Ginkgetin attenuates metastasis of breast cancer via inhibition of the Hedgehog signaling pathway

Objectives: To investigate the role and mechanism of Ginkgetin (GGT) in breast cancer metastasis. Methods: The anti-proliferative effects of GGT on breast cancer cell lines 4T1 and MDA-MB-231 were examined using the MTT assay and colony formation assay. The wound-healing assay and invasion assay were carried out to evaluate GGT's impact on the migration and invasion abilities of these cancer cells. The effect of GGT on the stemness of breast cancer cells was further analyzed by the tumorsphere assay. The expression levels of tumor stem cell (CSC) markers SOX2, OCT4, CD44, and ALDH1 were determined by qRT-PCR. The effect of GGT on breast cancer cell metastasis in vivo was studied in a mouse model. RNA sequencing analysis was performed to explore the mechanism of GGT action in breast cancer cells. Results: The MTT and colony formation assays showed that GGT exerted moderate anti-proliferative effects on the breast cancer cell lines 4T1 and MDA-MB-231. Moreover, the woundhealing and transwell invasion assays demonstrated that GGT significantly inhibited the migration and invasion capabilities of these cancer cells. Tumorsphere experiments further indicated that GGT substantially reduced the stemness of breast cancer cells. The qRT-PCR analysis results showed that GGT markedly suppressed the expression of cancer stem cell (CSC) markers, such as SOX2, OCT4, CD44, and ALDH1. In vivo experiments revealed that GGT significantly inhibited breast cancer cell metastasis and enhanced mouse survival. Preliminary mechanistic studies based on RNA sequencing analysis showed that GGT downregulated the Hedgehog (Hh) signaling pathway in breast cancer cells. Conclusion: The data suggest that GGT inhibits breast cancer stem cells by suppressing the Hh signaling pathway. This discovery provides a novel approach for breast cancer treatment.