CROI 2018 Abstract eBook

Abstract eBook

Poster Abstracts

and CD32a– cells, and placed in a quantitative viral outgrowth assay. Inducible, replication-competent latent HIV genomes were assayed at days 7, 14 and 21 using standard p24 ELISA (LOD=6.25 pg/mL) and Quanterix ultrasensitive p24 Simoa (LOD=0.003 pg/mL). Results: The mean age of participants was 16 years (range 15-23) and average duration of virologic suppression was 7.85 years (range 2.45-19). The proportion of CD32a+/CD4+ T cells ranged from 0.04%–0.31% (median=0.055, IQR=0.074), higher than reported in adults. Replication-competent virus was readily detected by standard ELISA in 50% of 4 participants with 7.2%, 14.3% and 21.4% of one million cell wells (mean wells cultured=7) in the CD32a–/ CD4+ T cells, at days 7, 14 and 21 respectively, but not in the CD32a+ fraction (mean cells cultured=19,880). Using Simoa, inducible latent HIV was identified in CD32a–/CD4+ T cells in 25% and 32.1% of one million cell wells at day 7 and 14, respectively; p24 was detected in CD32a+/CD4+ T cells in the same two participants. The concentration of p24 antigen was significantly higher in CD32a– cells than in CD32a+/CD4+ T cells at day 14 (p=0.04) and there was a significant increase in replication over time in CD32a–/CD4+ T cells only (p=0.03)(Figure 1). Conclusion: Despite high concentrations of CD32a+/CD4+ T cells in perinatal infection, the replication-competent CD4+ T cell reservoir resides predominantly in CD32a–/CD4+ T cells. However, while CD32a–/CD4+ T cells harbor a larger replication-competent proviral pool, CD32a+/CD4+ T cells are enriched 125-2,713-fold for cells producing virus with minimal replication- competence. Whether this inducible reservoir is capable of producing infectious virus under different stimulation conditions is unknown. The significance in perinatal infection and the implication for cure strategies requires further study.

