CROI 2016 Abstract eBook
17 Rare Host Genetic Variation Influencing Risk of Heterosexual HIV-1 Acquisition Romel D. Mackelprang 1 ; Mary Emond 1 ; Michael J. Bamshad 1 ; Xuanlin Hou 1 ; Jessica Chong 1 ; Kati Buckingham 1 ; Nelly R. Mugo 2 ; Jared M. Baeten 1 ; Connie M. Celum 1 ; Jairam R. Lingappa 1 ; for the Partners in Prevention HSV/HIVTransmission and Partners PrEP Studies 1 Univ of Washington, Seattle, WA, USA; 2 Kenya Med Rsr Inst, Thika, Kenya Background: Since CCR5-D32, no common host genetic variant has consistently been associated with HIV-1 acquisition. However, whole genome sequencing may identify rarer novel variation that alters HIV-1 acquisition risk. We applied this approach to samples from Africans with high HIV-1 exposure to increase statistical power to detect associations.
Methods: This analysis included Discovery and Replication stages. In the Discovery stage, whole genome sequences were compared between 50 HIV-1 seroconverting (SC) cases and 50 HIV-1 exposed seronegative (HESN) controls with the highest HIV-1 exposure among ~4000 eligible HESN. Exposure was quantified based on unprotected sex acts and plasma HIV-1 RNA of the infected partner. Comparisons were done for each gene by aggregating variants “by-gene” using RVT1 burden tests. Discovery phase genes were sequenced in 199 HESN with high HIV-1 exposure and 1030 HESN with low exposure; 179 of these HESN acquired HIV-1. We used survival analysis to determine if having ≥1 rare variants in the Discovery genes was associated with HIV-1 acquisition in the Replication sample. Results: Discovery: Rare variants in CD101 and UBE2V1 were most strongly associated with HIV-1 acquisition (p=3.6x10 -5 and 4.5x10 -5 , respectively). Specifically, 30 (60%) cases had 2-5 rare CD101 missense variants compared to 12 (24%) controls. Each additional rare variant raised HIV-1 infection risk by 2.6-fold (95% CI:1.5-4.5). For UBE2V1, 27 (54%) cases had 1-2 rare variants vs 10 (20%) HESN controls; each additional variant raised HIV-1 risk 3.7–fold (95% CI:1.7-8). Replication: Associations of rare variation in CD101 and UBE2V1 replicated in the high HIV-1 exposure subset (Fig 1). Specifically, ≥1 specific rare CD101 variants identified in the Discovery stage yielded a hazard ratio (HR) for SC of 3.2 (95% CI:1.7-6.3) among participants reporting unprotected sex at >40% of study visits, p=5.2x10 -4 . This association decreased with lower HIV-1 exposure among the entire cohort (HR=1.4, p=0.02). UBE2V1 variants were associated with a HR=3.2 (95% CI:1.2-8.2;p=0.02) in the increased exposure Replication sample.
Conclusions: Using whole genome sequencing, we discovered and replicated associations of rare genetic variation in CD101 and UBE2V1 with elevated HIV-1 acquisition risk. CD101 could influence HIV-1 susceptibility by altering regulatory T cell function, and UBE2V1 through immune activation. These findings may be valuable for HIV-1 vaccine development and therapeutics. 18LB HIV-1 Laboratory Contagion During Recombination Procedures With Defective Constructs Claudia Alteri 1 ; Alessandro Soria 2 ; Ada Bertoli 1 ; Alessandra Bandera 3 ; Gabriella Scarlatti 4 ; MonicaTolazzi 4 ; Emanuela Balestra 1 ; Andrea Gori 3 ; Francesca Ceccherini-Silberstein 1 ; Carlo Federico Perno 5 1 Univ of Rome Tor Vergata, Rome, Italy; 2 San Gerardo Hosp, Monza, Italy; 3 San Gerardo General Hosp, Monza, Italy; 4 San Raffaele Scientific Inst, Milan, Italy; 5 Univ of Rome Tor Vergata, Roma, Italy Background: An accidental contagion during the production of theoretically non-infectious HIV-1 laboratory-recombinant viruses, used only for research purpose, is here described. Methods: HIV-1 infection was diagnosed by ELISA assay, confirmed by Western Blot, and HIV-RNA/DNA test. Baseline plasma samples were used for routine pol and V3 sequences, PBMC for the whole HIV-1 genome sequencing and for in-vitro isolation. Phylogenetic analyses using Neighbor Joining (NJ) and Maximum Likelihood (ML) methods revealed the source of infection. Ethics committee approval and signed informed consent were obtained. Results: A lab worker resulted HIV-1 positive during a routine screening HIV test. In the 6 months preceding HIV