CROI 2016 Abstract eBook

Abstract Listing

Poster Abstracts

Methods: We conducted an observational study including 29 HIV-1 infected patients and 9 uninfected controls. Peripheral blood and fecal samples were collected from ART naïve patients at baseline and for 16 patients during follow up (FU, after introduction of ART). Levels of the TRP metabolites were evaluated by HPLC assay and microbiota composition was determined by 16s rRNA sequencing. Soluble markers were analyzed by ELISA. Results: We found significantly higher levels of KYNA in healthy controls compared to ART naive patients, p<0.05, and significantly increased levels of IDO-1 activity was detected during FU compared to healthy subjects, p<0.05. Levels of kyn (p<0.005), IDO-1 (p<0.0005), quinolinic acid (QA, p<0.0005), 3-hydroxykynurenine (3-HK, p<0.05) and Nicotinamide (NAM, p<0.005) were reduced in HIV patients at FU. Levels of kyn and IDO-1 were correlated to CD8+ Tcells (R-value=0.40; p<0.05). In ART naïve patients, kyn was associated to the genera: Granulicatella (0.48; p<0.05), Butyricimonas (0.43; p<0.05) and Escherichia (0.51; p<0.05). IDO-1 was associated to Granulicatella (0.43; p<0.05) and Lachnobacterium (0.38; p<0.05), while negatively correlated to Sutterella (-0.38; p<0.05). Butyricimonas was correlated to: HAA (0.61; p<0.05), XA (0.44; p<0.05), KA (0.42; p<0.05), QA (0.47; p<0.05) and NAM (0.43; p<0.05). After ART intro, new associations between gut microbiota composition and tryptophan catabolites were revealed. Thus, kyn was correlated to Blautia (0.63; p<0.05) and Rothia (0.59; p<0.05) and negatively associated to Heamophilus (-0.52; p<0.05) and Bilophila (-0.56; p<0.05). IDO-1 was correlated to Blautia (0.66; p<0.05), but negatively correlated to Oscillospira (-0.62; p<0.05) and Oxalobacter (-0.77; p<0.05). Conclusions: We show that ART decreases the levels of TRP catabolites in viremic patients. Several significant associations between TRP catabolites and gut microbiota were found, and changed at FU. Our findings imply close interplay between gut microbiota and TRP pathway during HIV-1 infection. 268 Chronic Semen Exposure Reduces SIVmac251 Vaginal Infection in Rhesus Macaques Shaheed A. Abdulhaqq 1 ; Melween I. Martinez 2 ; Guobin Kang 3 ; David Beaumont 4 ; Jan Münch 5 ; GeorgiaTomaras 4 ; Preston A. Marx 6 ; Qingsheng Li 7 ; Edmundo N. Kraiselburd 8 ; Luis J. Montaner 9 1 Oregon Hlth & Sci Univ, Beaverton, OR, USA; 2 Caribbean Primate Rsr Cntr, San Juan, PR, USA; 3 Univ of Nebraska, Lincoln, NE, USA; 4 Duke Univ, Durham, NC, USA; 5 Univ of Ulm, Ulm, Germany; 6 Tulane Univ Sch of PH and Trop Med, New Orleans, LA, USA; 7 Sch of Biological Scis and Nebraska Cntr for Virology, Univ of Nebraska, Lincoln, NE, USA; 8 Sch of Med, Univ of Puerto Rico, San Juan, PR, USA; 9 The Wistar Inst, Philadelphia, PA, USA Background: We tested the effects of prolonged intravaginal exposure to semen and/or defective SIV particles on a low-dose SIV challenge in nonhuman primates. We hypothesized that a decrease in infection would be documented based on cross-sectional studies of HIV negative human sexworker cohorts where chronic semen exposure was associated with an induced antiviral host response. Methods: We exposed 46 randomized Indian Rhesus macaques ( Macaca mulatta ) to twice-weekly intravaginal inoculations with (n=23) or without (n=23) human semen with/ without containing defective SIV particles for 20 weeks. A subset of animals (n=14) was euthanized and vaginal tissues taken to assess immune cell infiltrates by IHC. Remaining animals were given a low-dose 16-week intravaginal challenge with SIVmac251 while continuing respective semen exposures until infection. Peripheral immune cell status, mucosal activation, and SIVmac251 specific mucosal antibody were assessed at baseline and pre/post challenge. Differences between groups were tested using the Wilcoxon rank sum test, α=0.05. Log-rank test was used to determine the effect of semen with/without defective particle exposure on infection susceptibility. Results: During SIVmac251 challenge, animals receiving semen were less susceptible to SIVmac251 infection (Log-rank p=0.0332) with 50% infection not occurring until week 15 of the 16-week challenge series as compared to week 7 for those animals not receiving semen. Exposure to defective particles had no effect on infection. Animals receiving semen had increased levels of RANTES upon exposure to SIVmac251 (p=0.0280). 20-week exposure to semen increased cervical Mx1 protein expression (p=0.02), CD4+ T-cell infiltrate (p=0.0087), and HLA-DR expression (p=0.0027). Reduced expression of CCR5 on circulating CD4+ T-Cells (p=0.0167) was noted in semen-exposed animals yet no difference in activation markers HLA-DR and CD69. Conclusions: Chronic exposure to semen can modulate for reduced susceptibility to SIVmac251 in association with lower expression of CCR5 in peripheral CD4+ T-cells, an induction of RANTES, and an increase in Mx1. Chronic semen exposure may allow animal models to induce vaginal changes likely present in high-risk human cohorts otherwise not accounted for in pre-clinical studies. 269 Single-Cell Expression Profiling Reveals Immune Dysfunction in Acute SIV Infection Sama Adnan ; Premeela A. Rajakumar; James M. Billingsley; R. P. Johnson Emory Univ, Atlanta, GA, USA Background: Despite the correlation between the HIV set-point and disease progression, little is known about the immunologic determinants of the viral set-point. During the first weeks of wild-type SIV (SIV-wt) infection, the rate of viral clearance decreases steadily, culminating in viral set-point by week 8, whereas animals infected with nonpathogenic SIV∆nef maintain high virus clearance rates. We hypothesized that changes in the effector function of SIV-specific CD8 T cells would provide insights into the divergent virus clearance rates in the two models and applied a novel technique involving high-throughput expression profiling of SIV-specific CD8 T cells to address this hypothesis.

