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 sub-micromolar 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 strengthens the case for allosteric inhibition of HIV RNase H activity, providing a platform for designing improved antagonists for use in combination antiviral therapy.

Studies on Cycloheptathiophene‐3‐carboxamide Derivatives as Allosteric HIV‐1 Ribonuclease H Inhibitors

“…Several substituted phenyl rings were suitable for the C2 position. In agreement with previous reports on thienopyrimidinone derivatives, [19] the best substituent was the catechol moiety, represented by the most potent compound in this study, compound 33, which displayed an IC 50 value of 0.84 mm. O-Methylation of the hydroxy groups to give compound 31 resulted in a fivefold decrease in the potency.…”

Identification of Highly Conserved Residues Involved in Inhibition of HIV-1 RNase H Function by Diketo Acid Derivatives

“…Since the RNase H function is essential for viral replication (5), it has been explored as a drug target, and a number of RNase H inhibitors (RHIs) have been reported (6)(7)(8). RHIs can be classified based on their binding sites, i.e., (i) RHIs that coordinate the two Mg 2ϩ catalytic cofactors at the RNase H active site, such as N-hydroxyimides (9), hydroxytropolones (10), hydroxypyrimidines (11), naphthyridinones (12), nitrofuran-2-carboxylic acid carbamoylmethyl esters (13), hydroxyquinolinones (14), and thiocarbamates and triazoles (15), or (ii) allosteric RHIs, such as vinylogous ureas (16), thienopyrimidinones (17), hydrazones (18), anthraquinones (19), isatines (20,21), and propenones (22).…”

Discovery of diethyl 2,5-diaminothiophene-3,4-dicarboxylate derivatives as potent anticancer and antimicrobial agents and screening of anti-diabetic activity: Synthesis and in vitro biological evaluation. Part 1

“…IR spectra were recorded in KBr on a Shimadzu FT-IR 8400S spectrometer. The NMR spectra were recorded on a Varian 400-MR spectrometer (400 MHz for 1 H and 100 MHz for 13 C) using CDCl 3 as a solvent and TMS as an internal standard in 1 H NMR spectra. In 13 C NMR spectra chemical shifts were internally referenced to the solvent signals (CDCl 3 77.16 ppm related to TMS).…”

“…The structures of the newly synthesized compounds were appropriately confirmed by their spectral data. The 1 H NMR spectra of the compounds 2ael showed a characteristic singlets at a region 7.77e8.08 ppm accounting for N]CH proton and revealed carbon signals for N]CH at the range of 137.65e153.86 ppm in 13 C NMR spectra. Moreover, in both 1 Н and 13 С NMR spectra of compounds 2ael shifted downfield signals of the one ethoxy group.…”

“…Synthesized TP from 2-AT possess important biological properties, for example some of TP compound most potent nonsteroidal 17b-HSD1 inhibitors reported to date-inhibited the recombinant enzyme [12]. Other TP derivatives were combined and their biochemical characterization has been evaluated to wildtype HIV-1 RT, with rationally designed p66/p51 heterodimers exhibiting high-level sensitivity or resistance [13]. Dai et al discovered novel thienopyrimidine-based receptor tyrosine kinase inhibitors [14].…”