ALKYL PCDFS--INHIBITION OF MAMMARY CANCER

以前的研究表明，2，3，7，8-四氯二苯并-对- 二恶英（TCDD）抑制了广泛的雌激素诱导的反应， 雌性大鼠子宫和MCF-7人乳腺癌细胞中。 TCDD还 抑制大鼠和小鼠乳腺肿瘤的形成 MCF-7细胞。 6-甲基-1，3，8-三氯二苯并呋喃及相关物质 烷基多氯二苯并呋喃（PCDFs）是相对无毒的类似物， TCDD在体外和体内表现出相当的抗雌激素作用， 抗肿瘤活性。因此，烷基多氯二苯并呋喃代表了一类新的 通过芳香烃（Ah）受体信号起作用的抗雌激素 因此可能具有临床潜力， 用于治疗乳腺癌的化学治疗剂。TCDD和MCDF是 T47 D和MCF-7人乳腺癌细胞中的有效抗雌激素（ER-和 AhR阳性，ER+AhR+），但在ER-AhR- MDA-MB-231中表现出最小活性 细胞烷基PCDFs或TCDD对ER-AhR+的体内外影响 或ER+AhR-细胞以前没有研究过， 这些细胞系的不可用性。该项目的实验1将 彻底表征一系列ER-AhR+和ER+AhR-变异乳腺癌 癌细胞系。ER+AhR-细胞将通过培养MCF-7和 1微摩尔苯并[a]芘（BaP）和ER+AhR-变体中的T47 D细胞 将对细胞系进行充分表征。ER-AhR+变异细胞将被 分离自高传代T47 D细胞和MDA-MB-231细胞（ER-AhR-） 用人arnt基因稳定转染，该基因恢复Ah- 在这个细胞系中。所得稳定的转染子将 代表迄今为止尚未描述的ER-AhR+表型。 此外，由于MDA-MB-231细胞对内分泌和细胞毒性具有抗性， 在药物治疗中，ER-AhR+细胞将作为评估 AhR激动剂（例如烷基化的AhR激动剂）的抗肿瘤和抗增殖作用 PCDFs）在这些细胞中。虽然MDA-MB-231细胞的生长受到抑制， 雌激素非依赖性的各种生长因子（IGF-1，EGF，TGF α， 胰岛素）作为有丝分裂原，因此，生长因子诱导 增殖和ER-AhR+稳定的许多其他特征 将彻底研究转染细胞系。免疫缺陷 移植了乳腺癌细胞的小鼠将用于实验 2至（a）确定控制肿瘤发展的因素， 移植有野生型和变异型乳腺癌的无胸腺小鼠 在实验1和（B）中表征的细胞系评估了相对的 TCDD和烷基PCDFs作为化疗药物的效力， 体内模型。在平行研究中，烷基的相对效力 还将在雌性大鼠子宫中测定多氯二苯呋喃作为抗雌激素的作用。 这些短期研究将提供关键的相对效价数据， 这一系列的烷基多氯二苯并呋喃在一个公认的雌激素反应， 靶器官并优先考虑它们在体内研究中的应用（实验4）， 在DMBA诱导的大鼠肿瘤中选择的同源物的抗原致瘤性 模型这些拟议中的研究将提供关于 AhR激动剂作为抗雌激素和抗肿瘤的作用机制 试剂在两个体内模型，并确定哪些烷基PCDFs应该 进一步研究作为潜在的化疗剂用于治疗 乳腺癌

Previous studies have demonstrated that 2,3,7,8-tetrachlorodibenzo-p- dioxin (TCDD) inhibits a broad spectrum of estrogen-induced responses in the female rat uterus and in MCF-7 human breast cancer cells. TCDD also inhibits mammary tumor formation in rats and in mice transplanted with MCF-7 cells. 6-Methyl-1,3,8-trichlorodibenzofuran (MCDF) and related alkyl polychlorodibenzofurans (PCDFs) are relatively non-toxic analogs of TCDD which exhibit comparable in vitro and in vivo antiestrogenic and antitumorigenic activities. Thus, the alkyl PCDFs represent a new class of antiestrogens which act through the aryl hydrocarbon (Ah) receptor signal transduction pathway and thus may have clinical potential as chemotherapeutic agents for treatment of breast cancer. TCDD and MCDF, are potent antiestrogens in T47D and MCF-7 human breast cancer cells (ER- and AhR-positive, ER+AhR+) but exhibit minimal activity in ER-AhR- MDA-MB-231 cells. The in vitro or in vivo effects of alkyl PCDFs or TCDD on ER-AhR+ or ER+AhR- cells have not been previously investigated due to the unavailability of these cell lines. Experiment 1 of this project will thoroughly characterize a series of ER-AhR+ and ER+AhR- variant breast cancer cell lines. ER+AhR- cells will be isolated by culturing MCF-7 and T47D cells in 1 micromole benzo[a]pyrene (BaP) and the ER+AhR- variant cell lines will be fully characterized. ER-AhR+ variant cells will be isolated from high passage T47D cells and from MDA-MB-231 cells (ER-AhR-) stably transfected with the human arnt gene which restores Ah- responsiveness in this cell line. The resultant stable transfectants will represent an ER-AhR+ phenotype which hitherto has not been described. Moreover, since MDA-MB-231 cells are resistant to endocrine and cytotoxic drug therapy, the ER-AhR+ cells will serve as a model for assessing the antitumorigenic and antiproliferative effects of AhR agonists (e.g. alkyl PCDFs) in these cells. Although the growth of MDA-MB-231 cells are estrogen-independent, various growth factors (IGF-l, EGF, TGFalpha, insulin) act as mitogens and, therefore, growth factor-induced proliferation and a number of other characteristics of the ER-AhR+ stable transfectant cell lines will be thoroughly.investigated. Immune deficient mice transplanted with breast cancer cells will be utilized in Experiment 2 to (a) determine the factors which control the development of tumors in athymic mice transplanted with the wild-type and variant breast cancer cell lines characterized in Experiment 1 and (b) assess the relative potencies of TCDD and the alkyl PCDFs as chemotherapeutic agents in this in vivo model. In parallel studies, the relative potencies of the alkyl PCDF as antiestrogens will also be determined in the female rat uterus. These short-term studies will provide critical relative potency data for this series of alkyl PCDFs in a well-established estrogen-responsive target organ and prioritize their use in in vivo studies (Experiment 4) on the antitumorigenicity of selected congeners in the DMBA-induced rat tumor model. These proposed studies will provide critical new data on the mechanism of action of AhR agonists as antiestrogens and antitumorigenic agents in two in vivo models and determine which alkyl PCDFs should be further investigated as potential chemotherapeutic agents for treatment of mammary cancer.