Iminodipropionic acid as the leaving group for DNA polymerization by HIV-1 reverse transcriptase

Previous studies have demonstrated that some selected amino monoacids and amino diacids can function as leaving groups in the polymerase‐catalyzed incorporation of deoxynucleotides into DNA. Among these, the iminodiacetic acid phosphoramidate of deoxyadenosine monophosphate (IDA‐dAMP) represents an interesting example, as it could overcome some of the problems observed when using L ‐aspartic acid as the leaving group, that is, poor chain elongation. We have now synthesized and evaluated a series of IDA‐dAMP analogues that bear either an extended aliphatic chain in the amino acid function, or a phosphonic acid moiety (substituting for the carboxylic acid function). Among these compounds, the nucleotide with an iminodipropionic acid leaving group (IDP‐dAMP) was identified as the best substrate; the excellent single incorporation (91 % conversion to a P+1 strand at 50 μM ) was at a substrate concentration ten times lower than that used for IDA‐dAMP). This nucleotide also presented improved kinetics and elongation capability compared to IDA‐dAMP. The analogues with T, G, and C base moieties were also investigated for their incorporation ability with HIV‐1 RT. The incorporation efficiency was found to decrease in the order A>T>G>C. The properties of the iminodipropionic acid as the leaving group surpass those of previously evaluated leaving groups; this acid will be a prime candidate for in vivo testing.     

Selective inhibition of HIV-1 reverse transcriptase (HIV-1 RT) RNase H by small RNA hairpins and dumbbells

We present here the design of a novel class of RNA inhibitors of the RNase H domain of HIV-1 RT, a ribonuclease activity that is essential for viral replication in vivo. Specifically, we show that small RNA hairpins and dumbbells can selectively inhibit the RNase H activity of HIV-1 RT without affecting other cellular RNases H (e.g., E. coli and human RNase H). These results suggest that the inhibitors do not interact with the nucleic acid binding site of RT RNase H, as this region should be well conserved among the various enzymes. The most potent inhibitors displayed IC50 values in the 3-8 microM range. Remarkably, the DNA polymerase activity, an intrinsic property of HIV RT, was not inhibited by the hairpin and dumbbell aptamers, a property not previously observed for any nucleic acid aptamer directed against RT RNase H. The results described here suggest a noncompetitive binding mechanism, as outlined in the differential inhibitory characteristics of each of the nucleic acid aptamers against the bacterial, human, and viral RNase H homologues.     

Design, synthesis, biological evaluation, and molecular modeling studies of TIBO-like cyclic sulfones as non-nucleoside HIV-1 reverse transcriptase inhibitors

TIBO- and TBO-like sulfone derivatives 1 and 2 were designed, synthesized, and tested for their ability to block the replication cycle of HIV-1 in infected cells. The anti-HIV-1 activities of sulfones 3, which were intermediates in the syntheses of 1 and 2, were also evaluated. Surprisingly, the sulfone analogues of TIBO R82913 (compounds 1) were inactive, whereas interesting results were obtained for truncated derivatives 2. Compound 2 w was the most potent among this series in cell-based assays (EC50=0.07 microM, CC50>200 microM, SI>2857). It was twofold less potent than R82913, but more selective. An X-ray crystallographic analysis was carried out to establish the absolute configuration of 2 w and its enantiomer 2 x, which were obtained by semipreparative HPLC of 2 v, one of the most potent racemates. Compounds 1-3 were proven to target HIV-1 RT. In fact, representative derivatives inhibited recombinant HIV-1 RT in vitro at concentrations similar to those active in cell-based assays. 3D QSAR studies and docking simulations were developed on TIBO- and TBO-like sulfone derivatives to rationalize their anti-HIV-1 potencies and to predict the activity of novel untested sulfone derivatives. Predictive 3D QSAR models were obtained with a receptor-based alignment by docking of TIBO- and TBO-like derivatives into the NNBS of RT.