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Pilot Project 2

试点项目2

基本信息

批准号：
10000893
负责人：
Gayathri Devi
金额：
$ 2.54万
依托单位：
DUKE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
资助国家：
美国
起止时间：
至
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10000893
关键词：
Adult African American Age Apoptosis Archives Automobile Driving Biological Biological Assay Breast Cancer Cell Breast Cancer Model Breast Cancer Patient Breast Cancer cell line CRISPR/Cas technology Cancer Biology Cancer Control Cancer Patient Cancer cell line Cell Line Cell Proliferation Cell model Cells Cellular biology Cessation of life Clinic Collaborations Data Development Disadvantaged Disease Down-Regulation Drug resistance Embryonic Development Erinaceidae Exhibits FRAP1 gene GLI gene Gene Amplification Gene Expression Gene Expression Profile Gene Expression Profiling Genes Genetic Genetic Transcription Goals Growth Human Immunohistochemistry Incidence Income Joints Knock-out Lead Link Malignant Neoplasms Mediating Minority Mission Modeling Molecular Outcome Pathway interactions Patients Pharmacology Phenotype Pilot Projects Population-Based Registry Postdoctoral Fellow Prevalence Public Health Quantitative Reverse Transcriptase PCR Race Relapse Reporting Research Reverse Transcriptase Polymerase Chain Reaction Role Sampling Signal Transduction Testing Therapeutic Time Tissue Sample Translating Translational Research Underrepresented Minority United States National Institutes of Health Validation Woman Work Xenograft procedure advanced breast cancer anticancer research cancer health disparity cancer therapy cell growth cell motility clinical practice differential expression genetic signature genetically modified cells health care availability inflammatory breast cancer inhibitor/antagonist innovation insight knock-down malignant breast neoplasm migration mortality novel novel strategies novel therapeutic intervention novel therapeutics programs repository response screening small hairpin RNA therapeutic target transcription factor transcriptome sequencing tumor tumorigenesis

项目摘要

PROJECT ABSTRACT Inflammatory breast cancer (IBC), one of the most lethal forms of breast cancer, accounts for ~15% of all breast cancer deaths. IBC is particularly devastating in disadvantaged minority women who have both a higher incidence and poorer survival. Gene expression profiling of IBC tumors has identified the hedgehog(Hh)/GLI1 pathway as higher in patients with aggressive and relapsing IBC. We have previously demonstrated that GLI1 is highly expressed in IBC cells derived from an aggressive primary IBC tumor, and that down-regulation of GLI1 in these cells decreased cell proliferation, increased apoptosis and reduced migration. The objective of the proposed research is to identify those genes that are critical for GLI activation and function, including those that are both up- and downstream in of GLI1, to provide insight into the molecular mechanisms underlying or driving IBC. The hypothesis to be explored is that GLI1 and the genes that it regulates promote the growth and survival of IBC, and that identifying these genes may lead to the elucidation of novel targets. The long-term goal is to inform novel approaches for anti-tumor efficacy in IBC. This objective will be met by applying an innovative approach that includes utilization of high-throughput transcriptome sequencing (RNA-Seq), genetically engineered cell lines and human IBC tissue samples. This will become accomplished by the following Aims: 1) To identify gene expression differences between low and high GLI1 expressing IBC models including drug resistant and GLI1-antagonist treated IBC models. Differences in gene expression patterns will be determined by high-throughput transcriptome sequencing (RNA-Seq). 2) To confirm and validate those gene expression differences by qRT-PCR analysis including a comparison of IBC and non-IBC cell lines. To assess overlap of those differentially expressed genes with African American patient derived non-IBC cell lines. We will assess GLI1 expression and the expression of a subset of differentially expressed target genes identified in Aim 1 in archival IBC patient and control tissue samples accessed from the Duke Biospecimen Repository and Processing Core by immuno-histochemistry, and gene expression analysis. To define functional roles for these target genes, a subset will be knocked down by shRNA or knocked out using CRISPR/Cas9 and effects on IBC growth and survival will be determined. 3) Selective pharmacological inhibitors will be used to assess the role of previously identified pathways in activating GLI1 in IBC and non- IBC. We assess effects on IBC cell biology in a panel of functional and phenotypic assays. This proposed research will further the collaboration between Duke and NCCU on IBC and will support the development of an under-represented minority postdoctoral fellow performing research focused on transcriptional profiling of GLI1 activation pathways in IBC. Completion of these Aims will provide mechanistic insights into the consequences of GLI1 activation and antagonism in IBC, the downstream pathways involved and inform novel approaches for anti-tumor efficacy in IBC with the long term goal to translate new therapies for IBC into the clinic.

项目摘要炎性乳腺癌 (IBC) 是最致命的乳腺癌形式之一，约占所有乳腺癌的 15% 乳腺癌死亡。 IBC 对处境不利的少数族裔女性尤其具有破坏性，因为她们都拥有较高的收入。发生率和较差的生存率。 IBC 肿瘤的基因表达谱鉴定出刺猬蛋白 (Hh)/GLI1 侵袭性和复发性 IBC 患者的通路较高。我们之前已经证明了 GLI1 在源自侵袭性原发性 IBC 肿瘤的 IBC 细胞中高表达，并且下调这些细胞中的 GLI1 减少了细胞增殖，增加了细胞凋亡并减少了迁移。的目标拟议的研究是为了识别那些对 GLI 激活和功能至关重要的基因，包括那些位于 GLI1 的上游和下游，以深入了解潜在的分子机制或驾驶IBC。要探索的假设是 GLI1 及其调节的基因促进生长和 IBC 的存活，并且识别这些基因可能会导致新靶标的阐明。长期来看目标是提供 IBC 抗肿瘤功效的新方法。这一目标将通过应用创新方法，包括利用高通量转录组测序（RNA-Seq），基因工程细胞系和人类 IBC 组织样本。这将通过以下目标：1) 鉴定低表达 GLI1 和高表达 GLI1 的 IBC 模型之间的基因表达差异包括耐药和 GLI1 拮抗剂治疗的 IBC 模型。基因表达模式的差异将通过高通量转录组测序（RNA-Seq）来确定。 2) 确认并验证这些通过 qRT-PCR 分析基因表达差异，包括比较 IBC 和非 IBC 细胞系。到评估这些差异表达基因与非裔美国患者来源的非 IBC 细胞的重叠线。我们将评估 GLI1 表达和差异表达靶基因子集的表达在从杜克生物样本获取的档案 IBC 患者和对照组织样本中在目标 1 中鉴定出通过免疫组织化学和基因表达分析建立存储库和处理核心。定义这些目标基因的功能作用，其中一个子集将被 shRNA 敲除或使用将确定 CRISPR/Cas9 以及对 IBC 生长和存活的影响。 3）选择性药理抑制剂将用于评估先前确定的途径在 IBC 和非 IBC 中激活 GLI1 的作用国际散文委员会。我们通过一组功能和表型测定评估对 IBC 细胞生物学的影响。这个提议研究将进一步推进杜克大学和国立政治大学之间在 IBC 方面的合作，并支持开发代表性不足的少数族裔博士后研究员，从事 GLI1 转录谱研究 IBC 中的激活途径。完成这些目标将为后果提供机制性的见解 GLI1 在 IBC 中的激活和拮抗作用、涉及的下游途径并为新方法提供信息 IBC 的抗肿瘤功效，长期目标是将 IBC 的新疗法转化为临床。