Tuning spacer length improves the functionality of the nanobody-based VEGFR2 CAR T cell

• Published in: BMC biotechnology Vol. 24; no. 1; p. 1
• Main Authors: Taheri, Fatemeh Hajari, Hassani, Mahmoud, Sharifzadeh, Zahra, Behdani, Mahdi, Abdoli, Shahryar, Sayadi, Mahtab, Bagherzadeh, Kowsar, Arashkia, Arash, Abolhassani, Mohsen
• Format: Journal Article
• Language: English
• Published: England BioMed Central Ltd 04.01.2024; BioMed Central; BMC
• Subjects: Antibodies; Antigens; Cancer; Cancer immunotherapy; Care and treatment; CD25 antigen; CD28 antigen; CD69 antigen; Cell Line, Tumor; Cells; Chimeric antigen receptor (CAR); Chimeric antigen receptors; Complications and side effects; Cytokines; Cytolytic activity; Cytotoxicity; Health aspects; Hinge; Humans; IgG1 CH2-CH3; Immunoglobulin G; Immunotherapy; Immunotherapy, Adoptive - methods; Intracellular; Intracellular signalling; Ligands; Lymphocytes; Lymphocytes T; Nanobodies; Patient outcomes; Receptors, Antigen, T-Cell; Solid tumors; Spacer domain; T cells; T-Lymphocytes; Tumor necrosis factor; Vascular endothelial growth factor; VEGFR2; γ-Interferon; Iran; VEGFR2; Chimeric antigen receptor (CAR); Spacer domain; Hinge; IgG1 CH2-CH3
• ISSN: 1472-6750; 1472-6750
• DOI: 10.1186/s12896-023-00827-0

The chimeric antigen receptor-expressing T (CAR-T) cells for cancer immunotherapy have obtained considerable clinical importance. CAR T cells need an optimized intracellular signaling domain to get appropriately activated and also for the proper antigen recognition, the length and composition of the extracellular spacer are critical factors. We constructed two third-generation nanobody-based VEGFR2-CARs containing either IgG1 hinge-CH2-CH3 region or hinge-only as long or short extracellular spacers, respectively. Both CARs also contained intracellular activating domains of CD28, OX40, and CD3ζ. The T cells from healthy individuals were transduced efficiently with the two CARs, and showed increased secretion of IL-2 and IFN-γ cytokines, and also CD69 and CD25 activation markers along with cytolytic activity after encountering VEGFR2 cells. The VEGFR2-CAR T cells harboring the long spacer showed higher cytokine release and CD69 and CD25 expression in addition to a more efficient cytolytic effect on VEGFR2 target cells. The results demonstrated that the third-generation anti-VEGFR2 nanobody-based CAR T cell with a long spacer had a superior function and potentially could be a better candidate for solid tumor treatment.