were evaluated in rCD4 cells from both HIV-1-positive and negative donors. In parallel, flow cytometric evaluation of T cell activation markers was carried out to assess for co-expression of CD25, CD69 or HLA-DR with CD32. Based on strong correlations between HLA-DR and CD32 expression, total CD4 cells from aviremic HIV-1-positive participants (n=3) underwent flow sorting into populations expressing either HLA-DR, CD32, both markers or neither to evaluate for proviral enrichment. Viral reactivation was quantified in sorted populations. Results: rCD4 cells from aviremic HIV-1-positive donors expressing CD32 did not contain significantly more proviral DNA than the CD32-negative population or total rCD4 cells (repeated measures one-way ANOVA, P=0.48). CD32 expression increased significantly with T cell activation and de novo HIV-1 infection in vitro. CD32 expression did correlate with expression of the activation marker HLA-DR (R2=0.99, P<0.001), but not with CD69 or CD25. Viral reactivation did not differ between CD32+, CD32- and total CD4 sorted cell populations (P=0.49). Conclusion: CD32 expression on CD4+ T cells from aviremic HIV-1-positive participants was associated with the activation marker HLA-DR but did not identify a population of cells enriched for replication-competent proviral DNA. 390 PRODUCTIVE HIV-1 INFECTION UPREGULATES CD32 IN VITRO AND IN VIVO Carla Serra Peinado 1 , Judith Grau-Expósito 1 , Meritxell Genescà 1 , Laura Luque- Ballesteros 1 , Cristina Gálvez 2 , Josep Castellví 3 , Rein Willekens 1 , Luis Montaner 4 , Vicenç Falcó 3 , Javier Martinez-Picado 2 , María J. Buzón 1 1 Vall d’Hebron Research Institute, Barcelona, Spain, 2 IrsiCaixa Institute for AIDS Research, Badalona, Spain, 3 Hospital Universitario de la Vall d’Hebron, Barcelona, Spain, 4 Wistar Institute, Philadelphia, PA, USA Background: The cell surface receptor protein CD32a has recently been postulated as a marker of a CD4+ T-cell HIV reservoir harboring replication- competent proviruses in ART-suppressed subjects. The expression of CD32 in different CD4+ T-cell subsets in HIV-infected patients and expression in persistent infected cells retained within lymphoid tissue after ART are important questions that still need to be evaluated. Methods: The expression of CD32 in different CD4+ T-cell subpopulations was measured in 9 ART-treated patients by fluorescence activated cell sorting (FACS) immunophenotyping using antibodies that recognized CD3, CD4, CD27, CD45RO, CD32 and HLA-DR. HIV and CD32a RNA expression in tissue was measured by fluorescent in situ hybridization (FISH) in lymph node sections from 2 aviremic and 4 viremic HIV-infected patients. Ex vivo HIV-1 infection kinetics was measured by the RNA/flow technique during 6 days after the initial infection of healthy donor unstimulated PBMCs, and in a cervico-vaginal tissue explant model of HIV infection. Wilcoxon signed rank and spearman’s rank correlation tests were used for statistical analysis. Results: Terminally-differentiated CD4+ T cells expressed significantly higher proportions of CD32 receptor compared to other CD4+ T subpopulations (p<0.05 for all comparisons). The majority of CD4+CD32+ cells had a naïve phenotype (CD45RO–CD27+) followed by central memory (CD45RO+CD27+) and terminally-differentiated phenotype (CD45RO–CD27–). In vivo, the vast majority of productively infected cells in lymph node tissues from HIV aviremic and viremic patients also co-expressed the CD32a marker (90% of all infected cells), while most of CD32a single positive cells were absent from the B cell follicle, one of the major reservoirs for HIV. HIV infection of unstimulated PBMCs and cervico-vaginal histocultures upregulated the expression of CD32 in approximately 10-20% of all infected cells. CD32+ infected cells expressed more frequently the activation marker HLA-DR compared to CD32- infected cells (92% vs 77%) and the immune check-point PD-1 (46% vs 25%). Conclusion: CD32 is preferentially detected in HIV transcriptionally active cells in tissues after ART and was identified to be co-expressed with activation markers in in vitro infected cells suggesting its expression is also associate with active HIV-infected cells 391 HIV-1 BURDEN IN PERIPHERAL BLOOD AND GUT CD4+ T CELLS EXPRESSING CD30 AND CD32 Louise Hogan 1 , Joshua Vasquez 1 , Kristen Hobbs 1 , Emily Hanhauser 1 , Brandon Aguilar-Rodriguez 1 , Cassandra Thanh 1 , Erica Gibson 1 , Alexander Carvidi 1 , Louis Smith 1 , Ma Somsouk 1 , Daniel R. Kuritzkes 2 , Steven G. Deeks 1 , Timothy J. Henrich 1 1 University of California San Francisco, San Francisco, CA, USA, 2 Harvard University, Cambridge, MA, USA

Poster Abstracts

389 CD4+ T CELLS EXPRESSING CD32 FROM HIV-1+ PATIENTS ARE NOT ENRICHED FOR PROVIRAL DNA AdamM. Spivak 1 , Racheal Nell 1 , McKenna L. Coletti 1 , Luis Montaner 2 , Vicente Planelles 1 1 University of Utah, Salt Lake City, UT, USA, 2 Wistar Institute, Philadelphia, PA, USA Background: A recent publication described enrichment of the latent proviral reservoir in resting CD4+ T cells expressing the low-affinity Fc receptor CD32. This finding has not yet been reproduced. Identification of a cell surface marker that distinguishes the minority of cells harboring latent HIV-1 provirus from the larger pool of circulating, uninfected CD4+ T cells would represent a crucial scientific advance with potential therapeutic implications for HIV-1 eradication. Using cells obtained from aviremic HIV-1-positive participants, we hypothesized that CD32 expression would identify a CD4+ T cell population enriched for proviral DNA. Methods: Resting CD4+ T (rCD4) cells were purified via negative magnetic bead isolation from peripheral blood mononuclear cells obtained from aviremic HIV-1-positive study participants (n=6). CD32-positive rCD4 cells were purified via magnetic bead isolation, and quantitative proviral DNA PCR was carried out on CD32-negative, CD32-positive, and total rCD4 cell populations. Changes in CD32 expression upon T cell activation, de novo or reactivated HIV-1 infection

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