AML-MutationCounter, a tool to detect residual and recurrent leukemia

摘要 Asys Bio LLC计划推出一种名为AML-MutationCounter的分子工具包，用于计算体细胞突变 在促进急性髓系白血病(AML)发展的基因中。我们已经开发了一套 用于应用下一代测序来计算AML基因突变的试剂和计算机程序。 该工具包用途广泛，可与两个主要DNA测序平台Illumina Hi- SEQ和ThermoFisher离子质子。每年有20,000名美国人罹患急性髓细胞白血病，其中近10,000人死于 这种疾病。许多患者对初级治疗有反应，并获得临床缓解。减刑目前正在 由骨髓中的白血病母细胞减少到细胞的5%来确定。一个更敏感和更具体的 缓解试验可以指导临床医生实现更持久的缓解，并改善患者的分层 关于复发的可能性。最近的一项研究发现，48%的缓解期患者保留了AML 2.5%的骨髓或血细胞中存在基因突变，这些患者的存活率平均为25% 见于缓解期基因突变清除至2.5%的患者。因此残留物的存在或不存在 缓解期的疾病具有很高的预后价值。缓解后接受骨髓移植的患者 有时会出现宿主骨髓的克隆，称为骨髓嵌合体。重要的是要 尽早确定这些出现的克隆是否代表复发白血病 抢救疗法。我们在北卡罗来纳大学和北卡罗来纳纪念医院的肿瘤学家顾问小组 医院热衷于对残留和复发进行敏感、准确、快速和低成本的检测 白血病。在这一阶段的STTR赠款申请中的研究将涉及Asys Bio LLC和 北卡罗来纳大学教堂山分校生产一种用于测量急性髓系白血病的分子工具包- 骨髓标本中相关的体细胞基因突变。目标1的研究将确定精确度， 用模拟数据定量突变等位基因频率试剂盒的灵敏度和重复性 一种标准的参考细胞系。AIM 2中的研究将把该工具箱应用于收集的10个患者样本 在白血病随后复发之前的缓解时间。选择样本以包括 原发癌中NPM1白血病驱动基因突变的研究缓解期NPM1基因突变的研究 样本是一个不好的预后指标。我们将展示我们的试剂盒检测到NPM1和其他AML的突变- 注定要复发的患者缓解时的相关基因。此第一阶段项目演示了 AML-MutationCounter检测急性髓系白血病微小残留和复发的可行性我们的第二阶段 商业化计划将包括一项临床试验，以确定检测试剂盒的实用性，以及积极营销 将检测试剂盒运往学术医学中心和商业检测实验室。

Abstract AsystBio LLC proposes to market a molecular tool kit called AML-MutationCounter to count somatic mutations in genes that contribute to the development of acute myeloid leukemia (AML). We have developed a set of reagents and computer programs for application of next-generation sequencing to count AML gene mutations. The tool kit is versatile and can be used with either of the two major DNA sequencing platforms, Illumina Hi- Seq and ThermoFisher Ion Proton. Every year AML afflicts 20,000 Americans with nearly 10,000 dying from the disease. Many patients respond to primary therapy and achieve clinical remission. Remission is currently determined by a reduction of leukemic blasts in bone marrow to <5% of cells. A more sensitive and specific test for remission may guide clinicians to achieve more durable remission and improve stratification of patients for likelihood of relapse. One recent study found that forty eight percent of patients in remission retained AML gene mutations in >2.5% of marrow or blood cells and these patients’ survival was on average 25% of that seen in patients who cleared mutations to <2.5% during remission. Thus the presence or absence of residual disease at remission is highly prognostic. Patients who receive a bone marrow transplant after remission sometimes display emerging clones of host bone marrow, termed bone marrow chimerism. It is important to determine whether these emerging clones represent recurrent leukemia as early as possible to implement salvage therapies. Our advisory panel of oncologists at the University of North Carolina and NC Memorial Hospital expresses enthusiasm for a sensitive, accurate, rapid and low-cost test for residual and recurrent leukemia. Studies in this Phase I STTR grant application will involve a collaboration between AsystBio LLC and the University of North Carolina at Chapel Hill to produce a molecular tool kit for measurement of AML- associated somatic gene mutations in bone marrow specimens. Studies in Aim 1 will establish the accuracy, sensitivity and reproducibility of the kit for quantification of mutant allele frequencies using simulated data and a standard reference cell line. Studies in Aim 2 will apply the tool kit to 10 patient samples that were collected at the time of remission before a subsequent recurrence of leukemia. Samples are selected to include the NPM1 leukemia-driver gene mutation in the primary cancer. The presence of mutations in NPM1 in remission samples is a poor prognostic sign. We will show that our kit detects mutations in NPM1 and other AML- associated genes at remission in patients that were destined to relapse. This phase I project demonstrates the feasibility of AML-MutationCounter for detection of minimal residual and recurrent disease in AML. Our phase II commercialization plan will include a clinical trial to establish the utility of the test kit as well as active marketing of the test kit to academic medical centers and commercial test laboratories.