Generation of human ILC3 from allogeneic and autologous CD34 + hematopoietic progenitors toward adoptive transfer

• Published in: Cytotherapy (Oxford, England) Vol. 26; no. 2; pp. 136 - 144
• Main Authors: Van der Meer, Jolien M R, Bulder, Ingrid, Kuijk, Carlijn, Kleijer, Marion, Verheij, Myrddin W, Omar, Said Z, Haverkate, Nienke J E, Dolstra, Harry, Blom, Bianca, Hazenberg, Mette D, Voermans, Carlijn
• Format: Journal Article
• Language: English
• Published: England 01.02.2024
• Subjects: hematopoietic stem and progenitor cells; stroma-free; type 3 innate lymphoid cells; adoptive transfer; interleukin-22; human
• ISSN: 1465-3249; 1477-2566
• DOI: 10.1016/j.jcyt.2023.11.011

Type 3 innate lymphoid cells (ILC3) are important in tissue homeostasis. In the gut, ILC3 repair damaged epithelium and suppress inflammation. In allogeneic hematopoietic cell transplantation (HCT), ILC3 protect against graft-versus-host disease (GvHD), most likely by restoring tissue damage and preventing inflammation. We hypothesize that supplementing HCT grafts with interleukin-22 (IL-22)-producing ILC3 may prevent acute GvHD. We therefore explored ex vivo generation of human IL-22-producing ILC3 from hematopoietic stem and progenitor cells (HSPC) obtained from adult, neonatal and fetal sources. We established a stroma-free system culturing human cord blood-derived CD34 HSPC with successive cytokine mixes for 5 weeks. We analyzed the presence of phenotypically defined ILC, their viability, proliferation and IL-22 production (after stimulation) by flow cytometry and enzyme-linked immunosorbent assay (ELISA). We found that the addition of recombinant human IL-15 and the enhancer of zeste homolog 1/2 inhibitor UNC1999 promoted ILC3 generation. Similar results were demonstrated when UNC1999 was added to CD34 HSPC derived from healthy adult granulocyte colony-stimulating factor mobilized peripheral blood and bone marrow, but not fetal liver. UNC1999 did not negatively impact IL-22 production in any of the HSPC sources. Finally, we observed that autologous HSPC mobilized from the blood of adults with hematological malignancies also developed into ILC3, albeit with a significantly lower capacity. Together, we developed a stroma-free protocol to generate large quantities of IL-22-producing ILC3 from healthy adult human HSPC that can be applied for adoptive transfer to prevent GvHD after allogeneic HCT.