title: Exploiting drug-resistant enzymes as tools to identify thienopyrimidinone inhibitors of human immunodeficiency virus reverse transcriptase-associated ribonuclease H.
abstract: The thienopyrimidinone 5,6-dimethyl-2-(4-nitrophenyl)thieno[2,3-d]pyrimidin-4(3H)-one (DNTP) occupies the interface between the p66 ribonuclease H (RNase H) domain and p51 thumb of human immunodeficiency virus reverse transcriptase (HIV RT), thereby inducing a conformational change incompatible with catalysis. Here, we combined biochemical characterization of 39 DNTP derivatives with antiviral testing of selected compounds. In addition to wild-type HIV-1 RT, derivatives were evaluated with rationally designed, p66/p51 heterodimers exhibiting high-level DNTP sensitivity or resistance. This strategy identified 3',4'-dihydroxyphenyl (catechol) substituted thienopyrimidinones with submicromolar in vitro activity against both wild type HIV-1 RT and drug-resistant variants. Thermal shift analysis indicates that, in contrast to active site RNase H inhibitors, these thienopyrimidinones destabilize the enzyme, in some instances reducing the Tm by 5 °C. Importantly, catechol-containing thienopyrimidinones also inhibit HIV-1 replication in cells. Our data strengthen the case for allosteric inhibition of HIV RNase H activity, providing a platform for designing improved antagonists for use in combination antiviral therapy.
authors: RT Biochemistry Section, HIV Drug Resistance Program, National Cancer Institute , Frederick, Maryland 21702, United States.
authors: Takashi
authors: T
authors: Masaoka
authors: Suhman
authors: S
authors: Chung
authors: Pierluigi
authors: P
authors: Caboni
authors: Jason W
authors: JW
authors: Rausch
authors: Jennifer A
authors: JA
authors: Wilson
authors: Humeyra
authors: H
authors: Taskent-Sezgin
authors: John A
authors: JA
authors: Beutler
authors: Graziella
authors: G
authors: Tocco
authors: Stuart F J
authors: SF
authors: Le Grice
mesh_terms: D000998:Antiviral Agents / Q000737:chemistry / Q000494:pharmacology; D024882:Drug Resistance, Viral / Q000187:drug effects* / Q000235:genetics; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D004795:Enzyme Stability / Q000187:drug effects; D054303:HIV Reverse Transcriptase / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics; D015497:HIV-1 / Q000187:drug effects / Q000201:enzymology / Q000235:genetics; D006801:Humans; D008958:Models, Molecular; D015394:Molecular Structure; D009154:Mutation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011744:Pyrimidinones / Q000737:chemistry / Q000494:pharmacology*; D018894:Reverse Transcriptase Inhibitors / Q000737:chemistry / Q000494:pharmacology; D054309:Ribonuclease H, Human Immunodeficiency Virus / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics; D013696:Temperature
chemical_list: D000998:Antiviral Agents; D004791:Enzyme Inhibitors; D011744:Pyrimidinones; D018894:Reverse Transcriptase Inhibitors; D054303:HIV Reverse Transcriptase; D054309:Ribonuclease H, Human Immunodeficiency Virus
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