Abstract 6311: From bench to bedside: GMP manufacturing and testing of an LGR5-targeting CAR-T against colorectal cancer

Abstract

Abstract Colorectal cancer is the third most diagnosed cancer worldwide, and is responsible for the second largest number of cancer related deaths, warranting the need for new treatments. The successful use of chimeric antigen receptor T (CAR-T) cells against B-cell malignancies has inspired the search for CAR-T cell therapies against solid tumours such as colorectal cancer. Cancer stem cells (CSC) are a small population of cells within a tumour with the capacity to self-renew, differentiate, and initiate new tumours. CSC are resistant to chemotherapy and radiotherapy and are enriched in residual disease. Targeting CSC is a strategy to reduce the tumours’ ability to generate new cells. Leucine-rich G protein-coupled receptor 5 (LGR5) is a marker of CSC and LGR5+ cancer cells are implicated in tumour progression and metastasis in colorectal cancer. We designed and tested a chimeric antigen receptor (CAR) targeting the extracellular domain of LGR5. LGR5-targeting CAR-T cells showed significant antigen-specific cytotoxicity and IFNγ release against colorectal cancer (CRC) cell lines in vitro, and in vivo efficacy in a CRC xenograft mouse model. We confirmed the specificity and safety of the LGR5 targeting CAR-T cells by testing against normal human tissues. A GMP ready protocol for clinical scale manufacture of LGR5-targeting CAR-T cells have been developed using CD3+ T-cells from healthy donors. We have successfully tested this protocol using CRC patient-derived CD3+ T-cells and were able to generate clinically relevant cell numbers (80-fold expansion) with high CAR expression. Cells manufactured using this GMP protocol expressed markers of self-renewing naïve-like and central memory phenotype and showed very low percentage of cells that were triple-positive for PD1, TIM3 and LAG3. This protocol has been successfully tested and validated in a GMP manufacturing facility for the production of LGR5 targeting CAR-T cells, in preparation for an FDA approved first in man clinical trial against metastatic colorectal cancer, starting in 2023 (NCT05759728). Citation Format: Veronika Bandara, Batjargal Gundsambuu, Silvana Napoli, Jade Foeng, Dylan McPeake, Timona Tylis, Emma Thompson, Stuart Mills, Allison Cowin, Claudine Bonder, Timothy Sadlon, Shaun McColl, Simon Barry. From bench to bedside: GMP manufacturing and testing of an LGR5-targeting CAR-T against colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6311.