Seung Kwon Koh1, Jeehun Park2, Seong-Eun Kim3, Yuree Lim4, Minh-Trang Thi Phan5, Jinho Kim1, Ilwoong Hwang6, Yong-Oon Ahn7, Sue Shin8,9, Junsang Doh2,10,11, Duck Cho1,4,5,12. 1. Department of Health Sciences and Technology, Samsung Advanced Institute for Health Sciences and Technology (SAIHST), Sungkyunkwan University, Seoul, Korea. 2. Research Institute of Advanced Materials (RIAM), Seoul National University, Seoul, Korea. 3. Department of Mechanical Engineering, Pohang University of Science and Technology, Pohang, Korea. 4. Department of Biopharmaceutical Convergence, Sungkyunkwan University (SKKU), Suwon, Korea. 5. Cell and Gene Therapy Institute (CGTI), Samsung Medical Center, Seoul, Korea. 6. Department of Emergency Medicine, Soonchunhyang University Gumi Hospital, Gumi, Korea. 7. Department of Pediatrics, Columbia University Irving Medical Center, New York, NY, USA. 8. Department of Laboratory Medicine, Seoul National University, Seoul, Korea. 9. Department of Laboratory Medicine, Seoul Metropolitan Government-Seoul National University (SMG-SNU) Boramae Hospital, Seoul, Korea. 10. Department of Materials Science and Engineering, Seoul National University, Seoul, Korea. 11. Institute of Engineering Research, Bio-MAX Institute, Seoul National University, Seoul, Korea. 12. Department of Laboratory Medicine and Genetics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea.
Abstract
Background: Adoptive cell therapy using umbilical cord blood (UCB)-derived allogeneic natural killer (NK) cells has shown encouraging results. However, because of the insufficient availability of NK cells and limited UCB volume, more effective culture methods are required. NK cell expansion and functionality are largely affected by the culture medium. While human serum is a major affecting component in culture media, the way it regulates NK cell functionality remains elusive. We elucidated the effects of different culture media and human serum supplementation on UCB NK cell expansion and functionality. Methods: UCB NK cells were cultured under stimulation with K562-OX40L-mbIL-18/21 feeder cells and IL-2 and IL-15 in serum-containing and serum-free culture media. The effects of the culture media and human serum supplementation on NK cell expansion and cytotoxicity were evaluated by analyzing the expansion rate, activating and inhibitory receptor levels, and the cytotoxicity of the UCB NK cells. Results: The optimal medium for NK cell expansion was Dulbecco's modified Eagle's medium/Ham's F12 with supplements and that for cytotoxicity was AIM V supplemented with Immune Cell Serum Replacement. Shifting media is an advantageous strategy for obtaining several highly functional UCB NK cells. Live cell imaging and killing time measurement revealed that human serum enhanced NK cell proliferation but delayed target recognition, resulting in reduced cytotoxicity. Conclusions: Culture medium supplementation with human serum strongly affects UCB NK cell expansion and functionality. Thus, culture media should be carefully selected to ensure both NK cell quantity and quality for adoptive cell therapy.
Background: Adoptive cell therapy using umbilical cord blood (UCB)-derived allogeneic natural killer (NK) cells has shown encouraging results. However, because of the insufficient availability of NK cells and limited UCB volume, more effective culture methods are required. NK cell expansion and functionality are largely affected by the culture medium. While human serum is a major affecting component in culture media, the way it regulates NK cell functionality remains elusive. We elucidated the effects of different culture media and human serum supplementation on UCB NK cell expansion and functionality. Methods: UCB NK cells were cultured under stimulation with K562-OX40L-mbIL-18/21 feeder cells and IL-2 and IL-15 in serum-containing and serum-free culture media. The effects of the culture media and human serum supplementation on NK cell expansion and cytotoxicity were evaluated by analyzing the expansion rate, activating and inhibitory receptor levels, and the cytotoxicity of the UCB NK cells. Results: The optimal medium for NK cell expansion was Dulbecco's modified Eagle's medium/Ham's F12 with supplements and that for cytotoxicity was AIM V supplemented with Immune Cell Serum Replacement. Shifting media is an advantageous strategy for obtaining several highly functional UCB NK cells. Live cell imaging and killing time measurement revealed that human serum enhanced NK cell proliferation but delayed target recognition, resulting in reduced cytotoxicity. Conclusions: Culture medium supplementation with human serum strongly affects UCB NK cell expansion and functionality. Thus, culture media should be carefully selected to ensure both NK cell quantity and quality for adoptive cell therapy.