CROI 2016 Abstract eBook

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Oral Abstracts

Modeling data suggest the need for combination high-impact prevention in order to significantly reduce HIV incidence in this population. Uptake of and adherence to PrEP has been limited by community concerns about potential drug-drug interactions between ART and hormone therapy; and preliminary unpublished data suggest associations between ART adherence and co-location of gender-related care with HIV services. Current NIH-funded studies include behavioral, self-testing, PrEP adherence, and telemedicine interventions. However, more research is needed. Data on HIV prevalence in transgender populations is missing from the African continent as well as Eastern Europe/Central Asia. Little is known about the sexual partnerships and networks of transgender people; nor is there data on co-morbidities and co-infections among HIV-infected transgender people taking exogenous hormones. Implementation science studies are needed to inform how best to implement and scale up multi-component, high impact, prevention care and treatment interventions that address structural drivers, reduce HIV incidence, and improve the health and longevity of transgender people living with HIV. 80 Investigating the Mechanisms that Control HIV Transcription and Latency In Vivo Steven A. Yukl 1 ; Philipp Kaiser 1 ; Peggy Kim 1 ; Sunil K. Joshi 1 ; Nicholas Kim 1 ; Peilin Li 1 ; Harry Lampiris 1 ; Hongbing Liu 2 ; Andrew Rice 2 ; Joseph K.Wong 1 ; for the DARE Study Group 1 San Francisco VA Med Cntr, San Francisco, CA, USA; 2 Baylor Coll of Med, Houston, TX, USA Background: It is unclear what mechanisms maintain HIV latency in vivo. We hypothesized that the levels of different HIV RNA transcripts could be used to infer the sites of transcriptional blockade. Methods: CD4+T cells from blood of 9 ART-suppressed individuals were isolated using negative selection and frozen or activated for 2 days with anti-CD3/CD28 beads. RNA was extracted using Trireagent. Aliquots from common RT reactions were used in ddPCR assays specific for different HIV transcripts indicative of transcriptional interference (U3-U5; “read-through”), initiation (TAR), elongation (R-U5-tRNA; “long”), completion (3’LTR-polyA; “polyA”), and multiple splicing (tat-rev). HIV RNA levels were expressed as absolute levels (normalized to cell counts) and ratios to total (TAR) and processive (long) transcripts. Results: In unstimulated CD4+T cells, the relative abundance of HIV transcripts was: TAR (median 18,060 copies/10 6 cells) > long (1816) > polyA (257) and read-through (227) > tat-rev (4.7) [p≤0.05], indicating blocks to proximal elongation, completion, and splicing. Most HIV transcripts were prematurely-terminated transcripts that had not elongated past the TAR loop (median long/TAR=0.073). Of processive transcripts, most had not completed transcription (polyA/long=0.15) and very few were spliced (tat-rev/long=0.0045). Read-through transcripts were detected in all individuals, but levels were modest in relation to processive transcripts (read through/long=0.15) and total transcripts (read- through/TAR=0.013). Activation increased all transcripts, but the increases in read-through (median d2/d0=3.1) and TAR (5.5x) did not exceed the global increase in cellular transcription, while “long” tended to increase more (7.4x) and the greatest increases (p<0.05) were seen in polyA (45.8x) and tat-rev (116x). Activation did not change read- through/TAR, tended to increase long/TAR (median d2/d0=3.4), and resulted in larger increases (p<0.05) in polyA/TAR (11.8x), tat-rev/TAR (61.8x), polyA/long (3.4x), and tat-rev/ long (11.7x). Conclusions: In unstimulated CD4+T cells, transcriptional interference plays a modest role in limiting HIV transcription, while blocks to elongation, completion, and splicing dominate. Activation selectively increased distal and spliced transcripts but had less effect on read-through or total transcripts, suggesting that the main reversible blocks to HIV expression are not interference or lack of initiation, but rather inhibition of elongation/polyadenylation and splicing. 81 Human Galectin-9 Is a Potent Mediator of HIV Transcription and Reactivation Mohamed Abdel-Mohsen 1 ; Leonard Chavez 1 ; Glen M. Chew 2 ; Xutao Deng 1 ; Ali Danesh 1 ; Sheila M. Keating 1 ; Rebecca Hoh 3 ; Steven G. Deeks 3 ; Lishomwa C. Ndhlovu 2 ; Satish Pillai 1 1 Blood Systems Rsr Inst, San Francisco, CA, USA; 2 Univ of Hawaii, Honolulu, HI, USA; 3 Univ of California San Francisco, San Francisco, CA, USA Background: Identifying host determinants governing HIV transcription and latency is critical to developing an HIV cure. Based on our recent finding that the host factor p21 regulates HIV transcription during antiretroviral therapy (ART), and published data demonstrating that the glycan-binding protein galectin-9 (Gal-9) regulates p21, we hypothesized that Gal-9 modulates HIV transcription. Methods: Plasma Gal-9 levels were examined in relation to measures of the latent HIV reservoir in 72 HIV-infected ART-suppressed individuals. The ability of a recombinant, stable form of Gal-9 (rGal-9) to reactivate latent HIV was evaluated in the J-Lat 5A8 HIV latency model, and in primary CD4+ T cells isolated from 13 HIV-infected ART-suppressed individuals. Enzymatic and chemical deglycosylation was used to explore the requirement of glycans in viral reactivation by rGal-9. Effects of rGal-9 on the host transcriptome were evaluated using RNA-seq. Results: Endogenous levels of plasma Gal-9 were associated with levels of CD4+ T cell-associated HIV RNA (p<0.02) and with the quantity and binding avidity of anti-HIV antibodies (p<0.009) in vivo during ART. Administration of rGal-9 reactivated virus in J-Lat cells (15.1%) more potently than anti-CD3/CD28 stimulation (4.8%, p<0.0001). In CD4+ T cells from HIV-infected individuals, rGal-9 induced a mean 7.3-fold increase in intracellular HIV RNA levels, as compared to DMSO negative control (p=0.002). Induction was significantly higher than vorinostat (p=0.02, 3.2 fold). Dosages of rGal-9 required to reverse HIV latency ex vivo were well-tolerated in vivo in Lewis rats. Cell surface N-linked oligosaccharides and O-linked hexasaccharides were essential for rGal-9-induced HIV reactivation, mediated by key transcription initiation, promoter proximal-pausing, and chromatin remodeling factors (FDR<0.05). rGal-9 induced expression of the host antiviral deaminase APOBEC3G up to 29-fold in vitro and ex vivo (FDR<0.006), resulting in 6.7-fold reduction in infectivity of progeny virus. Conclusions: rGal-9 potently reactivates latent HIV and induces APOBEC3G expression in vitro and ex vivo . rGal-9-induced virus will likely be rendered replication incompetent as a result of APOBEC3G induction in the producer cell, ensuring that the reservoir will not be replenished when latency is reversed therapeutically, even in the setting of suboptimal ART suppression. Our data suggest that gal-9 and the glycosylation machinery should be explored as a foundation for novel HIV cure strategies.

Oral Abstracts

31

CROI 2016