Myeloid STAT3 promotes formation of colitis-associated colorectal cancer in mice

Myeloid cells crosstalk with T cells and prevent anti-tumor T cell responses in a STAT3dependent manner. We investigated the role of STAT3 in myeloid stromal cells of murine autochthonous colorectal cancers (CRCs). Conditional ablation of STAT3 in myeloid cells (STAT3 ) strongly suppressed formation of Azoxymethane/Dextransulfate (AOM/DSSinduced CRCs. Tumors that succeeded to develop in STAT3 M mice displayed enhanced stromalization with altered composition of stromal immune cells. Transplantation of MC38 tumor cells into STAT3 M host mice indicated that growth of CRCs was preferentially controlled by aberrant T cell activities. Gene expression profiling of autochthonous STAT3 M CRCs demonstrated that aberrant T cell activation was maintained in the tumor stroma and not suppressed by tumor derived factors. Moreover, STAT3 M CRCs displayed profound activation of STAT3, Myc and TGFß signaling in tumor cells whereas NF B/p65 signaling was compromised. These tumor cell-autonomous molecular changes might contribute to editing and immune escape of CRCs. Therefore, targeting these pathways could improve the efficacy of immune therapies and prevent relapse of escaped cancers. Introduction Colorectal cancer (CRC) represents the third most common form of cancer in humans worldwide. Major mutations leading to CRC formation have been identified in genes for adenomatous polyposis coli (Apc), K-ras, components of the TGFß signalling pathway (Smad4 or TGFßRII) and p53. CRCs develop frequently in the context of inflammatory bowel disease demonstrating a tumor-promoting role of inflammation. Moreover, a robust infiltration of T cells, B cells, neutrophils, dentritic cells (DCs), natural killer cells (NKs), tumorassociated macrophages (TAMs) and myeloid derived suppressor cells (MDSCs) is observed in the CRC tumor stroma. Stromal immune cells support growth of CRC via production of cytokines that activate the oncogenic transcription factors NF B and STAT3. Moreover, the stroma contains immune cells with anti-tumor activities which force tumors to develop mechanisms of escape and immune suppression. One paradigm is the ability of tumors to modulate polarization of TAMs. Bacterial LPS and the Th1 cytokine IFN polarize macrophages towards the classical M1 phenotype which is responsible for innate immune attack of infectious pathogens but also neoplastic cells. Alternative activation by Th2 cytokines IL4 and IL13 polarizes macrophages towards the M2 phenotype which is characterized by low expression of M1 markers (IL12, iNos) but high expression of M2 markers (IL10, VEGF, matrix metalloproteases, IL1decoyR, Arginase-1, CCL17, CCL22) . M2 macrophages have reduced anti-tumorigenic activities but are considered to promote angiogenesis and tumor progression . Repolarization of M2 macrophages towards the M1 phenotype represents a promising option for stromal anti-tumor therapies which was recently demonstrated in a mouse model for glioblastoma. Another layer of complexity was recently discovered in breast tumors where an additional TAM population, distinguishably from M1 or M2 polarized macrophages, was identified. STAT3 phosphorylation at tyrosine residue 705 (pY-STAT3) via Janus kinases (JAKs) is induced by proand anti-inflammatory cytokines such as IL6, IL10, IL11, IL21 and IL23. The oncogenic role of STAT3 in CRC tumor cells has been demonstrated in several studies. However, STAT3 seems to have a tumor-suppressive role in advanced CRCs because Apc mice lacking STAT3 in tumor cells developed aggressive intestinal carcinomas that were not observed in Apc control animals. STAT3 promotes CRC formation via regulation of genes implicated in cell survival and cell proliferation 18 and modulates nuclear localization of ß-Catenin by post-transcriptional mechanisms . In addition, STAT3 activation represents an important immune escape mechanism as cancer cells exploit STAT3-induced expression of VEGF and IL10 to blunt maturation of stromal DCs. An immune suppressive action is further given by the STAT3 dependent repression of proinflammatory cytokines and chemokines including IFN , TNF , IL6 and IP10 . Apart from tumor-cell specific functions, STAT3 is considered to promote tumor formation when being activated in stromal cells of CRCs 17, . STAT3 activation is the result of cytokines produced by both, stromal and tumor cells. Thereby a reciprocal feedback loop between tumor cells and stromal cells is established that maintains STAT3 activation in both cellular compartments. Tumor transplantation experiments into host mice lacking STAT3 in the hematopoietic system demonstrated that STAT3 activation in immune cells prevents antitumorigenic activities of DCs, Th1 cells, NKs and neutrophils. This led to the concept of inhibiting stromal/hematopoietic STAT3 as potential strategy for cancer immunotherapy. Myeloid STAT3 is considered of particular importance for the modulation of Th1-like antitumor responses. STAT3 directly regulates the expression of the pro-tumorigenic cytokine IL23 whereas production of the Th1 cytokine IL12 by dentritic cells is suppressed via an indirect NF B-dependent mechanism. Consequently, inhibition of STAT3 in myeloid cells and B cells using CpG-STAT3 siRNA enhanced cytotoxic T cell (CTL)-mediated responses in a melanoma transplantation model. Here we demonstrate that ablation of myeloid STAT3 significantly interfered with development of autochthonous AOM/DSS-induced colorectal tumors in mice. This effect was mediated by an enhanced anti-tumor T cell response. Weighted gene correlation network analysis of STAT3 M tumor and stroma tissue uncovered sustained T cell-mediated immune surveillance in escaped tumors which was accompanied by deregulation of several inflammatory signaling pathways in tumors cells. The latter might be part of a signaling network that contributes to immune editing which implicates that targeting of these pathways could synergistically improve the efficacy of immune therapies.