Genomics and Proteomics of Head and Neck Cancer

头颈癌的基因组学和蛋白质组学

• 批准号: 7967002
• 负责人: CARTER VAN WAES
• 金额: $ 59.6万
• 依托单位: NATIONAL INSTITUTE ON DEAFNESS AND OTHER COMMUNICATION DISORDERS
• 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/7967002
• 关键词: Adverse effects; Affect; Alcohols; Apoptosis; Apoptotic; Attenuated; Binding; Bioinformatics; Biological Assay; Biological Markers; Biology; Biotechnology; Cancer cell line; Cell Nucleus; Clinical Trials; Cluster Analysis; Collaborations; Cytokine Gene; Deglutition; Development; Diagnosis; Epidermal Growth Factor Receptor; Epithelial; Event; Exposure to; Gefitinib; Gene Cluster; Gene Expression; Gene Expression Profiling; Gene Proteins; Gene Targeting; Genes; Genetic; Genetic Transcription; Genome; Genomics; Goals; Growth; Head and Neck Cancer; Head and Neck Squamous Cell Carcinoma; Human; Human Papillomavirus; In Situ Hybridization; Inflammatory; Laboratories; Larynx; Link; MAP Kinase Gene; Malignant - descriptor; Malignant Neoplasms; Manuscripts; Modeling; Mus; Mutation; NF-kappa B; Neoplasm Metastasis; Normal tissue morphology; Oral cavity; Paclitaxel; Pathogenesis; Pathology; Pathway interactions; Patients; Pharyngeal structure; Phenotype; Phosphoproteins; Play; Preparation; Prevalence; Prevention; Prevention therapy; Preventive Intervention; Protein Microchips; Proteins; Proteomics; Proto-Oncogene Proteins c-akt; Quality of life; Radiation; Radiosurgery; Recurrent Malignant Neoplasm; Regulation; Regulatory Element; Regulon; Reporting; Resistance; Reverse Transcription; Role; STAT3 gene; Signal Pathway; Signal Transduction; Site; Small Interfering RNA; Specimen; Speech; Systems Biology; TNF gene; TP53 gene; Technology; Therapeutic; Tobacco; Transforming Growth Factor beta; Tumor Suppressor Genes; Voice; Yang; activating transcription factor; cDNA Arrays; carcinogenesis; chemotherapy; cytokine; inhibitor/antagonist; malignant phenotype; member; novel; novel strategies; programs; promoter; protein expression; receptor; response; response to injury; small molecule; therapy resistant; transcription factor

We previously showed using microarray profiling biotechnology that stepwise transformation and metastatic progression of SCC in a murine model results in the expression of gene programs related to the signal transcription factor NF-kappaB. Inhibition of NF-kappaB modulated over half the up or down-regulated genes differentially expressed, and attenuated the malignant phenotype, indicating it may be a critical target for prevention or therapy of head and neck cancer. Gene expression profiling and bioinformatic analysis of the promoters of gene clusters differentially expressed in human HNSCC provided evidence for increased prevalence of binding motifs for NF-kappaB as well as other signal transcription factors, such as p53, AP-1, STAT3 and EGR-1 (Yan et al, Genome Biology, 2007). NF-kappaB, p53, AP-1, STAT3 and EGR-1 activation has previously been associated with pathogenesis and therapeutic resistance, and the subsets expressing wt or mt 53 have been reported to differ in response to chemotherapy. These observations suggested the hypothesis that key alterations in a network of signal transcription factors can interact in determining gene expression and development of HNSCC of differing malignant potential and sensitivity or resistance to therapy. Next, we took a systems biology approach to define NF-kappaB regulons, interacting signal pathways and networks in the malignant phenotype of head and neck cancer cell lines differing in p53 status(Yan, Genome Biology, 2008). Unique gene signatures expressed by human HNSCC lines were identified by cDNA microarray, principal components, and cluster analyses and validated by quantitative reverse transcription-PCR (RT-PCR) and in situ hybridization. An inverse relationship between activation of NF-kB, p53 mutation, and inactivation of TGFbeta pathway and target genes suggested the hypothesis that activation of NF-kB may be linked to alteration of p53 and TGFbeta signaling. We discovered that mt p53 represses TGFbeta receptor II expression, attenuating TGF beta signaling, enhancing inflammatory cytokine responsiveness and activation of NF-kB in HNSCC (Cohen et al, Cancer Res 2009). Another relationship identified was between NF-kB member c-REL and p53 members p53, p63 and p73. This led to demonstration that cytokine TNF induced cREL interacts with p63, displacing p73 from growth arrest and apoptotic genes and the nucleus of HNSCC (Lu et al, manuscript submitted). siRNA knockdown of p63 revealed that it down regulates p53 regulated tumor suppressor genes and upregulated numerous NF-kB related genes, through interaction with c-REL. Novel p63, cREL as well as classical p53 and NF-kB sites were defined in these genes and validated by ChIP assay (Yang et al, manuscript in preparation). ChIP sequencing of global gene expression regulated by cREL, p63 and p73 in HNSCC is underway in collaboration with Laboratory of Genetics, NCI. In collaboration with the NCI and Department of Pathology, new proteomic technology, using protein microarrays, was used to analyze EGFR, NF-kB, MAPK, AKT and STAT3 signal phosphoprotein inhibition, and markers of apoptosis in protein extracts from HNSCC lines and specimens from patients in a clinical trial with small molecule inhibitor gefitinib, taxol and radiation.

