Comparative Analysis of Clinical-Scale IFN-γ-Positive T-Cell Enrichment Using Partially and Fully Integrated Platforms

• Published in: Frontiers in immunology 2016-09, Vol.7, p.393-393
• Main Authors: Priesner, Christoph, Esser, Ruth, Tischer, Sabine, Marburger, Michael, Aleksandrova, Krasimira, Maecker-Kolhoff, Britta, Heuft, Hans-Gert, Goudeva, Lilia, Blasczyk, Rainer, Arseniev, Lubomir, Köhl, Ulrike, Eiz-Vesper, Britta, Klöß, Stephan
• Format: Article
• Language: English
• Subjects: CliniMACS Cytokine-Capture-System; Closed GMP-compliant systems; immuneaffinity cell selection; Immunology; single platform and multicolor flow cytometry; Virus-specific T-cells
• ISSN: 1664-3224; 1664-3224
• DOI: 10.3389/fimmu.2016.00393

The infusion of enriched CMV-specific donor T-cells appears to be a suitable alternative for the treatment of drug-resistant CMV reactivation or infection after both solid organ and hematopoietic stem cell transplantation. Antiviral lymphocytes can be selected from apheresis products using the CliniMACS Cytokine-Capture-System either with the well-established CliniMACS Plus (Plus) device or with its more versatile successor CliniMACS Prodigy (Prodigy). Manufacturing of CMV-specific T-cells was carried out with the Prodigy and Plus in parallel starting with 0.8-1 × 10 leukocytes collected by lymphapheresis (  = 3) and using the MACS GMP PepTivator HCMVpp65 for antigenic restimulation. Target and non-target cells were quantified by a newly developed single-platform assessment and gating strategy using positive (CD3/CD4/CD8/CD45/IFN-γ), negative (CD14/CD19/CD56), and dead cell (7-AAD) discriminators. Both devices produced largely similar results for target cell viabilities: 37.2-52.2% (Prodigy) vs. 51.1-62.1% (Plus) CD45 /7-AAD cells. Absolute numbers of isolated target cells were 0.1-3.8 × 10 viable IFN-γ CD3 T-cells. The corresponding proportions of IFN-γ CD3 T-cells ranged between 19.2 and 95.1% among total CD3 T-cells and represented recoveries of 41.9-87.6%. Within two parallel processes, predominantly IFN-γ CD3 CD8 cytotoxic T-cells were enriched compared to one process that yielded a higher amount of IFN-γ CD3 CD4 helper T lymphocytes. T-cell purity was higher for the Prodigies products that displayed a lower content of contaminating IFN-γ T-cells (3.6-20.8%) compared to the Plus products (19.9-80.0%). The manufacturing process on the Prodigy saved both process and hands-on time due to its higher process integration and ability for unattended operation. Although the usage of both instruments yielded comparable results, the lower content of residual IFN-γ T-cells in the target fractions produced with the Prodigy may allow for a higher dosage of CMV-specific donor T-cells without increasing the risk for graft-versus-host disease.