Plasmacytoid dendritic cells (pDCs) produce more type I interferons (IFN-I) than any other cell type and therefore have significant therapeutic potential for the treatment of viral infections, cancer, and autoimmune diseases. Following stimulation, pDCs initially produce exceptional amounts of IFN-I but subsequently become functionally “exhausted,” a phenotype observed in both mouse and human pDCs during infection, cancer, and autoimmunity. A major barrier to studying pDC function and exhaustion is the scarcity and short lifespan of primary pDCs.
To overcome these limitations, we generated an improved human pDC cell line (4C1) that, in contrast to currently available pDC cell lines, recapitulates key hallmarks of primary pDCs, including IRF7-dependent IFN-α production. In parallel, we established a human in vitro model of pDC exhaustion using persistent stimulation with Toll-like receptor (TLR) 7 or TLR9 agonists. RNA-seq analysis comparing functional and exhausted pDCs revealed that exhaustion may be facilitated by activation of nuclear factor kappa B (NF-κB) downstream of TLR7 signaling.
To test this hypothesis, we individually deleted NF-κB subunits and identified p50 as a suppressor of IFN-α and IFN-β production in both functional and exhausted pDCs. Finally, we leveraged the 4C1 cell line for genetic screens and identified several candidate regulators of pDC function and exhaustion that were validated in primary pDCs.
These findings provide a foundation for the development of targeted pDC-based therapies to modulate IFN-I production across multiple human diseases. Moreover, the new CRISPR-editable 4C1 cell line generated here represents a valuable resource for the future identification of genetic and chemical regulators of human pDC biology.