Discovery of potent TNIK inhibitors containing a 1H-pyrrolo[2,3-b]pyridine scaffold as promising therapeutics for colorectal cancer.

Traf2-and Nck-interacting kinase (TNIK), a downstream effector of the Wnt/β-catenin pathway and a key regulatory component of the TCF4/β-catenin transcriptional complex, has emerged as a potential therapeutic target for colorectal cancer. In this study, based on compound 1, a previously reported TNIK inhibitor, we developed a series of optimized inhibitors featuring a 1H-pyrrolo[2,3-b]pyridine scaffold. Among these, compound N15 exhibited the most potent activity, with exceptional TNIK inhibition in an in vitro enzymatic assay (IC50 = 0.49 nM) and favorable metabolic stability in human liver microsomes (T1/2 = 241 min). In HCT116 cells, N15 exhibited strong antitumor activity by suppressing proliferation, inducing apoptosis, and causing cell cycle arrest, while also downregulating Wnt pathway target gene expression. Furthermore, N15 significantly inhibited tumor growth in an HCT116 xenograft mouse model without inducing notable adverse effects. Collectively, these results identify N15 as a promising lead compound for further development of TNIK inhibitors.