title: Pre-steady-state kinetic studies establish entecavir 5'-triphosphate as a substrate for HIV-1 reverse transcriptase.
abstract: The novel 2'-deoxyguanosine analog Entecavir (ETV) is a potent inhibitor of hepatitis B virus (HBV) replication and is recommended for treatment in human immunodeficiency virus type 1 (HIV-1) and HBV-co-infected patients because it had been reported that ETV is HBV-specific. Recent clinical observations, however, have suggested that ETV may indeed demonstrate anti-HIV-1 activity. To investigate this question at a molecular level, kinetic studies were used to examine the interaction of 5'-triphosphate form of ETV with wild type (WT) HIV-1 reverse transcriptase (RT) and the nucleoside reverse transcriptase inhibitor-resistant mutation M184V. Using single turnover kinetic assays, we found that HIV-1 WT RT and M184V RT could use the activated ETV triphosphate metabolite as a substrate for incorporation. The mutant displayed a slower incorporation rate, a lower binding affinity, and a lower incorporation efficiency with the 5'-triphosphate form of ETV compared with WT RT, suggesting a kinetic basis for resistance. Our results are supported by cell-based assays in primary human lymphocytes that show inhibition of WT HIV-1 replication by ETV and decreased susceptibility of the HIV-1 containing the M184V mutation. This study has important therapeutic implications as it establishes ETV as an inhibitor for HIV-1 RT and illustrates the mechanism of resistance by the M184V mutant.
authors.affiliation: Department of Pharmacology, Yale University School of Medicine, New Haven, Connecticut 06520-8066, USA.
authors.forename: Robert A
authors.initials: RA
authors.lastname: Domaoal
authors.forename: Moira
authors.initials: M
authors.lastname: McMahon
authors.forename: Chloe L
authors.initials: CL
authors.lastname: Thio
authors.forename: Christopher M
authors.initials: CM
authors.lastname: Bailey
authors.forename: Julian
authors.initials: J
authors.lastname: Tirado-Rives
authors.forename: Aleksander
authors.initials: A
authors.lastname: Obikhod
authors.forename: Mervi
authors.initials: M
authors.lastname: Detorio
authors.forename: Kimberly L
authors.initials: KL
authors.lastname: Rapp
authors.forename: Robert F
authors.initials: RF
authors.lastname: Siliciano
authors.forename: Raymond F
authors.initials: RF
authors.lastname: Schinazi
authors.forename: Karen S
authors.initials: KS
authors.lastname: Anderson
mesh_terms: D019943:Amino Acid Substitution; D000998:Antiviral Agents / Q000494:pharmacology*; D002478:Cells, Cultured; D024882:Drug Resistance, Viral / Q000187:drug effects* / Q000235:genetics; D006147:Guanine / Q000031:analogs & derivatives* / Q000494:pharmacology / Q000627:therapeutic use; D015658:HIV Infections / Q000188:drug therapy / Q000201:enzymology* / Q000235:genetics; D054303:HIV Reverse Transcriptase / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D015497:HIV-1 / Q000201:enzymology* / Q000235:genetics; D006509:Hepatitis B / Q000150:complications / Q000188:drug therapy / Q000201:enzymology; D006515:Hepatitis B virus / Q000201:enzymology / Q000235:genetics; D006801:Humans; D007700:Kinetics; D008214:Lymphocytes / Q000821:virology; D020125:Mutation, Missense; D014779:Virus Replication / Q000187:drug effects
chemical_list: D000998:Antiviral Agents; C413685:entecavir; D006147:Guanine; C514824:reverse transcriptase, Human immunodeficiency virus 1; D054303:HIV Reverse Transcriptase