Pim-2抑制前列腺癌细胞凋亡信号通路的研究
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摘要
目的探讨癌基因Pim-2抑制前列腺癌细胞凋亡的机制及其信号通路。方法分别检测前列腺癌组织及前列腺增生组织中Pim-2及其下游信号通路因子的表达,通过RNAi技术分别沉默前列腺癌细胞株中Pim-2或XIAP表达,通过基因转染技术增强非肿瘤性前列腺上皮细胞中Pim-2或XIAP表达,检测eIF4B磷酸化水平及XIAP表达水平,测定细胞凋亡率。结果 Pim-2在前列腺癌组织及细胞株中的表达显著高于前列腺增生组织及非肿瘤性前列腺细胞株(P <0. 05);在非肿瘤性前列腺细胞株中转染Pim-2后,其Pim-2表达水平显著增高,eIF4B磷酸化水平及XIAP表达水平也同时显著增高,而细胞凋亡率则显著降低(P <0. 05);在前列腺癌细胞株中转染Pim-2 SiRNA后,Pim-2表达水平显著减低,eIF4B磷酸化水平及XIAP表达水平也同时显著降低,而细胞凋亡率则显著增高(P <0. 05);在高表达Pim-2及XIAP的前列腺癌细胞株中转染XIAP SiRNA沉默其表达后,细胞凋亡率显著增高(P <0. 05);在低表达Pim-2及XIAP的非肿瘤性前列腺细胞株中转染XIAP增强其表达后,细胞凋亡率无明显变化。结论 Pim-2在前列腺癌细胞中异常高表达; Pim-2可通过磷酸化eIF4B促进XIAP表达,抑制细胞凋亡;仅当Pim-2与XIAP同时高表达时才能抑制前列腺癌细胞凋亡。
Objective To explore the mechanism by which Pim-2 signal pathway inhibits apoptosis of prostate cancer cells. Methods We examined the expression levels of Pim-2 and the downstream factors in prostate cancer tissues,benign prostatic hyperplasia tissues,prostate cancer cell lines and prostatic epithelial cells. In prostate cancer DU-145 cell line with Pim-2 or XIAP silencing using RNA interference technique and in non-neoplastic prostatic epithelial cell line RWPE-1 over-expressing Pim-2 or XIAP via gene transfection,we detected the phosphorylation level of eIF4 B and the expression of XIAP and examined the apoptosis rate of the cells. Results Pim-2 was significantly over-expressed in prostate cancer tissues and prostate cancer cell lines as compared with benign prostatic hyperplasia tissue and non-neoplastic prostatic epithelial cells. RWPE-1 cells over-expressing Pim-2 showed significantly increased phosphorylation level of eIF4 B with enhanced XIAP expression and a significantly lowered apoptosis rate. DU-145 cells with Pim-2 silencing exhibited significantly lowered phosphorylation level of eIF4 B and XIAP expression with obviously increased cell apoptosis rate. XIAP silencing in DU-145 cells that over-expressed Pim-2 and XIAP caused a significant increase in the cell apoptosis rate; XIAP over-expression in RWPE-1 cells with low expressions of Pim-2 and XIAP did not cause significant changes in the cell apoptosis rate. Conclusion Pim-2 is over-expressed in prostate cancer cells. Pim-2 may inhibit the apoptosis of prostate cancer cells through phosphorylating eIF4 B to promote XIAP expression,and this inhibitory effect occurs only when both Pim-2 and XIAP are over-expressed in prostate cancer cells.
