title: Ex vivo response to histone deacetylase (HDAC) inhibitors of the HIV long terminal repeat (LTR) derived from HIV-infected patients on antiretroviral therapy.
abstract: Histone deacetylase inhibitors (HDACi) can induce human immunodeficiency virus (HIV) transcription from the HIV long terminal repeat (LTR). However, ex vivo and in vivo responses to HDACi are variable and the activity of HDACi in cells other than T-cells have not been well characterised. Here, we developed a novel assay to determine the activity of HDACi on patient-derived HIV LTRs in different cell types. HIV LTRs from integrated virus were amplified using triple-nested Alu-PCR from total memory CD4+ T-cells (CD45RO+) isolated from HIV-infected patients prior to and following suppressive antiretroviral therapy. NL4-3 or patient-derived HIV LTRs were cloned into the chromatin forming episomal vector pCEP4, and the effect of HDACi investigated in the astrocyte and epithelial cell lines SVG and HeLa, respectively. There were no significant differences in the sequence of the HIV LTRs isolated from CD4+ T-cells prior to and after 18 months of combination antiretroviral therapy (cART). We found that in both cell lines, the HDACi panobinostat, trichostatin A, vorinostat and entinostat activated patient-derived HIV LTRs to similar levels seen with NL4-3 and all patient derived isolates had similar sensitivity to maximum HDACi stimulation. We observed a marked difference in the maximum fold induction of luciferase by HDACi in HeLa and SVG, suggesting that the effect of HDACi may be influenced by the cellular environment. Finally, we observed significant synergy in activation of the LTR with vorinostat and the viral protein Tat. Together, our results suggest that the LTR sequence of integrated virus is not a major determinant of a functional response to an HDACi.
authors.affiliation: Department of Infectious Diseases, Monash University, Melbourne, Victoria, Australia; Centre for Biomedical Research, Burnet Institute, Melbourne, Victoria, Australia.
authors.forename: Hao K
authors.initials: HK
authors.lastname: Lu
authors.affiliation: Department of Infectious Diseases, Monash University, Melbourne, Victoria, Australia; Centre for Biomedical Research, Burnet Institute, Melbourne, Victoria, Australia.
authors.forename: Lachlan R
authors.initials: LR
authors.lastname: Gray
authors.affiliation: Department of Infectious Diseases, Monash University, Melbourne, Victoria, Australia; Centre for Biomedical Research, Burnet Institute, Melbourne, Victoria, Australia.
authors.forename: Fiona
authors.initials: F
authors.lastname: Wightman
authors.affiliation: Department of Infectious Diseases, Monash University, Melbourne, Victoria, Australia; Centre for Biomedical Research, Burnet Institute, Melbourne, Victoria, Australia.
authors.forename: Paula
authors.initials: P
authors.lastname: Ellenberg
authors.affiliation: Department of Infectious Diseases, Monash University, Melbourne, Victoria, Australia; Centre for Biomedical Research, Burnet Institute, Melbourne, Victoria, Australia.
authors.forename: Gabriela
authors.initials: G
authors.lastname: Khoury
authors.affiliation: Centre for Biomedical Research, Burnet Institute, Melbourne, Victoria, Australia.
authors.forename: Wan-Jung
authors.initials: WJ
authors.lastname: Cheng
authors.affiliation: Department of Infectious Diseases, Monash University, Melbourne, Victoria, Australia; Centre for Biomedical Research, Burnet Institute, Melbourne, Victoria, Australia; Department of Microbiology and Immunology, University of Melbourne, Melbourne, Victoria, Australia.
authors.forename: Talia M
authors.initials: TM
authors.lastname: Mota
authors.affiliation: Centre for Biomedical Research, Burnet Institute, Melbourne, Victoria, Australia; South Australian Health and Medical Research Institute, Adelaide, Australia.
authors.forename: Steve
authors.initials: S
authors.lastname: Wesselingh
authors.affiliation: Department of Infectious Diseases, Monash University, Melbourne, Victoria, Australia; Centre for Biomedical Research, Burnet Institute, Melbourne, Victoria, Australia; Department of Microbiology and Immunology, University of Melbourne, Melbourne, Victoria, Australia.
authors.forename: Paul R
authors.initials: PR
authors.lastname: Gorry
authors.affiliation: Department of Infectious Diseases, Monash University, Melbourne, Victoria, Australia; Centre for Biomedical Research, Burnet Institute, Melbourne, Victoria, Australia; Infectious Disease Unit, Alfred Hospital, Melbourne, Victoria, Australia.
authors.forename: Paul U
authors.initials: PU
authors.lastname: Cameron
authors.affiliation: Centre for Biomedical Research, Burnet Institute, Melbourne, Victoria, Australia; Department of Microbiology, Monash University, Clayton, Victoria, Australia; Department of Medicine, Monash University, Clayton, Victoria, Australia.
authors.forename: Melissa J
authors.initials: MJ
authors.lastname: Churchill
authors.affiliation: Department of Infectious Diseases, Monash University, Melbourne, Victoria, Australia; Centre for Biomedical Research, Burnet Institute, Melbourne, Victoria, Australia; Infectious Disease Unit, Alfred Hospital, Melbourne, Victoria, Australia; Peter Doherty Institute, Melbourne University, Melbourne, Victoria, Australia.
authors.forename: Sharon R
authors.initials: SR
authors.lastname: Lewin
mesh_terms: D000328:Adult; D000368:Aged; D019380:Anti-HIV Agents / Q000627:therapeutic use; D001549:Benzamides / Q000494:pharmacology; D002460:Cell Line; D015658:HIV Infections / Q000097:blood* / Q000188:drug therapy / Q000821:virology; D016325:HIV Long Terminal Repeat / Q000187:drug effects*; D006367:HeLa Cells; D056572:Histone Deacetylase Inhibitors / Q000494:pharmacology*; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D007211:Indoles / Q000494:pharmacology; D064887:Observational Studies as Topic; D000077767:Panobinostat; D010802:Phylogeny; D011725:Pyridines / Q000494:pharmacology; D013601:T-Lymphocytes / Q000187:drug effects / Q000821:virology*; D000077337:Vorinostat; D054322:tat Gene Products, Human Immunodeficiency Virus / Q000494:pharmacology
chemical_list: D019380:Anti-HIV Agents; D001549:Benzamides; D056572:Histone Deacetylase Inhibitors; D006877:Hydroxamic Acids; D007211:Indoles; D011725:Pyridines; D054322:tat Gene Products, Human Immunodeficiency Virus; C118739:entinostat; C012589:trichostatin A; D000077337:Vorinostat; D000077767:Panobinostat