Biomarkers Core

生物标志物核心摘要/摘要 Biomarkers核心为快速、高通量量化转录本提供集中资源， 蛋白质和磷蛋白。转录本将使用定量逆转录聚合酶链式反应进行量化。 (RT-qPCR)。此外，Biomarkers Core将使用Illumina提供全基因组微阵列分析 串珠站。下一代测序可与Illumina测序仪一起使用。蛋白质的定量 血液或组织裂解物中的水平将使用中尺度成像仪或反相蛋白质阵列进行测量 (RPPA)。纳米串多组学将用于同时进行RNA/DNA/蛋白质研究。生物标记物 CORE将提供：1)与增殖相关的已知基因的定量mRNA水平 在癌症进展中；2)在细胞或组织中鉴定独特的基因和表达谱 分子或药物操作；3)最初在 通过微阵列或rna-seq进行筛选；4)蛋白质水平的量化作为一项独立验证 技术和确定转录本水平和蛋白质水平之间的关系；5)评估 使用由RPPA验证的一组全面的抗体来改变蛋白质/磷蛋白水平。 具体目标1-4代表孢子项目和生物标志物核心之间的相互作用。具体目标1 预防和保守治疗复杂性不典型增生和非典型增生的新的靶向策略 1级子宫内膜样癌。该项目将在显微解剖腺体后使用rna-seq。 和间质细胞以确定CAH/1级子宫内膜样癌中对应于 孕激素反应性疾病与孕激素抵抗型疾病的对比，以及对依维莫司反应的标记物。QPCR 蛋白质组学(纳米串多组学或RPPA)将用于验证。具体目标2.CTNNB1 突变和Wnt通路激活定义了临床侵袭性子宫内膜样癌。这 该项目将创建至少7个包含特定突变的细胞系，并使用转录和miRNA图谱 用于识别目标基因和途径的稳定系。将使用定量聚合酶链式反应和蛋白质组学分析来验证 筛选研究中确定的候选成绩单。特异性靶向3EphA2在子宫内的靶向 癌症。这些研究将评估EphA2在癌症中的功能，并进行临床试验 EPHARNA，一种利用中性脂质体纳米载体系统地传递siEphA2的新方法。前置和 参加试验的患者的治疗后活组织检查将用RPPA和QPCR进行分析。具体目标4 子宫内膜癌新靶向治疗组合的鉴定框架。目标1和目标2 将使用RPPA分析(约500个样本)在PARP的临床试验中评估治疗效果 和PI3K抑制剂(目标1)，并绘制下游分子变化图，用于后续合理药物的设计 组合(目标2)。RNA-Seq或纳米串多组学也可用于分析活检标本。

Biomarkers Core SUMMARY/ABSTRACT The Biomarkers Core provides a centralized resource for rapid, high throughput quantification of transcripts, proteins, and phosphoproteins. Transcripts will be quantified using quantitative reverse transcriptase PCR (RT-qPCR). In addition, the Biomarkers Core will provide genome-wide microarray analysis using an Illumina Beadstation. Next-generation sequencing is available with Illumina sequencers. Quantification of protein levels in blood or tissue lysates will be done using a MesoScale Imager or by reverse-phase protein array (RPPA). NanoString multi-omics will be available for simultaneous RNA/DNA/protein studies. The Biomarker Core will provide: 1) quantitative mRNA levels for known genes that are involved in proliferation and implicated in cancer progression; 2) identification of unique genes and expression profiles in cells or tissues after a molecular or pharmacologic manipulation; 3) validation of the expression of genes that are initially identified in screening by microarrays or RNA-seq; 4) quantification of protein levels both as an independent validation technique and to determine the relationship between transcript levels and protein levels; 5) assessment of changes in protein/phosphoprotein levels using a comprehensive panel of validated antibodies by RPPA. Specific Aims 1-4 represent interactions between SPORE Projects and the Biomarkers Core. Specific Aim 1 Novel Targeted Strategies for Prevention and Conservative Management of Complex Atypical Hyperplasia and Grade 1 Endometrioid Endometrial Cancer. This project will use RNA-seq following microdissection of glands and stromal cells to identify transcripts in CAH/grade 1 endometrioid endometrial cancer that correspond to progestin-responsive versus progestin-resistant disease, as well as markers of response to everolimus. QPCR and proteomics (NanoString multi-omics or RPPA) will be used for validation. Specific Aim 2. CTNNB1 Mutation and Wnt Pathway Activation Define Clinically Aggressive Endometrioid Endometrial Carcinoma. This project will create at least 7 cell lines containing specific mutations and use transcript and miRNA profiles of the stable lines to identify target genes and pathways. QPCR and proteomic analysis will be used to validate candidate transcripts identified in the screening studies. Specific Aim 3 EphA2 Targeting in Uterine Carcinoma. These studies will evaluate the function of EphA2 in cancer and conduct a clinical trial of EPHARNA, a novel approach to deliver siEphA2 systemically using a neutral liposome nanovehicle. Pre- and post-treatment biopsies from patients enrolled in the trial will be analyzed by RPPA and QPCR. Specific Aim 4 A Framework for Identification of Novel Targeted Therapy Combinations in Endometrial Cancer. Aims 1 and 2 of this proposal will use RPPA analysis (~500 samples) to assess treatment efficacy in clinical trials of PARP and PI3K inhibitors (Aim 1) and to map downstream molecular changes for subsequent design of rational drug combinations (Aim 2). RNA-Seq or NanoString multi-omics can also be used to analyze biopsy specimens.