Department of Ageing Biology and Stem Cells, European Research Institute for the Biology of Ageing, University Medical Center Groningen, University of Groningen, Groningen, The Netherlands.
Oncode Institute, Utrecht, The Netherlands; Princess Máxima Center for Pediatric Oncology, Utrecht, The Netherlands.
Exp Hematol. 2020 Nov;91:46-54. doi: 10.1016/j.exphem.2020.09.188. Epub 2020 Sep 15.
Clonal heterogeneity fuels leukemia evolution, therapeutic resistance, and relapse. Upfront detection of therapy-resistant leukemia clones at diagnosis may allow adaptation of treatment and prevention of relapse, but this is hampered by a paucity of methods to identify and trace single leukemia-propagating cells and their clonal offspring. Here, we tested methods of cellular barcoding analysis, to trace the in vivo competitive dynamics of hundreds of patient-derived leukemia clones upon chemotherapy-mediated selective pressure. We transplanted Nod/Scid/Il2Rγ (NSG) mice with barcoded patient-derived or SupB15 acute lymphoblastic leukemia (ALL) cells and assessed clonal responses to dexamethasone, methotrexate, and vincristine, longitudinally and across nine anatomic locations. We illustrate that chemotherapy reduces clonal diversity in a drug-dependent manner. At end-stage disease, methotrexate-treated patient-derived xenografts had significantly fewer clones compared with placebo-treated mice (100 ± 10 vs. 160 ± 15 clones, p = 0.0005), while clonal complexity in vincristine- and dexamethasone-treated xenografts was unaffected (115 ± 33 and 150 ± 7 clones, p = NS). Using tools developed to assess differential gene expression, we determined whether these clonal patterns resulted from random clonal drift or selection. We identified 5 clones that were reproducibly enriched in methotrexate-treated patient-derived xenografts, suggestive of pre-existent resistance. Finally, we found that chemotherapy-mediated selection resulted in a more asymmetric distribution of leukemia clones across anatomic sites. We found that cellular barcoding is a powerful method to trace the clonal dynamics of human patient-derived leukemia cells in response to chemotherapy. In the future, integration of cellular barcoding with single-cell sequencing technology may allow in-depth characterization of therapy-resistant leukemia clones and identify novel targets to prevent relapse.
克隆异质性推动白血病的演进、治疗耐药和复发。在诊断时即检测到具有治疗耐药性的白血病克隆，可能有助于调整治疗并预防复发，但由于缺乏识别和追踪单个白血病起始细胞及其克隆后代的方法，这一目标受到了阻碍。在这里，我们测试了细胞条形码分析方法，以追踪数百个患者来源的白血病克隆在化疗介导的选择压力下的体内竞争动态。我们将带有条形码的患者来源或 SupB15 急性淋巴细胞白血病 (ALL) 细胞移植到 Nod/Scid/Il2Rγ (NSG) 小鼠中，并在九个解剖部位进行纵向和横向评估，以评估对地塞米松、甲氨蝶呤和长春新碱的克隆反应。我们表明，化疗以药物依赖性方式降低克隆多样性。在疾病晚期，与接受安慰剂治疗的小鼠相比，接受甲氨蝶呤治疗的患者衍生异种移植物的克隆数量明显减少（100±10 个与 160±15 个克隆，p=0.0005），而长春新碱和地塞米松治疗的异种移植物的克隆复杂性不受影响（115±33 个与 150±7 个克隆，p=NS）。我们使用开发的工具来评估差异基因表达，以确定这些克隆模式是由随机克隆漂移还是选择引起的。我们鉴定了 5 个在接受甲氨蝶呤治疗的患者衍生异种移植物中重复富集的克隆，提示存在预先存在的耐药性。最后，我们发现化疗介导的选择导致白血病克隆在解剖部位之间的分布更加不对称。我们发现，细胞条形码是一种强大的方法，可以追踪人类患者来源的白血病细胞对化疗的克隆动态。在未来，细胞条形码与单细胞测序技术的整合可能允许对治疗耐药性白血病克隆进行深入表征，并确定预防复发的新靶点。
