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bioRxiv · 10.1101/2025.01.24.634731

Tunneling CARs: Increasing CAR T tumor infiltration through the overexpression of MMP7 and SPP1

Van Pelt, S. N.·White, M.·Tat, C.·Hooper, D.·Talbot, L.·Fernandes, R.·Rooney, C.·Omer, B.
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Abstract

Chimeric antigen receptor T cell (CART) therapy has demonstrated remarkable efficacy in hematologic malignancies but has struggled to achieve comparable success in solid tumors. A key obstacle is the extracellular matrix (ECM) in solid tumors, which significantly impedes CART cell infiltration. In clinical trials, neuroblastoma (NB) has shown responsiveness to GD2-directed CART therapy, however, the failure of GD2.CARTs to effectively clear bulky disease - characterized by dense ECM - highlights the critical challenge of infiltration. In this study, we demonstrate that GD2.CARTs exhibit a unique infiltration-restriction compared to other CARTs and endogenous T cells. A separate analysis of clinical datasets identified MMP7 and SPP1 (OPN) as candidate genes to improve the infiltration of GD2.CARTs as these were upregulated in tumor-infiltrating leukocytes. MMP-7 and OPN overexpression enhanced CART extravasation (p < .001) and interstitial movement (p < .05) in ECM-dense environments in vitro. Overexpression of either OPN (p < .0001) or MMP-7 (p < .001) improved tumor infiltration in a xenograft model of NB. This resulted in improved tumor control (94% reduction in tumor burden, p < .05) and a survival extension in OPN-GD2.CART treated mice compared to unmodified GD2.CARTs (median of 148 days, p < .05). OPN overexpression did not increase off-target infiltration into healthy tissues or promote tumor metastasis, highlighting its potential for safe therapeutic application. Our study provides a framework for further exploration of gene modifications to improve CART infiltration and efficacy in solid tumors and identifies SPP1 as a candidate gene to improve GD2.CART treatment of bulky tumors.

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Van Pelt, S. N., White, M., Tat, C., Hooper, D., Talbot, L., Fernandes, R., Rooney, C., Omer, B.. 2025-01-27. Tunneling CARs: Increasing CAR T tumor infiltration through the overexpression of MMP7 and SPP1. https://doi.org/10.1101/2025.01.24.634731

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