Bryostatin activates HIV-1 latent expression in human astrocytes through a PKC and NF-ĸB-dependent mechanism

Abstract

Multiple studies have shown that HIV-1 patients may develop virus reservoirs that impede eradication; these reservoirs include the central nervous system (CNS). Despite an undetectable viral load in patients treated with potent antiretrovirals, current therapy is unable to purge the virus from these latent reservoirs. To broaden the inhibitory range and effectiveness of current antiretrovirals, the potential of bryostatin was investigated as a latent HIV-1 activator. We used primary astrocytes, NHA cells, and astrocytoma cells U-87. Infected cells with HIV-1NL4.3 were treated with bryostatin alone or in combination with different inhibitors. HIV-1 production was quantified by using ELISA. Transcriptional activity was measured using luciferase reporter gene assays by using lipofectin. We performed cotransfection experiments of the LTR promoter with the active NF-κB member p65/relA. To confirm the NF-κB role, Western blot and confocal microscopy were performed. Bryostatin reactivates latent viral infection in the NHA and U87 cells via activation of protein kinase C (PKC)-alpha and -delta, because the PKC inhibitors rottlerin and GF109203X abrogated the bryostatin effect. No alteration in cell proliferation was found. Moreover, bryostatin strongly stimulated LTR transcription by activating the transcription factor NF-κB. Bryostatin could be a beneficial adjunct to the treatment of HIV-1 brain infection.

In vitro effects of the small-molecule protein kinase C agonists on HIV latency reactivation

“…Prostratin not only induces HIV expression from latently infected cells through phosphorylation and degradation of IκBα, leading to the rapid nuclear translocation of NF-κB35, but also inhibits HIV entry by interacting with a cellular target necessary for viral entry, displaying potent antiviral activity against different strains of HIV-1 and SIV36373839. Likewise, PKC agonist bryostatin-1, isolated from the marine invertebrate Bugula neritina 4041, also shows excellent potential as a therapeutic agent that acts through modulation of PKC signal transduction, as indicated its potency to revert HIV-1 latency via the adenosine monophospate (AMP)-activated protein kinase (AMPK) pathway254243444546. However, recent study of bryostatin-1 treatment shows inhibitory effects on the HIV-specific CD8+ T cells47.…”

Strategies to target non-T-cell HIV reservoirs

“…Bryostatin has been shown to have anti-HIV-1 activity by downregulating CXCR4 receptors or indirectly inhibiting HIV-1 Tat function by dephosphorylating a kinase necessary for RNA polymerase-II functional initiation. Furthermore, Bryostatin has been shown to have moderate HIV latency reversing agent (LRA) activity in astrocyte cell lines in vitro and in cultured primary astrocytes by inducing HIV-1 expression through NF-kB activation 29 .…”

Bryostatin-1 for latent virus reactivation in HIV-infected patients on antiretroviral therapy

“…More recently, we have shown that bryostatin-1 also reactivates latent HIV-1 in astrocytes [21]. Bryostatin-1 has attracted renewed attention after the recent study by Bullen et al where a modified viral outgrowth ex-vivo assay was used to compare the efficacy of different leading LRA candidates [22].…”

“…In-vitro studies carried out by our group demonstrated that bryostatin-1 synergizes with HDAC inhibitors such as valproic acid to antagonize HIV-1 latency, and downregulates the expression of the HIV-1 receptors CD4 þ and CXCR4, preventing HIV-1-induced cytotoxicity [12,21].…”