Tractable In Vivo Reprogramming of Tumor Cells to Type 1 Conventional Dendritic Cell-like Cells

The efficacy of cancer immunotherapy relies on the recruitment and activation of cytotoxic T cell responses against solid tumors by type 1 conventional dendritic cells (cDC1s). However, the generation of cDC1s for cancer immunotherapy faces significant limitations, including poor cell yield, functional heterogeneity, and susceptibility to immunosuppression in the tumor microenvironment (TME). We recently developed an immunotherapy modality based on in vivo reprogramming of cancer cells into immunogenic cDC1-like cells, which enabled cancer cells to present tumor antigens as cDC1s and elicited polyclonal cytotoxic T cell responses and durable systemic anti-tumor immunity. Here, we describe a tractable protocol to generate cDC1-like cells within the TME by overexpressing the minimal cDC1-specific gene regulatory network-PU.1, IRF8, and BATF3 (collectively referred to as PIB)-in cancer cells, followed by subcutaneous implantation of a mixture of transduced and parental cells. PIB overexpression drives the gradual acquisition of the hematopoietic marker CD45 and the professional antigen presentation complex MHC class II on tumor cells, serving as cell surface readouts for in vivo cDC1 reprogramming. When compared to the reprogramming process in vitro, reprogramming of the YUMM1.7 mouse melanoma model in vivo demonstrated faster kinetics and higher efficiency. cDC1-like cells induced rapid remodeling of the TME by recruiting host immune cells within the first 3 days and leading to the formation of tertiary lymphoid structure by day 9. Reprogrammed cDC1-like cells persisted in tumors for at least 9 days but were undetected at day 15. The in vivo cDC1 reprogramming protocol described here provides a tractable and robust method to effectively transform "immune-cold" tumors into "immune-hot". Overall, it offers a powerful platform to study the mechanisms underlying cDC1-mediated anti-tumor immunity and uncover synergistic combinations with other cancer immunotherapy modalities.