A Study On Endocrine Treatment Of Tanshinone ⅡA Against Human Breast Cancer

Objective Breast cancer nowadays is a major public health problem in the United States and in most industrialized countries. It is the most common cancer in women and the leading cause of cancer death among women 35-54 years of age. As we know, more than half of breast cancer is hormone-dependent, and estrogen is the key stimulating factor to the forming and development of the tumor. The mechanism of endocrine therapy of breast cancer is to decrease or diminish the estrogen level and preventing the growth and proliferation of the cancer. But the target of endocrine therapy is ER, which restrict the application of these endocrine drugs, which cant make sense to ER negative breast cancer. Tanshinoneâ…¡A is a derivative of phenanthrene-quinone isolated from Danshen. Many studies in vitro have shown that tanshinoneâ…¡A can inhibited the proliferation of multiple tumor cells. On the year of 2005, Wang et al. report-ed that Tanâ…¡A can inhibit the growth of ER positive breast cancer cell in vitro and ER negative breast cancer cell in vivo. Based on its this foundation, our experiment study is plan to observe the Tanâ…¡A's function on both ER positive and ER negative breast cancer cells in vitro, and use the endocrine drugs Tamoxifen as positive control drug, to explore the possibility of endo-crine treatment of Tanâ…¡A.Methods Human breast cancer cell lines (ER positive MCF-7 and ER negative MDA-MB-231) were acquired and routinely cultured. Different concentrations of TanllA and Tamoxifen were used to treat both cell lines separately. MTT assay, Brdu incorporation, immunohistochemistry method and PI flow cytometry were used to detect the proliferation and apoptosis associated changes after the treatment of TanllA and Tamoxifen.Results MTT assay shows that TanllA inhibit both ER positive breast cell MCF-7 and ER negative breast cell MDA-MB-231, and this function act as dose-time-dependent relations. The IC₅₀ of Tanâ…¡A on both cell lines is 0.25ug/ml; the positive control drug Tamoxifen can only inhibit ER positive breast cell prominently but no definite inhibitory function on ER negative breast cancer cell MDA-MB-231. And there exists significant difference between the two drugs.Brdu incorporation in vitro shows that, Brdu labeled cells of MCF-7 among Tanâ…¡A-treated cells is 23.8±4. 3%, lower in the number in Tamoxifen and untreated controls (40.8±6.9% and 54.8±2.3% separately), which were statistically significant(P＜0.05); Brdu labeled cells of MDA-MB-231 among TanllA-treated cells is 21.1±2.8%, also lower than Tamoxifen-treated cells and untreated controls(37.9±3. 9% and 43. 9±5.1%), which were statistically significant(P＜0.05).Immunohistochemistry method showed that ER and PR protein on MDA-MB-231 breast cancer cells treated 72h by Tanâ…¡A were weak positive, and the positive score were 0.263±0.069 and 0.121+0.030, which higher than the score of untreated controls (0 and 0.043±0.069), which were statistically significant(P＜0.05). Furthermore, after treated with TanllA, the positive expression of p53(positive score 0.376±0.065 and 1.233±0.132 separately) of both cancer cells were upregulated prominently, with contrast to untreated controls of 0.150±0.030 and 0.357±0.086 separately, which have significant diference(P＜0.05). With TanllA treating, the CerbB-2 expression of ER positive cell MCF-7 is upregulated, contrast with control group and TAM group(P＜0. 05), but there is no significant change CerbB-2 expression of MDA-MB-231 after treating with TanllA and TAM(P＞0.05). TanllA have no significant influence on Bcl-2 expression of both cell lines(P＞0.05). Flow cytometry examination showed that with Tanâ…¡A-treated and Tamoxifen-treated, apoptosic fiction of TanllA on MCF-7 and MDA-MB-231 were 29.9±5.5% and 23.3±5.9% relatively, and Tamoxifen group were 13.4±1.1% and 9.5±6.8% separately, and untreated controls were 6.72.8% and 6.1±5.0% relatively. According to these data, we can conclude that Tanâ…¡A can promote the apoptosis of MCF-7 and MDA-MB-231 breast cancer cells, which were statistically significant with untreated controls (P＜0.001), but Tamoxifen had no such apoptosis function on MDA-MB-231 beast cancer cell, which had no significant difference with untreated control group.Conclusion Tanâ…¡A can inhibit both ER positive breast cancer cell MCF-7 and ER negative breast cancer cell MDA-MB-231, and it act as adose-time dependent relations. And this inhibitory function is better than positive control drugs Tamoxifen. On the other hand, TanllA can upregulated the positive expression of ER and PR on ER negative breast cancer cell MDA-MB-231; finally, TanllA can also upregulate the expression of p53 on both MCF-7 and MDA-MB-231 breast cancer cells, remarkably decrease the percentage of S phase cancer cells, promote cell apoptosis.