Targeting Cys328 in the STAT3 DNA-binding domain with the germacrane-type sesquiterpene lactone SCP-7 inhibits colorectal Cancer growth in vitro and in vivo.

Signal transducer and activator of transcription 3 (STAT3) is a critical oncogenic driver hyperactivated in approximately 70% of human cancers, especially colorectal cancer (CRC), making it an attractive therapeutic target. Currently, the discovery of STAT3 inhibitors is primarily focused on the SH2 domain, however, there are still problems such as insufficient inhibitory activity and poor clinical efficacy. Targeting the DNA-binding domain (DBD) is a direct and effective strategy for inhibiting STAT3 function; but due to the lack of suitable targeting sites and design methods, reported compounds and their mechanisms of action are limited. Here, we identified a germacrane-type sesquiterpene lactone, SCP-7, from a natural product library through multi-dimensions progressive screening. SCP-7 exhibited potent binding affinity to the STAT3-DBD, with a Kd of 795 nM. Mechanistically, SCP-7 covalently modifies Cys328 and Cys367 of STAT3, as confirmed by mass spectrometry and mutational analysis, with Cys328 serving as the primary binding site and effectively inhibit the DNA-binding activity of STAT3. In CRC cells (HCT116 and HT29), SCP-7 effectively reduced p-STAT3 and c-Myc levels, and inhibited proliferation, migration, and colony formation. Moreover, SCP-7 induced G2/M cell cycle arrest, triggered mitochondrial dysfunction, elevated reactive oxygen species levels, and promoted apoptosis via modulation of BAX, Bcl-2, Mcl-1 and survivin. In a mouse xenograft model, SCP-7 significantly suppressed tumor growth, exhibiting activity comparable to that of oxaliplatin but with a markedly improved safety profile. Collectively, our findings identify SCP-7 as a novel covalent inhibitor targeting the STAT3-DBD with potent anti-CRC activity in vitro and in vivo, providing a promising lead compound for STAT3-targeted cancer therapy.