Bevirimat (3- O-(3'3'-dimethylsuccinyl)betulinic acid) was developed by activity-directed derivatization of betulinic acid, a plant-derived natural product. Bevirimat (BVM), previously called DSB, PA-457, or YK-FH312 potently inhibits HIV-1 replication in tissue culture and is efficacious in HIV-1 infected patients. Therefore, new anti-HIV-1 agents are constantly needed. Information gained from this study sheds light on the mechanisms by which the virus develops resistance to this class of drug and may aid in the design of next-generation maturation inhibitors.ĭespite the significant progress in the development of therapeutics against Human Immunodeficiency Virus type 1 (HIV-1), resistance to existing drugs is a continuing challenge for clinical management. This is the first demonstration of a direct interaction between the maturation inhibitor, bevirimat, and its target, Gag. Extensive prior genetic evidence suggests that the MHR is critical for virus assembly. Unexpectedly, a second region of interaction was found within the Major Homology Region (MHR). Bevirimat analogs were found to crosslink to sequences overlapping, or proximal to, the CA-SP1 cleavage site, consistent with previous biochemical data on the effect of bevirimat on Gag processing and with genetic data from resistance mutations, in a region predicted by NMR and mutational studies to have α-helical character. In this study, photoaffinity analogs of bevirimat and mass spectrometry were employed to map the binding site of bevirimat to Gag within immature virus-like particles. In contrast to inhibitors that target the active site of the viral protease, bevirimat specifically inhibits a single cleavage event, the final processing step for the Gag precursor where p25 (CA-SP1) is cleaved to p24 (CA) and SP1. Bevirimat, the prototype Human Immunodeficiency Virus type 1 (HIV-1) maturation inhibitor, is highly potent in cell culture and efficacious in HIV-1 infected patients.