CROI 2016 Abstract eBook

Abstract Listing

Poster Abstracts

Conclusions: Using systems biology, we identified proteome signatures linked to inflammaging in ART-treated, virally suppressed HIV + persons. Inflammaging was associated with proteins involved in a number of different biological networks including oxidative stress and cell death. Understanding the mechanistic pathways driving inflammaging may provide new targets for anti-inflammatory treatments and identify novel biomarkers to further interrogate the effects of chronic HIV infection and aging on inflammation. 280 IInflammasome and Th17 Activation in HIV+ Immunological Nonresponders Michela Masetti 1 ; Massimiliano Fabbiani 2 ; DariaTrabattoni 1 ; Antonio Muscatello 3 ; Mara Biasin 1 ; Nicola Squillace 4 ; Irma Saulle 5 ; Mario Clerici 1 ; Andrea Gori 4 ; Alessandra Bandera 4 1 Univ of Milan, Milan, Italy; 2 Inst of Clinical Infectious Diseases, Catholic Univ of Sacred Heart, Rome, Italy; 3 San Gerardo Hosp, Monza, Italy; 4 San Gerardo General Hosp, Monza, Italy; 5 Univ of Milan, Brescia, Italy Background: Inflammasomes are multimeric protein platforms involved in the regulation of inflammatory responses. Inflammasome activity results in the production of proinflammatory cytokines and in Th17 activity (Mills K.H.G. et al, JLB 2013; Peelen E. et al, Molecular Immunology 2015); its role in immune reconstitution in HIV+ patients is nevertheless unclear. We analyzed possible associations between inflammasomes or Th17 activation and the degree of immune reconstitution in HIV+ patients. Methods: Cross-sectional, single-site study enrolling HIV-infected patients on antiretroviral therapy for ≥24 months and plasma HIV-RNA<50cp/mL for ≥12 months. Exclusion criteria: presence of actual opportunistic AIDS-related diseases, HBV or HCV coinfection, chronic inflammatory disorders, ongoing immunosuppressive therapy. Patients were classified as immunological responders (IR) or non responders (INR) if CD4 count was ≥500 or ≤350 cells/µL, respectively. Immune activation markers (HLA-DRII, CD38 and Ki-67), Th17 activity and expression of genes involved in the inflammasome pathway were measured in unstimulated or LPS- and AT2-stimulated cells. Results: Thirty-nine patients (22 IR and 17 INR, 76.9%males, medians: age 47 years, time from HIV diagnosis 10 years, time with HIV-RNA<50cp/mL 57 months) were enrolled. INR patients were older (median 60 vs. 43 years, p<0.001) and had a higher prevalence of past AIDS-defining illnesses (76.5% vs. 18.2%, p<0.001) compared to IR patients. Median CD4 count was 840 (IQR 718-1131) cells/µL in IR vs. 295 (IQR 256-343) cells/µL in INR. LPS-stimulated inflammasomes (NLRP3 and NLRP1) and pro-inflammatory cytokines expression (IL-1β, IL-18, TNFα, type-I IFNs, CCL3, IL-6) were significantly increased in INR patients. Higher median levels of Th17 lymphocytes (CD4/IL17A/RORgT) were also seen in INR in unstimulated (0.32% vs. 0.19%, p=0.045), as well as in LPS- (0.60% vs 0.22%, p=0.011), and AT2- (0.67% vs 0.17%, p=0.006) stimulated conditions. Finally, HLADRII/ CD4 (12.48% vs. 5.86%, p=0.025) were significantly increased in unstimulated cell cultures of INR. Immune recovery was independently associated with lower Th17 levels (mean change -0.42%, p=0.005) after adjustment for age and past AIDS. Conclusions: Higher levels of inflammasome factors, an increased percentage of Th17 and immune activation characterize the immune scenario of ART-treated INR patients. These alterations are likely to contribute to the lack of CD4 recovery seen in INR. 281 Derangement in Protein S and C4b Binding Protein Levels in HIV-Infected Adults Fatai O. Bello 1 ; Suleiman Akanmu 2 ;Titilope Adeyemo 2 ; Funmi Lesi 2 ; Prosper Okonkwo 3 ; Sade Ogunsola 2 ; Phyllis Kanki 4 1 Lagos Univ Teaching Hosp, Lagos, Nigeria; 2 Coll of Med, Univ of Lagos, Lagos, Nigeria; 3 AIDS Prevention Initiative in Nigeria, Abuja, Nigeria; 4 Harvard Sch of PH, Boston, MA, USA Background: The introduction of highly active antiretroviral therapy (HAART) has significantly improved survival among HIV infected persons. Chronic complications of the disease and long term toxicities of the antiretroviral therapy are now important issues that confront the health care providers managing HIV infected patients.HIV infected patients have an increased risk of developing thrombotic disorders.Protein S is a vitamin K-dependent plasma glycoprotein that exists in a free form and one bound to complement protein (C4BP). Protein S is critical in the anticoagulation pathway functioning with Protein C in the inactivation of Factors Va and VIIIa. We evaluated this with the Euglobin clot lysis test and measured serum levels of free protein S and C4BP (an acute phase reactant that serves as carrier protein for Protein-S in plasma), to determine their role in the increased risk of thrombosis. Methods: The study population consisted of 61 HIV infected adults on HAART that had achieved virological suppression, 58 HIV infected adults not yet on treatment with ART and 59 HIV negative healthy adults as controls. Blood samples were collected and serum levels of free protein S,C4b binding protein beta and the Euglobin clot lysis time were determined. Results: There was a significant decrease in serum free Protein S level in untreated HIV infected adults(75.7%±27.3) compared to the control group (94.9%±7.9).