Selective inhibition of HIV-1 reverse transcriptase-associated ribonuclease H activity by hydroxylated tropolones

Budihas, S. R.; Gorshkova, I. I.; Gaidamakov, S. A.; Wamiru, A.; Bona, M. K.; Parniak, M. A.; Crouch, R. C.; McMahon, J. B.; Beutler, J. A. and LeGrice, S. F. J..: Nucl. Acids Res. 33: 1249-1256, 2005.

Abstract:

High-throughput screening of a National Cancer Institute library of pure natural products identified the hydroxylated tropolone derivatives ß-thujaplicinol (2,7-dihydroxy-4-1(methylethyl)-2,4,6-cycloheptatrien-1-one) and manicol (1,2,3,4-tetrahydro-5-7-dihydroxy-9- methyl-2-(1-methylethenyl)- 6H-benzocyclohepten-6-one) as potent and selective inhibitors of the ribonuclease H (RNase H) activity of human immunodeficiency virus-type 1 reverse transcriptase (HIV-1 RT). ß-Thujaplicinol inhibited HIV-1 RNase H in vitro with an IC50 of 0.2 µM, while the IC50 for Escherichia coli and human RNases H was 50 µM and 5.7 µM, respectively. In contrast, the related tropolone analog ß-thujaplicin (2-hydroxy-4-(methylethyl)-2,4,6-cycloheptatrien-1-one), which lacks the 7-OH group of the heptatriene ring, was inactive, while manicol, which possesses a 7-OH group, inhibited HIV-1 and E.coli RNases H with IC50 = 1.5 µM and 40 µM, respectively. Such a result highlights the importance of the 2,7-dihydroxy function of these tropolone analogs, possibly through a role in metal chelation at the RNase H active site. Inhibition of HIV-2 RT-associated RNase H indirectly indicates that these compounds do not occupy the nonnucleoside inhibitor-binding pocket in the vicinity of the DNA polymerase domain. Both ß-thujaplicinol and manicol failed to inhibit DNA-dependent DNA polymerase activity of HIV-1 RT at a concentration of 50 µM, suggesting that they are specific for the C-terminal RNase H domain, while surface plasmon resonance studies indicated that the inhibition was not due to intercalation of the analog into the nucleic acid substrate. Finally, we have demonstrated synergy between ß-thujaplicinol and calanolide A, a nonnucleoside inhibitor of HIV-1 RT, raising the possibility that both enzymatic activities of HIV-1 RT can be simultaneously targeted.

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