我们先前使用微阵列分析生物技术表明，在小鼠模型中SCC的逐步转化和转移进展导致与信号转录因子NF-κ B相关的基因程序的表达。抑制NF-kappaB可调节超过一半的上调或下调基因的差异表达，并减弱恶性表型，表明其可能是预防或治疗头颈癌的关键靶点。在人HNSCC中差异表达的基因簇的启动子的基因表达谱分析和生物信息学分析提供了NF-κ B以及其他信号转录因子如p53、AP-1、STAT 3和EGR-1的结合基序的普遍性增加的证据（Yan等人，Genome Biology，2007）。NF-κ B、p53、AP-1、STAT 3和EGR-1的激活以前已与发病机制和治疗抗性相关，并且已报道表达wt或mt 53的亚群在对化疗的反应中不同。这些观察结果表明，信号转录因子网络中的关键改变可以在决定不同恶性潜能和对治疗的敏感性或抗性的HNSCC的基因表达和发展中相互作用。 接下来，我们采用系统生物学方法来定义NF-κ B调节子，在p53状态不同的头颈癌细胞系的恶性表型中相互作用的信号通路和网络（Yan，Genome Biology，2008）。通过cDNA微阵列、主成分和聚类分析鉴定了人HNSCC系表达的独特基因特征，并通过定量逆转录-PCR（RT-PCR）和原位杂交进行了验证。NF-kB的激活、p53突变和TGF β通路和靶基因的失活之间的反向关系提示了NF-kB的激活可能与p53和TGF β信号传导的改变有关的假设。我们发现mt p53抑制TGF β受体II表达，减弱TGF β信号传导，增强HNSCC中炎性细胞因子反应性和NF-κ B的活化（Cohen等人，Cancer Res 2009）。 另一个确定的关系是NF-kB成员c-REL和p53成员p53，p63和p73之间。这证明了细胞因子TNF诱导的cREL与p63相互作用，从生长停滞和凋亡基因以及HNSCC的细胞核中置换p73（Lu等人，提交的手稿）。siRNA敲低p63后，通过与c-REL的相互作用，下调p53调控的肿瘤抑制基因，上调大量NF-κ B相关基因。在这些基因中定义了新的p63、cREL以及经典的p53和NF-kB位点，并通过ChIP测定法进行了验证（Yang等人，手稿在准备中）。与NCI遗传学实验室合作，正在对HNSCC中由cREL，p63和p73调控的全局基因表达进行ChIP测序。 与NCI和病理学系合作，使用蛋白质微阵列的新蛋白质组学技术，用于分析EGFR，NF-kB，MAPK，AKT和STAT 3信号磷蛋白抑制，以及来自HNSCC细胞系的蛋白质提取物和来自小分子抑制剂吉非替尼，紫杉醇和放射的临床试验中患者的标本中的细胞凋亡标志物。