Targeted proteomics of MUC16 to enable early detection of ovarian cancer recurrence

MUC16 的靶向蛋白质组学可实现卵巢癌复发的早期检测

• 批准号: 10619871
• 负责人: Rebecca Jean Whelan
• 金额: $ 21.91万
• 依托单位: UNIVERSITY OF KANSAS LAWRENCE
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-01-01 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10619871
• 关键词: Anxiety; Area; Biological Assay; Biological Markers; CA-125 Antigen; Cancer Patient; Cancer Survivor; Caregivers; Charge; Clinical; Collaborations; Community Clinical Oncology Program; Core Protein; Data; Detection; Diagnostic; Diagnostic tests; Disease; Early Diagnosis; Epitopes; FDA approved; Funding; Future; Glycopeptides; Glycoproteins; Goals; Gynecologic Oncology; Immunoassay; Individual; Institution; Intervention; Lesion; Link; Malignant Neoplasms; Malignant neoplasm of ovary; Maps; Mass Spectrum Analysis; Messenger RNA; Methods; Modernization; Molecular; Monitor; Mucin 1 protein; Mucins; Parents; Patients; Peptide Library; Peptides; Periodicity; Peritoneal Fluid; Polysaccharides; Population; Post-Translational Protein Processing; Process; Proteins; Proteomics; Protocols documentation; Publications; RNA Splicing; Reaction; Recovery; Recurrence; Recurrent disease; Reporting; Research; Research Personnel; Resources; Risk; Sampling; Schedule; Screening for Ovarian Cancer; Serous; Serum; Site; Tandem Repeat Sequences; Testing; Tumor Debulking; United States National Institutes of Health; Ursidae Family; Variant; Visit; Woman; base; cancer biomarkers; cancer cell; cancer recurrence; chemotherapy; computerized tools; diagnostic biomarker; diagnostic tool; early detection biomarkers; effective therapy; experimental study; fighting; follow-up; glycosylation; high reward; high risk; improved; in silico; novel; novel diagnostics; ovarian neoplasm; overexpression; treatment strategy; tumor

High-grade serous ovarian cancer (HGSOC) is a deadly disease, in large part because most cases recur, and little can be done after that point. New diagnostic tools are urgently needed to improve HGSOC management and detect recurrence before clinical presentation. MUC16 is overexpressed on ovarian cancer cells and bears the CA125 epitope that is currently detected in clinical assays. The value of CA125 titers for surveillance is an active area of debate within the gynecologic oncology community. Some studies report no survival benefit, while others point to higher quality secondary cytoreductive surgery if action is taken quickly. Successfully fighting HGSOC requires that new assays are developed for more reliable and sensitive detection of recurrent disease. Our goal is to develop a new biomarker for HGSOC recurrence by fundamentally rethinking MUC16. Instead of considering MUC16 as the carrier of the CA125 epitope, we recognize that MUC16 itself is present as multiple diverse proteoforms. Because of variation in mRNA splicing, post-translational cleavage, and post- translational modifications, the population of MUC16 molecules present in an individual is heterogeneous. MUC16 is abundantly glycosylated, and glycan profiles of MUC16 derived from cancer cells is an untapped wealth of diagnostic information that is now lost using the existing peptide-based CA125 immunoassay. The goal of this project is to develop a method to enrich MUC16 from the serum of women undergoing treatment for HGSOC and analyze MUC16 using targeted (glyco)proteomics. We hypothesize that the molecular diversity of MUC16 revealed using modern bioanalytical and computational tools will enable novel diagnostics to detect HGSOC resurgence earlier than is currently possible. This hypothesis will be investigated by pursuing three research aims. In Aim 1 (Develop methods to quantitate MUC16 (glyco)peptide libraries using mass spectrometry), MUC16 from banked peritoneal fluid of HGSOC patients will be used to develop an optimized novel targeted mass spectrometry method for identification of MUC16 glycopeptides and peptides before and after deglycosylation. A parallel reaction monitoring (PRM) inclusion list will be generated by examination of in silico digest data and experimental MS/MS data. In Aim 2 (Develop a microscale separation method to enrich MUC16 from serum), we will adapt an immunoaffinity-free protocol that we developed to enrich MUC16 from peritoneal fluid to isolate the mucin from serum samples. A cartridge-based format will be used to capture MUC16 based on its negative charge and specific glycosylation. Finally, in Aim 3: (Develop pilot data on longitudinal variations in MUC16 (glyco)peptide libraries in patients with HGSOC before and during treatment), longitudinal serum samples from HGSOC patients will be enriched for MUC16 using the microscale protocol. Samples will be proteolyzed and analyzed by nLC-PRM. Quantitative (glyco)peptide maps for MUC16 from each patient will be compared longitudinally. The (glyco)peptides that deviate most between samples will be determined as potential diagnostic markers for early detection of recurring HGSOC in a future study.

高级别浆液性卵巢癌(HGSOC)是一种致命的疾病，很大程度上是因为大多数病例复发，并且 在那之后，我们几乎无能为力。迫切需要新的诊断工具来改善HGSOC管理 并在临床表现前发现复发。MUC16在卵巢癌细胞和熊身上过表达 目前在临床检测中检测到的CA125表位。CA125滴度对监测的价值是 妇科肿瘤界的活跃辩论领域。一些研究报告称没有生存益处， 而其他人则指出，如果行动迅速，可以进行质量更高的二次细胞减少术。成功 抗击HGSOC需要开发新的检测方法，以更可靠和更灵敏地检测复发 疾病。我们的目标是通过从根本上重新考虑MUC16来开发一种新的HGSOC复发的生物标志物。 我们不认为MUC16是CA125表位的载体，而是认识到MUC16本身是存在的 作为多种不同的蛋白质形式。由于mRNA剪接、翻译后切割和翻译后切割的差异， 翻译修饰，个体中存在的MUC16分子群体是不同的。 MUC16是大量糖基化的，而来源于癌细胞的MUC16的糖链是一个未被开发的 丰富的诊断信息，这些信息现在使用现有的基于多肽的CA125免疫分析而丢失。这个 该项目的目标是开发一种从接受治疗的妇女的血清中提纯MUC16的方法。 HGSOC，并使用靶向(糖)蛋白质组学分析MUC16。我们假设这些生物的分子多样性 使用现代生物分析和计算工具发现的MUC16将使新的诊断方法能够检测到 HGSOC的复苏比目前可能的更早。这一假设将通过以下三个方面进行研究： 研究目的。在目标1中(开发使用质量法定量MUC16(Glyco)多肽文库 光谱)，从HGSOC患者的腹腔液中提取的MUC16将被用于开发一种优化的 一种新的靶向质谱法鉴定MUC16糖肽和多肽 在脱糖后。将通过检查In生成平行反应监测(PRM)包含列表 硅胶消化数据和实验MS/MS数据。在目标2中(开发一种微型分离方法来浓缩 MUC16)，我们将采用我们开发的一种无免疫亲和力的方法来从 腹腔液从血清样本中分离粘蛋白。将使用基于盒式磁带的格式来捕获 MUC16基于其负电荷和特异性糖基化作用。最后，在目标3：(开发关于以下方面的试点数据 HGSOC患者治疗前和治疗期间MUC16(糖基化)肽库的纵向变化)， HGSOC患者的纵向血清样本将使用微型方案进行MUC16的浓缩。 样品将被蛋白质分解，并用NLC-PRM进行分析。MUC16的定量(糖基化)肽图来自 每个病人都会被纵向比较。样品之间偏离最大的(糖基)多肽将是 被确定为未来研究中早期发现复发HGSOC的潜在诊断标记物。