The Reactivation And Eradication Effect Of BET Inhibitor Apabetalone On Latent HIV-1 Reservoirs And Its Molecular Mechanism

Although cART suppresses HIV to undetectable levels and partially restores immune function in infected individuals,it produces serious side effects,such as lipid metabolism disorders and cardiovascular diseases.Moreover,an interruption of cART causes the virus to rapidly rebound to its pre-treatment levels.The main cause of treatment failure is the existence of latent HIV-1 reservoirs.Recent studies have explored a strategy named "shock and kill",which would eradicate the HIV-1 in latent HIV-1 reservoirs by activating HIV-1 transcription and viral antigen expression in the presence of cART.When attempting to use this strategy,the first challenge is to find latency reversing agents(LRAs)to efficiently reactivate latent HIV-1.Several agents have been explored,and two types of LRAs have reached human testing.The findings from these studies showed some promise but failed to result in significant clearance of residual HIV reservoirs.Some possible mechanisms for this failure include themodest induction of HIV by this earlier generation of latency reversing agents(LRAs)when used singly,as well as immune defects which may have resulted in a failure to fully eradicate the infected cells,despite the reactivation of viral expression.Those studies demonstrate the urgent need to develop new strategies for disrupting HIV latency and facilitating the elimination of infected cells after HIV expression is reactivated.Apabetalone is a highly selective BET inhibitor that targets the BD2 domain of BET proteins and is being developed by Resverlogix Corporation for treatment of cardiovascular diseases and lipid metabolism disorders.Notably,low HDL cholesterol,lipid metabolism disorders and cardiovascular diseasesare all known as side effects of cART,and might be prevented by Apabetalone.In the current study,we evaluated the ability of Apabetalone to reactivate HIV latent reservoirs.Our data indicated that Apabetalone could significantly promote HIV expression in various types of HIV latency cell models in vitro and ex vivo.CCK8 assay,flow cytometry and ELISA assay were used to investingate the cytotoxicity and the expression of T cell surface activation biomarkers,pro-inflammatory cytokines and HIV receptor/co-receptor.Results showed that Apabetalone had no effect on T cell activation and pro-inflammatory cytokines,showed no evidence of toxicity,and significantly reducedthe expression of the HIV co-receptors CCR5 and CXCR4.Besides,Apabetalone significantly promoted the reactivation effect of SAHA or Prostratin,and the combination of cART with apabetalone does not interfere with cART drugs’ antiretroviral activity.Moreover,the combination of apabetalone with cART drugs,a rapid decrease of HIV-1 expression could be evaluated by detecting the expression of luciferase reporter gene,which indicated that virus reactivated by apabetalone could be inhibited by cART drugs.To substantiate the machanism of action of Apabetalone,various methods including Western Blot and chromatin immunoprecipation were used.We demonstrate that Tat plays an important role in Apabetalone-mediated HIV latent reactivation,and Apabetalone acts by activating P-TEFb via dissociating BDR4 from the HIV-1 promoter,recruiting Tat for stimulating HIV-1 elongation and induced transcription initiation by increasing histone acetylation.Furthermore,by using flow cytometry and Western Blot,we demonstrate that Apabetalone can downregulate cyclin D1 expression,upregulate p21waf21/Cip1,and induce G1/G0 phase cell cycle arrest.It was particularly interesting that Apabetalone induced the preferential apoptosis of HIV-1 latent cells,and further promoted the death of reactivated reservoir cellsFinally,we used cell and mouse model to verify that Apabetalone could inhibit T cell activation and inflammatory storm induced by Prostratin by inhibiting MAPK and NF-kB pathway.In conclusion,its low degree of cytotoxicity,coupled with its abilities to reactivate latent HIV-1 reservoirs,induce HIV-1 latent cell apoptosis,and reduce the side effects of cART,all make Apabetalone worth investigating for development as a possible latency reversing agent for use in accelerating HIV-1 eradication.