Jaya Lakshmi Thangaraj1,2,3, Minh-Trang Thi Phan4, SoonHo Kweon4,5, Jinho Kim4,6, Jong-Min Lee7, Ilwoong Hwang8, Jeehun Park9, Junsang Doh10, Seung-Hwan Lee11, Manh-Cuong Vo1,2, Tan-Huy Chu1,3, Ga-Young Song1,2, Seo-Yeon Ahn1,2, Sung-Hoon Jung1,2, Hyeoung-Joon Kim2, Duck Cho12,13,14, Je-Jung Lee15,16,17. 1. Research Center for Cancer Immunotherapy, Gwangju, South Korea. 2. Department of Hematology-Oncology, Chonnam National University Hwasun Hospital, 322 Seoyangro, Hwasun, Jeollanamdo, 519-763, Republic of Korea. 3. Department of Molecular Medicine, Chonnam National University, Gwangju, South Korea. 4. Stem Cell & Regenerative Medicine Institute, Samsung Medical Center, Seoul, South Korea. 5. School of Interdisciplinary Bioscience and Bioengineering (I-Bio), POSTECH, Pohang, South Korea. 6. Department of Health Sciences and Technology, SAIHST, Sungkyunkwan University, Seoul, South Korea. 7. Bio Research Center, Lugensci Co. Ltd., Bucheon, South Korea. 8. Department of Emergency Medicine, Konkuk University Chungju Hospital, Chungju, South Korea. 9. Research Institute of Advanced Materials, Seoul National University, Seoul, South Korea. 10. Department of Materials Science and Engineering, Seoul National University, Seoul, South Korea. 11. Department of Biochemistry, Microbiology and Immunology, Faculty of Medicine, University of Ottawa, Ottawa, ON, Canada. 12. Stem Cell & Regenerative Medicine Institute, Samsung Medical Center, Seoul, South Korea. duck.cho@skku.edu. 13. Department of Health Sciences and Technology, SAIHST, Sungkyunkwan University, Seoul, South Korea. duck.cho@skku.edu. 14. Department of Laboratory Medicine and Genetics, Samsung Medical Center, Sungkyunkwan University School of Medicine, 81 Irwon-dong, Gangnam-gu, Seoul, 06351, Republic of Korea. duck.cho@skku.edu. 15. Research Center for Cancer Immunotherapy, Gwangju, South Korea. drjejung@chonnam.ac.kr. 16. Department of Hematology-Oncology, Chonnam National University Hwasun Hospital, 322 Seoyangro, Hwasun, Jeollanamdo, 519-763, Republic of Korea. drjejung@chonnam.ac.kr. 17. Department of Molecular Medicine, Chonnam National University, Gwangju, South Korea. drjejung@chonnam.ac.kr.
Abstract
BACKGROUND: Natural killer (NK) cell-based immunotherapy is a promising treatment approach for multiple myeloma (MM), but obtaining a sufficient number of activated NK cells remains challenging. Here, we report an improved method to generate ex vivo expanded NK (eNK) cells from MM patients based on genetic engineering of K562 cells to express OX40 ligand and membrane-bound (mb) IL-18 and IL-21. METHODS: K562-OX40L-mbIL-18/-21 cells were generated by transducing K562-OX40L cells with a lentiviral vector encoding mbIL-18 and mbIL-21, and these were used as feeder cells to expand NK cells from peripheral blood mononuclear cells of healthy donors (HDs) and MM patients in the presence of IL-2/IL-15. Purity, expansion rate, receptor expression, and functions of eNK cells were determined over four weeks of culture. RESULTS: NK cell expansion was enhanced by short exposure of soluble IL-18 and IL-21 with K562-OX40L cells. Co-culture of NK cells with K562-OX40L-mbIL-18/-21 cells resulted in remarkable expansion of NK cells from HDs (9,860-fold) and MM patients (4,929-fold) over the 28-day culture period. Moreover, eNK cells showed increased expression of major activation markers and enhanced cytotoxicity towards target K562, U266, and RPMI8226 cells. CONCLUSIONS: Our data suggest that genetically engineered K562 cells expressing OX40L, mbIL-18, and mbIL-21 improve the expansion of NK cells, increase activation signals, and enhance their cytolytic activity towards MM cells.
BACKGROUND: Natural killer (NK) cell-based immunotherapy is a promising treatment approach for multiple myeloma (MM), but obtaining a sufficient number of activated NK cells remains challenging. Here, we report an improved method to generate ex vivo expanded NK (eNK) cells from MM patients based on genetic engineering of K562 cells to express OX40 ligand and membrane-bound (mb) IL-18 and IL-21. METHODS: K562-OX40L-mbIL-18/-21 cells were generated by transducing K562-OX40L cells with a lentiviral vector encoding mbIL-18 and mbIL-21, and these were used as feeder cells to expand NK cells from peripheral blood mononuclear cells of healthy donors (HDs) and MM patients in the presence of IL-2/IL-15. Purity, expansion rate, receptor expression, and functions of eNK cells were determined over four weeks of culture. RESULTS: NK cell expansion was enhanced by short exposure of soluble IL-18 and IL-21 with K562-OX40L cells. Co-culture of NK cells with K562-OX40L-mbIL-18/-21 cells resulted in remarkable expansion of NK cells from HDs (9,860-fold) and MM patients (4,929-fold) over the 28-day culture period. Moreover, eNK cells showed increased expression of major activation markers and enhanced cytotoxicity towards target K562, U266, and RPMI8226 cells. CONCLUSIONS: Our data suggest that genetically engineered K562 cells expressing OX40L, mbIL-18, and mbIL-21 improve the expansion of NK cells, increase activation signals, and enhance their cytolytic activity towards MM cells.
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