Blood biopsy test for pan-cancer mutations with error-checking probes

血液活检检测泛癌基因突变的错误检测探头机密PI：Shafer，David A.，PhD 项目总结 GeneTAG Technology(www.genetagtech.com)开发了一系列分子诊断分析方法，用于 基于DNA扩增和专有检测方法的癌症和传染病。我们的初选 系统，内部DDS(IDDS)探针，包括两个相互作用的组件，一个氟标记的探针和一个 猝灭剂标记的反探针几乎与探针互补。在没有预定目标的情况下，配对 探针和反探针结合在一起，猝灭荧光，防止偏离目标的检测。这个独一无二的 该系统在很宽的温度范围内(10-30°C)提供高度特定的单基座分辨率。最近， 我们将IDDS探针技术与Wild Terminator(WTx)方法相结合，以实现对低丰度的检测 通过阻断野生型序列的扩增而产生突变体(专利申请中)。通过结合这些技术， 我们可以可靠地在跟踪水平(0.01%)检测到大量目标的单碱基变体 型基因组DNA。这种增强的qPCR灵敏度达到或超过了专业平台的灵敏度 (例如水滴数字聚合酶链式反应、下一代测序[NGS]和MALDI-TOF)，它们已被用于 研究癌症患者血浆中罕见的突变。我们的主要目标是开发泛癌症筛查 基于这种组合方法的测试。这种新的方法应该能够检测到低频诊断 血液中的突变来自于身体其他部位的躯体肿瘤。我们的探测器可以 可靠地检测表明特定癌症或耐药变异的特定突变。通过 在一次检测中涵盖10个或更多常见的癌症特异性突变，我们可以确定存在 在早期阶段，未知的癌症，在某些情况下，我们可以准确地指出哪种癌症可能发生。具体的 这项第一阶段拨款的目的是：1、测试和比较用于检测低噪声的选择放大方法。 丰度，泛癌驱动因素突变和2，开发和测试泛癌驱动因素的原型测试 基于IDDS探针和选择性扩增的血浆突变。为了证明可行性，我们将 研究早期和晚期癌症患者血浆样本中的泛癌突变 高水平的ctDNA，如神经母细胞瘤、前列腺、卵巢、结直肠、肝细胞和乳房 癌症。我们的长期目标是到第二阶段结束时，推出一种针对约100例循环癌症的通用检测方法 生物标志物(可作用的和抗药性的变体)，基于WTx、IDDS、NGS和 统计方法。这些信息将指导医生进行后续测试并选择合适的 一线靶向治疗。反过来，这将通过实现早期癌症检测来为公众健康带来好处 通过一种非侵入性方法。

Blood biopsy test for pan-cancer mutations with error-checking probes Confidential PI: Shafer, David A., PhD PROJECT SUMMARY GeneTAG Technology (www.genetagtech.com) has developed a series of molecular diagnostic assays for cancer and infectious diseases based on DNA amplification and proprietary detection methods. Our primary system, internal DDS (iDDS) probes, comprises two interacting components, a fluor-labeled probe and a quencher-labeled antiprobe nearly complementary to the probe. In the absence of the intended target, paired probes and antiprobes bind together, quenching fluorescence and preventing off-target detection. This unique system provides highly specific single-base discrimination over a wide temperature range (10–30ºC). Recently, we merged iDDS probe technology with Wild Terminator (WTx) methods to enable detection of low-abundance mutants by blocking amplification of the wild-type sequence (patent pending). By combined these technologies, we can reliably detect single-base variants of numerous targets at trace levels (0.01%) in a background of wild type genomic DNA. This enhanced qPCR sensitivity meets or exceeds the sensitivity of specialized platforms (such as droplet digital PCR, next-generation sequencing [NGS], and MALDI-TOF), which have been used to study rare mutations in the plasma of cancer patients. Our primary goal is to develop a pan-cancer screening test based on this combined method. This new method should enable the detection of low frequency diagnostic driver mutations in the blood that derive from a somatic tumor located elsewhere in the body. Our probes can reliably detect specific mutations that are indicative of a particular cancer or of a drug resistant variant. By covering ten or more of the common cancer-specific mutations in one assay we can identify the presence of an unknown cancer at an early stage, and in some cases, we can pin-point which cancer is likely. The Specific Aims of this Phase I grant are: 1, to test and compare selective-amplification methods for detecting low- abundance, pan-cancer driver mutations and 2, to develop and test a prototype assay for pan-cancer driver mutations in plasma based on iDDS probes and selective amplification. To demonstrate feasibility, we will study pan-cancer mutations in plasma samples from early- and late-stage patients with cancers that show high levels of ctDNA, such as neuroblastoma, prostate, ovarian, colorectal, hepatocellular, and breast cancer. Our long-range goal by the end of Phase II is to market a generalized test for ~100 circulating cancer biomarkers (actionable and drug-resistant variants), based on a synthesis between WTx, iDDS, NGS, and statistical methods. Such information would guide physicians in follow-up testing and selecting appropriate first-line targeted therapy. This, in turn, would provide a public health benefit by enabling early cancer detection through a non-invasive approach.