CROI 2015 Program and Abstracts

Abstract Listing

Poster Abstracts

chemical extraction methods to fractionate collagen molecules based on solubility in guanidine HCl. Guanidine-soluble collagen represents more recently synthesized, less cross- linked collagen. Incorporation of the 2 H 2 O tracer into collagen in GALT was quantified by liquid-chromatography-mass spectrometry and fractional synthesis rate (FSR; per week) was calculated for guanidine-soluble and -insoluble GALT collagen. Results: Outpatient administration of 2 H 2 O was well tolerated. After 28 days of 2 H 2 O intake, subjects reached the target goal of 1-2% 2 H 2 O enrichment in TBW. The median FSR of guanidine-insoluble collagen in GALT was 3.2% per week (IQR 2.9%-3.9%). Guanidine-soluble collagen in GALT turned over much more quickly, with a median FSR of 12.8% per week (IQR 12.3%-16.1%). In comparison with other human studies where collagen FSR was measured by 2 H 2 O labeling, the FSRs in GALT were 5-fold higher than the FSRs of skin collagen in healthy volunteers but similar to the FSR of liver collagen in patients with fibrotic liver disease. Conclusions: The relatively high rates of collagen turnover observed in the GALT during treated HIV disease (guanidine soluble collagen half-life ~ 5 weeks) suggest that collagen deposition and turnover is ongoing and dynamic in this setting, and may therefore be amenable to interventions. The 2 H 2 O labeling technique provides an assessment of collagen turnover rates, in contrast to static metrics from traditional histologic techniques, and may be useful for evaluating the effects of future anti-fibrotic therapies.

THURSDAY, FEBRUARY 26, 2015 Session P-C7 Poster Session

Poster Hall

2:30 pm– 4:00 pm HIV/CMV Interactions in Transmission and Pathogenesis 300 Effect of CMV and HIV Replication on T-Cell Exhaustion and Senescence During ART Jennifer M. Dan 1 ; Marta Massanella 1 ; David M. Smith 1 ; Eric S. Daar 2 ; Michael P. Dube 3 ; Richard Haubrich 1 ; Sheldon Morris 1 ; Sara GianellaWeibel 1

