CROI 2020 Abstract eBook

Abstract eBook

Poster Abstracts

killing of infected cells or the production of β-chemokines that block HIV-co- receptor usage. However, it remains unclear whether effector HIV-specific CD8+T cells expressing cytolytic molecules are present within the airway, a site of HIV persistence. Methods: We recruited 80 HIV-uninfected healthy controls and 80 asymptomatic ART-naïve HIV-infected adults who were followed up for a year on ART. We collected paired bronchoalveolar lavage (BAL) fluid and peripheral blood samples on all participants. We performed flow cytometry-based characterization of CD8+T cell phenotypes and also quantified HIV (Gag, Nef and Pol)-specific IFN-g-producing CD8+T cells in BAL and blood cells. Results: CD8+ T cells expressing Perforin and Granzyme B were found predominantly in the blood compared to the airway, regardless of HIV infection status. The frequency of Eomes+CD8+ T cells was higher in blood-derived cells compared to those from the airway lumen. PD1 or 2B4-expressing CD8+ T cells were higher in airway-derived cells compared to blood. Untreated HIV-infected adults had more Eomes+PD1+CD8+T cells compared to healthy controls; predominately in the airway compared to the systemic circulation. HIV-specific (Gag, Nef and Pol) CD8+T cells exhibited a higher breadth in the airway than in blood. There was no correlation between HIV-specific CD8+T cell responses in the airway and peripheral blood. HIV-specific CD8+ T cells did not express Perforin and Granzyme B but expressed high levels of PD1 and Eomes, markers associated with immune regulation and exhaustion in HIV infection. Conclusion: We demonstrate that airway-derived HIV-specific CD8+T cells poorly express cytolytic molecules (Granzyme B and Perforin) and possess markers consistent with high immune regulation (PD1 and Eomes). The poor cytolytic potential and highly regulated phenotype of airway HIV-specific CD8+T cells could promote the persistence of HIV-infected cells in the airway in individuals on long term ART. 269 HIGH FREQUENCY OF CD8 ESCAPE MUTANTS IN ELITE CONTROLLER AS NEW OBSTACLE TO HIV CURE María Ángeles Navarrete 1 , Marcial García 1 , Ricardo Ramos 2 , Africa Holguin 3 , Clara Restrepo 1 , Alfonso Cabello 4 , Juan Carlos López-Bernaldo 5 , Francisco Javier De La Hera 4 , Carlos Barros 6 , Manuel Fernández 4 , Vicente Estrada 7 , Miguel Górgolas Hernández-Mora 4 , José M. Benito 1 , Norma Rallón 1 1 Instituto de Investigación Sanitaria Fundación Jiménez Díaz, Madrid, Spain, 2 Parque Científico de Madrid, Madrid, Spain, 3 Instituto Ramón y Cajal de Investigación Sanitaria, Madrid, Spain, 4 Fundacion Jimenez Diaz, Madrid, Spain, 5 Hospital General Universitario Gregorio Marañón, Madrid, Spain, 6 Hospital Universitario de Móstoles, Madrid, Spain, 7 Hospital Universitario Clínico San Carlos, Madrid, Spain Background: The shock and kill strategy to purge the reservoir has failed likely as a consequence of several obstacles. The existence of escape mutations in regions of the HIV proviral sequence coding for epitopes of CD8 immune response has been postulated as one of the reasons for this failure. Herein, we have analyzed the frequency of these mutations in two different groups of HIV patients with complete viral suppression: patients on successful cART and elite controller patients. Methods: Twenty HIV patients were included: 7 elite controllers (EC) and 10 non-controller patients on successful cART (TX). CD4 resting memory cells were immunomagnetically purified and total genomic DNA was extracted. The entire Gag gene was amplified by nested PCR and a pair-end sequencing run on a MiSeq systemwas performed. Sequences were mapped and aligned to the consensus HXB2 sequence. Optimal Gag epitopes of CD8 immune response were predicted for each patient based on their HLA class I haplotype (A, B, C). The prevalence of mutated epitopes as well as its impact on HLA recognition were calculated for each patient. Results: EC and TX groups were matched for age, years of HIV diagnosis and CD4 T-cell counts. TX patients had been on cART for a median of 12[9-16] years. The whole HIV-Gag sequence was successfully amplified and sequenced in all patients. The median number of CD8 Gag epitopes predicted for EC and TX patients were 7[4-7] and 7[6-12], respectively. Of note, the prevalence (%) of mutated CD8 epitopes was 75[46-100] and 54[48-74] in EC and TX respectively (p=0,432). Moreover the frequency (%) of mutated peptides with a significant impact reducing HLA recognition was similar in both groups (50[33-50] in EC and 41[19-52] in TX, p=0,552). Conclusion: Our results show a high prevalence of mutations in HIV-Gag epitopes of CD8 T-cell response not only in the HIV reservoir of patients with successful cART-mediated control, but also in patients with spontaneous HIV

