title: Accelerated production of antigen-specific T cells for preclinical and clinical applications using gas-permeable rapid expansion cultureware (G-Rex).
abstract: The clinical manufacture of antigen-specific cytotoxic T lymphocytes (CTLs) for adoptive immunotherapy is limited by the complexity and time required to produce large numbers with the desired function and specificity. The culture conditions required are rigorous, and in some cases only achieved in 2-cm wells in which cell growth is limited by gas exchange, nutrients, and waste accumulation. Bioreactors developed to overcome these issues tend to be complex, expensive, and not always conducive to CTL growth. We observed that antigen-specific CTLs undergo 7 to 10 divisions poststimulation. However, the expected CTL numbers were achieved only in the first week of culture. By recreating the culture conditions present during this first week-low frequency of antigen-specific T cells and high frequency of feeder cells-we were able to increase CTL expansion to expected levels that could be sustained for several weeks without affecting phenotype or function. However, the number of 24-well plates needed was excessive and cultures required frequent media changes, increasing complexity and manufacturing costs. Therefore, we evaluated novel gas-permeable culture devices (G-Rex) with a silicone membrane at the base allowing gas exchange to occur uninhibited by the depth of the medium above. This system effectively supports the expansion of CTL and actually increases output by up to 20-fold while decreasing the required technician time. Importantly, this amplified cell expansion is not because of more cell divisions but because of reduced cell death. This bioprocess optimization increased T-cell output while decreasing the complexity and cost of CTL manufacture, making cell therapy more accessible.
authors.affiliation: Center for Cell and Gene Therapy, Baylor College of Medicine, Houston, TX 77030, USA. jfvera@txccc.org
authors.forename: Juan F
authors.initials: JF
authors.lastname: Vera
authors.forename: Lara J
authors.initials: LJ
authors.lastname: Brenner
authors.forename: Ulrike
authors.initials: U
authors.lastname: Gerdemann
authors.forename: Minhtran C
authors.initials: MC
authors.lastname: Ngo
authors.forename: Uluhan
authors.initials: U
authors.lastname: Sili
authors.forename: Hao
authors.initials: H
authors.lastname: Liu
authors.forename: John
authors.initials: J
authors.lastname: Wilson
authors.forename: Gianpietro
authors.initials: G
authors.lastname: Dotti
authors.forename: Helen E
authors.initials: HE
authors.lastname: Heslop
authors.forename: Ann M
authors.initials: AM
authors.lastname: Leen
authors.forename: Cliona M
authors.initials: CM
authors.lastname: Rooney
mesh_terms: D000941:Antigens / Q000276:immunology*; D018929:Cell Culture Techniques / Q000295:instrumentation / Q000379:methods; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D006801:Humans; D016219:Immunotherapy, Adoptive / Q000379:methods; D020014:K562 Cells; D018655:Lymphocyte Count; D015203:Reproducibility of Results; D013602:T-Lymphocytes, Cytotoxic / Q000166:cytology* / Q000276:immunology* / Q000637:transplantation; D013997:Time Factors
chemical_list: D000941:Antigens
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