文章：

高通量药物筛选发现ATR-CHK1通路是CALR突变造血细胞的治疗靶点

High-throughput drug screening identifies the ATR-CHK1 pathway as a therapeutic vulnerability of CALR mutated hematopoietic cells

原文发布日期：2021-07-31

DOI: 10.1038/s41408-021-00531-2

类型: Article

开放获取: 是

英文摘要：

Mutations of calreticulin (CALR) are the second most prevalent driver mutations in essential thrombocythemia and primary myelofibrosis. To identify potential targeted therapies for CALR mutated myeloproliferative neoplasms, we searched for small molecules that selectively inhibit the growth of CALR mutated cells using high-throughput drug screening. We investigated 89 172 compounds using isogenic cell lines carrying CALR mutations and identified synthetic lethality with compounds targeting the ATR-CHK1 pathway. The selective inhibitory effect of these compounds was validated in a co-culture assay of CALR mutated and wild-type cells. Of the tested compounds, CHK1 inhibitors potently depleted CALR mutated cells, allowing wild-type cell dominance in the co-culture over time. Neither CALR deficient cells nor JAK2V617F mutated cells showed hypersensitivity to ATR-CHK1 inhibition, thus suggesting specificity for the oncogenic activation by the mutant CALR. CHK1 inhibitors induced replication stress in CALR mutated cells revealed by elevated pan-nuclear staining for γH2AX and hyperphosphorylation of RPA2. This was accompanied by S-phase cell cycle arrest due to incomplete DNA replication. Transcriptomic and phosphoproteomic analyses revealed a replication stress signature caused by oncogenic CALR, suggesting an intrinsic vulnerability to CHK1 perturbation. This study reveals the ATR-CHK1 pathway as a potential therapeutic target in CALR mutated hematopoietic cells.
 

摘要翻译： 

钙网蛋白（CALR）突变是原发性血小板增多症和原发性骨髓纤维化中第二常见的驱动突变。为寻找针对CALR突变型骨髓增殖性肿瘤的潜在靶向疗法，我们通过高通量药物筛选，寻找能选择性抑制CALR突变细胞生长的小分子化合物。我们使用携带CALR突变的同基因细胞系对89,172种化合物进行了研究，发现靶向ATR-CHK1通路的化合物具有合成致死效应。这些化合物的选择性抑制作用在CALR突变型与野生型细胞的共培养实验中得到验证。在测试的化合物中，CHK1抑制剂能有效清除CALR突变细胞，使得野生型细胞在共培养体系中随时间推移逐渐占据主导。无论是CALR缺失细胞还是JAK2V617F突变细胞，均未表现出对ATR-CHK1抑制的超敏反应，这表明该效应特异针对突变CALR的致癌激活作用。CHK1抑制剂在CALR突变细胞中诱导了复制应激，表现为γH2AX的全核染色增强和RPA2的过度磷酸化。与此同时，由于DNA复制未完成，细胞周期停滞在S期。转录组学和磷酸化蛋白质组学分析揭示了致癌CALR引发的复制应激特征，表明其对CHK1扰动存在内在脆弱性。本研究揭示了ATR-CHK1通路作为CALR突变造血细胞的潜在治疗靶点。

原文链接：

High-throughput drug screening identifies the ATR-CHK1 pathway as a therapeutic vulnerability of CALR mutated hematopoietic cells