p53 Acetylation as a Mechanism in Chemoprevention by Aspirin

p53 乙酰化作为阿司匹林化学预防的机制

基本信息

批准号：
7751188
负责人：
Jayarama B Gunaje
金额：
$ 7.43万
依托单位：
TEXAS TECH UNIVERSITY HEALTH SCIS CENTER
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-07-17 至 2011-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7751188
关键词：
Abnormal DNA Repair Acetylation Affect Aflatoxins Agreement Alkylating Agents Antineoplastic Agents Apoptosis Apoptosis Regulator Aspirin Attention Binding Biological Assay Breast Breast Cancer Cell Camptothecin Cancer cell line Cardiovascular Diseases Cell Cycle Cell Cycle Arrest Cell Cycle Progression Cell Death Cell Survival Cells Chemoprevention Chemopreventive Agent Chemosensitization Cisplatin Clinical Clinical Research Colon Carcinoma DNA Damage Degenerative polyarthritis Development Dose Down-Regulation Doxorubicin Drug usage Electrophoretic Mobility Shift Assay Epidemiology Epithelial Epithelial Cells Exposure to Flow Cytometry Fluorouracil Food Gene Expression Gene Targeting Genes Genetic Transcription Headache Hepatocyte Heterocyclic Amines Human Hydrocarbons In Vitro Injury Intake Kinetics Leucovorin Literature Lung Lysine MCF7 cell Malignant Neoplasms Mass Spectrum Analysis Measures Mediating Messenger RNA Minor Mitochondria Modeling Molecular Nitrosamines Normal Cell Outcome Oxidation-Reduction Oxidoreductase Pathway interactions Patients Pattern Peptide Hydrolases Peripheral Blood Mononuclear Cell Pharmaceutical Preparations Pilot Projects Preventive Property Prostaglandin-Endoperoxide Synthase Prostate Protein p53 Proteins Proto-Oncogenes Reaction Reporting Resistance Reverse Transcriptase Polymerase Chain Reaction Running Site Staging Stress TP53 gene Time Tissues Transferase Tumor Suppressor Proteins base cancer cell cancer therapy carcinogenesis cell type cigarette smoking cytotoxic design environmental chemical improved irinotecan neoplastic cell novel oncoprotein p21 pre-clinical preclinical study prevent promoter prophylactic protective effect research study response transcription factor treatment strategy tumor

项目摘要

DESCRIPTION (provided by applicant): A vast amount of epidemiological, preclinical and clinical studies have revealed aspirin as a promising chemopreventive agent, particularly in epithelial carcinogenesis. Despite the wide attention inhibition of cyclooxygenases has received, it is clear that aspirin elicits a myriad of molecular effects that counteract the carcinogenic episodes. Since aspirin's protective effect was mainly observed in epithelial cell types which are more resistant to chemotherapeutic efforts, an urgent need exists to dissect and identify the primary targets and cancer preventive pathways affected by aspirin. In preliminary studies, we have obtained the first and strong evidence for a dose- and time-dependent acetylation of p53 tumor suppressor protein by aspirin in MDA-MB-231 human breast cancer cells, several cancer cells belonging to different tumor types and also in normal liver cells. In MDA-MB-231 cells, aspirin induced the levels of p53 target genes namely p21CIP1, a protein involved in cell cycle arrest, and Bax, a proapoptotic protein; however, p21 induction was transient (1-12h); where as, induction of Bax was sustained (24 h). Interestingly, in DNA damaged cells (induced by camptothecin), aspirin treatment (24 h) inhibited the p21 induction, while the Bax induction was unaffected. Built on these findings, the central hypothesis of this R03 pilot project is that aspirin-induced multi-site acetylation of p53 alters its transcription factor function by shifting the gene expression spectrum from those that elicit cell cycle arrest / prosurvival properties to those that promote and drive cell death. Since deletion of p21 gene has been previously shown to increase the sensitivity of cells towards apoptosis, our observation that aspirin inhibits p21 suggests a potential mechanism by which it may exert anti-cancer effects in DNA damaged cells. The studies proposed in this application will determine the mechanisms by which aspirin regulates apoptosis in DNA damaged cells via inhibition of p21. We will use MDA-MB-231 and MCF-7 breast cancer cells as well as normal human Peripheral Blood Mononuclear Cells in our study. The experiments in Aim 1 will investigate the molecular basis of aspirin-mediated inhibition of p21 using real time RT-PCR, electrophoretic mobility shift assays, and run on transcription assays. We will also identify aspirin-induced acetylation sites on p53. In Aim II, we will determine the ability of aspirin to augment apoptosis in cells exposed to DNA damaging drugs by clonogenic cell survival assays and flow cytometry. In addition to camptothecin, all studies will be extended to include doxorubicin and cisplatin, to determine if aspirin also modulates p21 / Bax expression by these DNA damaging drugs. These studies will provide a novel mechanism by which aspirin may exert anticancer effects in DNA damaged cells via acetylation of p53, induction of Bax and inhibition of p21.

描述（由申请人提供）：大量的流行病学，临床前和临床研究表明，阿司匹林是一种有希望的化学预防剂，特别是在上皮癌中。尽管已广泛关注环加氧酶，但显然阿司匹林引起了无数的分子作用来抵消致癌事件。由于阿司匹林的保护作用主要是在对化学治疗努力更具抵抗力的上皮细胞类型中观察到的，因此存在迫切需要剖析和鉴定受阿司匹林影响的主要靶标和癌症预防途径。在初步研究中，我们获得了阿司匹林在MDA-MB-231人类乳腺癌细胞中对p53肿瘤抑制蛋白的剂量和时间依赖性乙酰化的第一个且有力的证据，这是几种属于不同肿瘤类型以及正常肝细胞中的癌细胞。在MDA-MB-231细胞中，阿司匹林诱导p53靶基因的水平，即p21CIP1，一种参与细胞周期停滞的蛋白质，而Bax是促凋亡蛋白的Bax。但是，p21诱导是瞬态（1-12H）；在其中，持续的BAX诱导（24小时）。有趣的是，在DNA受损的细胞（由Camptothecin诱导）中，阿司匹林治疗（24小时）抑制了P21诱导，而Bax诱导不受影响。建立在这些发现的基础上，该R03试点项目的中心假设是，阿司匹林诱导的p53的多站点乙酰化通过将基因表达谱从引起细胞周期停滞 / prosurvival特性转移到促进和驱动细胞死亡的人来改变其转录因子的功能。由于先前已证明p21基因的缺失会增加细胞对凋亡的敏感性，因此我们对阿司匹林抑制p21的观察结果表明，它可能在DNA受损细胞中发挥抗癌作用的潜在机制。在本应用中提出的研究将确定阿司匹林通过抑制P21调节DNA受损细胞中凋亡的机制。在我们的研究中，我们将使用MDA-MB-231和MCF-7乳腺癌细胞以及正常的人外周血单核细胞。 AIM 1中的实验将使用实时RT-PCR，电泳迁移率转移测定法研究阿司匹林介导的p21抑制的分子基础，并在转录测定中运行。我们还将在p53上鉴定阿司匹林诱导的乙酰化位点。在AIM II中，我们将确定阿司匹林通过克隆性细胞存活分析和流式细胞术中暴露于DNA损害药物的细胞中凋亡的能力。除了甲状腺激素外，所有研究还将扩展到包括阿霉素和顺铂，以确定阿司匹林是否还通过这些DNA损害药物调节p21 / bax表达。这些研究将提供一种新的机制，通过该机制，阿司匹林通过p53的乙酰化，诱导BAX和抑制p21在DNA受损细胞中发挥抗癌作用。