MECHANISM OF GLUCARATE INHIBITION OF MAMMARY CANCER

本提案的主要目的是检验假设 β-葡糖醛酸酶抑制剂D-葡糖醛酸的形成， D-葡萄糖二酸钙中的1，4-内酯（1，4-GL）是至关重要的 D-葡萄糖二酸抑制乳腺癌的机制， 1，4-GL介导的促乳腺激素水平和DNA调节 合成至少部分负责有效的 D-葡萄糖二酸盐的化学预防作用。 建立良好的模型， 在雌性Sprague-Dawley大鼠中诱导乳腺癌发生， 单次皮下将使用一定剂量的N-甲基亚硝基脲（MNU）。 具体 目的包括：（1）建立D-葡萄糖酸 酸/D-葡糖二酸/D-葡糖内酯家族，1，4-GL是最终的 致癌抑制剂，通过测试其在MNU诱导的 大鼠乳腺癌发生模型，使用非β- 葡萄糖醛酸酶，D-葡糖醛酸-6，3-内酯（6，3-GL）作为阴性对照。 在 为了克服已知的1，4-GL在体内的不稳定性，一些 将使用1，4-GL和6，3GL的衍生物。 不同 这些衍生物的浓度将被并入半- 纯化的AIN-76A饮食。 改良后的饲料将喂给大鼠 MNU后持续一周开始，以避免任何饮食影响 在开始之前乳腺的发育。 建议的作用 D-葡萄糖二酸钙作为1，4-GL的前体将得到进一步证实 通过研究代谢、药代动力学和处置， 使用非放射性和14C标记的大鼠中D-葡萄糖二酸钙 化合物. (ii)确定全身和局部效应 1，4-GL，连续饲喂幼龄雌性Sprague-Dawley大鼠， 在MNU治疗前早期，即在促乳腺激素水平上， 雌激素受体与乳腺增生状态的关系 时间点和最终致瘤反应。 的组合 [3H]胸腺嘧啶核苷放射自显影和免疫组化染色 β-葡萄糖醛酸酶或雌激素受体，将用于研究 B-葡萄糖醛酸酶和/或雌激素受体之间的潜在相关性 表达和DNA合成。(iii)证明了荷尔蒙 置换对大鼠D-glucaro-1，4-lactone抑制作用无影响 乳腺癌 患有MNU诱导的乳腺肿瘤的大鼠将被 切除垂体并用雌二醇和催乳素或赋形剂处理， 同时饲喂含和不含D-葡糖-1，4-内酯的饲料。 变化 肿瘤的大小将与激素受体状态相关。

The primary objective of this proposal will be to test the hypothesis that formation of the beta-glucuronidase inhibitor, D-glucaro- 1,4-lactone (1,4-GL) from calcium D-glucarate, is of critical importance for the mechanism of D-glucarate inhibition of mammary cancer and that 1,4-GL-mediated modulation of mammotropic hormone levels and DNA synthesis is at least partially responsible for the potent chemopreventive effect of D-glucarate. The well established model of mammary carcinogenesis induced in female Sprague-Dawley rats with a single s.c. dose of N-methylnitrosourea (MNU) will be used. Specific Aims include: (i) Establishment of the theory that in D-glucaric acid/D-glucarate/D-glucarolactones family, 1,4-GL is the ultimate inhibitor of carcinogenesis by testing its efficacy in the MNU-induced rat mammary carcinogenesis model, with a non-inhibitor of beta- glucuronidase, D-glucaro-6,3-lactone (6,3-GL) as negative control. In order to overcome the known instability of 1,4-GL in vivo, some derivatives of 1,4-GL and 6,3GL will be utilized. Different concentrations of these derivatives will be incorporated to the semi- purified AIN-76A diet. The modified diets will be fed to rats continually beginning one week after MNU to avoid any dietary effects on the development of mammary gland prior to initiation. The suggested role of calcium D-glucarate as a precursor of 1,4-GL will be further confirmed by investigation of the metabolism, pharmacokinetics and disposition of calcium D-glucarate in the rat using both non-radioactive and 14C-labeled compounds. (ii) Determination of the systemic and local effects of 1,4-GL, fed continually to young female Sprague-Dawley rats beginning early before MNU treatment, i.e. on mammotropic hormones levels, the estrogen receptor and proliferative status of mammary gland at different time-points, and on the final tumorigenic response. A combination of [3H]thymidine autoradiography and immunohistochemical staining for beta-glucuronidase or estrogen receptor, will be used to investigate a potential correlation between B-glucuronidase and or estrogen receptor expression and DNA synthesis. (iii) Demonstration that hormone replacement has no effect on D-glucaro-1,4-lactone inhibition of rat mammary cancer. The rats with MNU-induced mammary tumors will be hypophysectomized and treated with estradiol and prolactin or vehicle, while fed diets with and without D-glucaro- 1,4-lactone. Changes in the size of tumors will be correlated with the hormone receptor status.