CROI 2016 Abstract eBook

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Poster Abstracts

Conclusions: Soluble inflammatory biomarkers decreased after 1 year of ART and subsequently remained mostly stable in controls; greater increases in cases prior to a non-AIDS event may reflect subclinical disease processes. Controls with greater pre-ART VL levels or lower pre-ART CD4 levels had higher biomarker levels at year 1 after ART initiation while also experiencing the greater biomarker declines in the first year of ART. Results support earlier initiation of ART in HIV+ infected adults.

294 Increased Innate Function Following HIV-1 Suppression Is Linked to Epigenetic Changes

Eileen P. Scully 1 ; Ainsley Lockhart 2 ;Wilfredo Garcia-Beltran 2 ; Christine D. Palmer 2 ; Chelsey Musante 2 ; Eric S. Rosenberg 1 ;Todd M. Allen 2 ; J. Judy Chang 3 ; Ronald Bosch 4 ; Marcus Altfeld 5 1 Massachusetts General Hosp, Boston, MA, USA; 2 Ragon Inst of MGH, MIT, and Harvard, Cambridge, MA, USA; 3 Doherty Inst for Infection and Immunity, Univ of Melbourne, Melbourne, Australia; 4 Harvard Sch of PH, Boston, MA, USA; 5 Heinrich Pette Inst, Hamburg, Germany Background: HIV-1 infection leads to both profound immune suppression and pathologic inflammation. While adaptive immune dysfunction has been extensively studied, less is known about innate immunity in the context of HIV-1. Based on the broad pathogen susceptibility in progressive HIV-1, we hypothesized that there would be aberrant innate responses. We tested innate function and investigated the underlying molecular mechanisms, analyzing subset distribution, transcription factor translocation, and epigenetic modifications. Methods: Pre and post-antiretroviral therapy (ART) peripheral blood mononuclear cells from HIV-1 infected subjects were tested for ex vivo responses to a panel of innate stimuli (TLR2, TLR4, Dectin-1, and NOD) by culture supernatant analysis and intracellular cytokine staining (ICS). Cell activation and subsets and plasma levels of cytokines, sCD163, LPS, and sCD14 were measured. Associations between innate responses and plasma factors were assessed using resampled Spearman correlations and generalized estimating equations to account for repeated sampling. Monocytes were tested for NFκB nuclear translocation, and epigenetic modifications at the TNFα promoter were investigated by chromatin immunoprecipitation (ChIP) qPCR. Results: Declining viral load correlated with increasing magnitude of monocyte and mDC responses to innate stimuli. This inverse relationship with viral load was most striking for TLR4(LPS) stimulation; culture supernatant: TNFα r=-0.37 (p<0.01), ICS: monocyte TNFα r=-0.57, mDC TNFα r=-0.54, mDC IL-1β r=-0.58, (p<0.001 for all). There was no difference in monocyte subset distribution or in baseline or LPS-induced nuclear translocation of NFκB between the pre and post-ART samples. In contrast, histone 4 acetylation and H3K4 trimethylation increased after viral suppression, connoting a poised transcriptional status. Conclusions: Suppression of HIV-1 viral load with ART correlated with increased ex vivo innate immune responses. The altered monocyte function was not associated with subset redistribution or changes in NFκB signaling, but was associated with epigenetic modifications at the TNFα promoter. The high viral load environment conditions an epigenetic program associated with decreased innate function that is reversed with virus suppression. Further work is warranted to elucidate additional epigenetic determinants of innate immune deficiency and excessive inflammation in HIV-1 infection and their functional recovery following viral control. 295 Inhibition of P2X7 Enhances T-Cell Potential of CD34 in HIV+ c-ART Nonresponders Inna Menkova-Garnier 1 ; Clarisse Benne 1 ; Hakim Hocini 2 ; PascalineTisserand 2 ; Constance Delaugerre 3 ; Emile Foucat 4 ; Laure Bourdery 4 ;Yves Lévy 1 ; Jean-Daniel Lelièvre 1 1 Inst Mondor de Recherche Biomédicale INSERM U955, Univ Paris Est Créteil, Créteil, France; 2 Inst Mondor de Recherche Biomédicale INSERM U955, Vaccin Rsr Inst (VRI), Créteil, France; 3 Hosp Saint Louis et Univ Paris Diderot, Paris, France; 4 Univ Paris Est-Créteil, Créteil, France Background: Although many factors are known to limit CD4 T-cell recovery during HIV infection, the impact of alteration in T-cell lymphopoiesis remains poorly understood. We hypothesize that deficient T-cell differentiation from CD34 hematopoietic progenitors (HP) could, at least partially, explain poor CD4 levels restoration under c-ART in HIV+ patients. Methods: PBMC were obtained from HIV+ immune responders under c-ART (IR, n=16, CD4=1086/mm 3 , CD4/CD8=1.7) and HIV+ c-ART nonresponders (INR, n=16, CD4=380/ mm 3 , CD4/CD8=0.58) followed in CHU Henri Mondor (Creteil, France) and from HIV- controls (n=18, CTS, Creteil). Ethical committee approval and written informed consent from all subjects were obtained before study initiation. T-cell precursor’s frequency was determined using limiting dilution assay (LDA) on coculture of HP on OP9-DLL1 cell line. LDA results were generated using ELDA webtool. Flow cytometry analysis was performed on LSR2 cytometer, and FlowJo v8.2. Statistical analyses were performed using appropriate test in Prism software. Results: We have previously shown a specific T-cell, but not B-cell differentiation impairment in CD34 HP from INR patients (see table 1). Now we extended our knowledge by demonstrating that CD34 HP from INR appeared extremely sensitive in regard of extracellular ATP known to induce caspase-1 mediated pytoptosis. We found abnormally high P2X7 levels (ATP recognition) and absent CD73 (ATP hydrolysis) expression. Importantly, caspase-1 inhibitor, VX-765, disturbed normal lymphopoiesis in vitro , thus P2X7 antagonist, PPAD, restored T-cell lymphopoiesis in INR: T-cell frequency of 1/314.7 (1/186.3-532.3) without PPAD vs 1/145.3 (1/98.3-214.7) with PPAD ( p <0.05). Transcriptomic analysis revealed a general alteration in cellular death pathway in INR as compared with IR individuals. Conclusions: Our results suggest that CD34 HP from INR are more prone to cellular death via extracellular ATP that affects normal T-cell lymphopoiesis and impacts CD4 T-cell restoration under c-ART. Currently we are evaluating reasons of this dysregulation and possible ways to interfere with it in therapeutic perspective. Such complementary to c-ART strategies might dramatically improve outcomes in treated immune nonresponders.

Poster Abstracts

113

CROI 2016