Evodiamine inhibits ESCC by inducing M-phase cell-cycle arrest via CUL4A/p53/p21 axis and activating noxa-dependent intrinsic and DR4-dependent extrinsic apoptosis

Background: Esophageal squamous cell carcinoma (ESCC) is a malignancy with high incidence in several regions of China, and the prognosis of patients with ESCC is unfavorable. Evodiamine (Evo), a small molecule derived from the traditional Chinese herb Evodia rutaecarpa, has shown anti-cancer efficacy in numerous human malignancies but not in ESCC.

Purpose: To determine whether Evo induces cell-cycle arrest and Apoptosis in ESCC in vitro and in vivo and elucidate the underlying mechanisms.

Methods: ATPlite and colony formation assay were used to validate the inhibiting effect of Evo on three ESCC cells in vitro; Two subcutaneous tumor models of ESCC cells were used to evaluate the anti-ESCC effect of Evo and assess the biosafety of Evo in vivo; RNAseq and Database of KEGG pathway analysis provided a direction for the mechanistic study of Evo; FACS was used to detect Evo-induced cell cycle arrest and cell Apoptosis in ESCC cells; Western blot and QPCR were respectively used to detect the level of related genes and proteins in Evo-treated ESCC cells; SiRNA and Other experimental techniques were used to identify the molecular mechanism of Evo-induced ESCC cell cycle arrest and cell Apoptosis.

Results: Evo significantly suppressed the growth of ESCC both in vitro and in vivo. Mechanistically, Evo induced M-phase cell-cycle arrest by inactivation of CUL4A E3 Ligase, which mediates degradation blockage of p53 and transcriptional activation of p21. With the prolonged treatment time, Evo triggered both Noxa-dependent intrinsic and DR4-dependent extrinsic cell Apoptosis in two ESCC cell lines.

Conclusion: Our findings revealed the anti-tumor efficacy and mechanisms of Evo, providing a solid scientific basis for Evo as an attractive choice for ESCC treatment.

Apoptosis; Esophageal squamous cell carcinoma (ESCC); Evodiamine; M-phase cell-cycle arrest.