Tanshinone IIA induces apoptosis of ovarian cancer cells in vitro and in vivo through attenuation of PI3K/AKT/JNK signaling pathways

Abstract

Ovarian cancer is one of the most common gynecological tumors and is the second most common cause of gynecological cancer-associated mortality worldwide. Tanshinone IIA (Tan-IIA) possesses anticancer activities through inducing the apoptosis of tumor cells. The purpose of the present study was to analyze the ability of Tan-IIA to induce apoptosis of human ovarian cancer cells in vitro and in vivo, and to examine the potential mechanism underlying its activity. Western blot analysis, immunohistochemistry and flow cytometry were used to analyze the therapeutic effects of Tan-IIA on ovarian cancer. It was demonstrated that Tan-IIA significantly inhibited the growth and aggressiveness of human ovarian cancer cells. Tan-IIA significantly increased the apoptosis of human ovarian cancer cells through cleavage activation of caspases-3, caspase-8 and caspases-9. In addition, Tan-IIA treatment decreased the expression of mitochondrial-protective B-cell lymphoma 2-like protein 2 (Bcl-w) and myeloid cell leukemia-1 long (Mcl-1L) in ovarian cancer cells. Tan-IIA also reduced the expression of phosphoinositide 3-kinase (PI3K), AKT and c-Jun N-terminal kinase (JNK) in human ovarian cancer cells. A specific PI3K inhibitor (LY294002) enhanced the Tan-IIA-inhibited expression of AKT and JNK. The overexpression of PI3K negated the Tan-IIA-inhibited expression of AKT and JNK, and eliminated the Tan-IIA-induced apoptosis of human ovarian cancer cells. Additionally, the in vivo assay showed that Tan-IIA treatment inhibited the growth of ovarian cancer through increasing the apoptosis of tumor cells. In conclusion, these findings suggested that the induction of apoptosis by Tan-IIA involves the PI3K/AKT/JNK signaling pathways in ovarian cancer.

Keywords: apoptosis; ovarian cancer; phosphoinositide 3-kinase/AKT/c-Jun N-terminal kinase; tanshinone IIA.

Figure 1.

Tan-IIA significantly inhibits ovarian cancer cell growth and aggressiveness through inhibition of target PI3K. (A) Tan-IIA (50, 100, 150, 200 and 250 µM) inhibited A2780 cell growth. Tan-IIA (150 µM) inhibited the (B) migration and (C) invasion of A2780 cells, compared with PBS-treated group. (D) Tan-IIA (150 µM) downregulated the expression of PI3K in A2780 cells, compared with the control. PI3K inhibitor (LY294002; PI3KIR) promoted Tan-IIA-inhibited (Tan-IIA+PI3KIR) A2780 cell (E) growth, (F) migration and (G) invasion. Overexpression of PI3K inhibited Tan-IIA-inhibited (H) growth, (I) migration and (J) invasion (J) of A2780 cells. Tan-IIA, tanshinone IIA; PI3K, phosphoinositide 3-kinase; pPI3K, pLentivirus-PI3K; NS, not significant. *P<0.05 and **P<0.01 vs. control.

Figure 2.

Tan-IIA significantly induces the apoptosis of ovarian cancer cells through inhibition of target PI3K. (A) Tan-IIA significantly increased the apoptosis of human A2780 ovarian cancer cells, compared with the control. (B) Tan-IIA upregulated the expression of caspases-3, caspase-8 and caspases-9 in A2780 cells. (C) Tan-IIA treatment decreased the expression of mitochondrial protective Bcl-w and Mcl-1L in A2780 cells. (D) PI3K inhibitor (LY294002) enhanced Tan-IIA-induced apoptosis of A2780 cells. (E) Overexpression of PI3K eliminated Tan-IIA-induced apoptosis of A2780 cells. Tan-IIA, tanshinone IIA; PI3K, phosphoinositide 3-kinase; Bcl-1, B-cell lymphoma 2-like protein 2; Mcl-1L, myeloid cell leukemia-1 long; pPI3K, pLentivirus-PI3K; ns, not significant. *P<0.05 and **P<0.01 vs. control.

Figure 3.

Tan-IIA regulates the expression of key proteins of PI3K/AKT/JNK signaling pathways in ovarian cancer cells. (A) Tan-IIA decreased the expression and phosphorylation of AKT and JNK in A2780 cells. (B) PI3K inhibitor (LY294002) enhanced the Tan-IIA-inhibited expression and phosphorylation of AKT and JNK in A2780 cells. (C) Overexpression of PI3K eliminated the Tan-IIA-inhibited expression and phosphorylation of AKT and JNK in A2780 cells. Tan-IIA, tanshinone IIA; PI3K, phosphoinositide 3-kinase; pPI3K, pLentivirus; JNK, c-Jun N-terminal kinase; pJNK, phosphorylated JNK.

Figure 4.

Tan-IIA exerts therapeutic effects in an ovarian cancer lesion model rats. (A) Tan-IIA treatment significantly inhibited ovarian cancer growth, compared with that in PBS-treated rats. (B) Rats in the Tan-IIA-treated ovarian cancer lesion model had increased apoptotic bodies in tumor tissues, compared with the PBS-treated rats. (C) Tan-IIA upregulated the expression levels of PI3K, AKT and JNK in tumor tissues, compared with those in the control. (D) Tan-IIA treatment prolonged survival of experimental rats. Tan-IIA, tanshinone IIA; PI3K, phosphoinositide 3-kinase; JNK, c-Jun N-terminal kinase; PBS, phosphate buffer solution. **P<0.01 vs. PBS.