Abstract B086: Mesothelin-targeted CAR Th1/17 cells are a novel cellular engineering platform for pancreatic cancer
Delaney K. Geitgey, Megan M. Wyatt, Maggie J. Phillips, Alexis F. Dixon, Erin E. Grundy, Megen C. Wittling, Carl H. June, Chrystal M. Paulos, Gregory B. LesinskiAbstract
Purpose:
Pancreatic ductal adenocarcinoma (PDAC) remains refractory to immunotherapies, including adoptive T cell therapy, necessitating evaluation of alternative T cell engineering platforms in immunosuppressive solid tumors. Human CD4+ T cells co-secreting IFN-γ and IL-17A (Th1/17 hybrids) have demonstrated enhanced antitumor activity when engineered with chimeric antigen receptors (CARs) compared with conventional CAR T cell products. However, that work was confined to immunodeficient mice and hematological malignancies, and CAR Th1/17 activity has not been evaluated in an immunocompetent model of orthotopic PDAC. We engineered a CAR targeting mesothelin (MesoCAR), a clinically relevant tumor-associated antigen in PDAC, into murine Th1/17 hybrids and hypothesized that MesoCAR Th1/17 cells would mediate antitumor activity in an immunocompetent orthotopic PDAC model.
Methods:
CD4+ T cells from murine splenocytes were Th17-polarized using our published cocktail, transduced with MesoCAR containing CD3ζ and 4-1BB co-signaling domains, and expanded with IL-12 to promote acquisition of Th1 characteristics. Cytokine production was profiled by intracellular flow cytometry and analyzed using a two-way ANOVA with Tukey’s multiple comparisons test, while impedance-based cytotoxicity assays assessed antigen-specific killing. In vivo efficacy and biodistribution studies for MesoCAR Th1/17 cells were conducted in mice bearing syngeneic mesothelin-expressing MT-5 PDAC cells orthotopically implanted into the pancreas. Survival was analyzed using Kaplan-Meier estimates and compared using the logrank (Mantel-Cox) test.
Results:
CD4+ T cells were reproducibly polarized into Th1/17 hybrids with a significantly higher frequency of double-positive IFN-γ+IL-17A+ cells (45%) than single-subset Th1 (<1%; P < 0.0001) or Th17 (11%; P = 0.0014) controls (mean values across n = 5 independent experiments). MesoCAR Th1/17 cells mediated potent antigen-specific killing of mesothelin-expressing PDAC cells in cytotoxicity assays. Following lymphodepletion, MesoCAR Th1/17 cells were well-tolerated as evidenced by stable body weight and prolonged survival compared with mice receiving lymphodepletion alone (median survival, 67 vs 48 days; logrank P = 0.0424; n = 10/group).
Conclusions:
MesoCAR Th1/17 cells maintain dual IFN-γ/IL-17A production after CAR engineering and expansion, demonstrate antigen-specific killing of PDAC cells in vitro, and prolong survival in mice bearing orthotopic PDAC tumors. To our knowledge, this is the first demonstration that CAR Th1/17 cells extend survival in an immunocompetent orthotopic PDAC model. Ongoing studies will define the contributions of endogenous immunity and MesoCAR CD8+ T cell cooperation to therapeutic efficacy, while parallel studies using human MesoCAR CD4+ T cell subsets will facilitate translation of this platform to optimize CAR T cell therapy for patients with PDAC.
Citation Format:
Delaney K. Geitgey, Megan M. Wyatt, Maggie J. Phillips, Alexis F. Dixon, Erin E. Grundy, Megen C. Wittling, Carl H. June, Chrystal M. Paulos, Gregory B. Lesinski. Mesothelin-targeted CAR Th1/17 cells are a novel cellular engineering platform for pancreatic cancer [abstract]. In: Proceedings of the AACR Conference on Pancreatic Cancer: New Frontiers in Biology and Therapeutic Development; 2026 Sep 25-28; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(18_Suppl_2):Abstract nr B086.