Effect of xy2004, a centchroman derivative, on proliferation of MCF-7 cells in vitro and the mechanism / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the effect of xy2004, a centchroman derivative, on the proliferation of MCF-7 cells and the mechanisms.</p><p><b>METHODS</b>The effects of xy2004 on MCF-7 cell proliferation and apoptosis were evaluated with MTT assay and flow cytometry, respectively. The expressions of the apoptosis-related proteins were examined with Western blotting. Competitive estrogen-receptor binding assay was used to investigate the affinity of xy2004 to estrogen receptors (ER).</p><p><b>RESULTS</b>xy2004 induced proliferation of MCF-7 cells at low concentrations but inhibited cell proliferation at high concentrations. The application of tamoxifen inhibited xy2004-induced proliferation of MCF-7 cells. The relative binding affinity of xy9906 to ERα and ERβ, presented as the IC50 value, was 7.38 × 10⁻³ mol/L and 4.12 × 10⁻⁷ mol/L, respectively. Treatment of MCF-7 cells with high-concentration xy2004 reduced the cellular expression of Bcl-2 protein and increased Bax protein expression.</p><p><b>CONCLUSION</b>At low concentrations, xy2004 directly stimulates the proliferation of MCF -7 cells through ligand-receptor binding, and at high concentrations, it inhibits the cell proliferation by regulating the expression levels of the apoptosis-related proteins.</p>

Effect of centchroman on cellular and humoral immunity.

Centchroman (Ormeloxifene) is a nonsteroidal selective estrogen receptor modulator that is used as once a week oral contraceptive agent. The effect of centchroman on the immune system was evaluated by using different experimental models such as carbon clearance test, cyclophosphamide induced neutropenia, neutrophil adhesion test, effect on serum immunoglobulins, mice lethality test and indirect haemagglutination test. The first three models namely carbon clearance test, cyclophosphamide induced neutropenia and neutrophil adhesion test were used to study cell mediated immunity while the latter three models were used to see the effect on humoral immunity. Centchroman was administered orally at a dose of 5 mg/kg and levamisole (2.5 mg/kg/ p.o) was used as standard drug. Centchroman significantly increased the levels of serum immunoglobulins and also prevented the mortality induced by bovine Pasteurella multocida in mice. It also increased significantly the circulating antibody litre in indirect haemagglunation test. However, it did not show any significant effect on phagocytic index in carbon clearance assay and nor did influence the adhesion of neutrophils in the neutrophil adhesion test. Centchroman was also not effective in preventing the cyclophosphamde induced neutropenia. Hence, it was concluded that centchroman increases humoral immunity with no significant effect on cell mediated immunity.

Evaluation of anti-tumor efficacy of injectable Centchroman in mice bearing Ehrlich ascites carcinoma.

Anti-tumor efficacy of Centchroman formulated as niosomes and gel implant was evaluated in Swiss albino mice bearing Ehrlich ascites carcinoma at 10 mg/kg body weight dose given subcutaneously. Median day of death, percentage increase in host life span and changes in body weight were studied. Centchroman significantly (P < 0.05) increased the median day of death both in free and formulated systems. Also, injectable formulations exhibited a significant (P < 0.05) increase in host life span compared to free drug, hence, enhanced anti-tumor efficacy against Ehrlich ascites carcinoma.

In-vitro effect of estrogen-antagonist on motility and penetration ability of human spermatozoa.

Antiestrogens affect spermatozoa through their action on Leydig and Sertoli cells. Direct effect of antiestrogens namely tamoxifen and centchroman in concentration of 1, 2.5, 5, 10 and 20 micrograms/ml in incubation medium was determined on motility and penetration ability of human spermatozoa. Motility (%) was invariably reduced after 15, 30 and 60 min. of incubation. Addition of 17 beta-estradiol to medium with antagonist caused inhibition of motility in dose related manner. The distance travelled by spermatozoa treated with tamoxifen or centchroman in media was reduced by 30% and addition of estradiol along with antiestrogen reduced it to 50% compared to that of untreated spermatozoa.

