Immunophenotypic characteristics of CD19 chimeric antigen receptor(CAR)-transfected Nalm-6 cells

Abstract

Objective \nTo investigate the immunophenotypic characteristics of potential leukemia cells transfected with CD19 antigen receptor(CAR) during CAR-T cell preparation. \n \n \nMethods \nMorphological changes in CD19 CAR-transfected cells were observed under inverted microscope. The transfection rate and immunophenotype of transfected Nalm-6 cells were analyzed by flow cytometry. Secretion of cytokines in the culture system was detected by chemiluminescence. \n \n \nResults \nThe transfection rate of Nalm-6 cells by CD19 CAR was (46.50±3.78) % and that of KG1a cells was (15.70±1.22) %. CD19 CAR-transfected Nalm-6 cells proliferated more rapidly than Nalm-6 cells (P values on 0 d, 4 d, 7 d and 12 d were 6.339, 3.447, 0.012 and 0.009). In the culture of CD19 CAR-transfected Nalm-6 cells, cell aggregation and adhesion were observed and they gradually gathered into a group. The rate of CD19 expression was only 1.19% in the CD19 CAR-transfected Nalm-6 cell culture system with the transfection rate of (46.50±3.78) %. After increasing the proportion of Nalm-6 cells in the culture system, CD19 expression was gradually increased, while the expression of CD22 remained stable. CD19 expressed by Nalm-6 cells cultured in the supernatant of CD19 CAR-transfected Nalm-6 cell culture system was decreased gradually. The levels of IL-10 and TNF-α secreted by CD19 CAR-transfected Nalm-6 cells were higher than those by Nalm-6 cells. \n \n \nConclusions \nResults of the immunophenotypic analysis of CD19 CAR-transfected leukemia cells suggested that CD22 CAR-T cell therapy could be used as a rescue or combination therapy for CD19 CAR transfection into leukemia cells. \n \n \nKey words: \nCD19 CAR;\xa0Nalm-6 cells;\xa0Transfection rate;\xa0Immunophenotype;\xa0CD22