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浙江大学学报（医学版）
浙江大学学报（医学版）  2019, Vol. 48 Issue (1): 50-57    DOI: 10.3785/j.issn.1008-9292.2019.02.09
原著     
肾母细胞瘤1基因表达及其对急性髓系白血病患者预后的预测价值
杜东芬1,2(),朱丽霞1,王云贵3,叶琇锦1()
1. 浙江大学医学院附属第一医院血液病高级病区， 浙江 杭州 310003
2. 浙江省绍兴市中心血站， 浙江 绍兴 312000
3. 浙江大学医学院附属第一医院血液科 浙江大学血液病研究所， 浙江 杭州 310003
Expression of WT1 gene and its prognostic value in patients with acute myeloid leukemia
DU Dongfen1,2(),ZHU Lixia1,WANG Yungui3,YE XiujinG1()
1.Department of Hematology, the First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou 310003, China
2.Shaoxing Central Blood Station, Shaoxing 312000, Zhejiang Province, China
3.Department of Hematology and Institute of Hematology, the First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou 310003, China
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摘要： 目的

分析急性髓系白血病（AML）患者肾母细胞瘤1（WT1）基因表达水平的差异，探讨WT1表达及其在核仁磷酸蛋白1（NPM1）或Fms样酪氨酸激酶3内部串联重复（FLT3-ITD）不同突变状态下与AML患者疗效和生存的相关性，评估其对于患者预后的预测价值。

 方法

以167例初发AML患者（除外M3亚型）为研究对象，根据初诊时WT1表达水平分为WT1高表达组和WT1低表达组，回顾性分析两组的临床及实验室检查资料，采用Kaplan-Meier法进行患者生存分析以明确WT1表达水平在预后评估中的价值，特别是在NPM1或FLT3-ITD不同突变状态AML患者中的预后评估价值。

 结果

WT1高表达组126例患者中治疗有效83例（65.9%），低表达组41例患者中治疗有效39例（95.1%），差异有统计学意义（P＜0.01）。WT1高表达组2年总存活率低于WT1低表达组（46.1%和75.2%，P＜0.05），2年无病存活率也低于WT1低表达组（43.5%和68.5%，P＜0.05）。诱导化疗前后WT1表达量下降≥1 log患者总反应率和2年总存活率优于下降＜1 log患者（均P＜0.05），但两者2年无病存活率差异无统计学意义（P＞0.05）。NPM1野生型患者中，WT1高表达组总反应率、2年总存活率较WT1低表达组低（均P＜0.05）；FLT3-ITD野生型患者中，WT1高表达组总反应率、2年总存活率和2年无病存活率均较WT1低表达组低（均P＜0.05）；而NPM1或FLT3-ITD突变患者中，不同WT1表达水平组疗效及生存分析结果差异无统计学意义（均P＞0.05）。

 结论

初发AML患者中WT1基因过表达提示预后不良；诱导治疗后WT1表达量下降≥1 log的患者预后优于下降＜1 log的患者；初诊WT1基因表达水平可以作为NPM1或FLT3-ITD野生型AML患者的预后判断指标。
关键词： 白血病，髓样，急性/病理学;  基因，肾母细胞瘤;  磷蛋白类/遗传学;  核蛋白质类/遗传学;  蛋白酪氨酸激酶类/遗传学;  fms样酪氨酸激酶3/遗传学;  突变;  预后    
Abstract: Objective

To investigate the expression of Wilms’ tumor 1 (WT1) gene in patients with acute myeloid leukemia (AML), and to explore its application in predicting prognosis of AML in patients with wild or mutated nucleophosmin 1(NPM1) and Fms-like tyrosine kinase 3-internal tandem duplication (FLT3-ITD).

 Methods

One hundred and sixty-seven newly diagnosed AML patients（exclued M3 type）were enrolled in this study. The survival of patients were analyzed with Kaplan-Meier method. The clinical data, laboratory findings and the survival of patients were analyzed and compared between patients with high WT1 expression (high-WT1 group) and those with low WT1 expression (low-WT1 group), as well as among the patients with NPM1 or FLT3-ITD wild type and mutants.

 Results

The overall response rates (ORR) in high-WT1 and low-WT1 groups were 65.9% (83/126) and 95.1% (39/41), respectively (P＜0.01). Compared with the low-WT1 group, the high-WT1 group had lower 2-y overall survival (OS) rate (46.1% vs. 75.2%，P＜0.05) and 2-y disease free survival (DFS) rate (43.5% vs. 68.5%，P＜0.05). After induction chemotherapy, the patients with decreased WT1 gene expression ≥1log was associated with higher ORR and 2-y OS rate (all P＜0.05), but the advantage of 2-y DFS rate was not shown (P＞0.05). In patients with NPM1 wild type, the high-WT1 group had inferior ORR and 2-y OS rate (all P＜0.05), while in the patients with FLT3-ITD wild type, the high-WT1 group had inferior ORR, 2-y OS rate and 2-y DFS rate (all P＜0.05). In patients with NPM1 or FLT3-ITD mutations, the WT1 expression had no significantly predicting values in treatment efficacy and survival (all P＞0.05).

