Small molecules inhibited dysregulated STAT3-proinflammatory axis and induced antitumor responses in pancreatic and breast cancer

Signal transducer and activator of transcription 3 (STAT3) is a promising anti-cancer therapeutic target that promotes tumor development and progression. To study the STAT3-specific mechanisms underpinning tumor formation and that underlie the responses to its targeting, we used the validated mutant Kras G12D (KC) mice that under high-fat diet (HFD) has a high pancreatic adenocarcinoma (PDAC) incidence and progression, human cancer-associated fibroblasts (CAFs), PDAC and triple-negative breast cancer (TNBC) cells and xenograft models and interrogated STAT3 activity with two small molecule inhibitors, H182 and H279. Human CAFs, PDAC and TNBC cells harboring aberrantly-active STAT3 responded to growth factor and cytokine stimulation by further enhancing phospho-tyrosine-STAT3 (pYSTAT3) levels. Likewise, upon feeding KC mice the HFD, pYSTAT3 was strongly increased in cells that tracked along a path of altered phenotype, including acinar-ductal-metaplasia (ADM) elements, neoplastic ducts, stromal and immune cells, but was not present in normal acinar cells, in parallel with the development of pancreatic intraepithelial neoplasia (PanIN) lesions . To define the contexts of the pYSTAT3 and the phenotype changes, we probed the cells and mouse models with the inhibitors. The pYSTAT3 levels in the cells were suppressed early by H182 and H279 treatment. In PDAC cells, prolonged treatments with H182 severely shifted the secretome, including decreased IL-6-stimulated production of IL-8, SERPINE1, CXCL1, CXCL12, CCL2 and MIF, repressed the expression of genes involved in immune evasion, angiogenesis, cell proliferation, migration, oxidative stress and tumor progression, and upregulated tumor suppressor genes. In PDAC or TNBC cells, similar prolonged treatments with H182 or H279 also suppressed the expression of anti-apoptotic Mcl-1, Bcl-xL and c-Myc, pro-invasive matrix metalloproteinase 9, and the epithelial-mesenchymal transition proteins, SNAIL, Twist, Snug and ZEB, while inducing E-cadherin and pro-apoptotic Bax, Bad, PUMA and BIM. Treatments of PDAC and TNBC cells with the inhibitors also led to DNA damage, gammaH2AX, ATM and Chk2 induction, mitochondrial oxygen consumption rate impairment, cell cycle arrest at the G2/M phase, decreased viability, migration and invasiveness in vitro , and apoptosis. H182 treatment prevented acinar cell de-differentiation into ADM/PanINs, in parallel with decreased pro-fibroinflammatory gene expression in the pancreas, such as CXCL12, CCL2, and CCL5 in HFD-fed KC mice, and inhibited growth of PDAC subcutaneous xenografts in mice. Together, the findings support a hyperactivated STAT3-pro-fibroinflammatory cytokines and chemokines axis in diverse cell types within tumors that is critical in tumor initiation and progression. Inhibiting STAT3 functions may be a preventative and therapeutic approach against cancer.