Large-scale Isolation of Highly Pure "Untouched" Regulatory T Cells in a GMP Environment for Adoptive Cell Therapy

• Published in: Journal of immunotherapy (1997) Vol. 38; no. 6; pp. 250 - 258
• Main Authors: Haase, Doreen, Puan, Kia Joo, Starke, Mireille, Lai, Tuck Siong, Soh, Melissa Yan Ling, Karunanithi, Iyswariya, San Luis, Boris, Poh, Tuang Yeow, Yusof, Nurhashikin, Yeap, Chun Hsien, Phang, Chew Yen, Chye, Willis Soon Yuan, Chan, Marieta, Koh, Mickey Boon Chai, Goh, Yeow Tee, Bertin-Maghit, Sebastien, Nardin, Alessandra, Ho, Liam Pock, Rotzschke, Olaf
• Format: Journal Article
• Language: English
• Published: United States 01-07-2015
• Subjects: Cell Separation - methods; Cell Survival; Cells, Cultured; Clinical Trials as Topic; Forkhead Transcription Factors - metabolism; Graft vs Host Disease - etiology; Graft vs Host Disease - immunology; Graft vs Host Disease - prevention & control; Humans; Immunosuppression; Immunotherapy, Adoptive; Integrin alpha4 - metabolism; Interleukin-7 Receptor alpha Subunit - metabolism; Leukemia - complications; Leukemia - immunology; Leukemia - therapy; Stem Cell Transplantation; T-Lymphocyte Subsets - immunology; T-Lymphocytes, Regulatory - immunology; T-Lymphocytes, Regulatory - transplantation
• ISSN: 1524-9557; 1537-4513
• DOI: 10.1097/CJI.0000000000000083

Adoptive cell therapy is an emerging treatment strategy for a number of serious diseases. Regulatory T (Treg) cells represent 1 cell type of particular interest for therapy of inflammatory conditions, as they are responsible for controlling unwanted immune responses. Initial clinical trials of adoptive transfer of Treg cells in patients with graft-versus-host disease were shown to be safe. However, obtaining sufficient numbers of highly pure and functional Treg cells with minimal contamination remains a challenge. We developed a novel approach to isolate "untouched" human Treg cells from healthy donors on the basis of negative selection using the surface markers CD49d and CD127. This procedure, which uses an antibody cocktail and magnetic beads for separation in an automated system (RoboSep), was scaled up and adapted to be compatible with good manufacturing practice conditions. With this setup we performed 9 Treg isolations from large-scale leukapheresis samples in a good manufacturing practice facility. These runs yielded sufficient numbers of "untouched" Treg cells for immediate use in clinical applications. The cell preparations consisted of viable highly pure FoxP3-positive Treg cells that were functional in suppressing the proliferation of effector T cells. Contamination with CD4 effector T cells was <10%. All other cell types did not exceed 2% in the final product. Remaining isolation reagents were reduced to levels that are considered safe. Treg cells isolated with this procedure will be used in a phase I clinical trial of adoptive transfer into leukemia patients developing graft-versus-host disease after stem cell transplantation.