雌激素硫酸转移酶诱导剂治疗乳腺癌的机理及其药物动力学/药效动力学研究

As a member of sulfotransferase superfamilies, estrogen sulfotransferase (EST) is a cytosolic enzyme that catalyzes the sulfation reaction of estrogens. Since the sulfated estrogens are not the active forms of estrogens any longer in the breast epithelial cells, sulfation reaction catalyzed by EST plays important roles in the regulation of estrogen levels in vivo. It has been known that an excessively high level of estrogens along with the excessively low level of EST expression in breast epithelial cells could lead to breast cancer proliferation and invasion. Consequently, the molecules that enhance the expression and enzymatic activity of EST (EST inducers) could be crucial for breast cancer treatment, and EST could be used as an effective tumor biomarker. However, applications of EST inducers to breast cancer therapy as well as their modes of actions in cells have not yet been well explored. Based on the promising outcomes of our recent preliminary studies of the inhibitory effects of EST inducers on the proliferation of the MCF-7 cells, we are planning to keep the major focuses of our future research on the studies of (i) cellular mechanisms that underlie the inhibitory effect of EST inducers on the proliferation of human breast cancer cells; and (ii) antitumor activities of an EST inducer on female BALB/c nude mice bearing MCF-7 cell xenografts. Our specific aims in the current research proposal accordingly are (1) to investigate a series of natural and synthetic compounds on their induction of EST in MCF-7 cells and HepG2 cells using real-time PCR, western blot assay, and HPLC as the experimental tools in order to find out the ones that show highly effective induction effect on EST; (2) to investigate the inhibitory effects of these EST inducers on proliferation of human breast cancer cells，and to evaluate the effects of one selected iducer on female BALB/c nude mice bearing MCF-7 cell xenografts as well as to evaluate the pharmacokinetic (PK) and pharmacodynamic (PD) characteristics of this inducer when it is used alone and used in combination with doxorubicin; (3) to elucidate the corresponding anticancer mechanisms on the basis of siRNA technique, flow cytometry analysis, immunohistochemical labelling, and so on; and (4) to establish a mechanism-based PK/PD model to quantitatively describe the correlations among EST levels, plasma concentrations of this inducer, and inhibitory efficiencies of the EST inducer on tumor growth. It is our expectation that the results from conducting the proposed research projects could contribute greatly to (i) the discovery of new tumor biomarke of breast cancer and (ii) the creation of new strategies for the treatments of breast cancer and other estrogen-dependent tumors. In addition, we wish that our future investigations on breast cancer chemotherapy based on EST induction could provide beneficial information to our understanding of the fundamental mechanisms of actions of cancer cells themselves.

雌激素硫酸转移酶(EST)是催化雌激素硫酸化反应以维持体内雌激素平衡的重要代谢酶。雌激素水平过高是乳腺癌产生的重要原因，通过EST诱导而降低雌激素水平对乳腺癌的治疗具有重要意义。EST诱导剂用于乳腺癌治疗的系统研究尚属空白。通过前期工作和文献调研提出假设：EST诱导剂可治疗雌激素依赖性乳腺癌或增加其他抗癌药物的抗癌活性；EST可能成为新的肿瘤生物标记物。本研究拟通过实时PCR等方法确认EST强诱导剂；考察它们单用和与阿霉素联用对乳腺癌细胞增殖的抑制作用；并选择一个强诱导剂用于乳腺癌荷瘤鼠模型，评价其药动学和药效学性质；利用RNA干扰等技术确证机理；建立基于机制的药动学/药效学模型来描述血中诱导剂浓度、EST水平与肿瘤抑制的定量关系，提供EST作为肿瘤标记物以及EST诱导剂抗癌增效的依据。课题将为乳腺癌等雌激素依赖性肿瘤的治疗提供新策略和新的肿瘤标记物，具有重要理论意义和广阔应用前景。

乳腺癌是全世界妇女发病率最高死亡人数最多的癌症。而乳腺癌的发生发展与体内雌激素水平密切相关。过多雌激素会导致乳腺癌细胞增殖和肿瘤生长，而且由雌激素硫酸转移酶(EST)介导的雌激素硫酸化是体内雌激素失活的主要途径，因此诱导EST可降瘤内活性雌激素水平，对乳腺癌的治疗与预防具有重要意义。.本课题研究内容如下：①筛选出具有明显EST诱导活性的两个化合物：地塞米松(DEX)和抗癌新化合物TM208，以此用作EST诱导剂的模型药物；②评价DEX和TM208治疗乳腺癌的体内外药效，并系统研究了EST相关的雌激素调节机制；③建立了测定裸鼠体内DEX和TM208的体内分析方法，并考察其药物动力学(PK)特征；④建立DEX和TM208单独治疗乳腺癌的药物动力学/药效动力学(PK/PD)模型；⑤将DEX和临床上常见化疗药吉西他滨(GEM)联合用于乳腺癌，并考察其联合作用机制，建立两药联用的PK/PD模型，定量说明两者的协同作用，并通过模型仿真对给药方案进行优化；⑥鉴于日益发现的雌激素受体及雌激素在肺癌发生发展中的重要性，我们将TM208和DEX用于非小细胞肺癌（NSCLC）NSCLC的移植瘤模型，也得到了理想疗效，同时验证了EST相关机制，并建立其PK/PD模型，并通过仿真对给药方案进行了优化。.结果证明和发现了雌激素可促进乳腺癌和肺癌得肿瘤生长，DEX和TM208可使细胞与肿瘤内EST从基因、蛋白到活性具有剂量依赖性的增加，而且照组中瘤内雌激素具有随时间增加、EST随时间下降的现象，给药后EST水平显著增加，而瘤内雌激素水平下降到正常乳腺中的激素水平，且循环系统中的雌激素水平也显著下降。这些结果证明了EST介导的雌激素代谢是内分泌治疗的重要组成部分，验证了该课题的研究假设。此外，我们还建立了TM208和DEX在两种肿瘤模型中单用和与化疗药联用的PK/PD模型，定量描述了血药浓度和肿瘤抑制之间的关系，以及通过模型仿真优化了给药方案。.总之，该研究证实了EST在乳腺癌和肺癌治疗及预防中的重要作用，EST诱导剂DEX和TM208对乳腺癌和NSCLC的潜在治疗价值及EST相关机制，以及它们和化疗药物联用的协同增效作用，通过PK/PD模型的建立和仿真，有利于给药方案的优化，从而增加药物疗效并降低其毒副作用。