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Development Project 2

开发项目2

基本信息

批准号：
8744896
负责人：
BETI THOMPSON
金额：
$ 10.81万
依托单位：
FRED HUTCHINSON CANCER RESEARCH CENTER
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-09-25 至 2018-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8744896
关键词：
Affect African American Aliquot Anthracyclines Antibodies Apoptosis Apoptotic Attention Biochemical Bioinformatics Biological Assay Biological Markers Breast Breast Cancer Cell Bromodeoxyuridine Bypass Cancer Biology Cancer Model Cancer Patient Cancer cell line Candidate Disease Gene Cardiotoxicity Cell Cycle Cell Cycle Arrest Cell Cycle Checkpoint Cell Cycle Deregulation Cell Cycle Progression Cell Cycle Regulation Cell Death Cell Line Cell Proliferation Cells Chromosomal Instability Cleaved cell Clinical Clinical Treatment Consult Coupled Cyclin A Cyclin B DNA DNA Damage DNA Repair DNA biosynthesis DNA repair protein DNA-PKcs DNA-dependent protein kinase Data Data Analyses Detection Development Disease Dose Double-Blind Method Doxorubicin Doxycycline Drug Efflux Drug Targeting Ensure Environment Epidermal Growth Factor Receptor Epithelial Cells Evaluation Event Fiber Fibroblasts Flow Cytometry Fluorescence Freezing Gene Expression Gene Silencing Gene Targeting Genes Genetic Grant Head and Neck Squamous Cell Carcinoma Histology Human Immunoblot Analysis Immunoblotting Immunodeficient Mouse Immunohistochemistry In Vitro Individual Kinetochores Knowledge Laboratories Latina Lead Length Lentivirus Vector Letters M Phase Arrest M cell Maintenance Malignant neoplasm of ovary Mammary Neoplasms Measures Microscopy Mitosis Mitotic Molecular Mus Mutate Mutation Necrosis Neuroblastoma Normal Cell Outcome P-Glycoproteins PRKDC gene Pathway interactions Patients Phosphotransferases Pre-Clinical Model Preclinical Testing Promega Propidium Diiodide Protein p53 Proteins Protocols documentation Publishing RNA Randomized Reader Reagent Receptor Signaling Recurrence Reporting Research Research Personnel Resected Resistance Resolution Role Running S Phase Scientist Signal Transduction Small Interfering RNA Staging Staining method Stains Stress System Testing Tetanus Helper Peptide The Cancer Genome Atlas Therapeutic Time Topoisomerase II Topoisomerase Inhibitors Toxic effect Transfection Translating Travel Treatment Failure Tumor Suppressor Proteins Validation Woman Xenograft procedure anticancer research arm base cancer cell cancer therapy cancer type career development caspase-3 caspase-9 cell type chemotherapy clinical care cytotoxicity drinking water early onset experience genome sequencing graduate student improved in vivo insight interest luminescence malignant breast neoplasm matrigel micronucleus mortality mutant novel outcome forecast overexpression penis foreskin prevent promoter protein expression research study response screening small hairpin RNA therapeutic target therapy design triple-negative invasive breast carcinoma tumor tumor growth

