Abstract B006: Intravesical CAR T Cell Therapy for Bladder Cancer Demonstrates Potent Antitumor Responses with Minimal Systemic Escape
Authors: Abrahimi Parwiz , Khan Jonathan F , Duren-Lubanski Alyssa , Cai Winson , Marouf Yacine , Chen Nan , Hirschhorn Daniel , Mammone Renata , Tallmann Jacob , Vela-Moreno Alejandra , Hamieh Mohamad , Faltas Bishoy , Al-Ahmadie Hikmat , Elemento Olivier , Hopkins Benjamin D , Scherr Douglas S , Brentjens Renier J , Wolchok Jedd D , Merghoub Taha 
Published: February 2025

Abstract
Intravesical therapies have been the mainstay of bladder cancer (BC) management; however, their efficacy is limited by toxicities, recurrences, and supply shortages. Consequently, many patients are recommended cystectomy, which is fraught with complications. Thus, bladder-sparing treatments present a major, unmet clinical need. Chimeric antigen receptor (CAR) T cell therapy, wherein T cells are engineered to express an artificial receptor to a target, is an immunotherapeutic approach with efficacy in hematologic malignancies. Similar success in solid tumors has been hindered by a lack of suitable targets, reduced T cell infiltration/activity, and toxicities, including on-target off-tumor effects and cytokine release syndromes. To overcome these limitations, locoregional delivery of CAR T cells is being investigated. Yet, to date, no studies have examined their potential as intravesical therapies. In this study, we used BC transcriptomics data to identify CAR targets which met criteria of high BC expression and minimal pan-tissue expression. We identified MUC16, a surface-bound glycoprotein, as a top candidate from this pipeline. Using MUC16 as a proof-of-principle target for BC-directed CAR T cell therapies, we designed second-generation CARs based on scFv derived from the anti-MUC16 antibody 3a5 (3a5-28z) and MUC16’s natural-ligand mesothelin (MSLN-28z) which were fused to CD28 co-stimulatory domains and CD3z chains of the T cell receptor. Using human BC lines expressing different MUC16 levels, we show MSLN-28z more effectively lyses MUC16+, but not MUC16-, cells than compared to 3a5-28z CAR T cells. Using gain- and loss-of-function studies in vitro, we show MUC16 is necessary and sufficient for MSLN-28z cytotoxicity and polyfunctionality, which requires signaling through CD28-CD3z endodomains. When delivered intravesically to orthotopic HT-1376 tumor bearing NSG mice, MSLN-28z CAR T cells confer superior tumor control in comparison to systemic intravenous adoptive transfers, which correlates with increased intratumoral T cell infiltrates. Despite this antitumoral effect, intravesical adoptive transfer in NSG mice results in limited systemic engraftment. To test the consequences of systemic escape after intravesical adoptive transfer, we corroborated these findings using the TRP-1 TCR autoimmune model, where intravenous adoptive transfer of TRP-1 T cells caused fur depigmentation within 30 days, whereas no mouse given the equivalent dose intravesically developed this toxicity. Similarly, using syngeneic MB49 BC cells modified to overexpress CD19, intravesical adoptive transfer of mCD19 CAR T cells significantly extends the survival of MB49-CD19 tumor-bearing C57BL/6 mice but does not result in peripheral B cell aplasia seen with intravenous transfers. Together, our findings substantiate MUC16 as a targetable antigen in BC and validate MSLN-based CAR T cells as an intravesical BCa therapy, with broad applicability to other target antigens and intravesical adoptive cell therapy platforms.
Citation Format: Parwiz Abrahimi, Jonathan F Khan, Alyssa Duren-Lubanski, Winson Cai, Yacine Marouf, Nan Chen, Daniel Hirschhorn, Renata Mammone, Jacob Tallmann, Alejandra Vela-Moreno, Mohamad Hamieh, Bishoy Faltas, Hikmat Al-Ahmadie, Olivier Elemento, Benjamin D Hopkins, Douglas S Scherr, Renier J Brentjens, Jedd D Wolchok, Taha Merghoub. Intravesical CAR T Cell Therapy for Bladder Cancer Demonstrates Potent Antitumor Responses with Minimal Systemic Escape [abstract]. In: Proceedings of the AACR IO Conference: Discovery and Innovation in Cancer Immunology: Revolutionizing Treatment through Immunotherapy; 2025 Feb 23-26; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Immunol Res 2025;13(2 Suppl):Abstract nr B006.