Characterization of the input material quality for the production of tisagenlecleucel by multiparameter flow cytometry and its relation to the clinical outcome

Martin Štach; Robert Pytlík; Kristýna Šmilauerová; Jana Rychlá; Martin Mucha; Jan Musil; Abhishek Koladiya; Matěj Nemec; Martina Petráčková; Iva Kaštánková; Pavla Pecherková; Lucie Šrámková; Kamila Polgárová; Marek Trněný; Petr Lesný; Jan Vydra; Pavel Otáhal

doi:10.3389/pore.2023.1610914

Characterization of the input material quality for the production of tisagenlecleucel by multiparameter flow cytometry and its relation to the clinical outcome

Pathol Oncol Res. 2023 Apr 20:29:1610914. doi: 10.3389/pore.2023.1610914. eCollection 2023.

Authors

Martin Štach^{1

2}, Robert Pytlík^{1

3}, Kristýna Šmilauerová^{1

2}, Jana Rychlá¹, Martin Mucha^{1

2}, Jan Musil¹, Abhishek Koladiya¹, Matěj Nemec¹, Martina Petráčková¹, Iva Kaštánková¹, Pavla Pecherková¹, Lucie Šrámková⁴, Kamila Polgárová^{3

5}, Marek Trněný^{3

5}, Petr Lesný¹, Jan Vydra¹, Pavel Otáhal^{1

3}

Affiliations

¹ Institute of Hematology and Blood Transfusion, Praha, Czechia.
² Faculty of Science, Charles University, Praha, Czechia.
³ First Faculty of Medicine, Charles University, Praha, Czechia.
⁴ Second Faculty of Medicine (2. LF UK), University Hospital in Motol, Praha, Czechia.
⁵ General University Hospital in Prague, Praha, Czechia.

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.

Keywords: B-cell lymphoma and leukemia; CAR-T cells; Kymriah; immunotherapy; tisagenlecleucel.

MeSH terms

CD8-Positive T-Lymphocytes / metabolism
Flow Cytometry
Humans
Immunotherapy, Adoptive / methods
Lymphoma, B-Cell*
Lymphoma, Large B-Cell, Diffuse* / pathology
Receptors, Antigen, T-Cell / metabolism

Substances

tisagenlecleucel
Receptors, Antigen, T-Cell

Grants and funding

This work was supported by the Czech health research council NU22-05-00374 and by grant OPVVV CZ.02.1.01/0.0/0.0/16_025/0007428.