{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,14]],"date-time":"2026-09-14T19:09:13Z","timestamp":1789412953753,"version":"build-2803163510"},"reference-count":44,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2026,6,2]],"date-time":"2026-06-02T00:00:00Z","timestamp":1780358400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Immunol."],"abstract":"<jats:sec>\n                    <jats:title>Background<\/jats:title>\n                    <jats:p>Gastric cancer is a leading cause of cancer-related mortality, with limited efficacy of immunotherapies due to intratumoral heterogeneity, an immunosuppressive microenvironment, and antigen-loss escape. The optimization of chimeric antigen receptor (CAR) T-cell therapy targeting PD-L1 and Claudin18.2 (CLDN18.2) is crucial for improving the antitumor response.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>\n                      We performed a stepwise optimization of CAR designs targeting PD-L1 and CLDN18.2, focusing on the costimulatory domains and intracellular signaling architecture. We engineered monospecific PD-L1 and CLDN18.2 CAR-T cells using CD28 or 4-1BB costimulatory domains and assessed their CAR expression, activation phenotype, subset composition, and cytotoxicity. We then engineered three bicistronic dual-target CARs with distinct CD3\u03b6 configurations and evaluated their functionality through\n                      <jats:italic>in vitro<\/jats:italic>\n                      assays and\n                      <jats:italic>in vivo<\/jats:italic>\n                      NUGC4-Luc gastric cancer xenograft models.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>\n                      The single-target CLDN18.2-CD28 CAR-T cells exhibited the strongest cytotoxicity\n                      <jats:italic>in vitro<\/jats:italic>\n                      . Among the bicistronic designs, the double-CD3\u03b6 CAR-T cells showed superior expansion, differentiation, and cytotoxicity compared to alternative bicistronic constructs and single-target CAR-T cells. Bulk RNA sequencing revealed enhanced immune activation and reduced exhaustion markers in the double-CD3\u03b6 CAR-T cells. In the NUGC4-Luc xenograft model, the double-CD3\u03b6 CAR-T cells achieved a tumor growth inhibition of 63.23%, outperforming the 48.06% inhibition observed with single CLDN18.2 CAR-T cells.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusions<\/jats:title>\n                    <jats:p>The study identifies a bicistronic dual-target, double-CD3\u03b6 CAR-T design that enhances T-cell functional fitness and antitumor efficacy in gastric cancer models. This approach offers a promising strategy for addressing antigen heterogeneity and improving the durability of CAR-T therapies in solid tumors.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.3389\/fimmu.2026.1805471","type":"journal-article","created":{"date-parts":[[2026,6,2]],"date-time":"2026-06-02T08:13:59Z","timestamp":1780388039000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":1,"title":["Systematic optimization of PD-L1 and CLDN18.2 CAR-T designs identifies a bicistronic dual-target, double-CD3\u03b6 architecture with enhanced antitumor activity in gastric cancer"],"prefix":"10.3389","volume":"17","author":[{"given":"Xiang","family":"Zhang","sequence":"first","affiliation":[{"name":"School of Medicine, Shanghai University","place":["Shanghai, China"]},{"name":"Shanghai Tenth People\u2019s Hospital","place":["Shanghai, China"]},{"name":"Institute of Artificial Intelligence and Biomanufacturing, School of Medicine, Shanghai University","place":["Shanghai, China"]},{"name":"State Key Laboratory of New Targets Discovery and Drug Development for Major Diseases","place":["Xi\u2019an, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yao-jie","family":"Kong","sequence":"additional","affiliation":[{"name":"School of Medicine, Shanghai University","place":["Shanghai, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jing-yao","family":"Li","sequence":"additional","affiliation":[{"name":"School of Medicine, Shanghai University","place":["Shanghai, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Heng-hui","family":"Li","sequence":"additional","affiliation":[{"name":"School of Medicine, Shanghai University","place":["Shanghai, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Liang","family":"Chen","sequence":"additional","affiliation":[{"name":"School of Medicine, Shanghai University","place":["Shanghai, China"]},{"name":"Shanghai Tenth People\u2019s Hospital","place":["Shanghai, China"]},{"name":"Institute of Artificial Intelligence and Biomanufacturing, School of Medicine, Shanghai University","place":["Shanghai, China"]},{"name":"State Key Laboratory of New Targets Discovery and Drug Development for Major Diseases","place":["Xi\u2019an, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2026,6,2]]},"reference":[{"key":"B1","doi-asserted-by":"publisher","DOI":"10.1007\/s10555-020-09925-3","article-title":"Gastric cancer: a comprehensive review of current and future treatment strategies","volume":"39","author":"Sexton","year":"2020","journal-title":"Cancer Metastasis Rev"},{"key":"B2","doi-asserted-by":"publisher","DOI":"10.1016\/j.trecan.2024.03.008","article-title":"Gastric cancer immunosuppressive microenvironment heterogeneity: implications for therapy 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