Novel Markers for Disease Outcome in Breast Cancer

乳腺癌疾病结果的新标志物

• 批准号: 8763263
• 负责人: Stefan Ambs
• 金额: $ 66.66万
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 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/8763263
• 关键词: Adrenergic beta-Antagonists; Affect; African; African American; American; Arachidonic Acids; Area; Baltimore; Biological Assay; Biological Markers; Biology; Breast; Breast Cancer Cell; Cancer Biology; Cancer Patient; Cancer Research Project; Cancerous; Carnitine; Catalytic Domain; Catecholamines; Categories; Cells; Cessation of life; Characteristics; Cities; Clinical Research; DNA Methylation; Data; Data Set; Development; Discrimination; Disease; Disease Marker; Disease Outcome; Disease Progression; ERBB2 gene; Employee Strikes; Enzymes; Epinephrine; Estrogen Receptor Status; Estrogen receptor negative; Estrogen receptor positive; Ethnic Origin; European; Excision; Exposure to; Freezing; Gene Expression; Gene Expression Profile; Gene Expression Profiling; Genes; Genetic Variation; Glioma; Goals; Histamine; Human; Infiltration; Isocitrate Dehydrogenase; Laboratories; Laboratory Research; Link; Lipids; Location; Mammary Neoplasms; Maryland; Mass Spectrum Analysis; Measurement; Measures; Medical Records; Medical center; Medicine; Metabolic; Methylation; Mitochondria; Modification; Molecular; Mutation; Neoplasm Transplantation; Norepinephrine; Normal tissue morphology; Nude Mice; Operative Surgical Procedures; Outcome; PECAM1 gene; Pathology; Pathway interactions; Patient Self-Report; Patients; Pattern; Phenotype; Population Study; Proteins; Proteome; Questionnaires; Race; Recruitment Activity; Services; Shapes; Signal Transduction; Social isolation; Source; Staging; Stem cells; Stress; Subgroup; System; TNM; Testing; Therapeutic Intervention; Tissue Sample; Tissues; Transplantation; Tumor Biology; Tumor Markers; Tumor Subtype; Universities; Validation; Woman; Work; base; clinically significant; college; density; design; experience; follow-up; genome-wide; health disparity; improved; indexing; interest; leukemia; macrophage; malignant breast neoplasm; mortality; multiple reaction monitoring; novel; novel marker; prognostic; response; stress management; tool; trait; tumor

In ongoing collaborative studies with the Sreekumar laboratory at the Baylor College of Medicine, our group has been investigating the metabolome of breast tumors. For discovery, we used the services of the company, Metabolon, to measure the abundance of 352 known and 184 unknown metabolites in 67 human breast tumors and 65 adjacent non-cancerous tissues with an untargeted mass spectrometry-based profiling approach. Quantitative differences for several key metabolites were validated in 70 estrogen receptor-negative tumors and 36 adjacent non-cancerous tissues with Multiple Reaction Monitoring-based targeted assays by the Sreekumar laboratory (validation set). Our current findings indicate that breast tumors have intrinsic metabolite signatures that persist with disease progression. We did not find that metabolites classify tumors into the gene expression-defined subtypes (luminal A & B, basal-like, HER2-enriched), suggesting that the breast tumor metabolome may describe different traits of the disease, as was suggested by others. Because our study population was ethnically diverse and was race/ethnicity-matched for tumor ER status and triple-negative/basal-like disease, we analyzed tissue metabolite differences comparing the African-American patients (n = 32) with European-American patients (n = 35). All patients were ancestry-typed with an average West African ancestry of 83.2% among the African-American patients and an average European ancestry of 97.6% among the European-American patients. Using unsupervised hierarchical clustering based on the abundance of 296 measured metabolites, both estrogen receptor (ER)-negative tumors and triple-negative tumors separated into subclusters that did not randomly represent the two patient groups. Instead, distinct clusters emerged that were enriched for either African-American or European-American patients, which was most striking for the triple-negative/basal-like tumor category. Notable, many mitochondrial metabolites, and also lyso(phospho)lipids, carnitines, and