Influence of 2-methoxyestradiol on cell morphology and Cdc2 Kinase activity in WHCO3 esophageal carcinoma cells

Abstract

The influence of 1 × 10 −6 M exogenous 2-methoxyestradiol (2ME) was investigated on nuclear and cytoplasmic morphology, as well as Cdc (cell division cycle) 2 kinase activity in WHCO3 esophageal carcinoma cells. Mitotic indices after 18 h of 2ME exposure revealed an increase in metaphase cells (9.0%) when compared to the vehicle-treated cells (0.9%). 2ME-treated cells showed apoptotic cells at 5.6% after 18 h of exposure to dimethyl sulphoxide, compared to 0.9% in vehicle-treated cells. Increased morphological characteristics of apoptosis were observed in 2ME-treated cells after 21.5 h of exposure. Twelve percent of cells were in apoptosis when compared to the 1.6% of vehicle-treated cells. Furthermore, 42.4% of cells were arrested in metaphase after 21.5 h of 2ME exposure compared to 2.9% of vehicle-control cells present in metaphase. Cdc2 kinase activity was statistically significantly increased (1.7-fold) (P < 0.005) after 18 h of 2ME exposure when compared to vehicle-treated controls. Although the mechanism of 2ME's action on esophageal carcinoma cells is not yet elucidated, the present study revealed that 2ME caused metaphase arrest, as well as an increase in Cdc2 kinase activity that culminated in the induction of apoptosis in these cells.2-Methoxyestradiol (2ME) is a naturally occurring physiological estrogen metabolite of 17beta-estradiol and considered as an antimitogenic compound and tubulin poison. Early research on 2ME's biological activity stated its effect on mitotic spindles and cell cycle progression (32). It has been shown to inhibit cell proliferation and to induce apoptosis in a large variety of tumor cells and is harmless to most normal cells (21). 2ME exerts both antiangiogenic and antitumor effects regardless of the cell's hormone receptor status (10,19,23,24,32,34). 2ME causes mitotic accumulation and abnormal mitotic spindle formation in both estrogen receptor (ER)-positive and negative cells (23,24,32).Recently, researchers have demonstrated the antiangiogenic influence of oral 2ME on a laser-induced murine model of choroidal neovascularization with no side effects of toxicity or weight loss being observed (9). investigated the efficacy on growth inhibition of 2ME on human hepatocellular carcinoma in vitro (31). 2ME caused a reduction in cell growth in these cells and it was concluded that the mechanism of action appears to be induction of apoptosis. SK-Hep1 hepatocellular carcinoma cells were the most sensitive to 2ME and an up-regulation of the p53 and p21 proteins was observed. However, normal human hepatocytes were not influenced when treated with 2ME (31). Roswall et al. (2006) demonstrated 2ME's antiproliferative effects in five human anaplastic thyroid carcinoma cell lines (HTh7, HTh 74, HTh83, C643, and SW1736) after treatment with 2ME. It was revealed that a G 2 /M-arrest was followed by an increased fraction of cells present in sub-G 1 (29). The

Docking, Synthesis, and in vitro Evaluation of Antimitotic Estrone Analogs

“…The GI 50 concentrations for the compounds are summarized in Table 1. In in vitro studies conducted in our laboratory, 2-ME showed an antiproliferative effect on HeLa, SNO, MCF-7, and MCF-12A cells in a concentration range of 1-2lM (43)(44)(45). Accordingly, Edsall et al (2004) demonstrated that the same compound inhibited MDA-MB-231 cancer cell proliferation by 50% at 1 lM (30).…”

In vitro effects of 2‐methoxyestradiol on cell numbers, morphology, cell cycle progression, and apoptosis induction in oesophageal carcinoma cells

“…32 Furthermore, we have previously demonstrated a significant increase in Cdc2 kinase activity in 2-ME-treated cells when compared to vehicle-treated controls in WHCO3 cells. 10 Cdc2 kinase activity was statistically significantly increased (1.7-fold) ( p < 0.005) after 2-ME exposure when compared to vehicle-treated controls. Our observation contributes to the elucidating of the mechanism of action in WHCO3 oesophageal carcinoma cells and reveals that 2-ME causes a metaphase arrest, disrupts mitotic spindle formation, enhances Cdc2 kinase activity leading to persistence of the spindle checkpoint, and thus prolonged metaphase arrest culminating in the induction of apoptosis in WHCO3 cells.…”

2-Methoxyestradiol blocks cell-cycle progression at the G&lt;sub&gt;2&lt;/sub&gt;/M phase and induces apoptosis in human acute T lymphoblastic leukemia CEM cells

“…The potential role of 2-ME2 as an anti-cancer agent has been intensively investigated. 2-ME2 was found to inhibit growth and induce apoptosis of tumors of Ewing sarcoma [6], chondrosarcoma [7], osteosarcoma [8], esophageal [9], hepatocellular [10], ovarian [11] and nasopharyngeal carcinomas [12], and prostate cancer [13] both in vitro and in vivo. 2-ME2 also showed anti-leukemic activity in vitro with therapeutic selectivity [14][15][16].…”

“…2-ME2 also showed anti-leukemic activity in vitro with therapeutic selectivity [14][15][16]. Several different mechanisms of action have been proposed, including the accumulation of cells in the G 2 /M phase [7][8][9], inhibition of tubule polymerization [17], inhibition of superoxide dismutase (SOD) [14], and accumulation of reactive oxygen species (ROS) [14,15], activation of c-jun N-terminal-activated kinase (JNK) [15], increased expression of p53 and p21 [7,8,10], Bcl-2 phosphorylation [11], and mitochondrial release of cytochrome c [14,15]. However, its exact mechanism on human acute T lymphoblastic leukemia remains unknown.…”