Indeed, the finding that IL-6 + sIL-6R elicits a stronger response than IL-6 alone suggests more gpl30 on the cell surface than IL-6R. In addition to activation of STAT3, a transcription factor that is of great importance in CRC establishment (18, 29), IL-6, sIL-6R, and IL-11 activate SFKs that support Hippo-independent YAP activation and thereby induce Notch receptors and ligands (17). remains an elusive goal. CRC pathogenesis is enhanced by inflammation (11). In the case of inflammatory bowel diseases, which greatly increase CRC risk, inflammation is caused by autoimmunity (12). However, even sporadic CRC, initiated by APC loss, depends on tumor-elicited inflammation, which originates from localized loss of the intestinal epithelial barrier (13). Barrier disruption results in invasion of early benign tumors (adenomas) by components of the colonic microbiota, which activate IL-23Csynthesizing myeloid cells and expand tumor-resident IL-17Cproducing T lymphocytes (13). Subsequent activation of IL-17 receptor A (IL-17RA) stimulates proliferation of early tumor progenitors and causes adenoma growth (14). Consistent with these experimental findings, epidemiological studies revealed that elevated IL-23 and IL-17 expression in low-grade human CRC predicts rapid progression to fatal metastatic disease (15). The normal function of the WntC-catenin pathway is to control the proliferation and differentiation of crypt-localized gastrointestinal epithelial stem cells (16). By activating ERK and NF-B, engagement of IL-17RA augments epithelial proliferation Bisdemethoxycurcumin and regeneration after injury (14). Other signaling pathways responsible for epithelial survival and injury repair rely on the key transcriptional regulators STAT3 and YAP (16, 17). Although the role of STAT3 in regeneration and colorectal tumorigenesis is unequivocal (18, 19), it is still debated whether YAP is a tumor suppressor (20) or an oncogenic driver (21). Here we show that Src, YAP, STAT3, and Notch are coordinately activated in mouse APC-deficient intestinal organoids and colonic tumors and in 64% of human CRC specimens. These pathways respond to the dramatic up-regulation of the IL-6 signal transducer (IL-6ST or gp130), a protein that serves as a co-receptor for IL-6, IL-11, and related cytokines. Constitutive gp130 activation in mouse intestinal epithelial cells (IEC) accelerates colorectal tumorigenesis initiated by APC loss. Conversely, inhibition of Src family Bisdemethoxycurcumin kinases (SFKs) and JAK tyrosine kinases that maintain YAP and STAT3 activation results in death of CRC progenitors and regression of established tumors. Results Concomitant Src, YAP, Bisdemethoxycurcumin Notch, and STAT3 Activation in Human CRC. SFKs, YAP, Notch, and STAT3 are critical mediators of inflammation-driven mucosal regeneration and are activated in inflammatory bowel diseases (16, 17), which increase CRC risk (12). To query their involvement in colorectal tumorigenesis, we stained a collection of human CRC surgical specimens (= 17) with antibodies to phosphorylated Src and STAT3, YAP, and HES1, a Notch target. Strikingly, 59% of the tumors exhibited concomitant activation and up-regulation of all four signaling molecules relative to nontumor tissue (Fig. 1= 27) were positive for all four markers, none of which were strongly expressed in normal tissue (Fig. S1 and and Table S2). Positivity of all four markers tended to be higher at advanced disease stages (Fig. S1= 17) and matched normal colon tissues (= 7) were stained with P-Src, YAP, P-STAT3, or HES1 antibodies. (= 3) and WT controls (= 3) were stained as above. (Scale bars, 100 m.) Open in a separate window Fig. S1. ( 0.05. (and = 27) and matched normal colon sections (= 27) were stained with P-Src, YAP, P-STAT3, or HES1 antibodies. Representative examples of one nontumor and one tumor sample are shown ( 0.05. (and Fig. S1mouse small intestinal (SI) organoids (enteroids) by transducing organoids with Adeno-Cre virus. APC-null enteroids exhibited increased STAT3 and Src Bisdemethoxycurcumin tyrosine (Y) phosphorylation (Fig. 2(Fig. 2 and organoids exhibited balloon-like morphology and up-regulation of mRNAs encoding the stem cell markers and (Fig. S2 and = 3). * 0.05. (SI organoids. (Scale bars, 100 m.) (SI organoids was analyzed by qRT-PCR for FANCB expression of the indicated mRNAs. Results are means SEM (= 3). * 0.05. ((gp130) and mRNAs and a more modest Bisdemethoxycurcumin increase in mRNA (Fig. 3enteroids led to a further increase in STAT3 phosphorylation without affecting gp130 expression (Fig. 3and mRNAs (Fig. S3and mRNAs, but lower expression of mRNA (Fig. S3organoids that express 4-hydroxytamoxifen (4-OHT)Cregulated Cre recombinase revealed strong induction of (gp130) mRNA was also up-regulated, but its induction peaked on day 4 and paralleled induction of the classic YAP targets.