{"doi":"10.1136/jitc-2022-sitc2022.0795","title":"795 STAT3 signaling as a checkpoint for TLR9-driven epigenetic reprogramming of acute myeloid leukemia into antigen-presenting cells","abstract":"<h3>Background</h3> Cultured AML blasts can differentiate into antigen-presenting cells, however, stimulating leukemic cell differentiation and immunogenicity in patients has not translated into therapeutic effects. We previously demonstrated that eliminating STAT3 signaling in <i>Cbfb/MYH11/Mpl</i> (CMM) leukemia using siRNA or decoy DNA strategies permits TLR9-induced AML cell differentiation and thereby leads to potent adaptive immune responses and leukemia regression.<sup>1,2</sup> <h3>Methods</h3> Here, we interrogated cellular and molecular underpinnings of leukemic cells differentiation following STAT3-inhibition and TLR9-activation (STAT3i/TLR9a). <h3>Results</h3> Our whole genome and single-cell transcriptomic analyses combined with immunophenotyping revealed that the combined STAT3i/TLRa had a dramatic effect on CMM cell differentiation. The treatment induced heterogenous leukemia cell clusters at different stages of myeloid cell differentiation towards macrophage and antigen-presenting cell phenotype. These were characterized by decreasing gene signatures of proliferation/survival genes with appearance of signature of leukocyte activation, antigen presentation, and in case of more differentiated AML-derived clusters, type I and type II IFNs. The more differentiated clusters 6, 12 and 15 were characterized by markers of antiviral/antimicrobial responses with pronounced expression of type I and II IFNs. The transcriptional profiling of CMM cells isolated from mice treated using inducible <i>STAT3</i> gene silencing or decoy STAT3 in combination with TLR9 stimulation, revealed the increased expression of genes regulating myeloid cell differentiation such as <i>Irf8, Cebpa, and Cebpe</i> with decreased expression of leukemia-promoting <i>Runx1</i>. The combination treatment also increased expression of genes regulating antigen-presentation such as <i>Gadd45A, Ciita, Il12a,</i> and<i> Ifng</i> in leukemic cells. We further confirmed that IRF8 expression is required for AML cell differentiation as shown by conditional <i>Irf8</i> silencing. The AML cell reprogramming was likely regulated epigenetically as indicated by the reduced gene methylation profile of crucial myeloid cell differentiation genes such as <i>Irf8</i>. These changes were associated with reduced expression of DNMT1 and DNMT3ab, known STAT3 target genes. <h3>Conclusions</h3> Our studies suggest that eliminating STAT3 checkpoint allows for TLR9-mediated reprogramming of leukemic cells into antigen-presenting cells capable of stimulating adaptive T cell immune responses against AML. Furthermore, these findings support further development of CpG-STAT3 inhibitors as a new bi-functional agent for AML immunotherapy. <h3>Acknowledgements</h3> This project was supported by the National Cancer Institute of the National Institutes of Health under award number R01CA213131 to M.K. <h3>References</h3> Hossain DMS<i>,</i> Dos Santos C<i>,</i> Zhang Q<i>,</i> Kozlowska A<i>,</i> Liu H<i>,</i> Gao C<i>,</i> Moreira D<i>,</i> Swiderski P<i>,</i> Jozwiak A<i>,</i> Kline J<i>, et al</i>. Leukemia cell-targeted STAT3 silencing and TLR9 triggering generate systemic antitumor immunity. <i>Blood.</i> 2014;<b>123</b>:15–25. PMCID: PMC3879904 Zhang Q<i>,</i> Hossain DMS<i>,</i> Duttagupta P<i>,</i> Moreira D<i>,</i> Zhao X<i>,</i> Won H<i>,</i> Buettner R<i>,</i> Nechaev S<i>,</i> Majka M<i>,</i> Zhang B<i>, et al</i>. (2016). Serum-resistant CpG-STAT3 decoy for targeting survival and immune checkpoint signaling in acute myeloid leukemia. <i>Blood.</i> 2016;<b>127</b>:1687–1700. PMCID: PMC4817311","journal":"Regular and Young Investigator Award Abstracts","year":2022,"id":304780,"datarank":0.0,"base_score":0.0,"endowment":0.0,"self_citation_contribution":0.0,"citation_network_contribution":0.0,"self_endowment_contribution":0.0,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":1,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9513,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2022-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":998772,"name":"Priyanka Duttagupta","orcid":null,"position":1,"is_corresponding":false},{"id":421917,"name":"Yu‐Lin Su","orcid":"0000-0002-8284-5845","position":2,"is_corresponding":false},{"id":281425,"name":"Haiqing Li","orcid":"0000-0001-5583-2240","position":3,"is_corresponding":false},{"id":634407,"name":"Mingye Feng","orcid":"0000-0002-3334-9917","position":4,"is_corresponding":false},{"id":230598,"name":"Bin Zhang","orcid":"0000-0003-4703-7524","position":5,"is_corresponding":false},{"id":478530,"name":"Ya‐Huei Kuo","orcid":"0000-0003-2595-0419","position":6,"is_corresponding":false},{"id":108574,"name":"Guido Marcucci","orcid":"0000-0002-3983-5908","position":7,"is_corresponding":false},{"id":421919,"name":"Marcin Kortylewski","orcid":"0000-0002-6003-1816","position":8,"is_corresponding":false},{"id":252189,"name":"Dongfang Wang","orcid":"0000-0003-1419-7118","position":0,"is_corresponding":true}],"reference_count":2,"raw_metadata":null,"created_at":"2026-07-19T00:32:37.185846Z","pmid":null,"pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}