AUTHOR=Bonifacius Agnes , Tischer-Zimmermann Sabine , Santamorena Maria Michela , Mausberg Philip , Schenk Josephine , Koch Stephanie , Barnstorf-Brandes Johanna , Gödecke Nina , Martens Jörg , Goudeva Lilia , Verboom Murielle , Wittig Jana , Maecker-Kolhoff Britta , Baurmann Herrad , Clark Caren , Brauns Olaf , Simon Martina , Lang Peter , Cornely Oliver A. , Hallek Michael , Blasczyk Rainer , Seiferling Dominic , Köhler Philipp , Eiz-Vesper Britta 

TITLE=Rapid Manufacturing of Highly Cytotoxic Clinical-Grade SARS-CoV-2-specific T Cell Products Covering SARS-CoV-2 and Its Variants for Adoptive T Cell Therapy

JOURNAL=Frontiers in Bioengineering and Biotechnology

VOLUME=10

YEAR=2022

URL=https://www.frontiersin.org/journals/bioengineering-and-biotechnology/articles/10.3389/fbioe.2022.867042

DOI=10.3389/fbioe.2022.867042

ISSN=2296-4185

ABSTRACT=<p><bold>Objectives:</bold> Evaluation of the feasibility of SARS-CoV-2-specific T cell manufacturing for adoptive T cell transfer in COVID-19 patients at risk to develop severe disease.</p><p><bold>Methods:</bold> Antiviral SARS-CoV-2-specific T cells were detected in blood of convalescent COVID-19 patients following stimulation with PepTivator SARS-CoV-2 Select using Interferon-gamma Enzyme-Linked Immunospot (IFN-γ ELISpot), SARS-CoV-2 T Cell Analysis Kit (Whole Blood) and Cytokine Secretion Assay (CSA) and were characterized with respect to memory phenotype, activation state and cytotoxic potential by multicolor flow cytometry, quantitative real-time PCR and multiplex analyses. Clinical-grade SARS-CoV-2-specific T cell products were generated by stimulation with MACS GMP PepTivator SARS-CoV-2 Select using CliniMACS Prodigy and CliniMACS Cytokine Capture System (IFN-gamma) (CCS). Functionality of enriched T cells was investigated in cytotoxicity assays and by multiplex analysis of secreted cytotoxic molecules upon target recognition.</p><p><bold>Results:</bold> Donor screening via IFN-γ ELISpot allows for pre-selection of potential donors for generation of SARS-CoV-2-specific T cells. Antiviral T cells reactive against PepTivator SARS-CoV-2 Select could be magnetically enriched from peripheral blood of convalescent COVID-19 patients by small-scale CSA resembling the clinical-grade CCS manufacturing process and showed an activated and cytotoxic T cell phenotype. Four clinical-grade SARS-CoV-2-specific T cell products were successfully generated with sufficient cell numbers and purities comparable to those observed in donor pretesting via CSA. The T cells in the generated products were shown to be capable to replicate, specifically recognize and kill target cells <italic>in vitro</italic> and secrete cytotoxic molecules upon target recognition. Cell viability, total CD3<sup>+</sup> cell number, proliferative capacity and cytotoxic potential remained stable throughout storage of up to 72 h after end of leukapheresis.</p><p><bold>Conclusion:</bold> Clinical-grade SARS-CoV-2-specific T cells are functional, have proliferative capacity and target-specific cytotoxic potential. Their function and phenotype remain stable for several days after enrichment. The adoptive transfer of partially matched, viable human SARS-CoV-2-specific T lymphocytes collected from convalescent individuals may provide the opportunity to support the immune system of COVID-19 patients at risk for severe disease.</p>