Development of a low-cost epigenetic screening assay for Pap specimen-based detection of early-stage ovarian cancer in high-risk women

开发一种低成本表观遗传筛查方法，用于基于巴氏标本的高危女性早期卵巢癌检测

• 批准号: 10678833
• 负责人: Thomas Russell Pisanic II
• 金额: $ 42.36万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-06-15 至 2026-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10678833
• 关键词: Aberrant DNA Methylation; Achievement; Address; Algorithms; BRCA mutations; Benign; Bioinformatics; Biological Assay; Biological Markers; Biometry; Cancer Biology; Carcinoma; Carcinoma in Situ; Case/Control Studies; Cervix Uteri; Clinical; Clinical Research; Clinical Sensitivity; Clinical assessments; Collaborations; Complex; Curative Surgery; Cytology; DNA; DNA Methylation; Detection; Development; Diagnosis; Discrimination; Disease; Early Diagnosis; Engineering; Epigenetic Process; Excision; Exhibits; Feces; Female Genital Diseases; Fluorescence; Germ-Line Mutation; Goals; Gynecologic; Gynecologic Pathology; High Risk Woman; Hypermethylation; Intercept; Laboratories; Lesion; Liquid substance; Malignant Epithelial Cell; Malignant Female Reproductive System Neoplasm; Malignant Neoplasms; Malignant neoplasm of ovary; Mammalian Oviducts; Mammography; Methods; Methylation; Microfluidic Microchips; Microfluidics; Morbidity - disease rate; Operative Surgical Procedures; Ovary; Pap smear; Pathology; Patients; Performance; Recording of previous events; Research; Research Personnel; Resolution; Sampling; Scheme; Sensitivity and Specificity; Serous; Side; Specificity; Specimen; Statistical Models; Survival Rate; Symptoms; Techniques; Technology; Testing; Time; Tissues; Training; Translating; Travel; Tumor stage; Ultrasonography; Vagina; Woman; Work; assay development; cancer type; cohort; cost; cost effective; detection assay; diagnostic technologies; digital; early detection biomarkers; epigenetic marker; genome wide methylation; high risk; imaging modality; improved; instrumentation; liquid biopsy; melting; methylation biomarker; mortality; multidisciplinary; multiplex assay; neoplastic cell; novel; parallelization; performance tests; ratiometric; screening; success; tumor; tumor progression; user-friendly; variant detection

PROJECT SUMMARY With a 5-year survival rate of only 47%, high-grade serous carcinoma (HGSC) remains the most lethal of all types of ovarian cancer. This is a particular concern for women with familial histories or BRCA mutations, who exhibit 20 to 35-fold higher risk of developing HGSC in their lifetime. The high mortality of the disease is largely attributable to the fact that HGSC almost always remains undiagnosed until advanced stages when curative surgery is no longer feasible. Recently, numerous studies have demonstrated that most HGSCs likely originate in the fallopian tubes as precursor lesions called serous tubal intraepithelial carcinomas (STICs) that progress over a period of 6-7 years before rapidly metastasizing to the ovary and surrounding tissues. This suggests there exists a critical window of time in which STIC lesions and early-stage tumors might be detected so that tumor progression can be effectively intercepted before reaching an incurable stage. Recent studies have held out hope by showing that STIC and tumor cells are able to freely travel from the fallopian tubes to the cervix where they can be readily-detected in routinely-collected Pap specimens. Nonetheless, two notable challenges to this approach remain: (1) issues associated with the sensitivity and specificity of current biomarkers that undermine test performance and (2) a cost-effective diagnostic technology with sufficient sensitivity to detect exceedingly-rare copies of biomarkers from early-stage HGSC in complex samples such as Pap specimens. In the present proposal, we seek to address both of these challenges by developing a new type of HGSC screening assay called PapDREAM that is based on the sensitive, cost-effective detection of a novel set of epigenetic biomarkers that we have recently shown to be prevalently and specifically hypermethylated in both early, precursor (STIC) lesions and later-stage HGSC tumors. The PapDREAM assay will employ a unique, digital methylation analysis technique called µ-DREAMing (microfluidic Discrimination of Rare EpiAlleles by Melt) that provides a simple, but highly-effective means of detecting and quantifying rare, aberrantly- methylated DNA, even in challenging samples such as stool, liquid biopsies and Pap specimens. A user- friendly workflow based on common laboratory instrumentation will be developed to enable PapDREAM to be readily translated and performed in various research and clinical settings at a cost of only a few dollars per assay. Ultimately, the PapDREAM assay can be used alone or in combination with other assays to provide an ideal screening method for women at high risk for developing HGSC. We have assembled a multi-disciplinary team with complementary expertise in gynecologic pathology, assay development and DNA methylation bioinformatics to accomplish this goal by achieving the following aims: (1) Evaluate and prioritize methylation biomarkers to be included in the PapDREAM assay, (2) Incorporate µ- DREAMing assays into PapDREAM: a simple, multiplexed digital methylation assay for the detection of early- stage HGSC, and (3) Assess the clinical performance of the PapDREAM assay in Pap specimens.

