Chk1在胶质母细胞瘤中的表达及其与肿瘤生物学行为和预后生存的关联性
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摘要
目的研究细胞周期检测点激酶1(Checkpoint kinase 1,Chk1)在胶质母细胞瘤细胞(Glioblastoma,GBM)中的表达及其与GBM细胞增殖、成瘤活性和预后生存的关联性。方法利用TCGA数据库和脑肿瘤分子数据库(Rembrandt)选择和分析GBM中Chk1的表达,并采用Western blot和Real-time PCR等分子生物学技术检测GBM细胞中Chk1的表达水平;通过慢病毒转染siRNA沉默Chk1的表达以探究其对于GBM细胞增殖和成球能力的影响;同时利用免疫组化染色及Rembrandt数据库对Chk1与GBM患者的预后生存关系进行分析。结果 TCGA和Rembrandt数据库分析结果表明Chk1在GBM组织中呈现高表达,同样Western blot和Real-time PCR检测结果显示Chk1在GBM细胞中呈现高表达;慢病毒转染siRNA特异性沉默Chk1能够显著抑制U87细胞的生长(P<0.01)和成球能力(P<0.05);预后生存分析结果显示Chk1低表达GBM患者的临床预后显著优于高表达患者(P<0.001)。结论 Chk1在GBM细胞中过量表达,表达上调能够促进GBM细胞的生长和增殖,并和GBM患者的不良预后相关。
Objective The aim of this study was to investigate the expression of cell cycle checkpoint kinase 1(Chk1)gene in glioblastoma cells(GBM)and its correlation with GBM cell proliferation,tumorigenic activity and prognosis.Methods The expression of Chk1 in GBM cells was selected and analyzed by TCGA database and brain tumor molecular database(Rembrandt),and the level of Chk1 expression in GBM cells was detected by molecular biology techniques such as Western blot and Real-Time PCR.The expression of Chk1 was silenced by siRNA to investigate its effect on proliferation and colony-forming ability of GBM cells.The prognosis survival of GBM patients accompanying with Chk1 expression was analyzed by immunohistochemical staining and Rembrandt database.Results The results of TCGA database and Rembrandt showed that Chk1 gene was highly expressed in GBM tissues.Western blot and Real-Time PCR also showed that Chk1 gene was highly expressed in GBM cells.Lentiviral transfection siRNA-specific silencing of Chk1 significantly inhibited proliferation and colony-forming ability of U87 cells(P<0.01 and P<0.05).Prognostic survival analysis showed that GBM patients with low expression of Chk1 gene had a significantly better clinical outcome than those of GBM patients with high expression of Chk1 gene(P<0.001).Conclusion Chk1 gene is overexpressed in GBM cells,up-regulation of Chk1 gene expression can promote the growth and proliferation of GBM cells,and Chk1 gene is associated with poor prognosis in GBM patients.
Objective The aim of this study was to investigate the expression of cell cycle checkpoint kinase 1(Chk1)gene in glioblastoma cells(GBM)and its correlation with GBM cell proliferation,tumorigenic activity and prognosis.Methods The expression of Chk1 in GBM cells was selected and analyzed by TCGA database and brain tumor molecular database(Rembrandt),and the level of Chk1 expression in GBM cells was detected by molecular biology techniques such as Western blot and Real-Time PCR.The expression of Chk1 was silenced by siRNA to investigate its effect on proliferation and colony-forming ability of GBM cells.The prognosis survival of GBM patients accompanying with Chk1 expression was analyzed by immunohistochemical staining and Rembrandt database.Results The results of TCGA database and Rembrandt showed that Chk1 gene was highly expressed in GBM tissues.Western blot and Real-Time PCR also showed that Chk1 gene was highly expressed in GBM cells.Lentiviral transfection siRNA-specific silencing of Chk1 significantly inhibited proliferation and colony-forming ability of U87 cells(P<0.01 and P<0.05).Prognostic survival analysis showed that GBM patients with low expression of Chk1 gene had a significantly better clinical outcome than those of GBM patients with high expression of Chk1 gene(P<0.001).Conclusion Chk1 gene is overexpressed in GBM cells,up-regulation of Chk1 gene expression can promote the growth and proliferation of GBM cells,and Chk1 gene is associated with poor prognosis in GBM patients.
引文
1 Qi L,Ding L,Wang S,et al.A network meta-analysis:the overall and progression-free survival of glioma patients treated by different chemotherapeutic interventions combined with radiation therapy(RT)[J].Oncotarget,2016,7(42):69002-69013.