control in whom control is reached at an early stage of infection. Indeed, many of these mutations have a potential negative impact on antigen recognition. These findings support the role of existence of escape mutations as another obstacle to purge the HIV reservoir. This could provide a proof of concept challenging the current HIV cure strategies based on reservoir reactivation. 270 ASSOCIATION OF POLYFUNCTIONAL CMV-SPECIFIC T CELLS WITH FRAILTY IN HIV-INFECTED MEN Weiying Zhang 1 , Jay Bream 2 , Sean X. Leng 3 , Tricia Nilles 1 , Huifen Li 1 , Joseph B. Margolick 2 1 Johns Hopkins University, Baltimore, MD, USA, 2 Johns Hopkins Bloomberg School of Public Health, Baltimore, MD, USA, 3 Johns Hopkins University School of Medicine, Baltimore, MD, USA Background: Cytomegalovirus (CMV) infection is associated with both HIV infection and frailty. CMV-specific T cell responses correlated differently with immune activation and inflammatory markers, depending on donor HIV and frailty status; and the proportion of CD4 T-cells producing IL-2 in response to CMV predicted onset of frailty. Here, we studied T-cell production of IFN- ᵞ and TNF- ⍺ as well as IL-2 as predictors of a) onset of frailty in nonfrail men, and b) stability of frailty in frail men, in HIV+ and HIV- men who have sex with men. Methods: CMV-specific T cell responses of 42 men (22 virologically suppressed HIV+, 20 HIV−; 21 frail, 21 non-frail) were assessed by flow cytometric analysis of production of IFN- ᵞ , TNF- ⍺ and IL-2 in response to overlapping peptide pools spanning 19 CMV open reading frames. Frailty was assessed semiannually using the Fried criteria. To explore the relationship between cytokine-producing T cells and onset (in nonfrail men) and stability (in frail men) of frailty, men were categorized into tertiles of percentages of these cells. Times to onset or loss of frailty were compared by tertiles using Kaplan-Meier estimators and the nonparametric log-rank test. Results: Cytokine production by T cells fell into three main patterns: IFN- ᵞ +TNF- ⍺ +IL-2- (median: 51% vs 58% of CD4 vs CD8 cytokine-producing cells), IFN- ᵞ +TNF- ⍺ -IL-2- (15% vs 34%), and IFN- ᵞ +TNF- ⍺ +IL-2+ (11% vs 5%). IFN- ᵞ - TNF- ⍺ +IL-2+ CMV-specific T cells were detected in only one man. Percentages of these subsets of cells did not differ significantly by HIV and frailty status. Over a median follow-up of 7 years, for HIV- men onset of frailty was associated with higher percentages of IL-2+ CD4 cells also producing IFN- ᵞ and/or TNF- ⍺ , and of IFN- ᵞ +TNF- ⍺ -IL-2- CD4 T cells (p<0.001). In contrast, for HIV+men, onset of frailty was associated with lower percentages of the latter cells (p<.05). Lower percentages of these cells were associated with remaining frail for all men (p<.05 for HIV- and p=0.06 for HIV+men). Conclusion: Percentages of IFN- ᵞ , TNF- ⍺ and IL-2-producing CMV-specific T cells did not differ significantly by HIV and frailty status. However, high percentages of IFN- ᵞ +TNF- ⍺ -IL-2- CD4 T cells predicted onset of frailty in HIV- men, and low levels of these cells predicted both onset of frailty among HIV+ men, and maintenance of frailty in both HIV- and HIV+men. Thus, this T cell subset may play different roles in onset and maintenance of frailty in HIV- and HIV+men. 271 VULNERABLE TARGETS IN HIV-1 POL FOR ATTENUATION-BASED VACCINE DESIGN Doty A. Ojwach 1 , Tarylee Reddy 2 , Daniel MacMillan 3 , Vladimir Novitsky 4 , Zabrina Brumme 3 , Mark Brockman 5 , Thumbi Ndungú 1 , Jaclyn Mann 1 1 University of KwaZulu-Natal, Durban, South Africa, 2 Medical Research Council, London, UK, 3 Simon Fraser University, Burnaby, BC, Canada, 4 Harvard T.H. Chan School of Public Health, Boston, MA, USA, 5 Simon Fraser University, Vancouver, BC, Canada Background: Identification of viral immune escape mutations that compromise HIV's ability to replicate may aid rational attenuation-based vaccine design. Focussing cytotoxic T cell (CTL) responses on several epitopes where CTL escape compromises viral replication may delay escape and or attenuate the virus. We investigated immune-mediated attenuation in Pol, specifically reverse transcriptase (RT)-integrase Methods: We generated 487 recombinant viruses encoding RT-integrase from individuals with chronic (n = 406) and recent (n = 81) HIV-1 subtype C infection and measured their in vitro replication capacities (RC) using a GFP-reporter T-cell assay. A codon-by-codon analysis was performed to identify amino acids associated with altered RC and mutagenesis experiments were performed to validate the effect of these mutations on RC.

Poster Abstracts

92

CROI 2020