项目摘要 5年生存率仅为47%，高级别浆液性癌（HGSC）仍然是所有恶性肿瘤中最致命的 卵巢癌的早期症状对于有家族史或BRCA突变的女性来说，这是一个特别值得关注的问题， 在他们的一生中表现出20至35倍的高风险发展HGSC。这种疾病的高死亡率主要是 这是由于HGSC几乎总是在晚期才被诊断出来， 手术已经不可行了。最近，许多研究表明，大多数HGSCs可能起源于 在输卵管中作为称为浆液性输卵管上皮内癌（STIC）的前驱病变， 在迅速转移到卵巢和周围组织之前，在6-7年的时间内。这表明 存在一个关键的时间窗口，在该时间窗口中可以检测到STIC病变和早期肿瘤， 在达到不可治愈的阶段之前可以有效地阻止肿瘤的发展。最近的研究表明， 通过显示STIC和肿瘤细胞能够自由地从输卵管移动到子宫颈， 在那里它们可以容易地在快速收集的巴氏标本中检测到。然而，两个显著的挑战 这种方法仍然存在：（1）与目前生物标志物的灵敏度和特异性相关的问题， 破坏测试性能和（2）具有足够灵敏度的成本效益诊断技术，以检测 复杂样本（如Pap标本）中早期HGSC生物标志物的罕见拷贝。 在本提案中，我们寻求通过开发一种新型的HGSC来解决这两个挑战 一种名为PapDREAM的筛查方法，该方法基于对一组新的 表观遗传生物标志物，我们最近已经表明，在这两个基因中， 早期前驱（STIC）病变和晚期HGSC肿瘤。PapDREAM测定将采用独特的， 名为µ-DREAMing的数字甲基化分析技术（稀有表位等位基因的微流体鉴别 熔融），提供了一种简单，但高度有效的手段，检测和量化罕见的，异常的- 甲基化的DNA，即使在挑战性的样品，如粪便，液体活检和巴氏标本。使用者─ 将开发基于通用实验室仪器的友好工作流程，使PapDREAM能够 易于翻译，并在各种研究和临床环境中进行，每个成本仅为几美元。 比色法最终，PapDREAM测定可单独使用或与其他测定组合使用，以提供对细胞的检测。 是一种理想的筛查方法。 我们组建了一支多学科团队，在妇科病理学、检测和诊断方面具有互补的专业知识。 发展和DNA甲基化生物信息学，以实现这一目标，通过实现以下目标：（1） 评价并优先考虑PapDREAM检测中包含的甲基化生物标志物，（2）将μ- 将DREAM检测转化为PapDREAM：一种用于检测早期乳腺癌的简单、多重数字甲基化检测方法。 HGSC期，和（3）评估PapDREAM测定在Pap标本中的临床性能。

项目成果

Multiplex Digital Methylation-Specific PCR for Noninvasive Screening of Lung Cancer.

• DOI: 10.1002/advs.202206518
• 发表时间: 2023-06
• 期刊: ADVANCED SCIENCE
• 影响因子: 15.1
• 作者: Zhao, Yang;O'Keefe, Christine M.;Hsieh, Kuangwen;Cope, Leslie;Joyce, Sonali C.;Pisanic, Thomas R.;Herman, James G.;Wang, Tza-Huei
• 通讯作者: Wang, Tza-Huei

Current and Emerging Methods for Ovarian Cancer Screening and Diagnostics: A Comprehensive Review.

• DOI: 10.3390/cancers14122885
• 发表时间: 2022-06-11
• 期刊: CANCERS
• 影响因子: 5.2
• 作者: Liberto, Juliane M.;Chen, Sheng-Yin;Shih, Ie-Ming;Wang, Tza-Huei;Wang, Tian-Li;Pisanic, Thomas R., II
• 通讯作者: Pisanic, Thomas R., II