Work

Antigen Presenting Cell-Mediated Ex Vivo Expansion of Human Umbilical Cord Blood Cells Yields Significant Expansion of Natural Killer Cells With Anti-Myeloma Activity

Year: 2011

Type: article

Abstract: Allogeneic natural killer (NK) cells are active in various hematologic malignancies and may have an important role in multiple myeloma (MM). Umbilical cord blood (UCB) is a potential source for allogeneic NK cells and ex vivo expanded UCB-derived NK (UCB-NK) cells demonstrate activity comparable to that of peripheral blood-derived NK cells. Here we demonstrate the ability to expand fresh and frozen UCB-NK cells to clinical grade by novel techniques using artificial antigen presenting feeder cells modified to express IL-21 (“K562-cl9-mIL21”) or IL-15 (“K562-mb-15-41BBL”). These UCB-NK cells demonstrate appropriate phenotype and are active against a variety of MM cell lines. UCB-NK cells were expanded from 1) fresh or 2) frozen cord blood units. 1) Fresh cord blood mononuclear cells (CBMCs) were cultured in 10% human serum albumin media with IL-2 (500 IU/ml) and irradiated K562-cl9-mIL21 feeder cells (2:1 feeder: CBMC ratio) for 21 days. Thereafter, cells were subjected to CD3-immunomagnetic depletion. 2) Frozen CBMC's were grown in a gas permeable culture flask with IL-2 (100 IU/ml) and irradiated K562-mb-15-41BBL feeder cells (2:1 feeder: CBMC ratio). Cells were CD3-depleted on day 7, replated with the same conditions and grown until day 14. CD3-negative cells were then used as effector cells in functional assays. Flow cytometry was used to confirm NK cell purity (C56+/CD3- cells) and a standard chromium-51 assay was performed to determine NK cell cytotoxicity. Targets included K562 cells and MM cell lines RPMI 8226, ARP-1 and U266. Expansion of fresh and frozen CBMCs yielded a > 2000 and > 200 fold expansion of NK cells, respectively, compared with only 47 fold expansion of fresh CD56-selected cells cultured with IL-2 alone. After CD3 depletion, fresh and frozen-derived UCB-NK cultures were comprised of 92% and 94% CD56+/CD3- cells respectively. APC-expanded UCB-NK cells from fresh and frozen cords demonstrated cytotoxicity against the classic NK cell target K562 as well as MM cell lines RPMI 8226, ARP-1 and U266. UCB-NK cells can be expanded ex vivo to clinically relevant doses for allogeneic NK cell therapy via co-culture with K562-cl9-mIL21 and K562-mb-15-41BBL feeder cells. Expanded UCB-NK cells are cytotoxic to K562 cells and various myeloma cell lines. Further study of fresh and frozen-derived UCB-NK cells as an adjunct therapy in stem cell transplantation for myeloma is warranted.

Source: Transplantation and Cellular Therapy

DOI: 10.1016/j.bbmt.2010.12.176