C-7. Adoptive Transfer of ZFN Mediated CCR5 Modified CD4 T-cells (SB-728-T) in HIV Subjects Leads to Long Term Engraftment of HIV Resistant T Memory Stem Cells and Decrease in Size of Latent Reservoir
Authors: Joumana Zeidan, Gary Lee, Remy Fromentin, Jacob Lalezari, Ronald Mitsuyasu, Shelley Wang, Martin A. Giedlin, Geoff Nichol, Winson Tang, Nicholas Chomont, Dale Ando, Rafick-Pierre Sekaly
Journal: Molecular Therapy (2015)

Background: Nine aviremic HIV+ subjects on ART with CD4 counts between 200-500 cells/mm3 received 10-30 billion SB-728-T cells. Sustained CD4 increases in peripheral blood (PB) were observed in all treated subjects (median +103 cells/mm3 at 1 year). CCR5-modified cells expanded and persisted (median = 1.6% in PB 3 years post infusion). Long-term persistence of CCR5-modified cells suggested their presence within long lived CD4 populations, such as central memory T-cells (TCM) and T memory stem cells (TSCM). Methods: TSCM phenotyping (CD95, CD58) was performed on cells at pre-infusion, 1 and 3 years post infusion. CCR5 modification level within CD4 subsets was determined by qPCR and deep sequencing. HIV DNA copy number was measured using digital droplet PCR, and integrated HIV DNA was measured using the Alugag PCR assay. Micro array analysis was performed to assess chronic inflammation in sorted T-cells. Results: We previously showed that TCM increased significantly post SB-728-T infusion and correlated with improved CD4 counts (r=0.87; p=0.0045). We have now identified a novel CD4 subset characterized by co-expression of low levels of CD45RA and CD45RO, as well as expression of CD27 and CCR7. This CD4 subpopulation was still detected at high levels 3 years post-infusion (19.7% of CD4 T-cells vs 8.7% pre-infusion; p=0.0156) and positively correlated with CD4 reconstitution (r=0.7904, p=0.0279). These cells expressed the memory stem cell marker CD95, and were highly enriched in CCR5-modified cells (23.2%±17.6 at 3 years), in contrast to TCM (2.4%±1.84 at 3 years). Higher frequencies of these cells post infusion correlated with decreased total HIV DNA in PBMCs (r=-0.85, p=0.016). In addition, the frequency of integrated HIV DNA at 3 year was low in TSCM as compared to other memory subsets (81 copies/1e6 TSCM vs 527-595 copies/1e6 TCM, transitional memory T-cells or effector memory T-cells). Upregulated expression of pro-inflammatory genes in T cells is a hallmark of HIV infection, even in subjects with controlled viremia. Transcriptional profiling of inflammatory pathways in T-cells showed decreased expression of interferon stimulated genes post SB-728-T infusion. Along with improvement in CD4 counts and CD4:CD8 ratios, these results provide additional mechanistic evidence for restoration global T-cell homeostasis. Conclusions: Generation of long-lived TSCM by SB-728-T is a potential mechanism by which CCR5-modified cells can persist for years post-infusion and reduce the reservoir. As TSCM have the capacity to self-renew and differentiate into other CD4 subsets, a protected CD4 TSCM population may reduce the latent reservoir by selection and/or dilution, and by providing sustained anti-HIV immune helper function to limit reservoir replenishment.