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bioRxiv · 10.1101/2024.02.23.581719

Induction of expansion of ex vivo NK cells using a new feeder cell built in Brazil. An extended flow cytometry evaluation of K562.mbIL21.4BBL

Watanabe, C. M.·Ishihama Suzuki, C.·Marins dos Santos, A.·Aloia, T. P. A.·Lee, G.·Wald, D.·Okamoto, O. K.·Azevedo, J. T. C. d.·Godoy, J. A. P. d.·Santos, F. P. S.·Weinlich, R.·Kerbauy, L. N.·Kutner, J. M.·Paiva, R. d. M. A.·Hamerschlak, N.
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Abstract

BackgroundNatural killer (NK) cells are lymphocytes from the innate immune system capable of promoting an antitumor response through activation and inhibition receptors. NK cells can better mediate cell engraftment after hematopoietic stem cell transplantation (HSCT), decreasing relapse rates and preventing viral diseases after HSCT. These characteristics make NK cells eligible for application in cell therapy by increasing the frequency of NK cells in peripheral blood. Manufacturing and using feeder cells is necessary to expand NK cells while keeping their cytotoxic characteristics. AimsWe aimed to develop feeder cells from a K562 leukemic cell line capable of promoting clonal expansion of NK cells ex vivo while preserving its antitumor potential. Methods and resultsFeeder cells, named K562.clone1, were produced by transduction of mbIL-21 and 4-1BBL proteins. Next, peripheral blood NK cells were co-cultivated with K562.clone1 and expanded more than 100 folds compared to NK cells co-cultivated with K562-WT (less than 10 folds). On the first day of cultivation, the average frequency of NK cells (CD3-CD56+CD16+/-) was 5.95% {+/-} 3.92%, increasing to 83.97% {+/-} 8.19% on the fourteenth day after co-cultivation with K562.Clone1, while the percentage of NK cells raised to 75.15% {+/-} 7.57% when co-cultivated with control. In addition, NK cells expanded with the feeder K562.Clone1 were potentially cytotoxic against acute myeloid leukemia (AML) blast, tumor cell lines of leukemia and glial origin. ConclusionWe successfully built a national feeder cell, named K562.Clone1. The co-culture with K562.Clone1 feeder cell, preserving their primordial functions such as missing-self, important to distinguish health and defective cell, and natural cytotoxicity against tumoral cells, and increasing the natural cytotoxicity.

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Watanabe, C. M., Ishihama Suzuki, C., Marins dos Santos, A., Aloia, T. P. A., Lee, G., Wald, D., Okamoto, O. K., Azevedo, J. T. C. d., Godoy, J. A. P. d., Santos, F. P. S., Weinlich, R., Kerbauy, L. N., Kutner, J. M., Paiva, R. d. M. A., Hamerschlak, N.. 2024-02-27. Induction of expansion of ex vivo NK cells using a new feeder cell built in Brazil. An extended flow cytometry evaluation of K562.mbIL21.4BBL. https://doi.org/10.1101/2024.02.23.581719

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