Abstract 2305: Comparative evaluation of peripheral blood T cells and resultant engineered anti-CD19 CAR T cell products from relapsed/refractory non-Hodgkin's lymphoma (NHL) patients (2016)

Abstract Introduction: Administration of autologous anti-CD19 CAR T cells prepared from peripheral blood mononuclear cells (PBMC) of patients with various B cell malignancies have mediated high rates of objective response (Kochenderfer et al. Blood 2012, J Clin Onc 2014, Mackall et al, Blood 2013). We analyzed characteristics of the starting material (PBMC) and resultant CAR T cells derived from 14 patients with relapsed/refractory NHL at the NCI Surgery Branch. Methods: After apheresis collection, the T cell-containing PBMC fraction was enriched, activated with anti-CD3 antibody and cultured in serum-free medium for 2 days. Activated T cells were transduced with a retroviral vector encoding the anti-CD19 CAR gene and further expanded with IL-2 to achieve a target dose of CAR T cells. The CAR T cells and starting PBMC population were cryopreserved and analyzed after thawing by flow cytometry. Differences in cell composition between the starting PBMC population and CAR T cells were evaluated with paired t-tests, adjusted for multiplicity. Results: Biologically active autologous CAR T cells were successfully prepared from all 14 NHL subjects. Each product was comprised of both CD4+ and CD8+ T cells, but showed considerable inter-subject variability. The CD4/CD8 ratio in the PBMC and product CAR T cell population were correlated (Pearson correlation coefficient = 0.74, p = 0.0027). While the starting T cell population generally showed a T cell profile comprised of similar proportions of effector T cells (Teff) ($\bar{x}$ = 20%, std = 16%) and naïve T cells (Tn) ($\bar{x}$ = 20%, std = 14%), there was a shift towards a less differentiated T cells population after CAR T cell production, with a decreased Teff (paired t test p = 0.0151) and elevated Tn (p = 0.0031). The percent of juvenile cells (Tn+Tcm) was higher in the final product ($\bar{x}$ = 64%, std = 21%) than in the starting T cell population ($\bar{x}$ = 43%, std = 20%, p* = 0.0031, *stepdown Bonferonni). These products were active in vitro and in subjects with NHL. Clinical responses occurred regardless of product characteristic differences such as CD4/CD8 T cell ratio and juvenile / differentiated T cells. Additional analyses are ongoing, including evaluation of PD-1 expression,% of myeloid cells and gene expression profile. Conclusions: CAR T cells were successfully prepared from all subjects enrolled in the study notwithstanding the diverse nature of the subject starting lymphocyte population. The CAR T cells showed a less differentiated profile, compared to the starting lymphocytes. CAR T cell production by essentially the same process described here is currently being utilized in company-sponsored multicenter trials (NCT02348216, NCT02601313). Citation Format: Timothy Langer, Emily Lowe, Arianne Perez, Steven Rosenberg, Steven A. Feldman, Lily Lu, John M. Rossi, Edmund Chang, Marika Sherman, Marc Better, James N. Kochenderfer, Adrian Bot. Comparative evaluation of peripheral blood T cells and resultant engineered anti-CD19 CAR T cell products from relapsed/refractory non-Hodgkin's lymphoma (NHL) patients. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2305.