PMID- 35101945 OWN - NLM STAT- MEDLINE DCOM- 20220317 LR - 20220317 IS - 2051-1426 (Electronic) IS - 2051-1426 (Linking) VI - 10 IP - 1 DP - 2022 Jan TI - Generation of cDC-like cells from human induced pluripotent stem cells via Notch signaling. LID - 10.1136/jitc-2021-003827 [doi] LID - e003827 AB - BACKGROUND: Dendritic cells (DCs) play critical roles in regulating the innate and adaptive immune responses, and have long been a major focus of cancer immunotherapy. Accumulating evidence suggests that conventional type 1 DCs (cDC1s) excel in cross-presentation of exogenous antigens on MHC-I molecules and induction of antitumor CD8(+) T cell immunity; however, obtaining large numbers of cDC1s is difficult. The use of reprogramming and differentiation technology is advantageous for obtaining unlimited numbers of autologous cDC1s especially for therapeutic interventions where repeated vaccinations are required. However, generation of cDC1s from human induced pluripotent stem cells (iPSCs) remains elusive. METHODS: Human iPSCs established from peripheral blood T cells and monocytes were differentiated to myeloid cells under on-feeder or feeder-free culture conditions in vitro. Phenotype, genomic and transcriptomic signature, and function of human iPSC-derived DCs were analyzed. The role of Notch signaling for the generation of HLA-DR(+) cells from human iPSCs was interrogated by a loss- and gain-of-function approach. RESULTS: Flow cytometric analyses and single-cell profiling of HLA-DR(+) cells revealed that human iPSCs gave rise to CD141(+)XCR1(+)CLEC9A(+) cells (cDC1s), CLEC4A(hi)CLEC10A(-)CD1c(+) cells (cDC2As), CLEC4A(lo)CLEC10A(+)CD1c(+) cells (cDC2Bs), CD163(-)CD5(+)CD1c(+) cells (CD5(+)cDC2s), and AXL(+)SIGLEC6(+) cells (AS-DCs) on OP9 feeder cells expressing the Notch ligand delta-like 1 (OP9-DL1) while the majority of iPSC-derived cells differentiated on OP9 cells were CD163(+)CD5(-)CD1c(+) cells (DC3s) and monocytes. Plasmacytoid DCs were not differentiated from iPSCs on either OP9 or OP9-DL1 cells. Inhibition of Notch signaling during co-culture of iPSC-derived CD34(+) hematopoietic progenitor cells with OP9-DL1 cells abrogated generation of cDC1s, cDC2As, cDC2Bs, CD5(+)cDC2s, and AS-DCs but increased frequency of DC3s. Notch-activated human iPSC-derived XCR1(+)CLEC9A(+)HLA-DR(+)CD11c(+) cells exhibited similar gene expression profile with peripheral blood cDC1s. Human iPSC-derived DCs have phagocytic, T-cell proliferative, and cytokine-producing functions. CONCLUSIONS: Our study demonstrates a critical role of Notch signaling in regulating developmental pathway of human cDCs. These findings provide insights into the future development of personalized treatment with unlimited numbers of autologous cDCs from human iPSCs. CI - (c) Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ. FAU - Makino, Kenichi AU - Makino K AD - Center for Immunotherapy, Roswell Park Comprehensive Cancer Center, Buffalo, New York, USA. AD - Department of Obstetrics and Gynecology, Akita University Graduate School of Medicine School of Medicine, Akita, Japan. FAU - Long, Mark D AU - Long MD AD - Department of Biostatistics and Bioinformatics, Roswell Park Comprehensive Cancer Center, Buffalo, New York, USA. FAU - Kajihara, Ryutaro AU - Kajihara R AD - Center for Immunotherapy, Roswell Park Comprehensive Cancer Center, Buffalo, New York, USA. FAU - Matsueda, Satoko AU - Matsueda S AD - Center for Immunotherapy, Roswell Park Comprehensive Cancer Center, Buffalo, New York, USA. FAU - Oba, Takaaki AU - Oba T AD - Center for Immunotherapy, Roswell Park Comprehensive Cancer Center, Buffalo, New York, USA. AD - Department of Surgery, Shinshu University Graduate School of Medicine School of Medicine, Matsumoto, Nagano, Japan. FAU - Kanehira, Kazunori AU - Kanehira K AD - Department of Pathology, Roswell Park Comprehensive Cancer Center, Buffalo, New York, USA. FAU - Liu, Song AU - Liu S AD - Department of Biostatistics and Bioinformatics, Roswell Park Comprehensive Cancer Center, Buffalo, New York, USA. FAU - Ito, Fumito AU - Ito F AUID- ORCID: 0000-0002-6866-671X AD - Center for Immunotherapy, Roswell Park Comprehensive Cancer Center, Buffalo, New York, USA fumito.ito@med.usc.edu. AD - Department of Immunology, Roswell Park Comprehensive Cancer Center, Buffalo, New York, USA. AD - Department of Surgical Oncology, Roswell Park Comprehensive Cancer Center, Buffalo, NY, USA. AD - Department of Surgery, University of Southern California, Los Angeles, CA, USA. LA - eng GR - K08 CA197966/CA/NCI NIH HHS/United States GR - P30 CA016056/CA/NCI NIH HHS/United States GR - R01 CA255240/CA/NCI NIH HHS/United States GR - U24 CA232979/CA/NCI NIH HHS/United States PT - Journal Article PT - Research Support, N.I.H., Extramural PT - Research Support, Non-U.S. Gov't PL - England TA - J Immunother Cancer JT - Journal for immunotherapy of cancer JID - 101620585 RN - 0 (Receptors, Notch) SB - IM MH - Animals MH - Cell Differentiation MH - Cells, Cultured MH - Dendritic Cells/*immunology MH - Humans MH - Induced Pluripotent Stem Cells/cytology/*immunology MH - Mice MH - Receptors, Notch/*immunology MH - Sequence Analysis, RNA MH - Signal Transduction MH - Single-Cell Analysis MH - Transcriptome PMC - PMC8804689 OTO - NOTNLM OT - cell engineering OT - cellular OT - dendritic cells OT - immune reconstitution OT - immunity OT - systems biology COIS- Competing interests: None declared. EDAT- 2022/02/02 06:00 MHDA- 2022/03/18 06:00 PMCR- 2022/01/31 CRDT- 2022/02/01 06:06 PHST- 2021/12/19 00:00 [accepted] PHST- 2022/02/01 06:06 [entrez] PHST- 2022/02/02 06:00 [pubmed] PHST- 2022/03/18 06:00 [medline] PHST- 2022/01/31 00:00 [pmc-release] AID - jitc-2021-003827 [pii] AID - 10.1136/jitc-2021-003827 [doi] PST - ppublish SO - J Immunother Cancer. 2022 Jan;10(1):e003827. doi: 10.1136/jitc-2021-003827.