1 University of California San Diego, La Jolla, CA, US; 2 Harbor–University of California Los Angeles Medical Center, Torrance, CA, US; 3 University of Southern California Keck School of Medicine, Los Angeles, CA, US Background: HIV-infected men who have sex with men (MSM) are nearly universally infected with CMV, and both viruses are associated with T-cell dysfunction and inflammation-related morbidities. The effect of asymptomatic CMV replication and persistent HIV transcription during suppressive ART on markers of T cell exhaustion and senescence is poorly defined. Methods: Paired seminal and blood samples from 45 asymptomatic chronically HIV-infected CMV-seropositive MSM on long term ART and with HIV RNA levels in blood plasma <50 copies/ml were studied. Levels of CMV DNA in semen and blood were measured by RT-PCR, and cell-associated HIV DNA and RNA transcripts (unspliced) were measured in PBMC by droplet digital PCR. Markers of T cell exhaustion (PD-1) and senescence (CD57) were measured in PBMC by flow cytometry for CD4 and CD8 T cells and subsets (naïve [CD45RA + CD27 + CD28 + ], central memory [CD45RA - CD27 + CD28 + ], effectors [CD4 + CD45RA - / + CD27 - CD28 + or CD8 + CD45RA - / + CD27 + CD28 - ] and terminally differentiated [CD45RA + / - CD27 - CD28 - ]). Associations between immunological markers and asymptomatic CMV and HIV replication, HIV DNA, CMV IgG, age, current and nadir CD4 and time on ART were determined using univariate and multivariate analysis. Results: CMV DNA was detected in 42% of seminal samples and 20% of PBMC. Detectable CMV DNA in semen but not blood was associated with increased PD-1 expression on circulating CD4 T cells compared to no CMV (P=0.01), particularly in the effector and terminally differentiated subsets (P<0.05). Similarly, higher levels of cellular HIV RNA (but not HIV DNA) were positively associated with greater PD-1 expression on total CD4 and central memory blood subset (P<0.01). There was no association between CMV DNA (blood and semen) or cellular HIV RNA with CD8 exhaustion or senescence or with markers of CD4 senescence. In multivariate analysis, detection of seminal CMV and higher cellular HIV RNA remained associated with increased PD-1 expression on total CD4 T cells (P<0.05). No other variable contributed significantly to the model. Conclusions: Our data suggest that detection of CMV in the genital tract may contribute to the activation of the PD-1 axis on circulating T cells during suppressive ART. Because increased PD-1 on T cells has been implicated in the maintenance of the HIV reservoir, HIV disease progression and the inability of the immune system to adequately control HIV infection, future studies should examine whether CMV-dependent mechanisms play a role in T cell exhaustion. 301 HIV Myeloid Derived Suppressor Cells Control Cytomegalovirus Inflammation by IL-27 Ankita Garg ; Stephen Spector University of California San Diego, La Jolla, CA, US Background: CMV is associated with persistent inflammation in HIV-infected persons. Here, we studied the effect of HIV expanded myeloid derived suppressor cells (MDSCs) in controlling CMV specific inflammation. Methods: PBMCs from HIV–/CMV-seropositive (CMV+) donors were cultured in presence of heat inactivated HIV. After 5 days, CD11b + CD33 + CD14 + HLA DR hi (DR hi monocytes) and CD11b + CD33 + CD14 + HLA DR -/lo (MDSCs) cell subsets were sorted by flow cytometry. B7H4 (a negative regulator of T cell function) was silenced using siRNA and cultured with/without autologous PBMCs in presence/absence of CMV pp65 peptides (pp65; 1 m g/ml). In some experiments, PBMCs were cultured with HIV/pp65 in presence/absence of neutralizing anti-IL-27 antibody. Enumeration of B7H4 on MDSCs, regulatory T-cells, intracellular IFN γ , activated forms phospho(p)-Zap70 and p-Akt were determined by flow cytometry; IFN γ and IL-27 were quantified by ELISA. Data were analyzed using two-tailed, paired Student’s t test. Results: MDSCs cultured with autologous PBMCs exposed to pp65 caused a decrease in IFN γ production vs. controls or DR hi monocytes (p=0.002). IFN γ release was restored when MDSCs were transfected with B7H4 siRNA and cultured with PBMCs in presence of pp65 (p=0.02). MDSCs cultured with PBMCs did not alter pp65 induced activation of proximal T-cell signaling molecule Zap70 but decreased activation of Akt; this was restored when B7H4 was knocked down in MDSCs cultured with PBMCs. Culture of MDSCs with pp65 produced more IL-27 vs. control MDSCs and DR hi monocytes (p=0.05). Culture of CMV+ PBMCs with pp65 increased the frequency of CD4 + IFN γ + cells and release of IFN γ in supernatants vs. controls (p=0.04). IFN γ was further increased when PBMCs were cultured in presence of anti-IL-27 and stimulated with pp65; CMV– PBMCs did not produce IFN γ when treated with pp65. Furthermore, culture of CMV+ PBMCs with pp65 led to expansion of FoxP3 + Tregs vs. controls (p=0.02) and CMV– (p=0.003); addition of anti-IL-27 had no effect on Treg expansion. Finally, HIV expanded MDSCs had increased expression of B7H4 when compared to DR hi monocytes (p=0.03) which was inhibited in the presence of anti-IL-27 neutralizing antibody (p=0.05). Conclusions: These findings suggest that IL-27 down regulates IFN γ during CMV infection. IL-27 induces B7H4 expression on HIV MDSCs which controls CMV induced T-cell IFN γ production by inhibiting p-Akt. IL-27 and B7H4 provide new therapeutic targets to control inflammation during HIV infection.

Poster Abstracts

246

CROI 2015