PMC10348479_01
PMC10348479_02
A schematic diagram depicting the clinical-grade manufacturing process of NK cell products from CB, as well as the schedules for CB-derived NK cell treatments and immunophenotype monitoring, is shown. (A) A schematic diagram depicting the process of CB-derived NK cell-based immunotherapy, which includes donor selection, NK cell manufacturing, quality control, adoptive transfer of NK cells to the patient, and in vivo NK cell augmentation. (B) A schematic diagram depicting the clinical-grade manufacturing process of NK cell products from CB, as well as the schedules for CB-derived NK cell treatments and immunophenotype monitoring, is shown. The patient was enrolled in February 2022 and received cell treatments beginning in March 2022. Two treatment courses were administered, each consisting of four infusions given over four days, with one course in March and the other in September. Immunophenotyping of the patient was performed before and after each round of NK cell treatments.
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CB-NK cells for EC. CB-NK immunotherapy requires multiple steps, including donor selection, NK cell manufacturing, quality control, adoptive transfer of NK cells to the patient, and NK augmentation in vivo to achieve therapeutic effects due to the scarcity of NK cells in peripheral circulation ( Figure 1A ). The NK cells were activated and expanded under good manufacturing practice (GMP) conditions ( Figure 1A ). The quality and quantity of the NK cell product were evaluated, including expansion, purity, potency, and safety ( Figure 1A ). Because the initial percentages of CD56+CD3- NK cells were 11.7 and 13.2 from donors 1 and 2, respectively, and the final percentages of NK cells in the expanded populations were 93.7 and 91.6, the overall NK cell expansions were 378.8 and 257.5, respectively ( Supplementary Data Table 1  and  Figure 1A ). We have previously observed a significant upregulation in the expression of activating receptors during culture, whereas most inhibitory receptors remained unchanged, except for PD-1 and LAG-3 that were slightly elevated ( Supplementary Data Figure 1B ). CB-NK cells had higher CD107a expression, suggesting more degranulation ( Supplementary Data Figure 1C ). Moreover, CB-NK cells efficiently lysed EC cell line-Ishikawan reaching 82.4% and 86.1% at effector/target ratio 10:1, respectively ( Supplementary Data Table 1  and  Figure 1D ). The therapeutic schedule and regimen for CB-NK cell immunotherapy are illustrated in  Figure 1B . Each course consisted of intravenous transfusion once daily for four consecutive days ( Figure 1B  and  Supplementary Data Table 1 ).