( p=0.000 ). There was no statistically significant difference between Protein S levels in HIV infected subjects on ART (86.9%±25.8) and the control group(94.9%±7.9)( p=0.119 ). C4b Binding Protein beta was significantly higher in untreated HIV infected subjects(mean:85.1mg/dl±52.6) compared to those on ART (mean:43.8mg/dl±42.4) and the control group(22.4mg/dl±18.4).( p=0.000 ). There was a statistically significant difference between C4BP levels in HIV infected subjects on ART and the control group ( p=0.012 ). Of the 24 HIV infected subjects with low protein-S levels, 19 (79.16%) had elevated C4B binding proteins. whereas in 95 HIV infected subjects with normal protein-S levels, 51(53.68%) had elevated C4B binding proteins. Protein S deficiency is more prevalent among the subjects with elevated C4b Binding Protein beta ( p=0.023 ). Mean Euglogin Clot Lysis Time (ECLT) was significantly longer among the (n=47) untreated HIVinfected subjects (241.9secs± 34.7)compared to the HIV uninfected control group (n=59) (189.5±40.7).( p=0.000 ). ECLT in HIV infected subjects on ART (210±61.9 sec n=50) was not significantly different from the control group.( p=0.064 ). Conclusions: HIV infection is associated with derangement in the levels of C4b Binding Protein beta and serum free Protein S.This may be responsible for the increased risk of thrombosis associated with HIV infection as demonstrated by the prolonged ECLT in treatment naïve HIV subjects.The risk of thrombosis as measured by these biomarkers was reduced with the use ARVs. 282 Monocytes From Treated HIV1 Patients Release ROS Causing DNA Damage and CD4 Cell Loss Background: Immune activation is a hallmark of HIV infection. Antiretroviral therapy reduces but doesn’t abolish it. Immune activation is linked to sub optimal immune response and to non-AIDS comorbidities, including various types of cancers. The aim of this project was to explore how immune activation could favor impaired CD4+ T cell restoration and oncogenesis. Methods: We tested the capability of various populations of peripheral blood mononuclear cells (PBMC), isolated by cell sorting from 70 HIV-1 aviremic patients under antiretroviral therapy for at least two years to induce DNA damage in bystander primary fibroblasts (BJ cells) or PBMC. DNA damage was measured in immunofluorescence by looking for the formation of γH2AX, 53BP1, and 8-hydroxy-2-deoxyguanosine foci. Reactive oxygen species (ROS) production in patients monocytes and apoptosis induced in CD4+ T cells was quantified in flow cytometry after annexin-V and 2’,7’ –dichlorofluorescin diacetate (DCFDA) labeling, respectively. Results: We observed that the PBMC from 38 out of 70 (54%) virologic responders induced gH2AX foci in bystander BJ cells. DNA double-strand breaks and DNA oxidation were detected in these BJ cells using anti-53BP1 and anti-8-hydoxy-2-deoxyguanosine, respectively. Positive and negative cell sorting provided evidence that among the HIV patients PBMC, the monocytes were responsible for this DNA damage. They cause DNA damage by secreting reactive oxygen species (ROS), since this effect was eliminated by the pretreatment of the monocytes with NADPH oxidase inhibitor Diphenyleneiodonium (DPI), or by the ROS scavenger N-acetylecysteine (NAC). Moreover, the intensity of the DNA damage induced by the monocytes in coculture was linked to the amount of ROS detected in these cells ex vivo by DCFDA staining (r =0.12±0.05, p ≤ 0.05). Monocytes also caused DNA damage in primary CD4+ T cells, thereby inducing apoptosis. Finally, the capability of the monocytes of HIV+ patient to damage DNA was inversely correlated with its CD4 slope during treatment. Conclusions: This study uncovers the genomic instability mediated by monocytes-derived ROS in about half of the aviremic patients under treatment. This genomic instability can lead to poor immune recovery and oncogenesis. Mehwish Younas 1 ;Yea Lih Lin 2 ; Christina Psomas 3 ; Sandrine Gimenez 1 ; Pierre Portales 3 ; Jacques Reynes 3 ; Philippe Pasero 1 ; Pierre Corbeau 1 1 Inst of Human Genetics, Montpellier, France; 2 Inst de Genetique Humaine, Montpellier, France; 3 Univ Hosp Montpellier, Montpellier, France

Poster Abstracts

108

CROI 2016