Evaluation of a new gene transfer strategy without cell activation for the production of CAR-T cells with higher antineoplastic potential

• Grant number: 24/04615-4
• Support Opportunities: Scholarships in Brazil - Master
• Start date: October 01, 2024
• End date: July 31, 2025
• Field of knowledge: Health Sciences - Medicine - Medical Clinics
• Principal Investigator: Lucas Eduardo Botelho de Souza
• Grantee: Millena Brandão
• Host Institution: Hemocentro de Ribeirão Preto. Hospital das Clínicas da Faculdade de Medicina de Ribeirão Preto da USP (HCMRP). Secretaria da Saúde (São Paulo - Estado). Ribeirão Preto , SP, Brazil
• Associated research grant: 20/07055-9 - Nucleus of Cellular Therapy - NuTeC, AP.NPOP

Abstract

Adoptive transfer of T lymphocytes expressing chimeric antigen receptors (CARs) has demonstrated impressive remission rates against B cell malignancies. However, many cancer patients experience refractoriness or disease relapse due to functional exhaustion of CAR-T cells. Therefore, it is desirable that the final cellular product is enriched with naïve T cells (TN), which have a greater capacity for expansion and persistence in vivo than more differentiated T cells. In the traditional manufacturing method, T cells are activated in vitro to allow transduction with lentiviruses pseudotyped with the vesicular stomatitis virus envelope (VSV-G). This process leads to the progressive differentiation of T cells and a consequent reduction in their cytotoxic activity. It has already been demonstrated that incubation with the cytokines IL-7 and IL-15 induces T cells to exit the quiescent state (G0 phase of the cell cycle) without cell activation or proliferation. Furthermore, two baboon retrovirus-based envelopes (BaEVTR and BaEVRLess) have been reported to transduce of non-activated T cells. Therefore, the hypothesis of this project is that the combination of IL-7 and IL-15 and transduction with lentiviruses pseudotyped with the BaEVTR and BaEVRLess envelopes will allow the production of CAR-T cells without the need for cell activation, resulting in a product with greater cytotoxic capacity. To test this hypothesis, non-activated T cells will be transduced with lentivirus carrying the gene encoding an anti-GD2 CAR pseudotyped with the BaEVTR, BaEVRLess and VSV-G envelopes. The transduction efficiency, frequency of TN and memory cells expressing CAR envelopes will be compared, as well as the antitumor activity in a human glioblastoma model in vitro and in vivo. We hope that the results obtained will allow the generation of more powerful CAR-T cells capable of benefiting patients currently refractory to treatment using cells produced according to the traditional manufacturing process.