The impact of ex vivo clinical grade activation protocols on human T-cell phenotype and function for the generation of genetically modified cells for adoptive cell transfer therapy.
Optimized conditions for the ex vivo activation, genetic manipulation, and expansion of human lymphocytes for adoptive cell therapy may lead to protocols that maximize their in vivo function. We analyzed the effects of 4 clinical grade activation and expansion protocols over 3 weeks on cell proliferative rate, immunophenotype, cell metabolism, and transduction efficiency of human peripheral blood mononuclear cells (PBMCs). Peak lentiviral transduction efficiency was early (days 2 to 4), at a time when cells showed a larger size, maximal uptake of metabolic substrates, and the highest level of proximal T-cell receptor signaling engagement. Anti-CD2/3/28 activation beads induced greater proliferation rate and skewed PBMCs early on to a CD4 phenotype when compared with the cells cultured in OKT3. Multicolor surface phenotyping demonstrated that changes in T-cell surface markers that define T-cell functional phenotypes were dependent on the time spent in culture as opposed to the particular activation protocol. In conclusion, ex vivo activation of human PBMCs for adoptive cell therapy demonstrate defined immunophenotypic and functional signatures over time, with cells early on showing larger sizes, higher transduction efficiency, maximal metabolic activity, and zeta-chain-associated protein-70 activation.
Tumeh
Paul C
PC
Division of Hematology-Oncology, Department of Medicine, University of California Los Angeles (UCLA), Los Angeles, California, USA.
Koya
Richard C
RC
Chodon
Thinle
T
Graham
Nicholas A
NA
Graeber
Thomas G
TG
Comin-Anduix
Begoña
B
Ribas
Antoni
A
D015703:Antigens, CD / Q000096:biosynthesis / Q000276:immunology; D024221:Antineoplastic Protocols; D018929:Cell Culture Techniques; D049109:Cell Proliferation; D002469:Cell Separation; D002478:Cells, Cultured; D005434:Flow Cytometry; D015316:Genetic Therapy; D006801:Humans; D016130:Immunophenotyping; D016219:Immunotherapy, Adoptive; D016086:Lentivirus / Q000235:genetics; D008213:Lymphocyte Activation; D015398:Signal Transduction; D013601:T-Lymphocytes / Q000276:immunology / Q000378:metabolism* / Q000473:pathology / Q000821:virology; D014161:Transduction, Genetic; D051746:ZAP-70 Protein-Tyrosine Kinase / Q000378:metabolism
D015703:Antigens, CD; D051746:ZAP-70 Protein-Tyrosine Kinase; C497261:ZAP70 protein, human