Lysyl Oxidase Propeptide: Breast Cancer Inhibitor

赖氨酰氧化酶前肽：乳腺癌抑制剂

• 批准号: 8211091
• 负责人: GAIL E. SONENSHEIN
• 金额: $ 50.56万
• 依托单位: TUFTS UNIVERSITY BOSTON
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-04-12 至 2015-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8211091
• 关键词: 1-Phosphatidylinositol 3-Kinase; Affect; African American; Agar; Alleles; American; Anabolism; Anchorage-Independent Growth; Anthracenes; Aromatic Polycyclic Hydrocarbons; Bioluminescence; Boston; Breast Cancer Cell; Cancer Etiology; Cancer cell line; Carcinogen exposure; Carcinoma; Case-Control Studies; Cell Line; Cell Proliferation; Cells; Cessation of life; Co-Immunoprecipitations; Collagen; Data; Databases; Deposition; Development; Disease Progression; E-Cadherin; ERBB2 gene; Effectiveness; Elastin; Environment; Enzymes; Estrogen Receptors; Estrogen receptor negative; Exposure to; Extracellular Matrix; Fatty acid glycerol esters; Fibroblasts; Frequencies; Gene Expression; Genes; Glutamine; Growth; HRAS gene; Heat shock proteins; Hypoxia; Image; In Vitro; Investigation; Knock-in Mouse; Lead; Length; MEKs; Malignant Neoplasms; Malignant neoplasm of lung; Malignant neoplasm of pancreas; Mammary gland; Mass Spectrum Analysis; Mediating; Minor; Modeling; Mutation; NIH 3T3 Cells; Neoplasm Metastasis; Normal Cell; Nude Mice; Oncogenic; Oxidases; Pancreatic carcinoma; Participant; Pathway Analysis; Pathway interactions; Phenotype; Pilot Projects; Protein Tyrosine Phosphatase; Protein-Lysine 6-Oxidase; Proteins; Proteolytic Processing; Ras Inhibitor; Risk; Risk Assessment; Risk Factors; Role; Screening procedure; Signal Pathway; Signal Transduction; Single Nucleotide Polymorphism; Snails; TP53 gene; Testing; Tissues; Treatment Protocols; Tumor Suppressor Proteins; United States National Institutes of Health; Universities; Up-Regulation; Variant; Vimentin; Woman; Women' s Health; Work; Xenograft Model; Yeasts; cancer cell; cohort; crosslink; dimethylbenzanthracene; effective therapy; epithelial to mesenchymal transition; extracellular; in vivo; inhibitor/antagonist; lung Carcinoma; malignant breast neoplasm; malignant phenotype; matrigel; migration; mouse model; mutant; novel; pancreatic cancer cells; protein function; public health relevance; ras Oncogene; receptor; research clinical testing; tool; transcription factor; tumor; tumor progression; tumorigenic; yeast two hybrid system

DESCRIPTION (provided by applicant): Breast cancer is the second leading cause of cancer deaths among American women, and thus additional, effective treatment protocols are needed. Lysyl oxidase (LOX) is an essential extracellular enzyme that controls matrix deposition. The LOX gene was isolated as the "ras recision" gene (rrg) with an ability to inhibit the transforming activity of the Ras oncogene in NIH 3T3 cells. Consistently many cancers display reduced LOX gene expression. The LOX gene encodes a 50 kDa pro-enzyme (termed Pro-LOX), which is secreted into the extracellular environment where it is processed by proteolytic cleavage to an 18 kDa amino terminal propeptide (LOX-PP) and a functional 32 kDa enzyme. The PI's group demonstrated that the rrg activity resides in the LOX-PP domain, whereas, the LOX enzyme itself has been found to promote a more aggressive phenotype and tumor metastasis under hypoxic conditions. Importantly, LOX-PP activity was shown by the PI's group to effectively inhibit breast cancer cells, driven by Her-2/neu which signals via Ras. In culture, LOX-PP suppressed the PI3K/Akt and MEK/Erk pathways and NF-?B. Oncogenic Her-2/neu induces epithelial to mesenchymal transition (EMT) and this was reverted by LOX-PP. LOX-PP induced E-cadherin, decreased levels of Snail, tumor migration and formation of branching colonies in Matrigel, and reduced tumor formation in nude mice. A single nucleotide polymorphism (SNP) (rs1800449) G473A, resulting in an Arg158Gln substitution in a highly conserved region in the LOX-PP domain, occurs with an average 24.6% 473A minor allele carrier frequency in the HapMap database. However when cancer cells were examined, the G473A SNP was present in the majority of lines and the LOX-PP Gln variant displayed substantially impaired tumor suppressor function in vitro and in vivo, and a reduced ability to oppose the pro-tumorigenic effects of LOX. In a pilot study of African-American women, the minor 473A allele was associated with increased risk of ER1 negative breast cancer. These findings lead to two related central hypotheses: 1. Wildtype LOX-PP represents an important tumor suppressor, the activity of which is compromised by an Arg to Gln substitution. 2. A G473A (rs1800449) SNP in the LOX gene leads to increased risk of more invasive cancers. In this new RO1 application, we propose to: (Aim 1) Elucidate the mechanisms whereby wildtype LOX-PP mediates its action as a suppressor of Ras signaling in breast cancer cells and determine how these are affected by the Arg158Gln substitution; (Aim 2) Test the ability of LOX-PP to impede the tumor promoting effects of the LOX enzyme in culture and in an orthotopic mammary fat pad model; (Aim 3) Test the hypothesis that the rs1800449 SNP represents a risk factor for more invasive breast cancer using a knock-in mouse that will be prepared. The proposed studies represent pre-clinical testing of LOX-PP as an inhibitor of Ras signaling and as a risk factor of disease progression. As the LOX-PP activity was also effective against lung and pancreatic cancers, our findings have broader implications for carcinomas driven by an activated Ras oncogene. PUBLIC HEALTH RELEVANCE: Overall, these studies represent pre-clinical testing of the LOX-PP protein, a novel suppressor of Ras signaling, in breast cancer. The rs1800449 single nucleotide polymorphism (SNP) results in an Arg-to-Gln variant in LOX-PP and in substantially reduced Ras suppressing activity and increased risk of ER1 negative breast cancer in African-American women. Our proposed work will delineate the mechanism of LOX-PP activity, the changes that result from the Gln variant and prepare a knock-in mouse model that allows for testing of the potential use of SNP in assessment of risk for breast cancer.

