Human epidermal growth factor receptor 2 as a target of 1,2,3,4,6-penta-O-galloyl-β-d-glucose in colon cancer

Although HER2 overexpression is observed in only 2%-3% of colorectal cancer (CRC) patients, its amplification can function as a compensatory mechanism that drives CRC resistance to targeted therapy. 1,2,3,4,6-Penta-O-galloyl-β-d-glucose (PGG) has been reported to possess anti-tumor and anti-metastatic activities in CRC. Nevertheless, the molecular targets through which PGG exerts its effects in CRC remain unclear. This study aimed to evaluate the role of HER2 as a potential molecular target of PGG in CRC.

Ultra performance liquid chromatography-mass spectrometer (UPLC-MS) was employed to determine the cellular uptake of PGG in HCT116 cells. Potential PGG targets were predicted using STITCH and molecular operating environment platforms. Functional rescue assays were performed with lapatinib, a HER2 inhibitor. Direct PGG-HER2 interactions were validated by drug affinity responsive target stability and thermal shift assays. Downstream signaling effects were examined by assessing HER2 expression and its downstream pathway by Western blotting.

UPLC-MS analysis confirmed the accumulation of PGG (204.5 ± 19.1 ng) in HCT116 cells after 40 μM PGG treatment for 12 h. Computational predictions suggested ErbB2 (HER2) as a potential binding target. Rescue experiments showed that the combination of lapatinib and PGG failed to further reduce cell viability when compared with lapatinib alone, implying HER2 as a molecular target of PGG. Both drug affinity responsive target stability and thermal shift assays verified that PGG protects HER2 from pronase-induced degradation and enhances its stability upon thermal challenge, indicating direct binding. Mechanistically, PGG suppressed HER2 expression and inhibited the PI3K-Akt-mTOR pathway, a canonical downstream pathway of HER2.

Our findings provide a strong preclinical rationale for future clinical trials of PGG as a HER2-targeted drug in CRC and highlight its considerable potential as a novel therapeutic for CRC treatment.