AUTHOR=Štach Martin , Pytlík Robert , Šmilauerová Kristýna , Rychlá Jana , Mucha Martin , Musil Jan , Koladiya Abhishek , Nemec Matěj , Petráčková Martina , Kaštánková Iva , Pecherková Pavla , Šrámková Lucie , Polgárová Kamila , Trněný Marek , Lesný Petr , Vydra Jan , Otáhal Pavel TITLE=Characterization of the input material quality for the production of tisagenlecleucel by multiparameter flow cytometry and its relation to the clinical outcome JOURNAL=Pathology and Oncology Research VOLUME=29 YEAR=2023 URL=https://www.por-journal.com/journals/pathology-and-oncology-research/articles/10.3389/pore.2023.1610914 DOI=10.3389/pore.2023.1610914 ISSN=1532-2807 ABSTRACT=

Tisagenlecleucel (tisa-cel) is a CD19-specific CAR-T cell product approved for the treatment of relapsed/refractory (r/r) DLBCL or B-ALL. We have followed a group of patients diagnosed with childhood B-ALL (n = 5), adult B-ALL (n = 2), and DLBCL (n = 25) who were treated with tisa-cel under non-clinical trial conditions. The goal was to determine how the intensive pretreatment of patients affects the produced CAR-T cells, their in vivo expansion, and the outcome of the therapy. Multiparametric flow cytometry was used to analyze the material used for manufacturing CAR-T cells (apheresis), the CAR-T cell product itself, and blood samples obtained at three timepoints after administration. We present the analysis of memory phenotype of CD4/CD8 CAR-T lymphocytes (CD45RA, CD62L, CD27, CD28) and the expression of inhibitory receptors (PD-1, TIGIT). In addition, we show its relation to the patients’ clinical characteristics, such as tumor burden and sensitivity to prior therapies. Patients who responded to therapy had a higher percentage of CD8+CD45RA+CD27+ T cells in the apheresis, although not in the produced CAR-Ts. Patients with primary refractory aggressive B-cell lymphomas had the poorest outcomes which was characterized by undetectable CAR-T cell expansion in vivo. No clear correlation of the outcome with the immunophenotypes of CAR-Ts was observed. Our results suggest that an important parameter predicting therapy efficacy is CAR-Ts’ level of expansion in vivo but not the immunophenotype. After CAR-T cells’ administration, measurements at several timepoints accurately detect their proliferation intensity in vivo. The outcome of CAR-T cell therapy largely depends on biological characteristics of the tumors rather than on the immunophenotype of produced CAR-Ts.