggest that NNRTI resistance mutations can lead to reductions in the efficiency of RNase H cleavage, which may contribute to a reduction in the replication fitness of HIV-1.
authors.affiliation: Department of Medicine, University of Rochester School of Medicine and Dentistry, Rochester, New York 14642, USA.
authors.forename: P
authors.initials: P
authors.lastname: Gerondelis
authors.forename: R H
authors.initials: RH
authors.lastname: Archer
authors.forename: C
authors.initials: C
authors.lastname: Palaniappan
authors.forename: R C
authors.initials: RC
authors.lastname: Reichman
authors.forename: P J
authors.initials: PJ
authors.lastname: Fay
authors.forename: R A
authors.initials: RA
authors.lastname: Bambara
authors.forename: L M
authors.initials: LM
authors.lastname: Demeter
mesh_terms: D020121:5' Untranslated Regions; D019380:Anti-HIV Agents / Q000494:pharmacology*; D002460:Cell Line; D004279:DNA, Viral / Q000378:metabolism*; D003673:Defective Viruses / Q000187:drug effects / Q000502:physiology*; D020008:Delavirdine / Q000494:pharmacology*; D004352:Drug Resistance, Microbial; D054303:HIV Reverse Transcriptase / Q000235:genetics; D015497:HIV-1 / Q000187:drug effects / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D016296:Mutagenesis; D012367:RNA, Viral / Q000378:metabolism*; D018894:Reverse Transcriptase Inhibitors / Q000494:pharmacology*; D016914:Ribonuclease H / Q000378:metabolism*; D014779:Virus Replication / Q000187:drug effects*
chemical_list: D020121:5' Untranslated Regions; D019380:Anti-HIV Agents; D004279:DNA, Viral; D012367:RNA, Viral; D018894:Reverse Transcriptase Inhibitors; D020008:Delavirdine; D054303:HIV Reverse Transcriptase; D016914:Ribonuclease H