Mechanistic Studies on Lung Cancer Prevention by Tea Polyphenols

我们的长期目标是了解茶叶成分的癌症预防活动及其在人类癌症预防中的应用。茶多酚的防癌作用已在动物模型中得到证实，但其作用机制尚不清楚。本项目旨在阐明(-)-表没食子儿茶素-3-没食子酸酯（EGCG）预防肺癌的机制，EGCG是主要的绿茶多酚。我们的目标是建立一个综合的体外和体内实验系统来验证我们的中心假设，即EGCG通过与对癌变至关重要的特定靶分子结合来抑制肺癌的发生，这种结合触发信号转导途径的改变，从而导致癌前和恶性细胞的生长抑制和凋亡。为了有效地验证我们的假设，我们计划同时使用体外和体内实验系统。具体目标如下：

Our long-term goal is to understand the cancer preventive activities of tea constituents and their applications to human cancer prevention. The cancer preventive activity of tea polyphenols has been demonstrated in animal models, but the mechanisms of action are not clearly understood. This project aims to elucidate the mechanisms of lung cancer prevention by (-)-epigallocatechin-3-gallate (EGCG), the major green tea polyphenol. Our goal is to establish an integrated in vitro and in vivo experimental system to test our central hypothesis that EGCG inhibits lung carcinogenesis by binding to specific target molecules which are vital to carcinogenesis and this binding triggers alterations in signal transduction pathways that lead to growth inhibition and apoptosis of premalignant and malignant cells. In order to effectively test our hypothesis, we plan to use both in vitro and in vivo experimental systems. The specific aims are as follows: 1. Establish and compare in vivo (xenograft & allograft) and in vitro experimental systems with lung cancer cell lines and study the mechanisms of action of EGCG. Lung cancer cell lines H1299 (human) and CL13 and CL30 (mouse) will be studied in culture and in xenografts/allografts. The effects of EGCG treatment on tumor growth, apoptosis, angiogenesis, and related molecular changes (e.g., ERK1/2, AKT, ASK1, JNK, VEGF, and oxidative stress parameters) will be compared in vivo vs. in vitro. 2. Identify direct molecular targets of EGCG by affinity-proteomics and functional studies. To identify direct molecular targets for EGCG, we will use affinity chromatography and mass spectrometry in collaboration with Dr. Zigang Dong at the University of Minnesota. The functional roles of the putative EGCG target proteins in mediating the action of EGCG will be studied with loss or gain of function experiments in engineered cells and in a xenograft/allograft model established in Aim 1. 3. Elucidate the mechanisms of cancer prevention by EGCG in a 4-(methylnitrosamino)-1-(3-pyridyl)-1- butanone (NNK)-induced lung carcinogenesis model in A/J mice. Adenoma-bearing mice will receive long- or short-term treatment with EGCG. The cellular and molecular changes related to the inhibition of tumor progression will be investigated using immunohistochemistry, Western blots, and other approaches. We will test specific hypotheses developed based on results from Aims 1 and 2 concerning putative EGCG molecular targets and related molecular events. A better understanding of the mechanisms of the cancer preventive action of EGCG and the dose- response relationship will help us to design future lung cancer prevention trials in humans, for example, in ex-smokers.