arachidonic acid, were more abundant in tumors from African-American patients while histamine was markedly decreased in them. Our data indicate that metabolic differences may exist between tumors from African-American and European-American patients. However, these findings need confirmation. We are currently collecting additional breast tumors from various sources for independent validation. Future research will perform a more in-depth analysis of the metabolite differences between the two patient groups, including a description of the key pathways that are affected by these differences, and how they may influence disease outcome and response to therapy. The Sreekumar laboratory has also access to human transplant tumors in nude mice. This system will be used to examine the relationship between the transplant metabolome and response to therapy and allows for experimental tissue metabolome modifications to test whether the changes improve the therapy response. Another interesting facet in our data was the elevated level of 2-hydroxyglutarate (2HG) in a subset of ER-positive and ER-negative tumors, with negligible levels in most adjacent noncancerous tissue samples. 2HG accumulates in gliomas and leukemias with hotspot mutations in either the cytosolic or mitochondrial isocitrate dehydrogenase (IDH1 and IDH2), which cause the enzyme to produce 2HG. Sequencing of the breast tumors did not reveal the presence of mutations in the catalytic domains of IDH1 and IDH2 mutations, consistent with other studies that failed to find these mutations in human breast tumors. Because aberrantly accumulated 2HG alters DNA methylation, we examined the global DNA methylation pattern in the 67 tumors using the Illumina Human Methylation 450 BeadChips. The analysis led to the discovery of a novel poor outcome tumor subtype with a high tissue 2HG concentration, a distinct genome-wide DNA methylation signature, and a stem cell-like transcriptional signature. Our data describe this molecular subtype as a class of tumors that tends to have high 2HG and includes a disproportionately high number of African-American patients. Thus, African-American patients may develop more frequently a methylation-defined subgroup III tumor than European-American patients, consistent with one earlier study that observed DNA methylation differences between these two patient groups. Our future research will concentrate on the relationship between 2HG and the development of the methylation-defined breast cancer subtype with poor outcome (subgroup III). In a second project, we will assess the influence of stress-related exposures on tumor biology in breast cancer patients, pursuing the hypothesis that exposures to stress, social isolation, and discrimination are associated with a prognostic gene expression signature and increased tumor catecholamine levels. We also hypothesize that this stress-induced signature is more prevalent in breast tumors of African-American than European-American patients and is a biomarker for catecholamine signaling that can be used to select patients for intervention therapy (beta-blocker, stress management). The cultural, race/ethnic, and sociodemographic diversity of Baltimore city and of the patients coming to the University of Maryland make the area an ideal study location to evaluate the influence of discrimination and stress on breast cancer biology, as proposed by our study, and to establish a link between stress-related exposures and the development of specific tumor characteristics (e.g., increased TAM infiltration and microvessel density in presence of a distinct poor outcome gene expression signature) that may affect African-American patients more frequently than European-American patients. We will analyze tumors from breast cancer patients who had surgical resection of their cancerous breast. These patients will complete the study questionnaire (evaluating stress) prior to the surgery. Per current design, we will initially analyze 80 fresh-frozen breast tumors from 40 African-American and 40 European-American breast cancer patients who have TNM stage I or stage II disease. We aim to include 40 ER-positive and 40 ER-negative tumors. The tumors will be selected from a projected patient pool