Suppression of c‐Myc enhances p21WAF1/CIP1‐mediated G1 cell cycle arrest through the modulation of ERK phosphorylation by ascochlorin

Numerous anti‐cancer agents inhibit cell cycle progression via a p53‐dependent mechanism; however, other genes such as the proto‐oncogene c‐Myc are promising targets for anticancer therapy. In the present study, we provide evidence that ascochlorin, an isoprenoid antibiotic, is a non‐toxic anti‐cancer agent that induces G1 cell cycle arrest and p21WAF1/CIP1 expression by downregulating of c‐Myc protein expression. Ascochlorin promoted the G1 arrest, upregulated p53 and p21WAF1/CIP1, and downregulated c‐Myc in HCT116 cells. In p53‐deficient cells, ascochlorin enhanced the expression of G1 arrest‐related genes except p53. Small interfering RNA (siRNA) mediated c‐Myc silencing indicated that the transcriptional repression of c‐Myc was related to ascochlorin‐mediated modulation of p21WAF1/CIP1 expression. Ascochlorin suppressed the stabilization of the c‐Myc protein by inhibiting ERK and P70S6K/4EBP1 phosphorylation, whereas it had no effect on c‐Myc degradation mediated by PI3K/Akt/GSK3β. The ERK inhibitor PD98059 and siRNA‐mediated ERK silencing induced G1 arrest and p21WAF1/CIP1 expression by downregulating c‐Myc in p53‐deficient cells. These results indicated that ascochlorin‐induced G1 arrest is associated with the repression of ERK phosphorylation and c‐Myc expression. Thus, we reveal a role for ascochlorin in inhibiting tumor growth via G1 arrest, and identify a novel regulatory mechanism for ERK/c‐Myc. Our results showed that ascochlorin specifically inhibited the synthesis of the c‐Myc protein regardless of proteasomal degradation. c‐Myc stability was regulated by phosphorylation of ERK, p70S6 kinase (p70S6K), and 4EBP1, and inhibition of ERK phosphorylation enhanced c‐Myc downregulation and p21WAF1/CIP1‐dependent G1 cell cycle arrest.