GENERATION AND CHARACTERIZATION OF "OFF-THE-SHELF" CHIMERIC ANTIGEN RECEPTOR ENGINEERED T CELLS (CAR-T) TO TARGET CANCER PATIENTS WITH HEMATOLOGICAL MALIGNANCIES

Abstract

Background: The usage of T cells engineered with a tumor-specific chimeric antigen receptor (CAR) for the therapeutic treatment of some types of hematological malignancies demonstrated great clinical success. However, the manufacturing of these medicinal products is commonly performed in an autologous setting, with a relatively lengthy and complex process. Therefore, not all the patients with the defined blood cancer can have access to this type of therapy. Allogenic CAR-T cells, due to their "off-the-shelf" availability can overcome some of the limitations associated with the use of the autologous approach. The principal aim of this study was to optimize the generation of "off-the-shelf" CAR-T cells utilizing the umbilical cord blood (UCB) as a source of T lymphocytes. Methods: UCBs were collected at Sidra Medicine from newborn babies at the time of birth. T lymphocytes where isolated by magnetic selection and then activated in vitro using anti-CD3/CD28 agonistic monoclonal antibodies. The activated T cells were then transduced with lentiviral vectors encoding for two types of CARs, CD19-CD28? and CD19-4-1BB? CARs. The characterization of the efficiency of transduction, the in vitro expansion, phenotype analyses and anti-tumor activity of these CD19-CAR-T cells was performed. PBMCs were utilized as source of CAR-T cells as reference of our experiments. Results: UCB-derived CD19-CAR-T cells showed an enrichment of CAR-T cells at early stage of differentiation (T stem cell memory/central memory T cells) associated with different activation markers. These cells demonstrated high in vitro expansion and exerted an efficient anti-tumor activity against target cell lines. Conclusions: Our results allowed to identify the method for the efficient isolation ex vivo of UCB-derived CD19-CAR-T cells. UCB can be considered an efficient source of T lymphocytes to generate "off the shelf" CAR-T cells endowed with anti-tumor activity and favorable phenotypic properties.