Robert A. Magarian

A comparison of the efficacy for antitumor activity of the non-steroidal antiestrogens analog II and tamoxifen in 7,12-dimethylbenz[a]anthracene-induced rat mammary tumors

Abstract Analog II is a cyclopropyl derivative of stilbene which has been shown to produce antiestrogenic activity with no latent estrogenic effects in the mouse and to reduce the growth of 7,12-dimethylbenz[ a ]anthracene (DMBA)-induced tumors in the rat. Thus, it was of interest to compare the antitumor activity of analog II to the antiestrogen, tamoxifen, which is a partial estrogen agonist in the rat and fully estrogenic in the mouse. The results indicate that analog II and tamoxifen were equally effective in reducing the growth of established DMBA-induced rat mammary tumors and that analog II produced a greater reduction in the occurrence of new tumors during the 7-week treatment period.

Synthesis and biological evaluation of 1,1-Dichloro-2,3-diarylcyclopropanes as antitubulin and anti-breast cancer agents

Z-1,1-Dichloro-2,3-diphenylcyclopropane (1) is an effective anti-breast cancer agent in rodents and in cell culture. We recently determined that 1 inhibits tubulin assembly in vitro and causes microtubule loss in breast cancer cells, leading to accumulation in the G2/M portion of the cell cycle. Aryl ring-halogenated, methoxylated and benzyloxylated derivatives of 1, as well as its E-isomer and the dichlorocyclopropyl derivative of diethylstilbestrol (DES), were synthesized and tested for their ability to inhibit, the assembly of tubulin into microtubules. Including 1, 17 cyclopropyl compounds were tested. One (Z-1,1-dichloro-2-(4-methoxyphenyl)-3-phenylcyclopropane (12)) was found to be more active than 1. In addition, E-1,1-dichlorocyclopropylDES (17) was more potent than DES. The E-isomer of 1 (16) was inactive. The cytostatic activities of the compounds against MCF-7 and MDA-MB231 human breast cancer cells, and their abilities to perturb microtubules in MCF-7 cells were also evaluated. Z-Dichloro-2-(4-fluorophenyl)-3-phenylcyclopropane (5), Z-1,1-dichloro-2-(4-fluorophenyl)-3-(4-methoxyphenyl)cyclopropane (11), and Z-1,1-dichloro-2-(4-methoxyphenyl)-3-phenylcyclopropane (12) were more potent than 1 against the breast cancer cells.

Methods of extraction and high-performance liquid chromatographic analysis of butylated hydroxytoluene from the tissues and serum of rats

Butylated hydroxytoluene (BHT) is a phenolic antioxidant which is widely used in foods and has been shown to inhibit chemical carcinogenesis in the mammary gland induced by 7,12-dimethylbenz(a)anthracene. However, its mechanism of action as a tumor inhibitor is unclear. The purpose of this work was first to develop a method for extracting and quantitating BHT and then to determine the amounts that accumulate in the tissues and serum of rats as a starting point for looking at mechanistic possibilities in the inhibition of mammary carcinogenesis. Methodology of extracting BHT from rat tissues and serum was developed using a modified lipid extraction procedure. The sensitive nature of reverse-phase high-performance liquid chromatography proved useful in detecting and quantifying BHT after its extraction from biological tissues. All tissues were taken from animals consuming semipurified diets with and without 0.3% BHT for various periods of time (weeks). BHT was found in much higher levels in mammary tissue than in the liver and serum of rats. The lipid content in mammary tissue appears to be predictive of the amount of BHT found in this tissue, presumably because of the lipophilic character of the antioxidant.

The interaction of dietary fat and antiestrogen treatment on DMBA-induced mammary tumors in the rat.

This study was designed to determine whether an estrogenic mechanism is involved in dietary fat-modulated tumor development and growth. Female Sprague-Dawley rats were placed on a semipurified low-fat (2% fat), high-saturated fat (20% fat), or high-polyunsaturated fat (20% fat) diet at 21 days of age. A single dose of 7,12-dimethylbenz[a]anthracene (DMBA, 10 mg) was administered intragastrically at 50 days of age. Two studies were performed. One tested the effectiveness of antiestrogen treatment (either tamoxifen or analog II) on tumor development when it was given one week prior to and one week after DMBA treatment in animals consuming a high-polyunsaturated fat diet. The second six-week study tested the antiestrogen effectiveness in arresting tumor growth and in producing regressions of established DMBA-induced tumors in rats consuming various levels and types of fat. The results of these studies indicate that both antiestrogens employed reduced the rate of growth and increased the number of regressions of established DMBA-induced tumors. In general, this was true in animals fed diets with a high content of either saturated or polyunsaturated fats and to a lesser extent in animals fed a low-fat diet. Tamoxifen produced a somewhat greater reduction in the growth of established tumor than did analog II. However, analog II, which is a more biologically pure antiestrogen, reduced the incidence of animals with mammary tumors and total tumor burden when administered one week before and one week after DMBA dosing. Tamoxifen, which is a partial estrogen-agonist, did not alter tumor incidence, but it did reduce the total tumor burden under these same experimental conditions.(ABSTRACT TRUNCATED AT 250 WORDS)

