4-Deoxyphorbol inhibits HIV-1 infection in synergism with antiretroviral drugs and reactivates viral reservoirs through PKC/MEK activation synergizing with vorinostat

Latent HIV reservoirs are the main obstacle to eradicate HIV infection. One strategy proposes to eliminate these viral reservoirs by pharmacologically reactivating the latently infected T cells. We show here that a 4-deox-yphorbol ester derivative isolated from Euphorbia amygdaloides ssp. semiperfoliata, 4 beta-dPE A, reactivates HIV-1 from latency and could potentially contribute to decrease the viral reservoir. 4 beta-dPE A shows two effects in the HIV replication cycle, infection inhibition and HIV transactivation, similarly to other phorboids PKC agonists such PMA and prostratin and to other diterpene esters such SJ23B. Our data suggest 4 beta-dPE A is non-tumorigenic, unlike the related compound PMA. As the compounds are highly similar, the lack of tumorigenicity by 4 beta-dPE A could be due to the lack of a long side lipophilic chain that is present in PMA. 4 beta-dPE activates HIV transcription at nanomolar concentrations, lower than the concentration needed by other latency reversing agents (LRAs) such as prostratin and similar to bryostatin. PKCO/MEK activation is required for the transcriptional activity, and thus, anti-latency activity of 4 beta-dPE A. However, CD4, CXCR4 and CCR5 receptors down-regulation effect seems to be independent of PCK/MEK, suggesting the existence of at least two different targets for 4 beta-dPE A. Furthermore, NF-Kb transcription factor is involved in 4 beta-dPE HIV reactivation, as previously shown for other PKC5 agonists. We also studied the effects of 4 beta-dPE A in combination with other LRAs. When 4 beta-dPE A was combined with another PKC agonists such as prostratin an antagonic effect was achieved, while, when combined with an HDAC inhibitor such as vorinostat, a strong synergistic effect was obtained. Interestingly, the latency reversing effect of the combination was synergistically diminishing the EC50 value but also increasing the efficacy showed by the drugs alone. In addition, combinations of 4 beta-dPE A with antiretroviral drugs as CCR5 antagonist, NRTIs, NNRTIs and PIs, showed a consistent synergistic effect, suggesting that the combination would not interefer with antiretroviral therapy (ART). Finally, 4 beta-dPE A induced latent HIV reactivation in CD4 + T cells of infected patients under ART at similar levels than the tumorigenic phorbol derivative PMA, showing a clear reactivation effect. In summary, we describe here the mechanism of action of a new potent deoxyphorbol derivative as a latency reversing agent candidate to decrease the size of HIV reservoirs.