CROI 2015 Program and Abstracts

Abstract Listing

Poster Abstracts

After interrupting the drug regimen (cART and Ing-B), VL stayed undetectable for 45 days before rebounding to pre-cART levels. Here we report results from Ing-B treatment in our consistent and accelerated SIV macaque model for HIV/AIDS and HAND. Methods: Three pigtailed macaques were dual inoculated with SIV∆B670 and SIV/17E-Fr, and treated at 12 days p.i. with CNS-penetrant cART (TNF, ATZ, RTN, L-870812). After 500 days of viral suppression, one animal was kept as control while two macaques received daily oral doses of Ing-B for 40 days. After a 2-week washout, the same animals received a 10-day treatment of Ing-B in combination with vorinostat (4 daily IV doses in 10 days). Animals were kept on cART until necropsy. SIV RNA was quantitated in plasma and CSF by ddPCR. SIV DNA was assessed in tissues by qPCR. In situ hybridization (ISH) for SIV RNA was performed in samples from brain and mesenteric lymph node. Resting CD4+ T cells were collected before and after latency reversing agents (LRA) treatment for quantitative viral outgrowth assay. Results: LRA induction caused a significant increase of plasma and CSF VL in one of the LRA-treated macaques. CSF viral load was 10x higher than in plasma, and the animal had to be euthanized due to encephalitis-related symptoms. SIV RNA could be detected by ISH in occipital cortex, despite undetectable levels of SIV DNA measured by qPCR. No change was observed in the other LRA-treated macaque. However, the number of SIV-infected resting CD4+ T cells was reduced after LRA-induction in both treated animals when compared to control. Conclusions: Treatment with LRAs led to a decrease in latent reservoirs in SIV-infected cART-treated macaques. In one animal, treatment activated viral genomes in basal ganglia, leading to CNS disease, indicating that the brain harbors latent virus and should be seriously considered when novel “Kick and Kill” strategies are designed for HIV eradication. 417 TLR7 Agonist GS-9620 Activates HIV-1 in PBMCs FromHIV-Infected Patients on cART Derek D. Sloan 1 ; Alivelu Irrinki 1 ; AngelaTsai 1 ; Jasmine Kaur 1 ; Jay Lalezari 2 ; Jeff Murry 1 ;Tomas Cihlar 1 1 Gilead Sciences, Inc., Foster City, CA, US; 2 Quest Clinical Research, San Francisco, CA, US Background: Pharmacologic activation of latent HIV reservoirs is considered to be a key part of the strategy towards eradicating HIV-1 infection. GS-9620 is a selective TLR7 agonist currently being evaluated in patients with chronic hepatitis B. Based on recent observations of induced plasma viral RNA and reduction of viral DNA in cART-suppressed SIV-infected rhesus monkeys (RM) treated with a close analog (Whitney et al ., CROI 2015 submitted), GS-9620 is now also being considered for evaluation in HIV-infected patients. Here we show that GS-9620 activates HIV gene expression ex vivo in PBMCs from HIV-infected patients on cART. Methods: PBMCs isolated from patients with plasma HIV RNA<50 copies/mL for >12 months were treated with GS-9620 in the presence of ARVs. HIV-1 RNA was quantified in culture supernatants by the AmpliPrep/COBAS ® TaqMan ® assay. GS-9620-induced cytokines were quantified by Luminex. In selected cultures, CD8 T cell-depleted PBMCs were treated with GS-9620. The effect on the inducible HIV reservoir was assessed by protein kinase C (PKC) agonist-mediated activation of CD4 T cells isolated from GS-9620-treated PBMCs. Results: In PBMCs from 11 of 12 donors tested, 0.01-1 m M GS-9620 activated HIV RNA expression compared to vehicle control with a 5.8-fold geometric mean maximal activation (range 2.0- to 26.8-fold across donors). Depletion of CD8 T cells increased overall HIV expression without affecting the fold activation induced by GS-9620. While there was no correlation between HIV activation levels and cytokines induced by GS-9620, blocking the type I IFN receptor reduced the maximal HIV expression in all donors assessed (n=4; 85% geometric mean reduction, p<0.05). GS-9620 treatment also reduced subsequent PKC agonist-mediated HIV activation (3 of 4 donors; 74% geometric mean reduction, p<0.05). Conclusions: GS-9620 activated HIV RNA expression in PBMCs isolated from patients on suppressive cART, a result consistent with observations in SIV-infected RM treated with a related TLR7 agonist. Type I IFN was required for maximal HIV activation by GS-9620. The HIV response to subsequent PKC activation was reduced in CD4 T cells isolated from GS-9620-treated PBMCs, suggesting a potential reduction in the inducible viral reservoir. Together with the results obtained from SIV-infected RM, these data support the clinical testing of GS-9620 in HIV-infected patients on suppressive cART for possible activation and reduction of the viral reservoir. 