Two novel bioisosteres of cabozantinib, 3 and 4, were designed and synthesized. The benzene ring in the center of the cabozantinib structure was replaced by trimethylpyridine (3) and pyridine (4), respectively. Surprisingly, the two compounds showed extremely contrasting mesenchymal–epithelial transition factor (c-Met) inhibitory activities at 1 μM concentration (4% inhibition of 3 vs. 94% inhibition of 4). The IC₅₀ value of compound 4 was 4.9 nM, similar to that of cabozantinib (5.4 nM). A ligand-based docking study suggested that 4 includes the preferred conformation for the binding to c-Met in the conformational ensemble, but 3 does not. The anti-proliferative activity of compound 4 against hepatocellular carcinoma (Hep3B and Huh7) and non-small-cell lung cancer (A549 and H1299) cell lines was better than that of cabozantinib, whereas 3 did not show a significant anti-proliferative activity. Moreover, the tumor selectivity of compound 4 toward hepatocellular carcinoma cell lines was higher than that of cabozantinib. In the xenograft chick tumor model, compound 4 inhibited Hep3B tumor growth to a much greater extent than cabozantinib. The present study suggests that compound 4 may be a good therapeutic candidate against hepatocellular carcinoma.

In the present study, we investigated the regulatory mechanisms underlying overexpression of EZH2, tryptophan hydroxylase 1 (TPH1), and 5-HT₇, in relation to gemcitabine resistance and CSC survival in PDAC cells. In aggressive PANC-1 and MIA PaCa-2 cells, knock-down (KD) of EZH2, TPH1, or HTR7 induced a decrease in CSCs and recovery from gemcitabine resistance, while preconditioning of less aggressive Capan-1 cells with 5-HT induced gemcitabine resistance with increased expression of EZH2, TPH1, and 5-HT₇. Such effects of the gene KD and 5-HT treatment were mediated through PI3K/Akt and JAK2/STAT3 signaling pathways. EZH2 KD or GSK-126 (an EZH2 inhibitor) inhibited activities of these signaling pathways which altered nuclear level of NF-kB, Sp1, and p-STAT3, accompanied by downregulation of TPH1 and 5-HT₇. Co-immunoprecipation with EZH2 and pan-methyl lysine antibodies revealed that auto-methylated EZH2 served as a scaffold for binding with methylated NF-kB and Sp1 as well as unmethylated p-STAT3. Furthermore, the inhibitor of EZH2, TPH1, or 5-HT₇ effectively regressed pancreatic tumor growth in a xenografted mouse tumor model. Overall, the results revealed that long-term exposure to 5-HT upregulated EZH2, and the noncanonical action of EZH2 allowed the expression of TPH1-5-HT₇ axis leading to gemcitabine resistance and CSC population in PDAC.

Inflammatory bowel disease (IBD) is a chronic relapsing inflammation in the gastrointestinal tract. Biological therapeutics and orally available small molecules like tofacitinib (a JAK inhibitor) have been developed to treat IBD, but half of the patients treated with these drugs fail to achieve sustained remission. In the present study, we compared the therapeutic effects of BJ-3105 (a 6-alkoxypyridin-3-ol derivative) and tofacitinib in IBD. BJ-3105 induced activation of AMP-activated protein kinase (AMPK) in the kinase activity measurement and recovery from cytokine-induced AMPK deactivation in HT-29 human colonic epithelial cells. Similar to tofacitinib and D942 (an AMPK activator), BJ-3105 inhibited IL-6-induced JAK2/STAT3 phosphorylation and TNF-α-stimulated activation of IKK/NF-κB, and consequently, stimulus-induced upregulations of inflammatory cytokines and inflammasome components. In addition, unlike tofacitinib or D942, BJ-3105 inhibited NADPH oxidase (NOX) activation and consequent superoxide production induced by activators (mevalonate and geranylgeranyl pyrophosphate) of the NOX cytosolic component Rac. In mice, oral administration with BJ-3105 ameliorated dextran sulfate sodium (DSS)-induced colitis and azoxymethane/DSS-induced colitis-associated tumor formation (CAT) much more potently than that with tofacitinib. Moreover, BJ-3105 suppressed the more severe form of colitis and CAT formation in mice with AMPK knocked-out in macrophages (AMPKα^fl/fl-Lyz2-Cre mice) with much greater efficacy than tofacitinib. Taken together, our findings suggest BJ-3105, which exerted a much better anti-colitis effect than tofacitinib through AMPK activation and NOX inhibition, is a promising candidate for the treatment of IBD.