Work

224 Preclinical evaluation of a claudin18.2-sensing, membrane-proximal mesothelin-targeted SNIPR CAR-T platform for pancreatic cancer and other solid tumors

Year: 2026

Type: conference-abstract

Abstract: Background Chemotherapy resistance in solid tumors has driven the development of various chimeric antigen receptor (CAR) T-cell therapies. However, clinical trials targeting the distal region of mesothelin (MSLN) on overexpressing tumor cells have yielded limited efficacy alongside notable toxicities. To overcome the neutralizing activity of shed MSLN and enhance efficacy, we developed CAR422, which recognizes the residual membrane-proximal ‘stump’ region of MSLN. To minimize off-tumor effects and enhance tumor specificity, we engineered an AND-gated synthetic intramembrane proteolysis receptor (SNIPR)-CAR system using Claudin18.2 (CLDN18.2)—a tight-junction protein aberrantly overexpressed in pancreatic cancers but restricted in normal tissues primarily to the gastric mucosa—as the sensor antigen. Upon CLDN18.2 recognition, the SNIPR receptor induces robust expression of the anti-MSLN stump CAR422 selectively within the tumor microenvironment, restricting CAR expression both spatially and temporally. Theoretically, this AND-gated CAR-T strategy enhances tumor specificity while mitigating off-tumor toxicity, thereby broadening the therapeutic window for CAR-T cells in pancreatic cancer.Methods The function and specificity of the CLDN18.2×CAR422 SNIPR system were evaluated in vitro using genetically engineered AsPC-1 pancreatic cancer cells expressing varying combinations of CLDN18.2 and MSLN. SNIPR kinetics and antigen-specific cytotoxicity were assessed in co-cultures via flow cytometry and IncuCyte live-cell killing assays. In vivo function and specificity were evaluated using subcutaneous AsPC-1 tumor models in wild-type and human MSLN knock-in NSG mice. Survival curves were statistically analyzed using the log-rank (Mantel-Cox) test.Results CLDN18.2 × CAR422 SNIPR CAR-T cells exhibited robust, antigen-dependent cytotoxic activity in vitro with minimal basal leakiness. In an in vivo pilot study utilizing human MSLN knock-in mice bearing subcutaneous CLDN18.2+/MSLN+ AsPC-1 tumors, administration of SNIPR CAR-T cells demonstrated an improved therapeutic safety profile and significantly enhanced survival compared to constitutively expressed CAR422 (P < 0.05). Dose-escalation studies for the CLDN18.2×CAR422 SNIPR CAR-T cells are currently underway in both subcutaneous pancreatic and peritoneal ovarian NSG mouse models.Conclusions Collectively, these findings demonstrate the capability of a CLDN18.2-sensing, membrane-proximal MSLN-targeted AND-gated SNIPR CAR-T platform to achieve spatially restricted CAR activation in solid tumors. This precise spatial control broadens the therapeutic window, potentially permitting expanded clinical dosing for MSLN-targeted CAR-T therapies. These results warrant further validation in larger, fully powered preclinical efficacy and safety studies.Ethics Approval All animal experiments in this study were reviewed and approved by the Institutional Animal Care and Use Committee (IACUC) of the University of Pennsylvania under approval number [804226].

Source: Journal for ImmunoTherapy of Cancer

DOI: 10.1136/jitc-2026-sitc2026.0224