Novel autophagy-promoted cancer therapy: Discovery of matrine-based selective TBC1D2 inhibitors by drug-target complex purification.

Autophagy strategies have seen clinical or preclinical studies underway to combat cancer therapeutic resistance, but the on-target effects in normal tissues necessitate new selective approaches. Based on the antitumor efficacy and low toxicity of matrine, a traditional Chinese medicine component, we designed and synthesized 24 matrine derivatives with higher in vitro activities. Among these compounds, A1 demonstrated the highest activity and selectivity against the HeLa cell line, with a half maximal inhibitory concentration of 0.43 μM and a selective index of 3.5. Subsequently, a label-free target fishing method using size exclusion chromatography was developed to separate proteins binding to A1. Surprisingly, TBC1 domain family member 2 (TBC1D2) emerged as the selective target of compound A1, which promotes Ras-related protein Rab-7a (RAB7A) accumulation on the lysosomal membrane and facilitates autophagy by inhibiting TBC1D2-mediated guanosine triphosphate (GTP) hydrolysis on RAB7A-GTP. Further mechanistic studies show that low-dose compound A1 induced reversible cell cycle arrest and anastasis through autophagy. Finally, A1 proved to be more efficient than matrine in HeLa-infected nude mice and was well-tolerated. These findings provide new insights into target identification techniques, ultimately leading to the discovery of first-in-class TBC1D2 inhibitors and paving the way for a novel therapeutic strategy for cervical carcinoma and beyond.