Epicatechins Purified from Green Tea (Camellia sinensis) Differentially Suppress Growth of Gender-Dependent Human Cancer Cell Lines

Abstract

The anticancer potential of catechins derived from green tea is not well understood, in part because catechin-related growth suppression and/or apoptosis appears to vary with the type and stage of malignancy as well as with the type of catechin. This in vitro study examined the biological effects of epicatechin (EC), epigallocatechin (EGC), EC 3-gallate (ECG) and EGC 3-gallate (EGCG) in cell lines from human gender-specific cancers. Cell lines developed from organ-confined (HH870) and metastatic (DU145) prostate cancer, and from moderately (HH450) and poorly differentiated (HH639) epithelial ovarian cancer were grown with or without EC, EGC, ECG or EGCG. When untreated cells reached confluency, viability and doubling time were measured for treated and untreated cells. Whereas EC treatment reduced proliferation of HH639 cells by 50%, EGCG suppressed proliferation of all cell lines by 50%. ECG was even more potent: it inhibited DU145, HH870, HH450 and HH639 cells at concentrations of 24, 27, 29 and 30 microM, whereas EGCG inhibited DU145, HH870, HH450 and HH639 cells at concentrations 89, 45, 62 and 42 microM. When compared with EGCG, ECG more effectively suppresses the growth of prostate cancer and epithelial ovarian cancer cell lines derived from tumors of patients with different stages of disease.

Figure 1

Morphology of Green Tea, Camellia sinensis.

Figure 2

Structure of epicatechin (EC), epigallocatechin (EGC), epicatechin gallate (ECG) and epigallocatechin gallate (EGCG) used in this investigation. All bioflavanoids have three rings; tea catechins are flavan-3-ols with an hydroxyl group at the 3, 5 and 7 positions.

Figure 3

Photomicrographs of cells grown in culture with 0, 25, 50, 75 or 100 µM of ECG or EGCG. Organ-confined prostate cancer cell line HH870 and primary and metastatic epithelial ovarian cancer cell lines (HH450 and HH639) were seeded (2.5 × 10⁵ cells per line) into flasks containing culture medium (RPMI-1640 with 9% FBS-antibiotics) with or without ECG or EGCG. When growth of untreated (control) cells reached confluency, cell monolayers in each flask were photographed under a light microscope. Both ECG and EGCG significantly affected the density of each cell line. Decrease in cell density was observed at higher concentrations of ECG and EGCG. Magnification: ×100.

Figure 4

Density of cancer cells seeded (2.5 × 10⁵ cells per line) into flasks containing culture medium (RPMI-1640 with 9% FBS-antibiotics) with or without catechins (50 µM) (n = 5 per treatment). When growth of untreated cells reached confluency, cells from each flask were harvested and viable/dead cells were counted. Mean and standard deviation are represented. P-values obtained with pairwise comparison and ANOVA are shown.

Figure 5

Doubling time of cancer cells seeded (0.25 × 10⁶ per line) into flasks containing culture medium (RPMI-1640 with 9% FBS-antibiotics) with or without catechins (50 µM) (n = 5 per treatment). When untreated cells reached confluency, cells from each flask were harvested and viable/dead cells were counted. Vertical bars refer to standard deviation. The mean doubling time was calculated from the mean of five viable cell counts. P-values were obtained with pairwise comparison and ANOVA.

Figure 6

Suppression of cell growth by ECG and EGCG. (A) DU145; (B) HH870; (C) HH450; (D) HH639. Cells (0.25 × 10⁶ per line) were seeded in flasks containing culture medium (RPMI-1640 with 9% FBS-antibiotics) with or without ECG or EGCG at concentrations of 0, 25, 50, 75 and 100 µM (three flasks for each dose). Mean (circles) and standard deviation (vertical lines) are represented. When untreated cells reached confluency, cell monolayers in each flask were photographed under a light microscope, harvested and counted. The suppressive effect on cell density was striking at higher concentrations of ECG and EGCG. At 25 µM of EGCG, cell counts for HH870 and DU145 were significantly higher than control values. P-values indicate significant differences between mean values of treated and untreated cells.