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Single-cell transcriptional profiling informs efficient reprogramming of human somatic cells to cross-presenting dendritic cells

ABSTRACT: Type-1 conventional dendritic cells (cDC1s) are rare immune cells critical for the induction of antigen-specific cytotoxic CD8+ T-cells. The genetic program driving human cDC1 specification remains largely unexplored. We have previously identified PU.1, IRF8 and BATF3 transcription factors as sufficient to induce cDC1 fate in mouse fibroblasts but reprogramming of human somatic cells was limited by low efficiencies. Here, we investigated single-cell transcriptional dynamics during human cDC1 reprogramming. Human induced cDC1s (hiDCs) generated from embryonic fibroblasts gradually acquire a global cDC1 transcriptional profile and activate antigen presentation signatures. Importantly, other DC subsets are not induced at the single-cell level. We extracted gene modules associated with successful reprogramming and identified inflammatory signaling and the cDC1-inducing transcription factor network as key drivers of the process. Combining IFN-γ, IFN-β and TNF-α with constitutive expression of cDC1-inducing transcription factors lead to improvement of reprogramming efficiency by 190-fold. hiDCs uptake dead cells, secrete inflammatory cytokines and perform antigen cross-presentation, a key cDC1 function. This approach allowed efficient hiDC1 generation from adult fibroblasts and mesenchymal stromal cells. Mechanistically, PU.1 shows dominant and independent chromatin targeting at early phases of reprogramming, recruiting IRF8 and BATF3 to shared binding sites. The cooperative binding at open enhancers and promoters leads to silencing of fibroblast genes and activation of a cDC1 program. These findings provide mechanistic insights into human cDC1 specification and reprogramming and represent a platform for generating patient-tailored cDC1s, a long-sought DC subset for vaccination strategies in cancer immunotherapy.

ORGANISM(S): Homo sapiens

PROVIDER: GSE162650 | GEO | 2022/03/05

REPOSITORIES: GEO

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Project description:Immune evasion is an important hallmark of cancer ensured by diverse strategies, including immunosuppression and downregulation of antigen presentation. Here, to restore immunogenicity of cancer cells, we employed the minimal gene regulatory network of highly immunogenic type 1 conventional dendritic cells (cDC1) to reprogram cancer cells into professional antigen presenting cells (APCs). We showed that enforced expression of PU.1, IRF8 and BATF3 (PIB) was sufficient to induce cDC1 phenotype in 33 cell lines derived from human and mouse hematological and solid tumors. PIB gradually modified the cancer cell transcriptional and epigenetic program imposing global antigen presentation and cDC1 gene signatures within 9 days. cDC1 reprogramming restored the expression of antigen presentation complexes as well as co-stimulatory molecules at the cell surface, leading to the presentation of endogenous antigens on MHC-I, and to CD8+ T cell mediated killing. Functionally, tumor- APCs acquired the ability to uptake and process exogenous proteins and dead cells, secreted inflammatory cytokines and cross-presented antigens to naïve CD8+ T cells. Importantly, tumor-APCs were efficiently generated at the single cell level from primary cancer cells of 7 solid tumors that presented antigens to memory and naïve T-cells, as well as to activated patient-specific intra-tumoral lymphocytes. Alongside antigen presentation, tumor-APCs harboring TP53, KRAS and PTEN mutations showed impaired tumorigenicity in vitro and in vivo. Finally, using in vivo mouse models of melanoma, we showed that intra-tumoral injection of tumor-APCs promoted lymphoid infiltration, delayed tumor growth and increased survival. The anti-tumor immunity elicited by tumor-APCs was synergistic with immune checkpoint inhibitors enabling tumor eradication. Our approach combines cDC1’s antigen processing and presenting abilities with endogenous generation of tumor antigens and serves as a platform for the development of novel immunotherapies based on endowed antigen presentation in cancer cells.

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Single-cell transcriptional profiling informs efficient reprogramming of human somatic cells to cross-presenting dendritic cells

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