Notch Signaling Facilitates In Vitro Generation of Cross-Presenting Classical Dendritic Cells

The IRF8-dependent subset of classical dendritic cells (cDCs), termed cDC1, is important for cross-priming cytotoxic T cell responses against pathogens and tumors. Culture of hematopoietic progenitors with DC growth factor FLT3 ligand (FLT3L) yields very few cDC1s (in humans) or only immature “cDC1-like” cells (in the mouse). We report that OP9 stromal cells expressing the Notch ligand Delta-like 1 (OP9-DL1) optimize FLT3L-driven development of cDC1s from murine immortalized progenitors and primary bone marrow cells. Co-culture with OP9-DL1 induced IRF8-dependent cDC1s with a phenotype (CD103+ Dec205+ CD8α+) and expression profile resembling primary splenic cDC1s. OP9-DL1-induced cDC1s showed preferential migration toward CCR7 ligands in vitro and superior T cell cross-priming and antitumor vaccination in vivo. Co-culture with OP9-DL1 also greatly increased the yield of IRF8-dependent CD141+ cDC1s from human bone marrow progenitors cultured with FLT3L. Thus, Notch signaling optimizes cDC generation in vitro and yields authentic cDC1s for functional studies and translational applications. [Display omitted] •DL1-Notch2 signaling induces differentiation of murine CD8α+ CD103+ cDC1s in vitro•Notch-induced cDC1s show improved expression profile and CCR7-dependent migration•Notch-induced cDC1s mediate superior T cell cross-priming and antitumor vaccination•DL1 signaling facilitates in vitro generation of human IRF8-dependent CD141+ cDC1s Dendritic cells (DCs) are critical inducers of immune responses, but current methods to generate them in vitro are suboptimal. Kirkling et al. report that Notch signaling facilitates the generation of DCs that closely resemble their in vivo counterparts and show superior capacity to vaccinate against tumors in vivo.