Alnus sibirica Compounds Exhibiting Anti-Proliferative, Apoptosis-Inducing, and GSTP1 Demethylating Effects on Prostate Cancer Cells

Abstract

Alnus sibirica (AS) is distributed in Korea, Japan, China, and Russia and has reported anti-oxidant, anti-inflammatory, and reducing activities on atopic dermatitis-like skin lesions, along with other beneficial health properties. In the present study, we tried to prove the cancer-preventive activity against prostate cancer. The extracted and isolated compounds, oregonin (1), hirsutenone (2), and hirsutanonol (3), which were isolated from AS, were tested for anti-proliferative activity. To do this, we used the MTT assay; NF-κB inhibitory activity, using Western blotting; apoptosis-inducing activity using flow cytometry; DNA methylation activity, using methylation-specific polymerase chain reaction in androgen-dependent (LNCaP) and androgen-independent (PC-3) prostate cancer cell lines. The compounds (1-3) showed potent anti-proliferative activity against both prostate cancer cell lines. Hirsutenone (2) exhibited the strongest NF-κB inhibitory and apoptosis-inducing activities compared with oregonin (1) and hirsutanonol (3). DNA methylation activity, which was assessed for hirsutenone (2), revealed a concentration-dependent enhancement of the unmethylated DNA content and a reduction in the methylated DNA content in both PC-3 and LNCaP cells. Overall, these findings suggest that hirsutenone (2), when isolated from AS, may be a potential agent for preventing the development or progression of prostate cancer.

Keywords: Alnus sibirica; DNA methylation; cancer prevention; hirsutenone; prostate cancer.

Figure 1

Structures of the three diarylheptanoids isolated from Alnus sibirica, oregonin (1); hirsutenone (2); hirsutanonol (3).

Figure 2

NF-kB inhibitory activities of oregonin (1), hirsutenone (2), and hirsutanonol (3) in PCa cell lines. A: PC-3 cell; B: LNCaP cell. The results were expressed as mean ± SD of triplicate experiments (n = 3). NC: Normal control group. *: p < 0.05, **: p < 0.01, ***: p < 0.001, compared with the normal control group.

Figure 3

Apoptosis-inducing activities of various concentrations of oregonin (1), hirsutenone (2), and hirsutanonol (3) in PCa cell line. (A): PC-3 cells; (B): LNCaP cell. The results are expressed as mean ± SD of triplicate experiments (n = 3). BAY: positive control. Letters a–o in the same column indicate significant differences, p < 0.05.

Figure 3

Apoptosis-inducing activities of various concentrations of oregonin (1), hirsutenone (2), and hirsutanonol (3) in PCa cell line. (A): PC-3 cells; (B): LNCaP cell. The results are expressed as mean ± SD of triplicate experiments (n = 3). BAY: positive control. Letters a–o in the same column indicate significant differences, p < 0.05.

Figure 4

DNA methylation activity of hirsutenone (2) in PCa cell lines. (A,C): relative unmethylated DNA contents in PC-3 and LNCaP cell lines; (B,D): relative methylated DNA contents in PC-3 and LNCaP cell lines. The results were expressed as the mean ± SD of triplicate experiments (n = 3). NC: Normal control group. *: p < 0.05, **: p < 0.01, compared with the normal control group.