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To identify mechanisms of therapeutic resistance, we conducted longitudinal single-cell RNA and TCR sequencing of cerebrospinal fluid (CSF) lymphocytes from DMG patients receiving intravenous followed by sequential intracerebral GD2-CAR therapy, with lymphodepleting chemotherapy administered once prior to the start of CAR T cell therapy (\n                  <jats:ext-link xmlns:xlink=\"http:\/\/www.w3.org\/1999\/xlink\" ext-link-type=\"clintrialgov\" xlink:href=\"NCT04196413\">NCT04196413<\/jats:ext-link>\n                  ). CSF GD2-CAR T cells manifested limited persistence and clonal expansion, while non-engineered CSF lymphocytes underwent significant clonal expansion and repertoire stabilization, ultimately dominating the CSF immune compartment. Concurrently, peripheral blood CD4\n                  <jats:sup>+<\/jats:sup>\n                  and CD8\n                  <jats:sup>+<\/jats:sup>\n                  T cells manifested anti-CAR immune reactivity targeting epitopes enriched within murine-derived and engineered junctional regions of the CAR construct. This was associated with appearance of circulating Human Anti-CAR Antibodies (HACAs) that bound cells expressing the GD2-CAR, as well as clonal expansion of CSF B cells which produced HACA which impeded the cytotoxic activity of GD2-CAR T cells. In several cases, appearance of circulating HACA temporally correlated with disease progression and across the patient population, and levels of circulating HACA inversely correlated with circulating CAR T cell persistence. These findings reveal robust induction of systemic and CNS adaptive T cell and B cell responses to GD2-CAR T cells following intravenous then sequential intracerebroventricular GD2-CAR therapy and provide strong evidence that anti-CAR immunity is a significant contributor to therapeutic resistance in this setting.\n                <\/jats:p>","DOI":"10.64898\/2026.06.25.26356492","type":"posted-content","created":{"date-parts":[[2026,7,9]],"date-time":"2026-07-09T19:05:16Z","timestamp":1783623916000},"source":"Crossref","is-referenced-by-count":1,"title":["Anti-CAR Immunity Drives Acquired Therapeutic Resistance to GD2-CAR T Cell Therapy in Diffuse Midline Glioma"],"prefix":"10.64898","author":[{"given":"Yiyun","family":"Chen","sequence":"first","affiliation":[{"name":"Stanford Center for Cancer Cell Therapy, Stanford Cancer Institute, Stanford University, Stanford, CA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kevin","family":"Reynolds","sequence":"additional","affiliation":[{"name":"Stanford Center for Cancer Cell Therapy, Stanford Cancer Institute, Stanford University, Stanford, CA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Maximilian R. 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