diagnosis, he/she exclusively worked on the production of a recombinant nef -defective HIV, starting from a nef / env -defective NL4.3 vector and JRFL env -encoding plasmid. In the same laboratory other HIV-1 constructs were handled by other researchers at the same time. A thorough investigation did not evidence any laboratory accident during the whole period. At diagnosis, CD4 were 392 cells/μL, HIV-RNA 3.30 log cps/mL, and HIV-DNA 157 cps/10 6 PBMC. Virus was R5-tropic. After 25 months of stable HIV-1 RNA and CD4-cells, a sudden 1-log HIV-RNA increase occurred; TDF/FTC/RPV cART was started, with full viremia control. NJ trees revealed a B-subtype virus, totally unrelated with other 629 HIV-1 clinical strains collected at the same hospital, but clustering exclusively with NL4.3 for pol sequence, and with JRFL for V3 (bootstrap: 96% and 100%, respectively). Genetic distance confirmed the homology of pol and V3 with NL4.3 and JRFL, respectively (0.002±0.002 and 0.000±0.000). Full-length viral sequence, performed in a different laboratory, confirmed the results and revealed a NL4.3/JRFL recombinant strain surprisingly expressing nef . A primary isolate obtained from his/her PBMC culture confirmed the nef -expressing NL4.3/JRFL recombinant strain. By inferring ML trees, the entire HIV-1 genome clustered again with NL4.3 (bootstrap>99%), with the exception of env (strongly linked with JRFL, bootstrap=99.7%). To date, how nef gene (absent in both vectors) entered into the recombinant virus, and mode of contagion, remain both unclear. Conclusions: This clinical case highlights that in-vitro recombination procedures with per se non-infectious vectors, in a laboratory handling multiple HIV constructs, may still represent a risk of HIV contagion notwithstanding increasing biosafety efforts. 19 Identification of a Highly Functional DC Subset in Controllers by Single-Cell RNA-Seq Enrique Martin-Gayo 1 ; Michael Cole 2 ; Kellie E. Kolb 3 ; Zhengyu Ouyang 1 ; SamW. Kazer 3 ; Bruce D.Walker 1 ; NirYosef 2 ; Alex K. Shalek 1 ; Xu G.Yu 1 1 Ragon Inst of MGH, MIT, and Harvard, Cambridge, MA, USA; 2 Electrical Engineering & Computer Sch, Berkeley, CA, USA; 3 MIT Inst for Med Engineering & Sci (IMES), Cambridge, MA, USA Background: Highly-efficient T cell-mediated immune responses are crucial for antiviral immune defense in HIV controllers, but the mechanisms supporting the development of T cell immunity in these persons are not well understood. One possibility is that conventional dendritic cells (cDCs) from these Elite Controllers (ECs) may contribute to antiviral immune defense through induction of potent type I IFN responses with improved antigen-presenting properties upon exposure to HIV-1. Here, we use single-cell transcriptional profiling to analyze cell-intrinsic immune responses to HIV-1 in isolated, individual cDCs from ECs. Methods: Single-cell RNA-seq profiling of HIV-1-exposed cDCs from an EC was used to identify gene expression programs associated with cell-intrinsic HIV-1 immune recognition and type I IFN signatures in cDCs. Potential surface markers for cDC subsets associated with improved functional properties were validated by flow cytometry. Functional properties of sorted cDC subsets were analyzed by co-culture with allogeneic T cells in mixed leukocyte reaction assays. Results: Single-cell RNA-seq identified three distinct cDC subpopulations from an EC after exposure to HIV-1. These cDC subsets differed in expression of genes related to IFN signaling, immune activation, cytokine signaling and HIV-1 replication, and were phenotypically distinguishable by two membrane markers. Importantly, among these cDC subsets, we identified a highly functional population, preferentially induced in ECs (n=8, p=0.007), in contrast to progressors (n=8) or healthy individuals (n=8), that had distinct and potent upregulation of interferon-stimulated genes and remarkably effective functional antigen-presenting properties. Importantly, induction of this highly functional cDC subset in response to HIV-1 exposure could be facilitated by specific TLR agonists in HIV-1 negative individuals (n=8, p=0.008).
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