Pre-analytical factors affecting ctDNA analysis in early and locally advanced breast cancer

影响早期和局部晚期乳腺癌 ctDNA 分析的分析前因素

• 批准号: 9893681
• 负责人: Muhammed Murtaza
• 金额: $ 44万
• 依托单位: TRANSLATIONAL GENOMICS RESEARCH INST
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-18 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9893681
• 关键词: Address; Advanced Malignant Neoplasm; Affect; Aliquot; Biological Assay; Blood; Blood Cells; Blood specimen; Breast Cancer Patient; Cancer Patient; Cell Extracts; Cells; Centrifugation; Clinical Research; Clinical Sensitivity; Collection; Cytolysis; DNA; DNA analysis; Detection; Detection of Minimal Residual Disease; Disseminated Malignant Neoplasm; Early Diagnosis; Edetic Acid; Enrollment; Future; Genotype; Gold; Hour; Individual; Localized Malignant Neoplasm; Malignant Neoplasms; Measures; Metastatic breast cancer; Methods; Molecular; Mutation; No Evidence of Disease; Noise; Nonmetastatic; Observational Study; Operative Surgical Procedures; Outcome; Patients; Peripheral; Plasma; Process; Prospective cohort; Protocols documentation; RNA; Residual Tumors; Sampling; Screening for cancer; Shipping; Ships; Speed; Temperature; Testing; Tube; Variant; Venipunctures; advanced breast cancer; base; cancer therapy; cell free DNA; cohort; digital; founder mutation; frontier; genotyped patients; healthy volunteer; improved; improved outcome; interest; magnetic beads; malignant breast neoplasm; oncology; prospective; success; tumor; tumor DNA

PROJECT SUMMARY Circulating tumor DNA (ctDNA) analysis has enabled noninvasive tumor genotyping for patients with advanced metastatic cancers. There is now growing interest in replicating this success in patients with early stage cancers. ctDNA analysis could enable blood-based minimal residual disease detection in patients receiving treatment and early detection of cancers in pre-symptomatic individuals. However, ctDNA levels are 10s-100s fold lower in localized cancers than in metastatic cancers, limiting the sensitivity of current assays. This is further confounded by pre-analytical variability such as differences in blood collection and processing and DNA extraction. For example, inappropriate or delayed blood processing can cause peripheral cell lysis and dilute ctDNA fraction in blood samples. In early stage cancer patients where ctDNA levels are already quite low, this can cause false-negative results. The effects of pre-analytical factors on ctDNA detection in patients with localized cancers are not well understood. To address this gap, we propose a study of 180 patients with early and locally advanced breast cancer to investigate three aspects of pre-analytical variation: 1) DNA extraction methods (Aim 1), 2) blood collection tubes and processing protocols (Aim 2) and 3) long-term storage of plasma and extracted DNA (Aim 3). We will investigate these factors using two assays we have recently developed: 1) a quality assessment assay that measures total cell-free DNA concentration and fragment size and 2) TARgeted DIgital Sequencing (TARDIS), an assay that simultaneously measures up to 30 patient-specific founder mutations in plasma DNA to quantify ctDNA levels. By leveraging multiple mutations for each patient, improving error suppression and minimizing loss of input DNA material, TARDIS enables sensitive detection and precise quantification of ctDNA in patients with localized cancers and improves limit of detection by 10-100 fold over current assays. Minimal residual disease detection and early detection using ctDNA analysis hold tremendous promise to individualize cancer treatment and to improve outcomes. Our study will clarify pre-analytical factors that could be critical to the success of future clinical studies across localized cancers of multiple subtypes.

项目摘要 循环肿瘤DNA（CTDNA）分析已使患有晚期患者的无创肿瘤基因分型 转移性癌症。现在，人们对在早期患者中复制这一成功的兴趣日益兴趣 癌症。 CTDNA分析可以使接受血液的最小残留疾病检测在接受的患者中 治疗和早期检测症状症状。但是，ctDNA水平为10s-100 局部癌症中的折叠比转移性癌症低，从而限制了当前测定的灵敏度。这是 进一步被分析前的变异性混淆，例如血液收集和加工和DNA的差异 萃取。例如，不适当或延迟的血液加工可能会导致外周细胞裂解并稀释 血液样本中的ctDNA分数。在早期的癌症患者中，ctDNA水平已经很低，这 可能导致虚假的结果。分析前因子对患者患者CTDNA检测的影响 局部癌症不太了解。 为了解决这一差距，我们建议对180例早期和局部晚期乳腺癌患者进行研究 研究预分析前变异的三个方面：1）DNA提取方法（AIM 1），2）血液收集 试管和加工方案（AIM 2）和3）血浆和提取的DNA的长期存储（AIM 3）。我们 将使用我们最近开发的两个测定法调查这些因素：1）质量评估测定法 测量总无细胞的DNA浓度和片段大小以及2）目标数字测序（TARDIS）， 一种在血浆DNA中同时测量多达30个患者特异性的创始人突变以量化的测定 ctDNA水平。通过利用每个患者的多个突变，改善误差抑制并最大程度地减少 输入DNA材料的丢失，TARDIS可以对患者进行敏感检测和精确定量CTDNA 与当前测定相比，随着局部癌症的局部癌症并将检测限制提高了10-100倍。 使用CTDNA分析的最小残留疾病检测和早期检测具有巨大的希望 个性化癌症治疗并改善预后。我们的研究将阐明可能 对于跨多种亚型局部癌症的未来临床研究的成功至关重要。