Cold Spring Harbor Laboratory, Cold Spring Harbor, NY 11724, USA.
The Feinstein Institute for Medical Research, Northwell Health, Manhasset, NY 11030, USA.
Nucleic Acids Res. 2020 Apr 17;48(7):e40. doi: 10.1093/nar/gkaa090.
Measuring minimal residual disease in cancer has applications for prognosis, monitoring treatment and detection of recurrence. Simple sequence-based methods to detect nucleotide substitution variants have error rates (about 10-3) that limit sensitive detection. We developed and characterized the performance of MASQ (multiplex accurate sensitive quantitation), a method with an error rate below 10-6. MASQ counts variant templates accurately in the presence of millions of host genomes by using tags to identify each template and demanding consensus over multiple reads. Since the MASQ protocol multiplexes 50 target loci, we can both integrate signal from multiple variants and capture subclonal response to treatment. Compared to existing methods for variant detection, MASQ achieves an excellent combination of sensitivity, specificity and yield. We tested MASQ in a pilot study in acute myeloid leukemia (AML) patients who entered complete remission. We detect leukemic variants in the blood and bone marrow samples of all five patients, after induction therapy, at levels ranging from 10-2 to nearly 10-6. We observe evidence of sub-clonal structure and find higher target variant frequencies in patients who go on to relapse, demonstrating the potential for MASQ to quantify residual disease in AML.
在癌症中检测微小残留病可用于预后、监测治疗和检测复发。用于检测核苷酸取代变异的简单序列方法的错误率（约为 10-3）限制了其灵敏性检测。我们开发并描述了 MASQ（多重精确敏感定量）的性能，这种方法的错误率低于 10-6。MASQ 通过使用标记物来识别每个模板，并要求多个读取的一致性，从而在存在数百万个宿主基因组的情况下准确地计数变异模板。由于 MASQ 方案对 50 个靶标进行了多重化，因此我们既可以整合来自多个变体的信号，又可以捕获对治疗的亚克隆反应。与用于变异检测的现有方法相比，MASQ 在急性髓细胞性白血病（AML）患者的初步研究中实现了灵敏度、特异性和产量的优异结合。在诱导治疗后，我们在所有五名患者的血液和骨髓样本中检测到白血病变体，水平从 10-2 到接近 10-6。我们观察到亚克隆结构的证据，并在继续复发的患者中发现更高的目标变体频率，表明 MASQ 有潜力对 AML 进行微量残留病定量。