Effect of centchroman and tamoxifen on protein patterns of fallopian tubes of rhesus monkeys, Macaca mulatta.

To study the effect of centchroman and tamoxifen on estrogen-dependent proteins of fallopian tubes of rhesus monkey, these antiestrogens were given with and without estradiol to ovariectomized monkeys. In absence of estradiol, both the compounds induced the synthesis of 130 and 95 K proteins. Concentration of 85 K protein was also increased markedly. These compounds, however, suppressed the estrogen stimulated synthesis of 130 K protein when administered with estradiol. The results show that both centchroman and tamoxifen possess estrogen agonistic as well as antagonistic properties and 130 K protein can be used as a marker protein to study estrogen action and for screening of antiestrogenic compounds in a primate model.

Effect of prolonged centchroman treatment on pituitary gonadotrophic activity in rhesus monkeys.

Daily oral administration of antiestrogen centchroman at 6.25, 12.5 and 25 mg/kg doses to adult female rhesus monkeys continuously for one year caused no significant effect on their total pituitary gonadotrophin content as evidenced by an almost similar extent of uterine weight gain, premature opening of vagina and cornification of vaginal epithelium in immature female mice treated with pituitary homogenates from control and centchroman treated monkeys.

Effects of centchroman, a synthetic estrogen antagonist on the fertility of female hamsters.

Centchroman (3, 4-trans-2, 2-dimethyl-3-phenyl-4-p-beta-pyrrolidinoethoxy-phenyl-7-methoxy-chroman) , a non-steroidal, estrogen antagonist, injected subcutaneously (2 mg/kg body wt) on days 1, 2 and 3 post-coitum in hamsters, prevented implantation in 70% of the animals. A significant decrease in the circulating levels of estradiol and progesterone was observed on day 4 post-coitum as compared to control animals following the treatment of centchroman. The activities of various lysosomal enzymes were also found diminished in the treated animals. This study shows that centchroman may act as an anti-implantation agent in hamsters indicating that estrogen plays a key role during the process of ovum implantation in this species.

Monoamine oxidase concentration in median eminence, pituitary and uterus of rat after estrogen and centchroman treatment.

Estradiol dipropionate (0.005 mg/kg) increased the monoamine oxidase levels in pituitary, median eminence and uterus of ovariectomized rats. Centchroman (3,4-trans-2, 2-dimethyl-3-phenyl-4-p-(-pyrrolidinethoxy)-phenyl-7 methoxy chroman, 1.25 mg/kg) decreased the enzyme levels in pituitary and median eminence but not in uterus. Centchroman, given to estradiol treated groups could not bring down the enhanced levels of the enzyme in the uterus.

Mode of action of centchroman at vaginal and ovarian level in immature rats.

Centchroman studied at various doses did not cause ovulation in immature rats as judged by morphological, histological and biochemical parameters whereas, 20 mg/kh dose caused early opening of vagina and cornification of the vaginal epithelial cells. However, no ovulation was detected by this regimen. No stimulatory effect was observed in the glycogen and cholesterol content in the ovaries of Centchroman treated rats. The effects on vagina persisted even in ovariectomised immature rats administered with Centchroman. The mode of various doses of Centchroman in immature rats as judged morphologically, histologically and biochemically has been discussed.

Monoamine oxidase activity in the pituitary of mice after estrogen, progesterone and centchroman treatment.

Monoamine oxidase activity (MAO) was estimated in the pituitary of ovariectomized mice after a single administration of estradiol-dipropionate (0.01 mg/kg; im), progesterone (1 mg/kg; im) and centchroman (1.25 mg/kg ip). Estrogen and progesterone were found to decrease the enzymic level, as compared to control, while centchroman remarkably increased it. The significance of dissimilarity in the pituitary threshold for steroidal and nonsteroidal molecule is discussed.