CROI 2019 Abstract eBook

Abstract eBook

Poster Abstracts

became undetectable in the urine and plasma on the subsequent follow-up visits while the recipient was continuously maintained on ART. Conclusion: Our study demonstrates that following HIV+ to HIV+ kidney transplantation viruses from the donor’s kidney are found in the urine of the recipient immediately following transplantation, suggesting that donor’s kidney as the source of these viruses. Our results warrant long termmonitoring of viral populations in the recipient to fully assess any clinical and virologic implications of this finding. 227 GUT MICROBES DRIVE EXPANSION AND PREFERENTIAL HIV INFECTION OF GUT CD4 CTL EX VIVO Stephanie Dillon , Sabrina Nesladek, Allison J. Christians, Christine Purba, Background: HIV infection is associated with disruption of gut homeostasis and changes in the gut microbiome (dysbiosis). During early infection, HIV replicates to high levels in gut CD4 T cells concurrent with epithelial barrier breakdown and onset of microbial translocation. In transcriptome profiling studies using primary human lamina propria mononuclear cells (LPMC), we identified Granzyme (GZ) B and GZA induction in microbe and microbe/HIV stimulated gut CD4 T cells ex vivo (PMID 28241075). Here, we profiled microbe- induced human gut GZB-expressing CD4 T cells (termed CD4 CTLs) to determine the specificity of this response, potential mechanisms driving expansion and relative levels of HIV infection. Methods: Jejunum LPMC (n=13 donors), peripheral blood mononuclear cells (n=5) or tonsil cells (n=5) were cultured with or without gut commensal Escherichia coli lysate as well as enteric pathogenic, probiotic or dysbiotic bacteria altered in people living with HIV (PLWH). LPMCs were pre-treated with HLA blocking/control antibodies prior to addition of bacteria (n=7). LPMC were infected with Transmitted/Founder HIV-1 strain CH40 (n=3). Cytolytic markers (GZB, perforin, CD107a), infection (intracellular p24) and proliferation (CFSE) were measured by flow cytometry. Paired t tests were used for analyses. Results: Percentages of gut CD4 T cells expressing GZB were low at baseline (mean, SEM 1.4±0.5%), but exposure to multiple enteric bacteria increased % of GZB+ CD4 CTLs (Table 1), with greatest increases with E. coli (733-fold) and S. typhimurium (376-fold). E. coli induced a 4-fold increase in % of blood GZB+ CD4 CTLs (p=0.008), but did not induce GZB expression in tonsil CD4 T cells. HLA-DR blockade decreased the % of E. coli-driven GZB+ CD4 CTLs by 33±10% (p=0.02), but not their proliferation. Following HIV infection of E coli- exposed LPMC, a greater fraction of GZB+ than GZB- CD4s were infected (p24+; p=0.058). GZB+ CD4 CTLs that expanded with HIV + E. coli exposure expressed perforin (23±9.5%) and of those, 25±7.7% had degranulated (CD107a+). Conclusion: Diverse enteric bacteria induced GZB+ gut CD4 CTLs that are preferentially infected by HIV-1 ex vivo. Microbe-driven GZB induction was prominent in the gut, but not blood and lymphoid tissue CD4 T cells and was partially MHC Class II dependent. Gut cytotoxic CD4 T cells may have evolved for antimicrobial defense, but in the setting of HIV infection, these cells may accelerate gut pathogenesis by enhancing overall HIV infection and CD4 T cell death. Kejun Guo, Martin McCarter, Mario Santiago, Cara Wilson University of Colorado Anschutz Medical Campus, Aurora, CO, USA

