深度分子反应（DMR）被定义为BCR::ABL1转录物通过RT-qPCR检测的水平≤0.01％，是慢性髓性白血病（CML）患者成功中断治疗的先决条件。然而，在治疗无重启期（TFR）研究中，约50％的患者在BCR::ABL1转录物水平升高到超过0.1％阈值后不得不恢复治疗。为了提高转录物检测的敏感性和准确性，可以使用数字PCR（dPCR）分析转录物水平。 dPCR可以使BCR::ABL1转录物的检测敏感性提高10-100倍；然而，它是否能够更好地选择成功的TFR患者尚不清楚。除了免疫系统的作用之外，复发可能是由于存在转录上静默的残余白血病干细胞（LSCs）所引起的。流式细胞术可用于鉴定和定量循环骨髓Ph+ LSCs CD34+/CD38-并共表达CD26（双肽酶IV）。到目前为止，在TFR中循环Ph+ LSCs的重要性尚不清楚。本研究的目的是比较和检查以三种不同方法（RNA（RT-qPCR和dPCR）和LSC（流式细胞术）水平）检测CML最小残留病变（MRD）的值在TFR或表现为DMR的患者中。所招募的27名患者接受了伊马替尼（12），达沙替尼（6），尼洛替尼（7），波瑟替尼（1）或干扰素（1）的治疗。其中12名患者处于TFR状态，其余的患者表现为DMR。TFR患者治疗停药时间分别为不到1年（3），<3年（2），6年（6）和17年（1）。采集并同时使用三种方法对血样进行测试。与RT-qPCR相比，dPCR和LSCs均显示出更高的敏感性，对RT-qPCR无法检测到的样本呈阳性结果（17/27）。没有患者在dPCR中呈阴性；然而，23/27患者低于0.468拷贝/μL的阈值，对应着稳定的DMR。结果被分成四分位数，最低分位数被定义为最低MRD。这些数据在15/27患者中强相关，对应于几乎一半的TFR患者。事实上，TFR患者中有些人持续时间长达17年，对应于最低可检测的DMR类别。据我们所知，这是首次尝试使用标准（RT-qPCR）和高灵敏度（dPCR和LSCs）方法分析和比较CML人群中的DMR。

A Deep Molecular Response (DMR), defined as a BCR::ABL1 transcript at levels ≤ 0.01% by RT-qPCR, is the prerequisite for the successful interruption of treatment among patients with Chronic Myeloid Leukemia (CML). However, approximately 50% of patients in Treatment-Free Remission (TFR) studies had to resume therapy after their BCR::ABL1 transcript levels rose above the 0.1% threshold. To improve transcript detection sensitivity and accuracy, transcript levels can be analyzed using digital PCR (dPCR). dPCR increases BCR::ABL1 transcript detection sensitivity 10-100 fold; however, its ability to better select successful TFR patients remains unclear. Beyond the role of the immune system, relapses may be due to the presence of residual leukemic stem cells (LSCs) that are transcriptionally silent. Flow cytometry can be used to identify and quantify circulating bone marrow Ph+ LSCs CD34+/CD38- co-expressing CD26 (dipeptidylpeptidase-IV). To date, the significance of circulating Ph+ LSCs in TFR is unclear. The aim of this work is to compare and examine the values obtained using the three different methods of detecting minimal residual disease (MRD) in CML at RNA (RT-qPCR and dPCR) and LSC (flowcytometry) levels among patients in TFR or exhibiting a DMR. The twenty-seven patients enrolled received treatment with either imatinib (12), dasatinib (6), nilotinib (7), bosutinib (1), or interferon (1). Twelve patients were in TFR, while the rest exhibited a DMR. The TFR patients had stopped therapy for less than 1 year (3), <3 years (2), 6 years (6), and 17 years (1). Blood samples were collected and tested using the three methods at the same time. Both d-PCR and LSCs showed higher sensitivity than RT-qPCR, exhibiting positive results in samples that were undetectable using RT-qPCR (17/27). None of the patients tested negative with d-PCR; however, 23/27 were under the threshold of 0.468 copies/μL, corresponding to a stable DMR. The results were divided into quartiles, and the lowest quartiles defined the lowest MRD. These data were strongly correlated in 15/27 patients, corresponding to almost half of the TFR patients. Indeed, the TFR patients, some lasting up to 17 years, corresponded to the lowest detectable DMR categories. To the best of our knowledge, this is the first attempt to analyze and compare DMRs in a CML population using standard (RT-qPCR) and highly sensitive (dPCR and LSCs) methods.