Objective To explore the mechanism by which Pim-2 signal pathway inhibits apoptosis of prostate cancer cells. Methods We examined the expression levels of Pim-2 and the downstream factors in prostate cancer tissues,benign prostatic hyperplasia tissues,prostate cancer cell lines and prostatic epithelial cells. In prostate cancer DU-145 cell line with Pim-2 or XIAP silencing using RNA interference technique and in non-neoplastic prostatic epithelial cell line RWPE-1 over-expressing Pim-2 or XIAP via gene transfection,we detected the phosphorylation level of eIF4 B and the expression of XIAP and examined the apoptosis rate of the cells. Results Pim-2 was significantly over-expressed in prostate cancer tissues and prostate cancer cell lines as compared with benign prostatic hyperplasia tissue and non-neoplastic prostatic epithelial cells. RWPE-1 cells over-expressing Pim-2 showed significantly increased phosphorylation level of eIF4 B with enhanced XIAP expression and a significantly lowered apoptosis rate. DU-145 cells with Pim-2 silencing exhibited significantly lowered phosphorylation level of eIF4 B and XIAP expression with obviously increased cell apoptosis rate. XIAP silencing in DU-145 cells that over-expressed Pim-2 and XIAP caused a significant increase in the cell apoptosis rate; XIAP over-expression in RWPE-1 cells with low expressions of Pim-2 and XIAP did not cause significant changes in the cell apoptosis rate. Conclusion Pim-2 is over-expressed in prostate cancer cells. Pim-2 may inhibit the apoptosis of prostate cancer cells through phosphorylating eIF4 B to promote XIAP expression,and this inhibitory effect occurs only when both Pim-2 and XIAP are over-expressed in prostate cancer cells.
引文
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[12] REN K,GOU X,XIAO M,et al. The over-expression of Pim-2 promote the tumorigenesis of prostatic carcinoma through phosphorylating e IF4B[J]. Prostate,2013,73(13):1462-1469. DOI:10. 1002/pros. 22693.
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[15] YE C,ZHANG C,HUANG H,et al. The natural compound myricetin effectively represses the malignant progression of prostate cancer by inhibiting PIM1 and disrupting the PIM1/CXCR4 interaction[J]. Cell Physiol Biochem,2018,48(3):1230-1244. DOI:10. 1159/000492009.
[16] MOLOGNI L,MAGISTRONI V,CASUSCELLI F,et al.The novel PIM1 inhibitor NMS-P645 reverses PIM1-dependent effects on TMPRSS2/ERG positive prostate cancer cells and shows anti-proliferative activity in combination with PI3K inhibition[J]. J Cancer,2017,8(1):140-145.
[17] QU Y,ZHANG C,DU E,et al. Pim-3 is a critical risk factor in development and prognosis of prostate cancer[J]. Med Sci Monit,2016,22:4254-4260. DOI:10. 12659/msm.898223.
[18] SHAHBAZIAN D,ROUX P P,MIEULET V,et al. The m TOR/PI3K and MAPK pathways converge on e IF4B to control its phosphorylation and activity[J]. EMBO J,2006,25(12):2781-2791. DOI:10. 1038/sj. emboj. 7601166.
[19] KROCZYNSKA B,KAUR S,KATSOULIDIS E,et al.Interferon-dependent engagement of eukaryotic initiation factor 4B via S6 kinase(S6K)-and ribosomal protein S6Kmediated signals[J]. Mol Cell Biol,2009,29(10):2865-2875. DOI:10. 1128/MCB. 01537-08.
[20] CHEN L S,REDKAR S,TAVERNA P,et al. Mechanisms of cytotoxicity to Pim kinase inhibitor,SGI-1776,in acute myeloid leukemia[J]. Blood,2011,118(3):693-702.DOI:10. 1182/blood-2010-12-323022.
[21] ZEMSKOVA M,SAHAKIAN E,BASHKIROVA S,et al.The PIM1 kinase is a critical component of a survival pathway activated by docetaxel and promotes survival of docetaxel-treated prostate cancer cells[J]. J Biol Chem,2008,283(30):20635-20644. DOI:10. 1074/jbc. M709479200.
