Evodiamine exerts anti-cancer activity including growth inhibition, cell cycle arrest, and apoptosis induction in human follicular thyroid cancers

Abstract

Thyroid cancer (TC) is one of the most prevalent endocrine malignancy with a steadily increasing incidence globally. Although standard treatments like thyroidectomy and radioiodine therapy effectively manage most cases of differentiated thyroid cancers (DTC), certain recurrent cases or those involving poorly differentiated thyroid cancers (PDTC) demand more specialized interventions. Follicular thyroid cancer (FTC) is the second most common type of DTC, and frequently metastasizes through the bloodstream to distant sites such as bones and lungs which is a leading cause of metastatic and recurrent DTC and significantly affects survival. However, existing drugs primarily address symptom management without offering a curative solution. Therefore, it is urgent to develop a new therapeutic agent for these challenging cases. Evodiamine (EVO), extracted from Evodia rutaecarpa, has shown potential as an anti-cancer agent in multiple types of human cancers including anaplastic thyroid cancer (ATC) and papillary thyroid cancer (PTC) cells. However, the anti-cancer effects of EVO on FTC have remained unclear. Therefore, the present study aims to investigate the anti-cancer effects of EVO in FTC cells. Our data showed that EVO effectively inhibits FTC cell growth, induces cell cycle arrest, and triggers apoptosis. Additionally, our study explored the underlying mechanisms through which EVO affects signaling pathways. To verify the anti-cancer effects of combination chemotherapy, EVO and doxorubicin were used together in FTC cells. In conclusion, this study demonstrates that EVO shows significant anti-human FTC activity, making it a promising therapeutic candidate for the treatment of follicular thyroid cancers.

Keywords: Evodiamine; anti-cancer therapy; apoptosis; follicular thyroid cancer.

Figure 1

Evodiamine inhibits cell proliferation in human follicular thyroid cancer cells. WRO cells were incubated with evodiamine, and (A) the cellular survival was examined with CCK-8 assay, and (B) colony formation was determined with colony formation assay. (C) The cytotoxicity was evaluated with LDH activity in the cells with evodiamine treatment. The control medium containing DMSO was used as a negative control. Three independent experiments were conducted.

Figure 2

Evodiamine induces cell cycle arrest at G2/M phase in human follicular thyroid cancer cells. WRO cells were incubated with evodiamine, and the cell cycle regulation was evaluated with (A) flowcytometry analysis, and (B) the cell cycle regulated proteins of G2/M phase were examined by Western blotting. The control medium containing DMSO was used as a negative control. GAPDH was used as the loading control. Three independent experiments were conducted.

Figure 3

Evodiamine induces cellular apoptosis in human follicular thyroid cancer cells. WRO cells were incubated with evodiamine, and (A) the apoptotic cells were assessed with flowcytometry analysis after PI/Annexin-V double staining. (B and C) The apoptotic related proteins expressions were examined with Western blotting in the cells under evodiamine treatment for 48 h. (D) DNA fragmentation assay was used to confirm the apoptosis induction in the cells after evodiamine treatment for 48 h. The control medium containing DMSO was used as a negative control. Z-VAD-FMK was used as the blocker of caspase activation. GAPDH was used as the loading control. Three independent experiments were conducted.

Figure 4

Evodiamine modulates Erk and p38 signaling pathways in human follicular cancer cells. WRO cells were incubated with control medium or evodiamine, and the expressions of signaling pathways including Erk, JNK and p38 were determined with Western blotting. The control medium containing DMSO was used as a negative control. GAPDH was used as the loading control.

Figure 5

The anti-cancer effect of doxorubicin in human follicular thyroid cancer cells was determined. (A) WRO cells were incubated with doxorubicin and the cellular survival was examined with CCK-8 assay. (B) WRO cells were incubated with doxorubicin and evodiamine or combination treatment with doxorubicin and evodiamine with the indicated concentrations for 48 h, and the cell apoptosis was examined with flowcytometry after PI/Annexin-V double staining. The control medium containing DMSO was used as a negative control. Three independent experiments were conducted.