CROI 2018 Abstract eBook

Abstract eBook

Poster Abstracts

persist over time. This suggests that an adult-like “set-point” in the T cell compartment is established early in life by cCMV infection. These results provide rationale for future studies to correlate immunologic findings with infant clinical outcomes, including cCMV clearance, vaccine responses, and rates of febrile illness. Collectively, these studies will add to our current understanding of immune development in the setting of congenital infection and provide critical information to inform CMV therapy and vaccine design in infants. 279 PHOSPHORYLATED HIV-1 AS POTENTIAL CYTOTOXIC T LYMPHOCYTE (CTL) TARGETS Fatema Z. Chowdhury 1 , Shivaali Maddali 1 , Matthew J. Szucs 2 , Rushdy Ahmad 2 , John Sidney 3 , Alessandro Sette 3 , Pratikkumar Rathod 4 , Emmanuel J. Chang 4 , Mathias Lichterfeld 1 , Xu G. Yu 1 1 Ragon Institute of MGH, MIT and Harvard, Cambridge, MA, USA, 2 Broad Institute of MIT and Harvard, Cambridge, MA, USA, 3 La Jolla Institute for Allergy and Immunology, La Jolla, CA, USA, 4 City University of New York, New York, NY, USA Background: As a small retrovirus, HIV-1 depends on the host cell machinery to support its life cycle. Host kinases can phosphorylate HIV-1, which can influence functional states of viral proteins and may influence viral replicative fitness. How HIV-1 phosphorylation affects viral immune recognition by cytotoxic T cells is unknown at present. Notably, recognition of phosphorylated epitopes by CD8 T cells is well documented in the context of lymphoma and melanoma. Methods: In vitro infected CD4 T cells (n=4) and cell-free HIV-1 isolated from culture supernatants (n=3) were subjected to LC-MS/MS mass spectrometry. Selected identified phosphorylated HIV-1 peptides were synthesized and tested for HLA class I binding. PBMCs from HIV-1 infected patients were stimulated with phosphorylated and non-phosphorylated peptides, followed by flow- phosphorylation in HIV-1, the majority of which were located in the Gag, Pol, and Rev proteins. Of 74 optimal CTL epitopes within Gag, 34 contained identified phosphorylated sites, suggesting that phosphorylation may influence immune recognition by CTLs. Out of six CTL epitopes with detectable phospho-residues selected for further analysis, three phosphorylated epitopes were recognized by CTLs from HIV-1 infected patients in direct ex-vivo assessments. Although CTL responses to these phosphorylated peptides were polyfunctional, the magnitude and cross reactivity of these phospho epitope-specific CTL varied with both epitope and HLA background (Fig A, B). Specifically, while both B*5701 and B*5703 patients responded to phosphorylated version of the immunodominant KF11 epitope, these responses were lower in magnitude and functional avidity when compared with the non-phosphorylated KF11 (Fig A). Cloning of KF11-specific CTLs indicated that cross-reactivity between the phosphorylated and the corresponding non-phosphorylated epitope were mediated by single TCRs (Fig C). Conclusion: Host kinases can phosphorylate HIV-1 at immunogenic regions. Multiple phosphorylated CTL epitopes can be recognized by HIV-1-specific CTLs, and TCRs able to cross-recognize phosphorylated epitopes are naturally recruited, suggesting that presentation of phosphorylated epitopes may occur in vivo. Phospho-epitope specific T cell responses may modulate the efficacy of antiviral cellular immune responses. cytometric analysis of antigen-specific effector response (Fig A, B). Results: We identified 30 unique and largely novel site-specific

Poster Abstracts

280 EVALUATING THE LINK BETWEEN MITOCHONDRIAL STATE & HIV- SPECIFIC CD8+ T CELL EXHAUSTION

Christian Deo T. Deguit 1 , Michelle A. Hough 2 , Rebecca Hoh 2 , Melissa Krone 2 , Christopher D. Pilcher 2 , Jeffrey N. Martin 2 , Steven G. Deeks 2 , Joseph M. McCune 2 , Peter W. Hunt 2 , Rachel L. Rutishauser 2 1 University of the Philippines Manila, Manila, Philippines, 2 University of California San Francisco, San Francisco, CA, USA Background: Antigen-specific CD8 T cell exhaustion, marked by poor proliferative and effector capacity, occurs in HIV disease and other settings of persistent antigen stimulation such as other chronic infections and cancer. Distinct metabolic features, such as increased mitochondrial mass (MM), decreased mitochondrial membrane potential (MMP), and increased reactive oxygen species (ROS) content are related to greater antigen-specific exhaustion. Yet, the contribution of these pathways to HIV-specific CD8 T cell exhaustion has not been explored. We hypothesized that HIV-specific CD8 T cells with features of exhaustion (e.g. cells from viremic individuals, with increased PD-1 expression and/or with poor proliferative capacity) have a distinct mitochondrial state that is linked to their dysfunction. Methods: Cryopreserved PBMC samples were obtained from 20 HIV-infected individuals in 3 clinical groups: Viremic (VL>2000 copies/mL, ART-naïve, n=6), ART-suppressed (VL<40 copies/mL on stable ART for a median of 11 years, n=8), and Controllers (VL<40 copies/mL, n=6). Using flow cytometry, we evaluated the MM, MMP, and ROS content of MHC Class I tetramer+ HIV-specific CD8 T cells in these individuals. We also characterized the tetramer+ cell expression of co- inhibitory receptors, effector molecules, and proliferative capacity after 6-day in vitro peptide stimulation. Results: Although the mitochondrial state of HIV-specific tetramer+ CD8 T cells did not vary by clinical group, we found significant differences within tetramer+ subsets identified by PD-1, CD127, and/or CD45RA expression. Total tetramer+ cells expressing PD-1 have greater ROS content (p<0.001) and MM (p=0.007) and lower MMP (p=0.027) than PD-1- cells, but for tetramer+ PD-1+ CD45RA- cells, CD127 expression was associated with decreased ROS content

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CROI 2018