描述（由申请人提供）：乳腺癌是美国妇女癌症死亡的第二大原因，因此需要其他有效的治疗方案。赖氨酸氧化酶（LOX）是控制基质沉积的必需细胞外酶。 LOX基因被分离为“ RAS恢复”基因（RRG），能够抑制NIH 3T3细胞中Ras癌基因的转化活性。一致地，许多癌症表现出降低的LOX基因表达。 LOX基因编码50 kDa的促酶（称为pro-lox），该酶被分泌到细胞外环境中，通过蛋白水解裂解为18 kDa氨基末端丙肽（LOX-PPP）和功能性32 kDa酶进行处理。 PI的组表明RRG活性驻留在LOX-PP结构域中，而LOX酶本身在低氧条件下促进了更具侵略性的表型和肿瘤转移。重要的是，PI组显示了LOX-PP的活性，以有效抑制乳腺癌细胞，这是由HER-2/NEU通过RAS发出信号的驱动的。在文化中，LOX-PP抑制了PI3K/AKT和MEK/ERK途径以及NF-？b。致癌性HER-2/NEU诱导上皮到间充质转变（EMT），并被LOX-PP恢复。 LOX-PP诱导的E-钙粘着蛋白，蜗牛的水平降低，肿瘤迁移和基质中分支菌落的形成以及裸鼠的肿瘤形成减少。单个核苷酸多态性（SNP）（RS1800449）G473A，在LOX-PP域中的高度保守区域中导致ARG158GLN取代，在HakpMap数据库中平均为24.6％473a次要小等位基因载体频率为24.6％。但是，当检查癌细胞时，大多数系列中存在G473a SNP，LOX-PPP GLN变体在体外和体内显示出显着损害肿瘤抑制器功能，并且降低了相反的LOX促肿瘤效应的能力。在一项针对非裔美国妇女的试点研究中，次要的473A等位基因与ER1乳腺癌阴性风险增加有关。这些发现导致了两个相关的中央假设：1。WildTypelox-PP代表了一个重要的肿瘤抑制剂，其活性因ARG替代而损害。 2。LOX基因中的A G473A（RS1800449）SNP导致增加侵入性癌症的风险增加。在这种新的RO1应用中，我们建议：（ AIM 1）阐明WildType Lox-PP介导其作为抑制乳腺癌细胞中Ras信号的抑制器的机制，并确定它们如何受到ARG158GLN取代的影响； （AIM 2）测试LOX-PP在培养物和原位乳腺脂肪垫模型中促进LOX酶促进肿瘤作用的能力； （AIM 3）检验以下假设：RS1800449 SNP代表了使用将要制备的敲入小鼠进行更具侵入性乳腺癌的危险因素。拟议的研究代表了LOX-PP作为RAS信号传导抑制剂的临床前测试，也是疾病进展的危险因素。由于LOX-PP活性也有效地针对肺和胰腺癌，因此我们的发现对由活化的Ras癌基因驱动的癌具有更大的影响。 公共卫生相关性：总体而言，这些研究代表了乳腺癌中LOX-PP蛋白的临床前测试，这是RAS信号的新型抑制剂。 RS1800449单核苷酸多态性（SNP）导致LOX-PP中的ARG-GLN变体，并且在非洲裔美国妇女中大大降低了RAS抑制活性和ER1阴性乳腺癌风险的大幅降低。我们提出的工作将描述LOX-PP活性的机制，LOX-PP活动是由GLN变体产生的变化，并准备了敲入小鼠模型，该模型允许测试SNP在评估乳腺癌风险中的潜在使用。