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3 Furusawa Y,Yamanouchi Y,Iizumi T,et al.Checkpoint kinase 2 is dispensable for regulation of the p53 response but is required for G2/M arrest and cell survival in cells with p53 defects under heat stress[J].Apoptosis,2017,22(10):1225-1234.
4 Gali-Muhtasib H,Kuester D,Mawrin C,et al.Thymoquinone triggers inactivation of the stress response pathway sensor CHEK1 and contributes to apoptosis in colorectal cancer cells[J].Cancer Res,2008,68(14):5609-56018.
5 Alcaraz-Sanabria A,Nieto-Jimenez C,Corrales-Sanchez V,et al.Synthetic lethality interaction between Aurora kinases and CHEK1 inhibitors in ovarian cancer[J].Mol Cancer Ther,2017,16(11):2552-2562.
6 Liu B,Qu J,Xu F,et al.MiR-195 suppresses non-small cell lung cancer by targeting CHEK1[J].Oncotarget,2015,6(11):9445-9456.
7 Lin WY,Brock IW,Connley D,et al.Associations of ATR and CHEK1 single nucleotide polymorphisms with breast cancer[J].PLoS One,2013,8(7):e68578.
8 Kim MK,Min DJ,Wright G,et al.Loss of compensatory pro-survival and anti-apoptotic modulator,IKKepsilon,sensitizes ovarian cancer cells to CHEK1 loss through an increased level of p21[J].Oncotarget,2014,5(24):12788-12802.
9 Kim MK,James J,Annunziata CM.Topotecan synergizes with CHEK1(CHK1)inhibitor to induce apoptosis in ovarian cancer cells[J].BMC Cancer,2015,15:196.
10 Bai X,Wang J,Huo L,et al.Serine/Threonine kinase CHEK1-dependent transcriptional regulation of RAD54L promotes proliferation and radio resistance in glioblastoma[J].Transl Oncol,2018,11(1):140-146.
11 Wang J,Cheng P,Pavlyukov MS,et al.Targeting NEK2 attenuates glioblastoma growth and radioresistance by destabilizing histone methyltransferase EZH2[J].J Clin Invest,2017,127(8):3075-3089.
12 Teo M,Martin S,Owusu-Agyemang K,et al.A survival analysis of GBM patients in the West of Scotland pre-and post-introduction of the Stupp regime[J].Br J Neurosurg,2014,28(3):351-355.
13 Duffau H.Glioblastoma in 2017[J].Rev Infirm,2017,66(228):16-18.
14 Sun X,Xiao D,Xu T,et al.miRNA-24-3p promotes cell proliferation and regulates chemosensitivity in head and neck squamous cell carcinoma by targeting CHD5[J].Future Oncol,2016,12(23):2701-2712.
15 Abdel-Fatah TM,Middleton FK,Arora A,et al.Untangling the ATR-CHEK1 network for prognostication,prediction and therapeutic target validation in breast cancer[J].Mol Oncol,2015,9(3):569-585.
16 Matsumoto K,Nagahara T,Okano J,et al.The growth inhibition of hepatocellular and cholangiocellular carcinoma cells by gemcitabine and the roles of extracellular signal-regulated and checkpoint kinases[J].Oncol Rep,2008,20(4):863-872.
17 Zuco V,Benedetti V,De Cesare M,et al.Sensitization of ovarian carcinoma cells to the atypical retinoid ST1926 by the histone deacetylase inhibitor,RC307:enhanced DNA damage response[J].Int J Cancer,2010,126(5):1246-1255.
18 Sankunny M,Parikh RA,Lewis DW,et al.Targeted inhibition of ATR or CHEK1 reverses radioresistance in oral squamous cell carcinoma cells with distal chromosome arm 11q loss[J].Genes Chromosomes Cancer,2014,53(2):129-143.
19 Lam MH,Rosen JM.Chk1 versus Cdc25:chking one′s levels of cellular proliferation[J].Cell Cycle,2004,3(11):1355-1357.
20 Kohn EA,Ruth ND,Brown MK,et al.Abrogation of the S phase DNA damage checkpoint results in S phase progression or premature mitosis depending on the concentration of 7-hydroxystaurosporine and the kinetics of Cdc25C activation[J].J Biol Chem,2002,277(29):26553-26564.
21 Levesque AA,Fanous AA,Poh A,et al.Defective p53 signaling in p53 wild-type tumors attenuates p21waf1 induction and cyclin B repression rendering them sensitive to Chk1 inhibitors that abrogate DNA damage-induced S and G2 arrest[J].Mol Cancer Ther,2008,7(2):252-262.
2 Signorovitch J,Li N,Ohashi E,et al.Overall survival(Os),quality of life(Qol),and Neurocognitive function(Nf)in recurrent glioblastoma multiforme(Gbm):a systematic literature review[J].Value Health,2015,18(7):433.
