Abstract 3182: Profiling of B7-H3 CAR T cells generated from patients with pancreatic ductal adenocarcinoma

Abstract Background: Chimeric antigen receptor (CAR) T cells targeting B7-H3 represent a promising treatment option for pancreatic ductal adenocarcinoma (PDAC), but their application in patients with advanced-stage disease is challenging. CAR T cell fitness is key for therapeutic success, and it correlates with the quality of T cells obtained via leukapheresis. T cell quality can be affected by several factors, including prior therapies and global T cell suppression driven by high tumor burden. In this study we aim to identify immune determinants of CAR T cell fitness in PDAC. Methods: Peripheral blood mononuclear cells (PBMC) were isolated from pre-operative blood of 9 PDAC patients and analyzed by flow cytometry. CAR T were generated from patients’ PBMC by retroviral transduction with a second generation B7-H3-CAR. Results are described as median values and interquartile range. Results: Of the 9 PDAC patients, 5 were treatment naïve (TN) and 4 had previously received neoadjuvant (NA) chemotherapy. Pathologic tumor size and CA 19-9 level were higher in NA patients than in TN patients (4.7cm [6.4-3.2] vs. 3.3cm [4.5-2.3] and 464 [700-110] vs. 54 [186-13], respectively). TN patients’ PBMC presented with a higher T cell count, as compared to the NA group (CD4: 1251 cells/ul [2356-274] vs. 675 cells/ul [1175-512]; CD8: 434 cells/ul [3124-255] vs. 225 cells/ul [1090-140]). The number of circulating Treg was similar between the two groups, while the level of circulating myeloid derived suppressor cells was higher in NA patients as compared to TN patients (875 cells/ul [1317-452] vs. 285 [1147-59]). T cells isolated from TN patients demonstrated a superior proliferation ability upon either acute or chronic in vitro activation with anti-CD3 and anti-CD28 antibodies, and were characterized by a higher presence of early memory T cell subsets, as compared to NA patients (percentage of CD45RA+CD45RO-: 17% [31-10] vs. 14% [17-12] for CD4 , and 44% [72-36] vs. 31 [33-28] for CD8). B7-H3 CAR T generated from either TN or NA patients demonstrated a good transduction efficiency (80% [84-73] vs. 73% [82-70]). However, NA-B7-H3 CAR T had a reduced in vitro expansion rate than TN-B7-H3 CAR T (18-fold [30-10] vs. 15-fold [21-8]). When cocultured with the human PDAC2 cell line, TN-B7-H3 CAR T demonstrated a significantly higher antitumor activity as compared to NA-B7-H3 CAR T (percentage of PDAC cell killing: 70% [82-65] vs. 50% [63-42], p=0.028). At the end of the coculture, NA-B7-H3 CAR T displayed a more differentiated phenotype, indicated by a higher presence of effector memory CD8 T. Conclusions: Our findings support the hypothesis that PDAC patients with a greater tumor burden present with a higher degree of T cell dysfunction and that this might hamper the fitness of the related CAR T product. This study provides the basis to further investigate mediators of CAR T cell biology to advance the therapeutic potential of CAR T cell therapy with PDAC. Citation Format: Giulia Cattaneo, Liti Zhang, 1\ Marco Ventin, Shahrzad Arya, Enrica Quattrocchi, Arsen Osipov, Andrew Hendifar, Russel W. Jenkins, Xinhui Wang, Cristina R. Ferrone. Profiling of B7-H3 CAR T cells generated from patients with pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 3182.