Inhibitors directed against two distinct points in the HIV-1 life cycle are being prepared. These inhibitors are intended to serve as potential new therapeutics and as pharmacological probes to investigate biochemical mechanisms of viral replication. The two areas of investigation are: (1) HIV-1 integrase (IN), where inhibitors may disrupt incorporation of viral cDNA into the host genome;(2) Binding of HIV p6Gag protein to human Tsg101 protein, where inhibitors may disrupt viral assembly and budding (1) HIV-1 IN inhibitors: This work is being done in collaboration with Drs. Yves Pommier (CCR, NCI) Steven Hughes (CCR, NCI) and Peter Cherepanov (Imperial College, London). A large number of IN inhibitors have been reported. Many of these exhibit common key structural features. These features include a co-planar arrangement of heteroatoms that chelate magnesium ions. Halogen-substituted aromatic functionality linked to the chelating portion of the inhibitors has also been shown to interact with a region formed between a viral DNA base and the protein in the IN-DNA complex. This class of IN inhibitors is thought to function by chelating Mg2+ ions within the IN catalytic site, where they selectively inhibit strand transfer (ST) reactions over 3-processing (3-P) reactions. Although an extensive body of scientific and patent literature exists concerning the development of HIV-1 integrase (IN) inhibitors, currently raltegravir represents the only FDA-approved IN inhibitor for the treatment of AIDS. The emergence of raltegravir-resistant strains of HIV-1 containing mutated forms of IN underlies the need for continued efforts to examine structural parameters that enhance efficacy or reduce sensitivity to mutant strains of IN. We have previously described bicyclic 6,7-dihydroxyoxoisoindolin-1-ones that show good IN inhibitory potency in vitro. More recently we have examined the effects of introducing substituents into the 4- and 5- positions of the parent 6,7-dihydroxyoxoisoindolin-1-one platform. As a result of this work we have developed several sulfonamide-containing analogues with enhanced potency in cell-based HIV assays. Several of these compounds exhibit better absolute efficacy than raltegravir against the clinically-relevant Y143R IN mutant. Our work in this area is being guided by co-crystal structures of our 4,5-dihydroxy-1H-isoindole-1,3(2H)-diones bound to the IN-DNA complex of the prototype foamy virus (PFV) integrase (done in collaboration with Dr. Peter Cherepanov, Imperial Colleage, London).(2) Tsg101-binding inhibitors: Binding of the HIV p6Gag protein to human Tsg101 protein has been shown to be necessary for viral budding and to involve a critical 9-mer P-E-P-T-A-P-P-E-E sequence of the p6 protein. In a collaboration with Dr. Eric Freed (CCR, NCI, NIH) we developed a fluorescently-labeled high affinity-binding antagonist that is currently being used for identification of Tsg101-binding antagonists by high-throughput screening of small compound libraries in the NIH Chemical Genomics Center under the direction of Dr. Doug Auld.
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