文章：

构建斑马鱼敲入模型以重现儿童ETV6::RUNX1阳性B细胞前体急性淋巴细胞白血病

Generation of a Zebrafish Knock-In Model Recapitulating Childhood ETV6::RUNX1-Positive B-Cell Precursor Acute Lymphoblastic Leukemia

原文发布日期：13 December 2023

DOI: 10.3390/cancers15245821

类型: Article

开放获取: 是

英文摘要：

Approximately 25% of children with B-cell precursor acute lymphoblastic leukemia (pB-ALL) harbor thet(12;21)(p13;q22)translocation, leading to theETV6::RUNX1 (E::R)fusion gene. This translocation occurs in utero, but the disease is much less common than the prevalence of the fusion in newborns, suggesting that secondary mutations are required for overt leukemia. The role of these secondary mutations remains unclear and may contribute to treatment resistance and disease recurrence. We developed a zebrafish model forE::Rleukemia using CRISPR/Cas9 to introduce the humanRUNX1gene into zebrafishetv6intron 5, resulting inE::Rfusion gene expression controlled by the endogenousetv6promoter. As seen by GFP fluorescence at a single-cell level, the model correctly expressed the fusion protein in the right places in zebrafish embryos. TheE::Rfusion expression induced an expansion of the progenitor cell pool and led to a low 2% frequency of leukemia. The introduction of targetedpax5andcdkn2a/bgene mutations, mimicking secondary mutations, in theE::Rline significantly increased the incidence in leukemia. Transcriptomics revealed that theE::R;pax5mut leukemias exclusively represented B-lineage disease. This novelE::Rzebrafish model faithfully recapitulates human disease and offers a valuable tool for a more detailed analysis of disease biology in this subtype.

摘要翻译： 

大约25%的B细胞前体急性淋巴细胞白血病（pB-ALL）患儿携带t(12;21)(p13;q22)易位，导致ETV6::RUNX1（E::R）融合基因形成。该易位发生于胎儿期，但疾病发生率远低于新生儿中该融合基因的检出率，提示显性白血病需要继发性突变参与。这些继发性突变的作用尚不明确，可能与治疗抵抗及疾病复发相关。我们利用CRISPR/Cas9技术将人类RUNX1基因插入斑马鱼etv6内含子5，构建了E::R白血病斑马鱼模型，使融合基因表达受内源性etv6启动子调控。单细胞水平的GFP荧光显示，该模型在斑马鱼胚胎的正确部位准确表达了融合蛋白。E::R融合表达诱导祖细胞池扩增，并导致2%的低白血病发生率。在E::R品系中引入模拟继发性突变的靶向pax5和cdkn2a/b基因突变，显著提高了白血病发病率。转录组学分析显示，E::R;pax5突变白血病完全表现为B系疾病。这一新型E::R斑马鱼模型准确再现了人类疾病特征，为深入解析该亚型疾病生物学机制提供了重要工具。

原文链接：

Generation of a Zebrafish Knock-In Model Recapitulating Childhood ETV6::RUNX1-Positive B-Cell Precursor Acute Lymphoblastic Leukemia