Pilot Project 2

试点项目2

• 批准号: 10000894
• 负责人: Gayathri Devi
• 金额: $ 1.72万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 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/10000894
• 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; Investigation; 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; x-linked inhibitor of apoptosis protein

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 炎症性乳腺癌（IBC）是乳腺癌中最致命的形式之一，占所有乳腺癌的15%。 乳腺癌死亡人数IBC对处于不利地位的少数民族妇女尤其具有破坏性， 发病率和生存率较差。IBC肿瘤的基因表达谱已经确定了hedgehog（Hh）/GLI 1 在侵袭性和复发性IBC患者中，我们之前已经证明GLI 1 在来源于侵袭性原发性IBC肿瘤的IBC细胞中高度表达， 这些细胞中的GLI 1降低细胞增殖，增加凋亡和减少迁移。的目标 拟议的研究是确定那些对GLI激活和功能至关重要的基因，包括那些 这两个上游和下游的GLI 1，提供深入了解的分子机制，或 驾驶IBC。有待探讨的假设是，GLI 1及其调节的基因促进了生长， IBC的生存，并确定这些基因可能导致新的目标的阐明。长期 目的是为IBC的抗肿瘤疗效提供新的方法。为实现这一目标， 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IBC组织样品。这将由 以下目的：1）鉴定低表达和高表达GLI 1的IBC模型之间的基因表达差异 包括耐药性和GLI 1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）确定。2)为了确认和验证那些 通过qRT-PCR分析的基因表达差异，包括IBC和非IBC细胞系的比较。到 评估这些差异表达基因与非裔美国人患者来源的非IBC细胞的重叠 线我们将评估GLI 1表达和差异表达的靶基因亚组的表达。 在目标1中，从杜克生物标本获取的存档IBC患者和对照组织样本中识别出 通过免疫组织化学和基因表达分析的储存库和处理核心。以限定 对于这些靶基因的功能作用，一个子集将被shRNA敲除或使用 将确定CRISPR/Cas9和对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GLI 1的作用。 IBC。我们在一组功能和表型测定中评估对IBC细胞生物学的影响。这一拟议 研究将进一步促进杜克和NCCU在IBC方面的合作，并将支持开发一种 一个代表性不足的少数民族博士后研究员，专注于GLI 1的转录谱研究 IBC中的激活途径。完成这些目标将提供对后果的机械见解 IBC中GLI 1激活和拮抗作用的下游途径涉及并为新方法提供了信息 IBC的抗肿瘤疗效，长期目标是将IBC的新疗法转化为临床。