细胞周期相关激酶：人类胶质母细胞瘤中的一种新型候选致癌基因。

Cell cycle-related kinase: a novel candidate oncogene in human glioblastoma.

作者信息

Ng Samuel S M, Cheung Yuen-Ting, An Xiao-Meng, Chen Yang Chao, Li Ming, Li Gloria Hoi-Yee, Cheung William, Sze Johnny, Lai Lihui, Peng Ying, Xia Harry H X, Wong Benjamin C Y, Leung Suet-Yi, Xie Dan, He Ming-Liang, Kung Hsiang-Fu, Lin Marie C

机构信息

Department of Chemistry, Open Laboratory of Chemical Biology, The University of Hong Kong, Pokfulam, Hong Kong, China.

出版信息

J Natl Cancer Inst. 2007 Jun 20;99(12):936-48. doi: 10.1093/jnci/djm011. Epub 2007 Jun 12.

DOI:10.1093/jnci/djm011

PMID:17565152

Abstract

BACKGROUND

Median survival for patients with glioblastoma multiforme, the most aggressive glioma, is only 12-15 months, despite multimodal treatment that includes surgery, chemotherapy, and radiotherapy. Thus, identification of genes that control the progression of glioblastoma multiforme is crucial for devising new therapies. We investigated the involvement of cell cycle-related kinase (CCRK), a novel protein kinase that is homologous to cyclin-dependent kinase 7, in glioblastoma multiforme carcinogenesis.

METHODS

We analyzed the expression levels of CCRK in 26 glioma patient samples (19 high-grade and seven low-grade) and normal brain by semiquantitative reverse transcription-polymerase chain reaction assays. CCRK expression was knocked down in human glioma U-373 MG and U-87 MG cells with small-interfering RNAs and short hairpin RNAs (siCCRK and shCCRK, respectively), and cell proliferation, cell cycle distribution, and cyclin-dependent kinase 2 (CDK2) phosphorylation were examined. A subcutaneous nude mouse xenograft model (n = 4 mice per group) was used to study the effect of CCRK knockdown and overexpression on tumorigenicity and growth of glioblastoma multiforme cells in vivo. All statistical tests were two-sided.

RESULTS

CCRK mRNA was elevated at least 1.5-fold and as much as 3.7-fold in 14 (74%) of 19 high-grade glioblastoma multiforme patient samples and in four (80%) of five glioma cell lines examined compared with normal brain tissue. Suppression of CCRK by siCCRK inhibited the proliferation of U-373 MG and U-87 MG glioblastoma cells in a time- and dose-dependent manner. The growth-inhibiting effect of siCCRK was mediated via G1- to S-phase cell cycle arrest and reduced CDK2 phosphorylation. CCRK knockdown statistically significantly suppressed glioma cell growth in vivo as indicated by the mean tumor volumes at week 6 after tumor cell injection (U-373-control = 1352 mm3, U-373-shCCRK = 294 mm3, difference = 1058 mm3, 95% confidence interval [CI] = 677 to 1439 mm3, P<.001; U-87-control = 1910 mm3, U-87-shCCRK = 552 mm3, difference = 1358 mm3, 95% CI = 977 to 1739 mm3, P<.001).

CONCLUSIONS

CCRK is a candidate oncogene in glioblastoma multiforme tumorigenesis.

摘要

背景

多形性胶质母细胞瘤是最具侵袭性的胶质瘤，尽管采用了包括手术、化疗和放疗在内的多模式治疗，但其患者的中位生存期仅为12 - 15个月。因此，鉴定控制多形性胶质母细胞瘤进展的基因对于设计新的治疗方法至关重要。我们研究了细胞周期相关激酶（CCRK），一种与细胞周期蛋白依赖性激酶7同源的新型蛋白激酶，在多形性胶质母细胞瘤致癌过程中的作用。

方法

我们通过半定量逆转录 - 聚合酶链反应分析，检测了26例胶质瘤患者样本（19例高级别和7例低级别）及正常脑组织中CCRK的表达水平。分别用小干扰RNA和短发夹RNA（分别为siCCRK和shCCRK）敲低人胶质瘤U - 373 MG和U - 87 MG细胞中的CCRK表达，并检测细胞增殖、细胞周期分布和细胞周期蛋白依赖性激酶2（CDK2）磷酸化情况。采用皮下裸鼠异种移植模型（每组n = 4只小鼠）研究CCRK敲低和过表达对多形性胶质母细胞瘤细胞体内致瘤性和生长的影响。所有统计检验均为双侧检验。

结果

与正常脑组织相比，19例高级别多形性胶质母细胞瘤患者样本中的14例（74%）以及所检测的5株胶质瘤细胞系中的4株（80%），CCRK mRNA升高至少1.5倍，高达3.7倍。siCCRK对CCRK的抑制以时间和剂量依赖性方式抑制了U - 373 MG和U - 87 MG胶质母细胞瘤细胞的增殖。siCCRK的生长抑制作用是通过G1期至S期细胞周期阻滞和降低CDK2磷酸化介导的。如肿瘤细胞注射后第6周的平均肿瘤体积所示，CCRK敲低在统计学上显著抑制了体内胶质瘤细胞的生长（U - 373 - 对照 = 1352 mm³，U - 373 - shCCRK = 294 mm³，差异 = 1058 mm³，95%置信区间[CI] = 677至1439 mm³，P <.001；U - 87 - 对照 = 1910 mm³，U - 87 - shCCRK = 552 mm³，差异 = 1358 mm³，95% CI = 977至1739 mm³，P <.001）。