Poster Abstracts

Methods: We isolated viable SIV-specific CD8 T cells sorted by expression of CD69 and CD137 after peptide stimulation followed by targeted qPCR transcriptional profiling. Cells were sorted from PBMC of 10 animals infected with wild-type SIV or SIV∆nef at weeks 3, 8 and 20. We then profiled the expression of 85 effector molecules in SIV-specific CD8 T cells using the Fluidigm Biomark platform. Single cell expression analysis of the same 85 effector molecule panel was conducted on SIV-specific CD8 T cells fromweek 8 using the Fluidigm C1 System. Results: At week 3, the expression profiles of CD8 T cell responses to SIV-wt and SIV∆nef were largely similar, although TGF-β1 expression, which inversely correlated with the magnitude of the CD8 T cell response, was significantly higher in SIV-wt infection, corresponding to a lower initial response magnitude in SIV-wt infection despite higher viral loads. By week 8, the CD8 T cell response in SIV-wt infection significantly increased the expression of pro-inflammatory cytokines (IL-1β, IL-1α, IL-18, IL-8) relative to week 3, even as responses in SIV∆nef infection downregulated pro-inflammatory cytokine expression. Single cell expression profiling at week 8 revealed a unique subset of SIV-specific CD8 T cells in wild-type SIV, but not SIV∆nef, infection (see Figure) that expressed CSF1 as well as lower levels of both chemotactic cytokines and cytolytic effector molecules. Conclusions: Our results demonstrate that, as early as week 3, the CD8 T cell response to SIV-wt is dampened relative to SIV∆nef, potentially due to TGF-β1 expression. By week 8, higher expression of pro-inflammatory cytokines, including CSF-1, in SIV-wt infection implicates myeloid derived suppressor cells, reportedly induced and recruited by persistent inflammation, in determining set-point viral load.

TNFSF4 TNFSF7 TNFSF8 IL_2 IL_23A CXCL13 TNFSF3 CXCL6 CCL23 IL_10 IL_14 TBP GZMA IL_18 GZMH Perforin GZMB TNFSF6 TNFSF9 IPO8 GZMK AIMP1 CCL3 CCL4 CCL4L1 IFNG XCL1 B2M GAPDH HU_PGK ACTB CCL25 XCL2 CSF1 TNFSF2 IL_1B CCL5 TNFSF1

CXCL11 CXCL16I L_16 TNFSF12 TGFBETA1 TNFSF13 CXCL9

Figure: Single-cell expression profiling of CD8 T cell response. K-means clustering of SIV-specific CD8 T cells from SIV∆nef (blue) and WT SIV (red) at week 8 reveals 4 distinct cell populations. The red cluster is largely composed of cells from WT SIV infection. WTTWTTWTTWTTWTTWT dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef WTTWTTWTTWTTWTTWTTWTTWTTWTTWTT dnef dnef WTTWTTWTTWTTWTTWTTWTTWTTWTTWTTWT dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef WTTWTTWTTWTTWTTWTTWT dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef dnef

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CROI 2016