Deciphering the molecular control of intratumoral dendritic cells

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PROJECT SUMMARY Dendritic cells (DC) have a major influence on tumor immunity. DC depletion abrogates tumor immunity and response to PD1 (αPD1) immunotherapy in many preclinical tumor models and DC infiltration in human tumors is a positive correlate of clinical outcome. Unfortunately, we still lack effective strategies for harnessing DC to stimulate tumor immunity and this is in large part because we do not fully understand the control of tumor DCs. To address this shortcoming, we performed scRNA-seq on human and mouse lung tumor lesions. Notably, we identified a distinct and nearly identical population of DC in both human and mouse lung tumors which upregulated genes associated with both DC maturation such as CD40 & IL12, and immunoregulation, including PD-L1 & CD200. This led us to annotate the cluster “mature DC enriched in immuno-regulatory molecules” (mregDC)(Maier et al. Nature 2020). Strikingly, mregDC were the DC carrying tumor antigen (Ag); meaning these DC are responsible for tumor Ag presentation. We hypothesize the immunostimulatory potential of tumor DC is dampened by genes upregulated in the mreg module and this thwarts induction of tumor immunity and response to αPD1. We propose that by targeting specific mreg genes we can decouple regulatory & stimulatory programs and enhance DC activation of tumor-reactive T cells and promote tumor immunity and αPD1 response. In support of our hypothesis, blocking signaling of IL4R, one of the upregulated genes in the mreg module, enhanced DC activation, expanded tumor-infiltrating T cells, and reduced tumor burden in a mouse model of NSCLC. Additionally, inhibition of Birc2/3, also upregulated in mregDC, led to substantially enhanced DC activation. To test our hypotheses and reach our objective, we will: (1) Determine the role of IL4R on induction of the mregDC state and tumor immunity. We will knockout IL4R in mouse & human DC and determine how this impacts mreg induction, Ag presentation, & tumor immunity. We will also combine anti-IL4R & anti-PD1 to assess synergy in controlling tumor growth in a preclinical model. (2) Evaluate Birc2/3 inhibition on the physiology, molecular state and immunostimulatory activity of intratumoral DC. We will test the hypothesis that pharmacological inhibition of Birc2/3 will enhance DC production of IL-12, as well co-stimulatory molecules, while facilitating cancer cell death and tumor Ag uptake, and result in robust tumor immunity. (3) Deconvolute the intrinsic regulators of DC phenotype and the mreg gene module. We will utilize a first-of-its-kind CRISPR genomics platform we developed to KO each of the 37 transcriptional related factor (TrF) genes upregulated by mregDC and determine how each impacts tumor DC activation and molecular state. The outcome of this project will provide a major advance in our understanding of intratumoral DC biology by determining the role of specific genes and pathways in dampening tumor DC functions, establish in preclinical models the therapeutic potential of compounds targeting two different pathways operating in mregDC, and identify additional molecules that could be targeted to enhance DC activity.