Tumor-spezifische CAR-NK-Zellen für die adoptive Krebs-Immuntherapie
Zusammenfassung der Projektergebnisse
Natural killer (NK) cells are innate lymphocytes which constitute important effector cells for adoptive cancer immunotherapy. Similar to T cells, their antitumor activity can be enhanced by expression of chimeric antigen receptors (CARs) that facilitate selective recognition and killing of malignant cells. Thereby, characteristics unique to NK cells such as their endogenous natural cytotoxicity and the fact that pure populations of allogeneic NK cells do not carry the risk of causing graft-versus-host disease favor the use of donor-derived NK cells as CAR-engineered effectors. While significant progress has been made in recent years to bring such approaches to the clinics, challenges like the limited expansion potential of primary NK cells and the reduced effectiveness of CAR gene transfer still remain. Within this project, we investigated directed ex vivo differentiation of mobilized hematopoietic stem and progenitor cells (HSCs) from healthy donors as a strategy for the reliable generation of mature NK cells. We established a novel two-step protocol based on sequential culture in cytokine-containing media that for most donors consistently resulted in differentiation and selective expansion of NK cells from HSCs, confirmed by detailed phenotypic and functional analysis. Furthermore, we developed optimized feeder cells expressing membrane-anchored IL-15, IL-21 and 4-1BB ligand. Addition of such feeder cells during NK-cell differentiation and expansion further increased NK cell yield, and overcame remaining donor-to-donor variability observed upon HSC differentiation only with cytokine cocktails. Subsequently, the principles established were applied to generate CAR-NK cells from HSCs transduced with a lentiviral CAR construct targeting ErbB2 (HER2). Thereby, despite efficient transduction of HSCs, only a small proportion of the resulting NK cells expressed the construct and showed selective cytotoxicity against otherwise NK-resistant but ErbB2-positive tumor cells. Hence, as an alternative means to redirect NK cells to tumor cells a bispecific NK-cell engager was developed which simultaneously binds to the activating NK-cell receptor NKG2D and the ErbB2 antigen. In conclusion, combination of optimized cytokine-containing media with genetically modified feeder cells constitutes a reliable and effective strategy to generate highly functional NK cells from hematopoietic stem and progenitor cells ex vivo, which upon further optimization of retargeting strategies may become valuable tools for adoptive cancer immunotherapy.
Projektbezogene Publikationen (Auswahl)
- Chimeric antigen receptor-engineered NK-92 cells: An off-the-shelf cellular therapeutic for targeted eliminationof cancer cells and induction of protective anti-tumor immunity. Front. Immunol. 8: 533, 2017
Zhang C, Oberoi P, Oelsner S, Waldmann A, Lindner A, Tonn T, Wels WS
(Siehe online unter https://doi.org/10.3389/fimmu.2017.00533) - CAR-engineered NK cells for the treatment of glioblastoma: Turning innate effectors into precision tools for cancer immunotherapy. Front. Immunol. 10: 2683, 2019
Burger MC, Zhang C, Harter PN, Romanski A, Strassheimer F, Senft C, Tonn T, Steinbach JP, Wels WS
(Siehe online unter https://doi.org/10.3389/fimmu.2019.02683) - Genetically engineered CAR NK cells display selective cytotoxicity against FLT3-positive B-ALL and inhibit in vivo leukemia growth. Int. J. Cancer 145: 1935-1945, 2019
Oelsner S, Waldmann A, Billmeier A, Röder J, Lindner A, Ullrich E, Marschalek R, Dotti G, Jung G, Große-Hovest L, Oberoi P, Bader P, Wels WS
(Siehe online unter https://doi.org/10.1002/ijc.32269) - Directed differentiation of mobilized hematopoietic stem and progenitor cells into functional NK cells with enhanced antitumor activity. Cells 9: 811, 2020
Oberoi P, Kamenjarin K, Villena Ossa JF, Uherek B, Bönig H, Wels WS
(Siehe online unter https://doi.org/10.3390/cells9040811)
