THE IN-VITRO EJECTION OF ZINC FROM HUMAN-IMMUNODEFICIENCY-VIRUS (HIV)TYPE-1 NUCLEOCAPSID PROTEIN BY DISULFIDE BENZAMIDES WITH CELLULAR ANTI-HIV ACTIVITY

Several disulfide benzamides have been shown to possess wide-spectrum antiretroviral activity in cell culture at low micromolar to submicrom olar concentrations, inhibiting human immunodeficiency virus (HIV) typ e 1 (HIV-1) clinical and drug-resistant strains along with HIV-2 and s imian immunodeficiency virus [Rice, W. G., Supko, J. G., Malspeis, L., Buckheit, R. W., Jr., Clanton, D., Bu, M., Graham, L., Schaeffer, C. A., Turpin, J. A., Domagala, J., Gogliotti, R., Bader, J. P., Halliday , S. M., Coren, L., Sowder, R. C., II, Arthur, L. O. & Henderson, L. E . (1995) Science 270, 1194-1197]. Rice and coworkers have proposed tha t the compounds act by ''attacking'' the two zinc fingers of HIV nucle ocapsid protein. Shown here is evidence that low micromolar concentrat ions of the anti-HIV disulfide benzamides eject zinc from HIV nucleoca psid protein (NCp7) in vitro, as monitored by the zinc-specific fluore scent probe N-(6-methoxy-8-quinoyl)-p-toluenesulfonamide (TSQ). Struct urally similar disulfide benzamides that do not inhibit HIV-1 in cultu re do not eject zinc, nor do analogs of the antiviral compounds with t he disulfide replaced with a methylene sulfide. The kinetics of NCp7 z inc ejection by disulfide benzamides were found to be nonsaturable and biexponential, with the rate of ejection from the C-terminal zinc fin ger 7-fold faster than that from the N-terminal. The antiviral compoun ds were found to inhibit the zinc-dependent binding of NCp7 to HIV psi RNA, as studied by gel-shift assays, and the data correlated well wit h the zinc ejection data. Anti-HIV disulfide benzamides specifically e ject NCp7 zinc and abolish the protein's ability to bind psi RNA in vi tro, providing evidence for a possible antiretroviral mechanism of act ion of these compounds. Congeners of this class are under advanced pre clinical evaluation as a potential chemotherapy for acquired immunodef iciency syndrome.