418 Baracitinib, Ruxolitinib, Dasatinib Block HIV Replication, Activation, Reactivation Christina Gavegnano Emory University, Atlanta, GA, US Background: HIV-induced activation/inflammation is associated with HIV persistence/reactivation, and is a predictor of morbidity in HIV-infected individuals. HIV-induced activation of myeloid cells promotes trafficking of infected cells to the CNS, and is associated with HIV-induced neurocognitive impairments and HIV-associated dementia. Jak-STAT pathway and tyrosine kinases are activated in macrophages and lymphocytes upon HIV-1 infection, representing attractive cellular targets. As ruxolitinib and baracitinib are Jak1/2 inhibitors either FDA approved or in late phase 3 clinical investigation for myelofibrosis or arthritis respectively, and dasatinib is FDA approved for myeloid malignancies, we evaluated their ability to block 1)HIV replication in primary human macrophages, 2)HIV-induced activation in primary human monocytes and macrophages, and 3)reactivation of latent HIV in T cells. Methods: Macrophages were treated with various concentrations of ruxolitinib, baracitinib, or dasatinib for 2hr prior to infection (HIV-1 baL ) . Cells were maintained for 6 days before viral quantification (p24-ELISA). Macrophages or monocytes were treated with various concentrations of drug prior to infection (HIV-1 baL ) , and maintained for 3 or 6 days before quantification of HLA-DR, CD163, CCR5 (macrophages), or CD14/CD16 (monocytes). Macrophages and monocytes were treated with various concentrations of drug for 6 days and stained with Near-IR live/dead dye and quantified by FACS. J-lat latent T cells were treated with various concentrations of drug plus TNF-a and reactivation was quantified by GFP reporter 24 hr post induction. Results: Ruxolitinib, baracitinib, and dasatinib were not cytotoxic (>50 m M for ruxolitinib/baracitinib; 20 m M) and demonstrated 1)antiviral potency ranging from 0.001-0.08 m M in macrophages, 2)inhibition of HIV-induced upregulation of CCR5, CD163, and HLA-DR (macrophages) and CD14/CD16 (monocytes), and inhibition of reactivation of latent HIV-1 in T cells (p<0.05 versus no drug controls). Conclusions: Ruxolitinib, baracitinib, and dasatinib inhibit HIV replication in macrophages, reactivation of latent HIV-1 (T-cells), and HIV-induced activation markers (monocytes/ macrophages), which are linked to trafficking of infected cells to the CNS, disease progression, and neurocognitive dysfunction. Selectively blocking Jak-STAT or tyrosine kinase signaling could reverse or prevent HIV-mediated immune activation both systemically and within the brain/CNS. 419 Ex Vivo Identification of Highly Effective Latency-Reversing Drug Combinations Gregory M. Laird 1 ; C Korin Bullen 1 ; Daniel I. Rosenbloom 2 ; Alyssa R. Martin 1 ; Alison L. Hill 3 ; Christine M. Durand 1 ; Janet D. Siliciano 1 ; Robert F. Siliciano 1 1 Johns Hopkins University School of Medicine, Baltimore, MD, US; 2 Columbia University, New York, NY, US; 3 Harvard University, Boston, MA, US Background: HIV-1 persists in a latent reservoir despite ART, and reactivation of this reservoir has been proposed as a cure strategy. Effective drug combinations that achieve high levels of HIV-1 latency reversal will likely be required for a cure. We set out to identify highly effective combinations using resting CD4 + T cells (rCD4s) from infected individuals and use our ex vivo measurements to predict in vivo efficacy. Methods: Combinations of latency reversing agents (LRAs) that reverse latency ex vivo to an extent approaching the benchmark of maximal T cell activation (~100 fold induction) have not yet been identified. We therefore measured intracellular HIV-1 mRNA levels and supernatant virion production following individual or combination LRA treatment in

Poster Abstracts

297

CROI 2015