of up to 150 breast cancer patients that are being recruited into the study at the University of Maryland Medical Center. Currently, we have completed recruitment of 18 patients. We will obtain clinicopathological information from pathology and medical records including tumor receptor status (ER, PR, HER2/neu) and survival follow up through the National Death Index. We will determine adrenaline and noradrenaline concentrations and perform gene expression profiling and immunohistochemical analysis of tumors and adjacent normal tissue. We will stratify patients by perceived stress, social isolation, and discrimination, will identify gene signatures associated with stress-related exposures and tumor catecholamine levels, and evaluate these gene signature(s) as predictor of patient survival and/or response to therapy using the many publically available datasets for breast cancer. We will also examine the relationship of stress-related exposures with selected protein markers (macrophage infiltration [CD68 as marker] and tumor microvessel density [CD31 as marker]). It is the strength of our design that we can use the measurement of tumor catecholamine levels as a validation tool for self-reported stress exposures. Our clinical studies will be supported by laboratory research that evaluates the influence of noradrenaline on cell phenotypes, gene expression, and DNA methylation patterns in human breast cancer cells. If we find that noradrenaline can shape DNA methylation patterns, we will analyze the relationship between DNA methylation and stress exposures in the breast tumors.

在与贝勒医学院的Sreekumar实验室进行的持续合作研究中，我们的小组一直在研究乳腺肿瘤的代谢组。为了发现，我们使用公司的服务，测量67种人类乳腺肿瘤中的352个已知和184个未知代谢产物的丰度，并使用基于质谱分析的方法来测量67种人体乳腺肿瘤和65个相邻的非癌组织。在70种雌激素受体阴性肿瘤和36个相邻的非癌组织中，通过Sreekumar实验室（验证集）对几种关键代谢产物的定量差异进行了验证。我们目前的发现表明，乳腺肿瘤具有固有的代谢产物特征，这些特征持续存在疾病进展。我们没有发现代谢产物将肿瘤分类为基因表达定义的亚型（腔A＆B，基底样，富含HER2的亚型），这表明乳腺肿瘤代谢组可能描述了该疾病的不同特征，正如其他人所建议的那样。由于我们的研究人群在种族上是多样的，并且在肿瘤ER状态和三阴性/基础样疾病的种族/种族匹配中，我们分析了比较非裔美国人患者（n = 32）与欧美患者的组织代谢物差异（n = 35）。在非裔美国人患者中，所有患者的祖先类型为83.2％，欧洲裔美国人的平均欧洲血统为97.6％。使用基于296种测量代谢产物的丰度，使用无监督的分层聚类，雌激素受体（ER）阴性肿瘤和三阴性肿瘤分离为分离为子群体的三阴性肿瘤，这些肿瘤并未随机代表这两个患者群体。取而代之的是，出现了针对非裔美国人或欧美患者富含的独特簇，这对于三阴性/基底样肿瘤类别最为惊人。值得注意的是，许多线粒体代谢产物，以及溶血（磷）脂质，肉碱和花生四烯酸在非裔美国人患者的肿瘤中更为丰富，而组胺的组胺显着降低。我们的数据表明，非裔美国人和欧美患者的肿瘤之间可能存在代谢差异。但是，这些发现需要确认。我们目前正在从各种来源收集其他乳腺肿瘤，以进行独立验证。未来的研究将对两个患者组之间的代谢物差异进行更深入的分析，包括对受这些差异影响的关键途径的描述，以及它们如何影响疾病结果和对治疗的反应。 Sreekumar实验室还可以进入裸鼠的人类移植肿瘤。该系统将用于检查移植代谢组和对治疗反应之间的关系，并允许实验性组织代谢组修饰以测试这些变化是否改善了治疗反应。我们数据中的另一个有趣的方面是在ER阳性和ER阴性肿瘤的子集中2-羟基戊二酸（2HG）的水平升高，在大多数相邻的非癌组织样品中，水平升高。 2HG在胞质或线粒体异位酸脱氢酶（IDH1和IDH2）中积聚在神经胶质瘤和白血病中，并在胞质或线粒体异位中积累突变，这会导致酶产生2Hg。对乳腺肿瘤的测序并未揭示IDH1和IDH2突变催化结构域中突变的存在，这与其他未在人类乳腺肿瘤中发现这些突变的研究一致。由于异常积累的2HG改变了DNA甲基化，因此我们使用Illumina Human甲基化450珠芯片检查了67个肿瘤中的全局DNA甲基化模式。该分析导致发现了新型较差的结果肿瘤亚型，具有高组织2HG浓度，全基因组DNA甲基化特征和干细胞样转录特征。我们的数据将这种分子亚型描述为一类肿瘤，这些肿瘤往往具有高2Hg，并且包括非裔美国人数量不成比例的肿瘤。因此，与欧美患者相比，非裔美国人患者可能更频繁地发展甲基化定义的亚组III肿瘤，这与一项早期的研究一致，该研究观察到这两个患者组之间的DNA甲基化差异。我们未来的研究将集中于2HG与甲基化定义的乳腺癌亚型的发展之间的关系，结果不佳（III亚组）。在第二个项目中，我们将评估与压力相关的暴露对乳腺癌患者中肿瘤生物学的影响，该假设假设压力，社会隔离和歧视与预后基因表达信号和肿瘤儿茶酚胺水平增加有关。我们还假设，这种压力引起的特征在非裔美国人的乳腺肿瘤中比欧美患者更为普遍，并且是儿茶酚胺信号传导的生物标志物，可用于选择患者进行干预疗法（β-阻滞剂，应力管理）。 The cultural, race/ethnic, and sociodemographic diversity of Baltimore city and of the patients coming to the University of Maryland make the area an ideal study location to evaluate the influence of discrimination and stress on breast cancer biology, as proposed by our study, and to establish a link between stress-related exposures and the development of specific tumor characteristics (e.g., increased TAM infiltration and microvessel density in presence of a distinct poor outcome gene expression签名）可能比欧美患者更频繁地影响非裔美国人患者。我们将分析对癌症乳房手术切除的乳腺癌患者的肿瘤。这些患者将在手术前完成研究问卷（评估应力）。根据目前的设计，我们最初将分析来自40名非裔美国人和40名患有TNM I期或II期疾病的欧美乳腺癌患者的80种新鲜乳房肿瘤。我们的目标是包括40个ER阳性和40个ER阴性肿瘤。这些肿瘤将从预计的患者库中选出，该肿瘤最多可在马里兰大学医学中心招募到研究中。目前，我们已经完成了18名患者的招聘。我们将从病理和病历中获得临床病理学信息，包括肿瘤受体状况（ER，PR，HER2/NEU）以及通过国家死亡指数的随访。我们将确定肾上腺素和去甲肾上腺素浓度，并对肿瘤和邻近正常组织进行基因表达分析和免疫组织化学分析。我们将通过感知的压力，社会隔离和歧视对患者进行分层，将确定与压力相关的暴露和肿瘤儿茶酚胺水平相关的基因特征，并评估这些基因特征作为患者生存和/或使用许多公共可用乳腺癌数据集的患者生存和/或反应的预测因素。我们还将检查与压力相关的暴露与选定的蛋白质标志物（巨噬细胞浸润[CD68作为标记]和肿瘤微血管密度[CD31作为标记]）的关系。我们设计的强度是，我们可以使用肿瘤儿茶酚胺水平的测量作为自我报告应力暴露的验证工具。我们的临床研究将得到实验室研究的支持，该研究评估去甲肾上腺素对人乳腺癌细胞中细胞表型，基因表达和DNA甲基化模式的影响。如果我们发现去甲肾上腺素可以塑造DNA甲基化模式，我们将分析乳腺肿瘤中DNA甲基化和应激暴露之间的关系。