DiverDC - Understanding dendritic cell diversity with cell reprogramming

Overview

Project Summary

Cancer immunotherapy has changed the paradigm for cancer treatment, but the majority of patients do not respond. Non-responsiveness to checkpoint blockade could result from many causes, including failure of dendritic cell (DC) priming of CD8+ T cells. Indeed, checkpoint blockade effectiveness has recently been shown to be dependent on the conventional type I subset of dendritic cells (cDC1). Recent studies have also highlighted a critical contribution of type 2 dendritic cells (cDC2s) and plasmacytoid dendritic cells (pDCs) within the tumor microenvironment. In contrast, the regulatory program of a subset of DCs limits anti-tumor immunity. Understanding how this diversity of DCs is generated is crucial to either predict or promote response to checkpoint blockade and overall patient survival. The transcription factors (TFs) underlying the instruction of DC sub-programs remains poorly understood and hinders rational design for cancer immunotherapy. We were first to identify that enforced expression of PU.1, IRF8 and BATF3 induce cDC1-like cells from fibroblasts. An exciting opportunity for understanding DC lineage diversity emerges with direct cell reprogramming approaches. The purpose of this project is to uncover TFs codes underlying DC diversity employing a forward screening approach at the single cell level. The project will be centered on testing the hypothesis that the combinatorial action of different TFs results in multiple DC subsets mirroring specification during development and specialization in tissues. This will allow understanding the drivers of DC diversification and their combinatorial interactions. Our first aim is to screen for DC-inducing TF combinations by transducing mouse fibroblasts with barcoded TFs followed by single-cell profiling. This approach allows the combined identification of the transcriptome and instructive TFs in the same cell. Productive TF combinations for cDC2 and pDC-like cells will be validated by ability to activate DC-specific reporters, immunophenotypic and gene expression characterization at several time-points to confer their similarity with “natural” DCs. We will then assess the functional properties such as secretory pattern and antigen presentation abilities of induced DCs using both in vitro and in vivo assays. To evaluate their impact in anti-tumor immunity we will use a panel of syngeneic mouse models representing a variety of molecular and phenotypic subtypes of human melanomas that exhibit a diverse range of responses to checkpoint blockade. Finally, we will evaluate whether the identified combinations of TFs are conserved in human by testing the ability of similar TF combinations to reprogram human dermal fibroblasts, supporting the translation of newly identified DC networks.

Main Goals

To test the hypothesis that combinatorial action of different TFs results in multiple DC subsets mirroring in vivo specification during development and specialization in tissues

Project Details