The Studies Of Amino Acid Derivatives Of Gossypol Anti-HIV-1Activities And Mechanism

During the past30years we fight against with HIV/AIDS, the greatest progresses we have made are the development of anti-HIV drugs, vaccines and gene therapies for HIV. Although we have invested so much money in anti-HIV vaccines, there is no one effective vaccine, hitherto. The enormous sequence diversity of HIV-1and the relative inaccessibility of conserved domains on Env decrease the elicitation of protective antibodies with global coverage. Moreover, the poor understanding of the immune responses that can control HIV-1replication makes the development of vaccines that induce such immune responses rather difficult. Up to the present, dozens more anti-HIV gene therapeutics having reached clinical and pre-clinical trials, but the efficacy, safety and especially affordability of gene therapy should be improved. Of the32anti-retroviral drugs currently used in the treatment of HIV infection, most are reverse transcriptase and protease inhibitors. Albeit the success of these drugs in improving and extending the lives of patients infected with HIV is remarkable, the toxicity and the emergence of drug-resistant strains of HIV greatly restrict the liberal usage of these drugs. Therefore, the development of new small molecule anti-HIV-1drugs but orally administered and less expensive is a pressing need for AIDS, especially patients in developing countries.Gossypol is a polyphenolic aldehyde extracted from cottonseed and has the biological activity of anti-fertility, anti-viral, anti-cancer, induction of apoptosis and regulation of immune. To develop potential drugs that can better inhibit HIV-1activity, decrease cellular cytotoxicity and avoid the male anti-fertility activity, a series of small molecules was covalently attached to gossypol.Our results show that mostly neutral, aliphatic amino acid derivatives of (-)gossypol demonstrate the strongest inhibitory effect on the laboratory adapted HIV-1strain. We speculate that this finding may be due to the increased hydrophobicity conferred on the molecule by the aliphatic amino acid moiety, as further mechanism study indicates that their target site is hydrophobicity. To further improve the activities of the derivatives and lower its cellular cytotoxicity, the gossypol is split into enantiomers and the aldehyde group is effectively blocked by the amino acid moiety. Then we harvest the efficacy with a100-fold improvement in selectivity index. The spermicidal of (-)gossypol and its amino acid derivatives were also discussed.It is widely accepted that gossypol and gossypol derivatives inhibit HIV-1replication by targeting reverse transcriptase. However, from the results of our time-of-addition assay, HIV-1-mediated cell fusion assay and VSV-G pseudotyped virus assay, we demonstrate that the alanine-(-) gossypol derivative ((-)G-Ala) is an effective HIV-1entry inhibitor. Further mechanistic analysis revealed that (-)G-Ala neither blocks gpl20-CD4binding nor interacts with the HIV-1cc-receptor CXCR4. Results from sandwich ELISA, native-PAGE and circular dichroism (CD) show that (-)G-Ala inhibits the cell fusion-activated gp41core domain. Moreover,(-)G-Ala binds to the HIV-5-Helix protein and blocking D-peptide (PIE7) binding to the hydrophobic pocket on the surface of the gp41internal trimeric coiled-coil domain. The contraceptive properties of (-) gossypol and amino acid derivatives of (-) gossypol are also discussed. Collectively, our results indicate that (-)G-Ala may bind to the gp41hydrophobic pocket and block the formation of the cell fusion-activated gp41core to inhibit HIV-1-mediated membrane fusion and subsequent viral entry.