3 Furusawa Y,Yamanouchi Y,Iizumi T,et al.Checkpoint kinase 2 is dispensable for regulation of the p53 response but is required for G2/M arrest and cell survival in cells with p53 defects under heat stress[J].Apoptosis,2017,22(10):1225-1234.
4 Gali-Muhtasib H,Kuester D,Mawrin C,et al.Thymoquinone triggers inactivation of the stress response pathway sensor CHEK1 and contributes to apoptosis in colorectal cancer cells[J].Cancer Res,2008,68(14):5609-56018.
5 Alcaraz-Sanabria A,Nieto-Jimenez C,Corrales-Sanchez V,et al.Synthetic lethality interaction between Aurora kinases and CHEK1 inhibitors in ovarian cancer[J].Mol Cancer Ther,2017,16(11):2552-2562.
6 Liu B,Qu J,Xu F,et al.MiR-195 suppresses non-small cell lung cancer by targeting CHEK1[J].Oncotarget,2015,6(11):9445-9456.
7 Lin WY,Brock IW,Connley D,et al.Associations of ATR and CHEK1 single nucleotide polymorphisms with breast cancer[J].PLoS One,2013,8(7):e68578.
8 Kim MK,Min DJ,Wright G,et al.Loss of compensatory pro-survival and anti-apoptotic modulator,IKKepsilon,sensitizes ovarian cancer cells to CHEK1 loss through an increased level of p21[J].Oncotarget,2014,5(24):12788-12802.
9 Kim MK,James J,Annunziata CM.Topotecan synergizes with CHEK1(CHK1)inhibitor to induce apoptosis in ovarian cancer cells[J].BMC Cancer,2015,15:196.
10 Bai X,Wang J,Huo L,et al.Serine/Threonine kinase CHEK1-dependent transcriptional regulation of RAD54L promotes proliferation and radio resistance in glioblastoma[J].Transl Oncol,2018,11(1):140-146.
11 Wang J,Cheng P,Pavlyukov MS,et al.Targeting NEK2 attenuates glioblastoma growth and radioresistance by destabilizing histone methyltransferase EZH2[J].J Clin Invest,2017,127(8):3075-3089.
12 Teo M,Martin S,Owusu-Agyemang K,et al.A survival analysis of GBM patients in the West of Scotland pre-and post-introduction of the Stupp regime[J].Br J Neurosurg,2014,28(3):351-355.
13 Duffau H.Glioblastoma in 2017[J].Rev Infirm,2017,66(228):16-18.
14 Sun X,Xiao D,Xu T,et al.miRNA-24-3p promotes cell proliferation and regulates chemosensitivity in head and neck squamous cell carcinoma by targeting CHD5[J].Future Oncol,2016,12(23):2701-2712.
15 Abdel-Fatah TM,Middleton FK,Arora A,et al.Untangling the ATR-CHEK1 network for prognostication,prediction and therapeutic target validation in breast cancer[J].Mol Oncol,2015,9(3):569-585.
16 Matsumoto K,Nagahara T,Okano J,et al.The growth inhibition of hepatocellular and cholangiocellular carcinoma cells by gemcitabine and the roles of extracellular signal-regulated and checkpoint kinases[J].Oncol Rep,2008,20(4):863-872.
17 Zuco V,Benedetti V,De Cesare M,et al.Sensitization of ovarian carcinoma cells to the atypical retinoid ST1926 by the histone deacetylase inhibitor,RC307:enhanced DNA damage response[J].Int J Cancer,2010,126(5):1246-1255.
18 Sankunny M,Parikh RA,Lewis DW,et al.Targeted inhibition of ATR or CHEK1 reverses radioresistance in oral squamous cell carcinoma cells with distal chromosome arm 11q loss[J].Genes Chromosomes Cancer,2014,53(2):129-143.
19 Lam MH,Rosen JM.Chk1 versus Cdc25:chking one′s levels of cellular proliferation[J].Cell Cycle,2004,3(11):1355-1357.
20 Kohn EA,Ruth ND,Brown MK,et al.Abrogation of the S phase DNA damage checkpoint results in S phase progression or premature mitosis depending on the concentration of 7-hydroxystaurosporine and the kinetics of Cdc25C activation[J].J Biol Chem,2002,277(29):26553-26564.
21 Levesque AA,Fanous AA,Poh A,et al.Defective p53 signaling in p53 wild-type tumors attenuates p21waf1 induction and cyclin B repression rendering them sensitive to Chk1 inhibitors that abrogate DNA damage-induced S and G2 arrest[J].Mol Cancer Ther,2008,7(2):252-262.