Generation of human CAR-T cells targeting tumor-associated macrophages

• Grant number: 24/04661-6
• Support Opportunities: Scholarships in Brazil - Master
• Start date: December 01, 2024
• End date: June 30, 2025
• Field of knowledge: Health Sciences - Medicine
• Principal Investigator: Vanderson Geraldo Rocha
• Grantee: Vinicius Alex Cano Pereira
• Host Institution: Faculdade de Medicina (FM). Universidade de São Paulo (USP). São Paulo , SP, Brazil
• Associated research grant: 13/08135-2 - CTC - Center for Cell-Based Therapy, AP.CEPID

Abstract

Immunotherapy has revolutionized cancer treatment in recent years, particularly adoptive cell therapy using chimeric antigen receptor (CAR) T cells for hematological tumors. However, for solid tumors, there are several ongoing preclinical and clinical studies showing a challenging scenario due to the lack of expansion and persistence of CAR T cells in vivo, difficulty in migration within solid tumors, and the immunosuppressive tumor microenvironment. Among the cells in the tumor microenvironment, macrophages are present in large quantities in various solid tumors, serving as a poor prognostic biomarker due to their strong immunosuppressive capacity to inhibit the cytotoxic activity of T lymphocytes. Therefore, aiming for a broad therapy for various solid tumors, we propose the development of two independent second-generation CAR receptors that target and eliminate tumor-associated macrophages (TAMs) through the CD163 and CD14 target molecules. To insert the CAR into T lymphocytes from peripheral blood mononuclear cells (PBMCs) of healthy donors, we will use electroporation with 2 plasmids. The first plasmid is a PiggyBac vector containing the CAR with a CMV enhancer and the chicken beta-actin promoter. The second vector contains the transposase sequence and inserts it into the target cell DNA. In addition to confirming CAR expression by flow cytometry, we will perform in vitro functional assays such as cytotoxicity of CAR-T cells against macrophages. Macrophages will be obtained by in vitro monocyte differentiation using cytokines according to a protocol already established in the laboratory. By the end of this project, we aim to obtain two new CARs with anti-macrophage activity for future in vivo testing with models for different types of solid tumors.