项目摘要

Doxorubicin is one of the most effective and widely used chemotherapy agents for breast cancer. However, resistance to this anthracycline agent is common, leading to treatment failure and poor prognosis. Triple negative breast cancer (TNBC), a clinical subtype type seen disproportionately in African American and Latina women (1, 2), is characterized by higher recurrence and lower overall survival following anthracycline treatment (1, 3). Developing targeted therapies for TNBC, especially those that do not respond to doxorubicin, is the most urgent priority for the clinical treatment of this aggressive disease (4). Several mechanisms have been implicated in doxorubicin resistance, including an increase in drug efflux pathways (5), epidermal growth factor receptor signaling (6), and mutations in the tumor suppressors ATM or p53 (7-9), but these findings have not been widely translated for clinical benefit. Recent studies from whole genome sequencing of over 1,000 breast cancers reveal that the tumor suppressor p53 is mutated in 43-62% (10, 11), making it one the most commonly mutated genes in this cancer type. p53 is required for DNA damage induced apoptosis, so therapies designed to increase the sensitivity of p53 mutant breast cancer cells to genotoxic therapy would be immensely beneficial. We recently reported that the DNA repair protein DNAPK regulates p53 independent apoptosis (12), pointing to a novel pathway to sensitize p53 mutant tumors. To identity additional genes that regulate p53 independent cell death, we performed a kinome wide siRNA doxorubicin sensitizer screen with p53 mutant cancer cells. The doxorubicin survival genes identified function in G2/M cell cycle regulation, DNA repair, and apoptosis. Based on these findings, we hypothesize that targeting these doxorubicin survival genes will lead to p53 independent apoptotic or mitotic cell death and will sensitize breast tumors to doxorubicin. Our partnership combines expertise in DNA repair, cancer biology, and functional genetics together with the outstanding research environments of the FHCRC and NMSU. Our objectives in this pilot proposal are: 1. Identify mechanisms leading to p53 independent cell death. We will determine if depletion of doxorubicin survival genes exacerbates DNA damage and increases apoptosis in doxorubicintreated p53 deficient triple negative breast cancer cells. We will determine if silencing doxorubicin survival genes affects cell cycle progression and DNA replication, and assess the mechanism of progression into mitotic cell death. 2. Validate candidate therapeutic targets in preclinical models of breast cancer. In parallel with Aim 1, we will determine if knockdown of doxorubicin survival genes in breast cancer xenografts improves response to doxorubicin. The outcome will be mechanistically and preclinically validated targets for doxorubicin resistant TNBC. Knowledge ofthe pathways that control sensitivity to doxorubicin will identity new candidate drug targets for breast cancer therapy, and will point to new biomarkers to stratify patients and inform clinical care. Because doxorubicin resistance leads to treatment failure and subsequent mortality, these findings will elucidate new strategies to treat these unresponsive aggressive tumors.

阿霉素是目前治疗乳腺癌最有效、应用最广泛的化疗药物之一。然而，对这种蒽环类药物的耐药性很常见，导致治疗失败和预后不良。三阴性乳腺癌(TNBC)是一种临床亚型，在非裔美国人和拉丁裔妇女中比例较高(1，2)，其特点是在使用蒽环类药物治疗后复发率较高，总存活率较低(1，3)。开发针对TNBC的靶向治疗，特别是那些对阿霉素无效的治疗，是这种侵袭性疾病临床治疗的最紧迫的优先事项(4)。阿霉素耐药有几个机制，包括药物外排途径增加(5)，表皮生长因子受体信号转导增加(6)，以及肿瘤抑制基因ATM或P53突变(7-9)，但这些发现尚未被广泛转化为临床益处。最近对1000多例乳腺癌全基因组测序的研究表明，抑癌基因P53在43-62%(10，11)中发生突变，使其成为乳腺癌中最常见的突变基因之一。P53是DNA损伤诱导细胞凋亡所必需的，因此提高P53突变的乳腺癌细胞对基因毒性治疗的敏感性的治疗将是非常有益的。我们最近报道了DNA修复蛋白DNAPK调节P53非依赖性细胞凋亡[12]，指出了一条使P53突变肿瘤增敏的新途径。为了确定其他调控p53非依赖性细胞死亡的基因，我们对p53突变的癌细胞进行了动态组广泛的siRNA阿霉素增敏剂筛选。阿霉素生存基因在G2/M细胞周期调控、DNA修复和细胞凋亡中发挥作用。基于这些发现，我们假设靶向这些阿霉素生存基因将导致p53非依赖性的凋亡或有丝分裂细胞死亡，并将使乳腺肿瘤对阿霉素敏感。我们的合作伙伴关系将DNA修复、癌症生物学和功能遗传学方面的专业知识与FHCRC和NMSU出色的研究环境结合在一起。我们在这个试点方案中的目标是：1.确定导致P53非依赖性细胞死亡的机制。我们将确定阿霉素生存基因的缺失是否会加剧阿霉素治疗的p53缺陷型三阴性乳腺癌细胞的DNA损伤和增加细胞凋亡。我们将确定沉默阿霉素生存基因是否会影响细胞周期进程和DNA复制，并评估进展为有丝分裂细胞死亡的机制。2.在乳腺癌临床前模型中验证候选治疗靶点。与目标1同时，我们将确定敲除乳腺癌异种移植瘤中的阿霉素生存基因是否改善了对阿霉素的反应。其结果将是对阿霉素耐药的TNBC进行机械和临床前验证的靶点。对控制阿霉素敏感性的途径的了解将确定乳腺癌治疗的新候选药物靶点，并将指出新的生物标记物来对患者进行分层并指导临床治疗。由于阿霉素耐药性导致治疗失败和随后的死亡，这些发现将阐明治疗这些无反应的侵袭性肿瘤的新策略。