1 National Institute of Allergy and Infectious Diseases, Bethesda, MD, USA, 2 Duke University, Durham, NC, USA, 3 University of Florida, Gainesville, FL, USA Background: Emergence of CXCR4-using HIV-1 (X4) expands host cell range and is associated with advanced stage disease in the absence of therapy. Yet, the developmental programmodulating X4 evolution remains elusive. This study tracked X4 evolution genetically during the natural history of pediatric HIV-1 infection to develop sequence profiles associated with functional characteristics of entry and tropism. Methods: Archived longitudinal blood samples were collected over 2-10 years from 8 untreated perinatally HIV-infected children. Disease progression was monitored by CD4 T-cell inflection point and CD4 T-cell decline to <15%. A total of 831 HIV-1 Env single genome sequences were generated. Env evolution was inferred by time-calibrated phylogenetic trees. CCR5 and CXCR4 coreceptor use was predicted by position specific scoring matrix and verified functionally using coreceptor indicator cells. Single-cycle viruses pseudotyped with Env V1-V5 were constructed to test tropism and entry efficiency into blood lymphocytes and monocyte-derived macrophages (MDM). Results: Infection was initiated by R5 variants in 7 cases or by X4 viruses in 1 case. R5 viruses persisted over years in 2 cases, while R5X4- and X4-predicted genotypes evolved from low frequency R5 viruses in 5 individuals prior to CD4 decline. Alignment of R5 and R5X4 Env sequences identified discontinuous nonsynonymous changes that altered neutralizing antibody epitopes initially in V1V2 and subsequently in V3. Single-cycle viruses generated using R5 and R5X4 Envs displayed entry into CD4 T-cells, but only R5X4 viruses infected MDM. In contrast to R5, R5X4 Envs were more sensitive to sCD4 (CD4 antagonist) or 447-52D (V3 antibody), indicating increased access to CD4 binding site and the V3-loop, but less sensitive to Maraviroc (anti-CCR5) or T20 (fusion inhibitor), consistent with increased CCR5-use and fusion efficiency. Conclusion: X4 evolution follows a complex developmental pathway that includes R5 ancestral strains and R5X4 intermediates, expands HIV-1 cell tropism, enhances viral entry via increased access to the CD4 binding site and the V3 loop and increase in fusion efficiency. Evolution of coreceptor preference accompanied by changes in neutralizing epitopes may reflect escape from immune response. 226 HIV+ TO HIV+ KIDNEY TRANSPLANT: TRACKING DONOR VIRUS IN RECIPIENT URINE AND BLOOD Maria Blasi 1 , Hannah Stadtler 2 , Jerry Chang 2 , Marion Hemmersbach Miller 2 , Feng Gao 2 , Cameron R. Wolfe 2 , Mary Klotman 2 1 Duke University School of Medicine, Durham, NC, USA, 2 Duke University, Durham, NC, USA Background: HIV-1 positive individuals have increased risk of end-stage kidney disease due the HIV-1 infection and associated treatments, yet now live longer. The HOPE Act allows individuals living with well-controlled HIV-1 to be eligible for organ transplant from HIV-1 positive donors that would have been otherwise discarded. One concern associated with HIV+ to HIV+ transplantation is the risk of superinfection and/or viral recombination resulting from the transmission of a genetically distinct HIV-1 strain from the donor to the recipient. In this study we used analysis of viral sequences derived from donor and recipient specimens to determine the source of virus in urine and blood specimens in the transplant recipient. Methods: Blood and urine specimens were obtained from both donor and recipient before transplantation and at different time points post- transplantation from the recipient. A renal biopsy from the donor kidney was obtained at time of procurement. We performed single genome amplification (SGA) of the full-length HIV-1 env gene with viral RNA extracted from urine, plasma and donor kidney biopsy as well as from viral DNA extracted from PBMC and urine derived renal cells. Neighbor-joining trees were constructed using the Kimura 2-parameter model. Results: Multiple HIV-1 env sequences were obtained from the samples collected from both donor and recipient. We found that all the env sequences from the recipient’s urine collected at 12 hours post-transplant were genetically similar to those in the donor while subsequent urine-derived sequences were genetically similar to the recipient virus. Furthermore, the majority of the urine derived sequences formed a separate cluster from donor-derived blood sequences, suggesting that the majority of urine-derived viruses were produced by infected cells within the donor kidney. Although the donor viruses could be readily amplified from the recipient’s urine soon after transplantation, it

Poster Abstracts

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CROI 2019