Antitumor mechanism of action of a cyclopropyl antiestrogen (compound 7b) on human breast cancer cells in culture

Abstractu2002Cyclopropyl compound 7b [(Z)-1,1-dichloro2-[4-[2-(dimethylamino)ethoxy] phenyl]-2-(4-methoxy-phenyl)-3-cyclopropane] has been shown to be a pure antiestrogen in mouse uterine tissue. Antitumor activity was examined by evaluating the influence of 7b on the proliferation, estrogen receptor (ER) affinity and cell-surface morphology of ER-positive and ER-negative human breast cancer cells in culture. The antiproliferative potency of 7b was found to be equal to tamoxifen in ER-positive MCF-7 human breast cancer cells. Further, the antiproliferative activities of 7b and tamoxifen were reversed by coadministration of estradiol. Accordingly, the antiproliferative activity of compound 7b appears to be estrogen-mediated since it did not influence the growth of either ER-negative MDA-MB-231 human breast cells or A-549 human lung cancer cells in culture. An ER-dependent mechanism of action is also supported by the specific binding affinity of 7b for ER in MCF-7 cells. Further, a study of cell surface morphology using scanning electron microscopy (SEM) revealed that 7b reduced the density and distribution of microvilli (MV) on MCF-7 cells, which was reversed by coadministration of estradiol. Compound 7b did not alter the cell surface morphology of ER-negative MDA-MB-231 cells. In conclusion, 7b inhibited the growth of ER-positive MCF-7 cells in an estradiol-reversible manner, and had no effect on either ER-negative MDA-MB-231 cells or A-549 lung cancer cells. The results of this study confirm an antiestrogenic mechanism of action for 7b as previously observed in vivo and suggest that 7b would be effective in the treatment of estrogen-dependent breast cancer or as a prophylactic treatment for women with a high risk of breast cancer development.

The influence of a novel cyclopropyl antiestrogen (compound 7a) on human breast cancer cells in culture

Compound 7a ([Z]-1,1,-dichloro-2,3-diphenyl-2-(4-(2-dimethylamino)ethoxy)phenyl) cyclopropane, dihydrogen citrate salt) is a novel cyclopropyl antiestrogen which was shown to be an estrogen antagonist without estrogen agonist activity. The antiproliferative activity of 7a was examined on estrogen receptor (ER)positive MCF-7 and ER-negative MDA-MB-231 human breast cancer cells and A-549 human lung cancer cells. Compound 7a inhibited the growth of MCF-7 cells in a dose-related manner over a concentration range of 10−9 to 10−5M, but did not alter the growth of MDA-MB-231 or A-549 cells. The antiproliferative activity of 7a (10−7M) on MCF-7 cells was reversed by co-administration of estradiol (10−8M). An ER-dependent mechanism of action is also supported by the specific ER binding of 7a in MCF-7 cells observed in this study. A study of cell surface morphology using scanning electron microscopy (SEM) revealed that compound 7a at 10−6M reduced the length and density of microvilli (MV) on MCF-7 cells, which was reversed by co-administration of estradiol (10−8M). Compound 7a did not alter the cell surface morphology of ER-negative MDA-MB-231 cells. In conclusion, 7a inhibited the growth of ER-positive MCF-7 cells in an estradiol-reversible manner, and had no effect on ER-negative MDA-MB-231 cells or A-549 lung cancer cells. The results of this study support the antiestrogenic action of 7a previously observedin vivo and suggest that 7a may be highly effective in the treatment of estrogen-dependent breast cancer and/or in the prophylactic treatment of women with a high risk of breast cancer development.

Biodistribution of a novel antiestrogen (Analog II) in the mouse and rat.

The biodistribution of a novel antiestrogen Analog II was determined in the mouse and rat. The tritiated product, [3H]-Analog II was prepared by New England Nuclear and was purified by preparative chromatography using silica gel and petroleum ether/methylene chloride (80:20). The fat tissue had the highest uptake due to the hydrophobic nature of Analog II. The second highest uptake was in the mouse uterine tissue which was greater than that observed in the rat. The differences in biodistribution between the mouse and rat may partially explain the differences in biological activity of Analog II previously observed in these two animal species.

Synthesis of Air and Light Sensitive Cis-phenolic Cyclopropane Derivatives

Abstract The synthesis of several cis-phenolic cyclopropanes, 6, 7, 8, and 13 is described. These cyclopropanes are new potential antiestrogens. A hydrogenolysis study of precursors, 5b, 5c, 5d, 10 11, and 12 and a novel method for the prevention of gem-dichlorocyclopropane ring-opening during hydrogenolysis are discussed.