CROI 2015 Program and Abstracts

Abstract Listing

Poster Abstracts

Results: As expected, both U3-Psi and gag assays detected HIV-1 DNA in >90% of the patients (44/48 and 46/48, respectively) with no significant quantitative bias between the assays, demonstrating the functionality of the U3-Psi assay. HIV-1 gag RNA was detected in 44/48 of these patients (92%) with the median copy number of 590 (interquartile range, 217-1194) copies/ m g total RNA. However, the detectability of readthrough RNA was only 40% (19/48 patients). In the 19 patients where the readthrough RNA was detected, its copy number was 49 (41-122) copies/ m g total RNA, representing only 8.3% (2.4%-11.2%) of the HIV-1 gag RNA. Notably, the real readthrough/ gag RNA ratio is much lower, as patients with undetectable readthrough RNA (60% of all patients) were excluded from this calculation.

Figure 1. Schematic representation of real-time PCR assays for the detection of readthrough and gag RNA. LTR, long terminal repeat; ORF, open reading frame; Ψ , HIV packaging signal (Psi). Conclusions: We observed only a minor contribution of host-HIV-1 readthrough transcripts to the level of HIV-1 gag RNA. The vast majority of HIV-1 gag RNA transcripts in ART- treated patients represent genuine HIV-1 unspliced RNA. 385 MicroRNA-155 Reinforces HIV Latency by Downregulating the TRIM32 Viral Activator Debbie S. Ruelas 2 ; Jonathan Chan 2 ; Eugene Oh 1 ; Amy Heidersbach 2 ; Andrew Hebbeler 2 ; Leonard Chavez 2 ; EricVerdin 2 ;Warner C. Greene 2 1 University of California San Francisco (UCSF), San Francisco, CA, US; 2 University of California Berkeley, Berkeley, CA, US Background: Achieving a cure for HIV will require both the complete suppression of active viral replication and the clearance of the transcriptionally silent proviral latent reservoir. Current drug treatments effectively target the active virus but leave the latent reservoir intact. We describe a cellular pathway involving miR-155 and one of its many cellular targets, TRIM32, that appears to promote a return to latency in reservoir cells transiently producing virus. Methods: We first assessed whether miRNAs play a role in maintenance of viral latency by separately knocking down the expression of two of the major enzymes involved in miRNA biogenesis, DGCR8 and Dicer, in J-Lat 5A8 cells. We next used miRNA TLDA analysis to identify specific miRNAs that alter the level of reactivation following stimulation. We focused on miR-155 because its reintroduction into Dicer-deficient cells was able to rescue the level of latent reactivation in J-Lat 5A8 cells to the greatest extent, suggesting that it plays a prominent role in reinforcing HIV latency. We confirmed TRIM32 as a novel target of miR-155 using luciferase binding assays. An IkB kinase (IKK) kinase assay revealed that TRIM32 acts downstream of the IKKs. Finally, our i n vitro ubiquitination assays demonstrate that TRIM32 is able to directly ubiquitinate I κ B α . Results: MiR-155, which is expressed at high levels in activated cells, impairs the expression of TRIM32, which normally serves as an HIV-activating agent. TRIM32 activates latent HIV by stimulating nuclear translocation of NF- κ B. However, our studies reveal that TRIM32 activates NF- κ B in a novel manner involving direct ubiquitination of I κ B α . Specifically, TRIM32 induction of NF- κ B proceeds independently of IKK activation within signalosomes. Conclusions: Our studies of the potential role of microRNAs in the regulation of HIV latency have led to the identification of miR-155 and its inhibition of TRIM32 activation of NF- κ B as events that promote the reestablishment of HIV latency in reservoir cells undergoing transient viral production. 386 Select Host Restriction Factors Are AssociatedWith HIV Persistence During Therapy Mohamed Abdel-Mohsen 1 ; Leonard Chavez 1 ; CharleneWang 1 ; Matt Strain 2 ; Xutao Deng 1 ; Christopher D. Pilcher 3 ;Teri Liegler 3 ; Douglas D. Richman 2 ; Steven Deeks 3 ; Satish Pillai 1 1 Blood Systems Research Institute, San Francisco, CA, US; 2 University of California San Diego, La Jolla, CA, US; 3 University of California San Francisco, San Francisco, CA, US Background: Although antiretroviral therapy (ART) results in sustained inhibition of HIV replication, the virus may continue to replicate at low levels. Moreover, ART has no effect on cells harboring integrated and silenced DNA (“latent reservoir”). A number of retroviral restriction factors have been identified which directly inhibit viral replication in vitro . However, the potential impact of these cell-intrinsic immune factors on viral persistence during ART is unknown. We investigated the relevance of a comprehensive panel of anti- HIV-1 host restriction factors to multiple virologic and immunologic measures of viral persistence in a cohort of 72 HIV-infected, ART-suppressed individuals. Methods: We measured the expression of 42 anti-HIV-1 host restriction factors, levels of cell-associated HIV-1 RNA, total pol and 2-LTR circle HIV-1 DNA, and immunophenotypes of CD4+ T cells from 72 HIV-1-infected subjects on suppressive ART (23 subjects initiated ART <6 months post-infection, and 49 subjects initiated >1 year post-infection). Data were analyzed using non-parametric tests. Results: The enhanced expression of three host restriction factors (p21, schlafen 11, and PAF1) was strongly associated with reduced CD4+ T cell-associated HIV RNA during ART (p<0.001). In contrast to previously published data on ART-untreated individuals, the expression of several restriction factors during ART exhibited negative correlations with frequencies of CD4+ T cells expressing markers of T cell activation and exhaustion (CD38, HLA-DR, PD-1) (p<0.01). Cell-intrinsic immune responses were significantly enhanced in subjects who initiated ART during early versus chronic infection (p=0.02). Conclusions: A few select intrinsic immune responses may modulate HIV persistence during suppressive ART, and therefore may be manipulated to both enhance the efficacy of ART and/or reverse latency in vivo . For example, inhibition of p21, which is known to suppress HIV transcription post-integration, may promote viral reactivation in the setting of the shock and kill eradication framework. Our data also suggest that ART perturbs the regulatory relationship between CD4+ T cell activation and restriction factor expression, which is pertinent to latency reversal agents that activate infected cells. The enhanced intrinsic immune response observed when ART is initiated early provides another possible justification for early administration of ART, and may contribute to the smaller reservoir size noted when ART is initiated early. 387 Selectively Eliminating HIV Latently Infected Cells Without Viral Reactivation Grant R. Campbell ; Rachel S. Bruckman;Yen-Lin Chu; Stephen A. Spector University of California San Diego, La Jolla, CA, US Background: Efforts to disrupt the establishment and maintenance of the latent reservoir have focused on the “shock-and-kill” therapeutic approach to reverse HIV latency from CD4 + T cells with subsequent killing of the infected cells. The X-linked inhibitor of apoptosis protein (XIAP) is up regulated in latently infected cell lines. In this study, we

Poster Abstracts

284

CROI 2015