TGF-β Signaling Activation Confers Anlotinib Resistance in Gastric Cancer

Gastric cancer (GC) has always been a great threat to human health due to its aggressiveness and lethality. Anlotinib, a novel multi-target tyrosine kinase inhibitor (TKI), has been certified its anti-tumor effects on various tumors. Nonetheless, there are few studies on applying anlotinib as a treatment for GC. The underlying mechanism of acquired resistance during anlotinib administration remains unclear.

We investigated the toxicologic effects of anlotinib on GC cells through CCK8, colony-forming, and flow cytometry assays in vitro and xenograft models in vivo. Anlotinib-resistant GC cells, AGS-R and MGC803-R, were generated and characterized by cell proliferation and apoptosis assays. The signaling pathways involved in anlotinib resistance were probed using Cignal™ Finder 10-Pathway Reporter Array. Western blot and dual-luciferase reporter assays were performed to confirm the relationships. The TGF-β inhibitor LY364947 was introduced to demonstrate the importance of TGF-β signaling in anlotinib resistance via a series of functional assays.

Anlotinib suppressed cell growth and induced apoptosis in vitro and inhibited tumorigenesis and metastasis in vivo, while its anti-tumor effects were impaired in anlotinib-resistant cells. The results of dual-luciferase reporter assays and western blot indicated TGF-β signaling was activated in anlotinib-resistant GC cells. LY364947 combined with Anlotinib exerted a better antineoplastic effect than monotherapy and considerably reversed the anlotinib resistance in GC.

Our findings suggested that TGF-β signaling may take a significant part in anlotinib resistance in GC. The suppression of TGF-β signaling may be a possible and promising approach for the GC oncotherapy when combined with anlotinib.