 Conclusions

WT1 gene overexpression indicated poor prognosis of AML patients; the patients with decreased WT1 gene expression ≥1 log after the first induction therapy show better prognosis than those with＜1 log. The WT1 gene expression level at diagnosis can be used as an unfavorable prognostic factor for AML patients with NPM1 or FLT3-ITD wild types.
Key words: Leukemia, myeloid, acute/pathology    Genes, Wilms tumor    Phosphoproteins/genetics    Nuclear proteins/genetics    Protein-tyrosine Kinases/genetics    fms-like tyrosine kinase 3/genetics    Mutation    Prognosis
收稿日期: 2018-09-18 出版日期: 2019-05-10
:  R557  
基金资助: 浙江省医药卫生科技计划(2017KY059)
通讯作者: 叶琇锦     E-mail: dongfen166@163.com;yxjsunny@zju.edu.cn
作者简介: 杜东芬（1980—），女，硕士研究生，主治医师，主要从事血液病学研究；E-mail： dongfen166@163.com； https://orcid.org/0000-0003-1429-3345|叶琇锦（1962—），女，博士，主任医师，硕士生导师，主要从事血液病学研究；E-mail： yxjsunny@zju.edu.cn； https://orcid.org/0000-0003-1264-0307
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引用本文:

杜东芬,朱丽霞,王云贵,叶琇锦. 肾母细胞瘤1基因表达及其对急性髓系白血病患者预后的预测价值[J]. 浙江大学学报（医学版）, 2019, 48(1): 50-57.

DU Dongfen,ZHU Lixia,WANG Yungui,YE XiujinG. Expression of WT1 gene and its prognostic value in patients with acute myeloid leukemia. J Zhejiang Univ (Med Sci), 2019, 48(1): 50-57.

链接本文:

http://www.zjujournals.com/med/CN/10.3785/j.issn.1008-9292.2019.02.09    或    http://www.zjujournals.com/med/CN/Y2019/V48/I1/50

基因或探针名称 序列（5′-3′）
WT1 正向：GATAACCACACAACGCCCATC
反向：CACACGTCGCACATCCTGAAT

WT1探针

 FAM-ACACCGTGCGTGTGTATTCTGTATTGG-TAMRA

ABL

正向：TGGAGATAACACTCTAAGCATAACTAAAGGT
反向：GATGTAGTTGCTTGGGACCCA
ABL探针 GATGTAGTTGCTTGGGACCCA

FLT3-ITD

正向：FAM-AGCAATTTAGGTATGAAAGCCAGCTA
反向：CTTTCAGCATTTTGACGGCAACC

NPM1

正向：GTTTCTTTTTTTTTTTTTCCAGGCTATTCAAG
反向：HEX-CACGGTAGGGAAAGTTCTCACTCTGC
表 1  PCR所用引物序列
组　别 n 年龄（岁） 性别（男/女） 白细胞(×109/L) 血红蛋白(g/L) 血小板(×109/L) 肌酐(μmol/L) 乳酸脱氢酶(IU/L)
WT1高表达组 126 48±15 71/55 14.2(3.8,53.1) 84±22 45.0(24.0,87.3) 64.0(53.0,75.0) 378(237,768)
WT1低表达组 41 46±15 19/22 9.3(3.5,31.2) 101±23 55.0(28.0,95.5) 69.0(57.0,84.0) 358(229,691)
t/χ 2/Z值 — -0.47 1.25 -0.99 4.30 -0.95 -1.79 -0.31
P值 — ＞0.05 ＞0.05 ＞0.05 ＜0.01 ＞0.05 ＞0.05 ＞0.05
组　别 n 骨髓原始细胞比例(%) 染色体核型异常 NPM1突变* FLT3-ITD突变*

FBA分型

(M0/M1/M2/M4/M5/M6)

 既往肿瘤史
WT1高表达组 126 62.0(39.9,80.0) 33(75.0) 30(25.6) 27(23.1) 7/8/67/6/38/0/14 14(82.4)
WT1低表达组 41 57.0(39.0,82.5) 11(25.0) 5(13.9) 2(5.6) 4/1/16/1/18/1/3 3(17.6)
t/χ 2/Z值 — -0.12 0.01 2.16 5.50 8.17 0.49
P值 — ＞0.05 ＞0.05 ＞0.05 ＜0.05 ＞0.05 ＞0.05
表 2  不同WT1基因表达水平急性髓系白血病患者临床及遗传学特征比较
图 1   WT1基因不同表达水平患者总生存和无病生存曲线比较
图 2   WT1基因表达量不同下降幅度患者总生存和无病生存曲线比较
图 3   WT1基因不同表达水平的NPM1野生型患者总生存和无病生存曲线比较
图 4   WT1基因不同表达水平的FLT3-ITD野生型患者总生存和无病生存曲线比较
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