Sommer Charlotte, Mack Hildegard I D, Killer Madeleine C, Ross Petra, Nist Andrea, Stiewe Thorsten, Neubauer Andreas, Brendel Cornelia, Mack Elisabeth K M
Department of Hematology, Oncology and Immunology, Philipps-University Marburg, and University Hospital Gießen and Marburg, Campus Marburg, Marburg, Germany.
Genomics Core Facility, Institute of Molecular Oncology, Universities of Gießen and Marburg Lung Center, Member of the German Center for Lung Research (DZL), Philipps-University Marburg, Marburg, Germany.
Sci Rep. 2025 Sep 23;15(1):32679. doi: 10.1038/s41598-025-20589-3.
Minimal/measurable residual disease (MRD) in Acute Myeloid Leukemia (AML) is defined as persistent leukemic cells below cytomorphological detection threshold. Next generation sequencing (NGS) of circulating cell-free DNA (cfDNA) to profile cancer-associated mutations has been shown to allow for quantification of disease burden in solid tumors and has also been suggested to enable minimally invasive follow-up of AML patients. In this pilot study we investigated the technical sensitivity and potential prognostic implications of cfDNA-based MRD monitoring in AML after allogeneic stem cell transplantation in comparison to donor chimerism analysis or, respectively, after consolidation chemotherapy. 75 cfDNA samples from 29 patients were analyzed by targeted NGS using a commercially available 10- or 37-gene hotspot panel (VariantPlex Core AML or Core Myeloid panel, ArcherDx). Patients' leukemias exhibited 1-7 mutations as determined by routine diagnostics. Only previously identified mutations were considered for MRD evaluation. cfDNA was isolated in sufficient amounts for NGS from all samples (total yield 24 ng-5.2 µg). The sensitivity of variant detection increased with higher overall read count and higher mutation-specific coverage (variant allele frequency [VAF] range 0.08-100%). At least one previously known mutation was identified in 32/55 samples (58%, VAF 0.08-78.04%) which were taken during hematological complete remission (CR) in both patients after allogeneic stem cell transplantation (aHSCT) and patients after consolidation chemotherapy. In patients after aHSCT (n = 25), at least one previously known mutation was detected in 16/29 cfDNA samples (55.1%, VAF 0.08-6.7%) obtained when donor chimerism was ≥ 90% and in 6/6 samples (100%, VAF: 0.88-63.77%) with reduced donor chimerism. Probability of progression-free survival 17 months after aHSCT in patients with donor chimerism ≥ 90% but mutation-positive cfDNA was 64% compared to 100% in patients with undetectable MRD. In patients after consolidation chemotherapy, cfDNA was positive in all samples taken during CR (n = 4; VAF 0.26-29.84%) and non-CR (n = 4; VAF 8.46-100%). Our results indicate that NGS of cfDNA is suitable for MRD monitoring in AML and offers higher sensitivity for detecting residual leukemic cells than chimerism analysis in patients after aHSCT. Further studies are needed to evaluate clinical relevance of MRD status as determined in cfDNA.
急性髓系白血病（AML）中的微小/可测量残留病（MRD）定义为低于细胞形态学检测阈值的持续性白血病细胞。循环游离DNA（cfDNA）的下一代测序（NGS）用于分析癌症相关突变，已被证明可用于定量实体瘤中的疾病负担，也有人建议其可对AML患者进行微创随访。在这项初步研究中，我们调查了基于cfDNA的MRD监测在异基因干细胞移植后AML中的技术敏感性和潜在预后意义，并与供体嵌合分析进行比较，或者分别在巩固化疗后进行比较。使用市售的10或37基因热点 panel（VariantPlex Core AML或Core Myeloid panel，ArcherDx）通过靶向NGS分析了来自29名患者的75份cfDNA样本。通过常规诊断确定患者的白血病表现出1 - 7个突变。MRD评估仅考虑先前鉴定的突变。从所有样本中分离出足够量的用于NGS的cfDNA（总产量24 ng - 5.2 μg）。变异检测的敏感性随着总读数增加和更高的突变特异性覆盖率而增加（变异等位基因频率[VAF]范围为0.08 - 100%）。在异基因干细胞移植（aHSCT）后和巩固化疗后的患者中，在血液学完全缓解（CR）期间采集的32/55个样本（58%，VAF 0.08 - 78.04%）中鉴定出至少一个先前已知的突变。在aHSCT后的患者（n = 25）中，当供体嵌合率≥90%时，在16/29份cfDNA样本（55.1%，VAF 0.08 - 6.7%）中检测到至少一个先前已知的突变，在供体嵌合率降低的6/6个样本（100%，VAF：0.88 - 63.77%）中也检测到。供体嵌合率≥90%但cfDNA突变阳性的患者在aHSCT后17个月无进展生存的概率为64%，而MRD不可检测的患者为100%。在巩固化疗后的患者中，CR期间采集的所有样本（n = 4；VAF 0.26 - 29.84%）和非CR期间采集的所有样本（n = 4；VAF 8.46 - 100%）的cfDNA均为阳性。我们的结果表明，cfDNA的NGS适用于AML中的MRD监测，并且与aHSCT后患者的嵌合分析相比，在检测残留白血病细胞方面具有更高的敏感性。需要进一步研究来评估cfDNA中确定的MRD状态的临床相关性。
