Hou Panpan, Wu Chao, Wang Yuchen, Qi Rui, Bhavanasi Dheeraj, Zuo Zhixiang, Dos Santos Cedric, Chen Shuliang, Chen Yu, Zheng Hong, Wang Hong, Perl Alexander, Guo Deyin, Huang Jian
State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, P.R. China.
Department of Pathology and Laboratory Medicine, Lewis Katz School of Medicine, Temple University, Philadelphia, Pennsylvania.
Cancer Res. 2017 Aug 15;77(16):4402-4413. doi: 10.1158/0008-5472.CAN-16-1627. Epub 2017 Jun 16.
Acute myeloid leukemia (AML) is a malignant hematopoietic disease and the most common type of acute leukemia in adults. The mechanisms underlying drug resistance in AML are poorly understood. Activating mutations in FMS-like tyrosine kinase 3 (FLT3) are the most common molecular abnormality in AML. Quizartinib (AC220) is a potent and selective second-generation inhibitor of FLT3. It is in clinical trials for the treatment of relapsed or refractory FLT3-ITD-positive and -negative AML patients and as maintenance therapy. To understand the mechanisms of drug resistance to AC220, we undertook an unbiased approach with a novel CRISPR-pooled library to screen new genes whose loss of function confers resistance to AC220. We identified SPRY3, an intracellular inhibitor of FGF signaling, and GSK3, a canonical Wnt signaling antagonist, and demonstrated reactivation of downstream FGF/Ras/ERK and Wnt signaling as major mechanisms of resistance to AC220. We confirmed these findings in primary AML patient samples. Expression of and was dramatically reduced in AC220-resistant AML samples, and SPRY3-deleted primary AML cells were resistant to AC220. Intriguingly, expression of was greatly reduced in knockout AML cells, which positioned SPRY3 downstream of GSK3 in the resistance pathway. Taken together, our study identified novel genes whose loss of function conferred resistance to a selective FLT3 inhibitor, providing new insight into signaling pathways that contribute to acquired resistance in AML. .
急性髓系白血病（AML）是一种恶性造血疾病，也是成人中最常见的急性白血病类型。AML中耐药的潜在机制尚不清楚。FMS样酪氨酸激酶3（FLT3）的激活突变是AML中最常见的分子异常。奎扎替尼（AC220）是一种强效且选择性的第二代FLT3抑制剂。它正在进行临床试验，用于治疗复发或难治性FLT3-ITD阳性和阴性AML患者以及作为维持治疗。为了了解对AC220耐药的机制，我们采用了一种无偏见的方法，利用新型CRISPR混合文库筛选功能丧失会导致对AC220耐药的新基因。我们鉴定出FGF信号的细胞内抑制剂SPRY3和经典Wnt信号拮抗剂GSK3，并证明下游FGF/Ras/ERK和Wnt信号的重新激活是对AC220耐药的主要机制。我们在原发性AML患者样本中证实了这些发现。在AC220耐药的AML样本中， 和 的表达显著降低，并且缺失SPRY3的原发性AML细胞对AC220耐药。有趣的是，在 敲除的AML细胞中， 的表达大大降低，这将SPRY3定位在耐药途径中GSK3的下游。综上所述，我们的研究鉴定出功能丧失会导致对选择性FLT3抑制剂耐药的新基因，为导致AML获得性耐药的信号通路提供了新的见解。
