Evaluation of the cytotoxicity of nanobodybasedPSMA-specific CAR-T cells against prostate cancercells

Introduction: Prostate cancer is one of the leading causes ofcancer-related deaths worldwide. Recently, chimeric antigenreceptor (CAR) T cell therapy has been a promising treatmentstrategy for cancer patients. As of now, six products have beenapproved by the FDA. CAR-T cell therapy provides a personalizedapproach to treating hematological and solid tumors.The antigen-targeting domain of CARs is conventionally the single-chain variable fragment (scFv) of a monoclonal antibody(mAb). Recently, nanobodies are employed for this due to theirsmall size, stability, specificity, strong affinity, and easy production.CAR-Ts based on nanobodies have been shown to beas effective as those based on scFvs in preclinical and clinicalsettings. In this study, we have developed a ۲nd generation nanobody-based CAR T cell targeting prostate-specific membraneantigen (PSMA) and evaluated its efficacy against prostate cancercells.Materials and Methods: The chimeric receptor construct wascomposed of CD۳ζ and co-stimulatory molecules (CD۸ and۴-۱BB) joined with a spacer (CD۸) to an anti-PSMA nanobody.T cells were isolated from healthy donor blood and activatedwith CD۳/CD۲۸ beads in the presence of rIL-۲ (۵۰U/ml). ActivatedT cells were transduced with lentiviral vectors encodingthe anti-PSMA CAR Then, the CAR surface expression, andengineered T cell’s activation and cytotoxicity were evaluatedin vitro.Results: The surface expression of activation markers CD۶۹and CD۲۵ was increased on CAR-T cells upon coculturing withPSMA expressing LNCaP tumor cells (۲۳% and ۲۱%, respectively),compared with non-expressing DU-۱۴۵ as a negativecontrol. Also, the cell surface expression of the degranulationmarker, CD۱۰۷a, was increased to ۴۰% on engineered T cellsafter coculturing with LNCaP cells.Conclusion: We developed a nanobody-based anti-PSMACAR T cell. Nanobodies are preferred to scFvs because oftheir small size, low immunogenicity, enhanced stability, andenhanced penetration into dense solid tumors. Therefore, theseengineered PSMA-CAR T cells may be developed for adoptiveT-cell immunotherapy of prostate cancer.

کلیدواژه ها:

CAR T Cell Therapy ، Personalized Medicine ، TargetedTherapy ، Prostate Cancer ، Nanobody