Next-Generation CMA/CMB Derivatives as Targeted Inhibitors of HIV Nef

Current combined antiretroviral therapies (cART) suppress viral levels in the blood but do not eradicate reservoirs of cells harboring integrated copies of HIV proviral genomes. These cells persist in part because the provirus maintains a latent state that evades the immune response and viral cytopathic effect. Approaches that would couple latency reactivation with strategies to eradicate the infected cells – such as by design and activation of more efficacious anti-HIV cytotoxic T lymphocytes (CTLs) would facilitate clearance of these cells. Because Nef inhibits the activity of anti-HIV CTLs, a potent Nef inhibitor would help achieve this goal. To date, no Nef inhibitor that achieves potent restoration of MHC-I in the presence of Nef has advanced to the clinic. The most potent agent(s) capable of inhibiting this activity of Nef are a class of compounds previously known to inhibit vacuolar ATPase (V-ATPase), which is necessary for lysosomal function. However, our studies clearly show that Nef inhibition is separable from effects of the drugs on the lysosome. In particular, concanamycin A (CMA) restores MHC-I to the surface of Nef-expressing cells at low nM concentrations that did not interfere with lysosomal acidification and demonstrated no observable toxicity in primary cell cultures after 24 hours. Moreover, we found that derivatives of CMA at the C-3’ and C-9 positions further isolate the Nef inhibitory activity from effects on lysosomal acidification. Strong preliminary data support the exciting hypothesis that this results from selective targeting of a V-ATPase isoform that resides in endosomal compartments. We are now poised to; (1) develop the next generation of these potent inhibitors by further isolating the anti-Nef effect from off-target activities, (2) initiate in vivo small animal model testing and (3) determine the mechanism by which the inhibitors selectively inhibit Nef so that optimization can be conducted using rational structure-activity relationships.