[22] KAPELKO-SOWIK K,URBANIAK-KUJDA D, WOOWIEC D,et al. Expression of Pim-2 and NF-κB genes is increased in patients with acute myeloid leukemia(AML)and acute lymphoblastic leukemia(ALL)and is associated with complete remission rate and overall survival[J].Postepy Hig Med Dosw(Online),2013,67:553-559.
[23] ASANO J,NAKANO A,ODA A,et al. The serine/threonine kinase Pim-2 is a novel anti-apoptotic mediator in myeloma cells[J]. Leukemia,2011,25(7):1182-1188.DOI:10. 1038/leu. 2011. 60.
[24] PARK Y H,SEO S Y,LEE E,et al. Simvastatin induces apoptosis in castrate resistant prostate cancer cells by deregulating nuclear factor-κB pathway[J]. J Urol,2013,189(4):1547-1552. DOI:10. 1016/j. juro. 2012. 10. 030.
[25] KIM H R,PARK C G,JUNG J Y. Acacetin(5,7-dihydroxy-4’-methoxyflavone)exhibits in vitro and in vivo anticancer activity through the suppression of NF-κB/Akt signaling in prostate cancer cells[J]. Int J Mol Med,2014,33(2):317-324. DOI:10. 3892/ijmm. 2013. 1571.
[26] RODRíGUEZ-BERRIGUETE G,TORREALBA N,ORTEGA M A,et al. Prognostic value of inhibitors of apoptosis proteins(IAPs)and caspases in prostate cancer:caspase-3forms and XIAP predict biochemical progression after radical prostatectomy[J]. BMC Cancer,2015,15:809. DOI:10.1186/s12885-015-1839-z.
[27] SELIGSON D B,HONGO F,HUERTA-YEPEZ S,et al.Expression of X-linked inhibitor of apoptosis protein is a strong predictor of human prostate cancer recurrence[J].Clin Cancer Res,2007,13(20):6056-6063. DOI:10.1158/1078-0432. CCR-07-0960.
[28] HWANG C,OETJEN K A,KOSOFF D,et al. X-linked inhibitor of apoptosis deficiency in the TRAMP mouse prostate cancer model[J]. Cell Death Differ,2008,15(5):831-840. DOI:10. 1038/cdd. 2008. 15.
[29] OWENS T W,FOSTER F M,VALENTIJN A,et al. Role for X-linked inhibitor of apoptosis protein upstream of mitochondrial permeabilization[J]. J Biol Chem,2010,285(2):1081-1088. DOI:10. 1074/jbc. M109. 072322.
[30] GOGADA R,PRABHU V,AMADORI M,et al. Resveratrol induces p53-independent,X-linked inhibitor of apoptosis protein(XIAP)-mediated bax protein oligomerization on mitochondria to initiate cytochrome crelease and caspase activation[J]. J Biol Chem,2011,286(33):28749-28760.DOI:10. 1074/jbc. M110. 202440.
[2] AZIZ A U R,FARID S,QIN K,et al. Pim kinases and their relevance to the pi3k/akt/mtor pathway in the regulation of ovarian cancer[J]. Biomolecules,2018,8(1):E7. DOI:10. 3390/biom8010007.
[3] ISHIKAWA C,SENBA M,HASHIMOTO T,et al. Expression and significance of Pim-3 kinase in adult T-cell leukemia[J]. Eur J Haematol,2017,99(6):495-504. DOI:10.1111/ejh. 12940.
[4] HIASA M,TERAMACHI J,ODA A,et al. Pim-2 kinase is an important target of treatment for tumor progression and bone loss in myeloma[J]. Leukemia,2015,29(1):207-217. DOI:10. 1038/leu. 2014. 147.
[5] REN K,ZHANG W,SHI Y,et al. Pim-2 activates API-5 to inhibit the apoptosis of hepatocellular carcinoma cells through NF-κB pathway[J]. Pathol Oncol Res,2010,16(2):229-237. DOI:10. 1007/s12253-009-9215-4.
[6] FOX C J,HAMMERMAN P S,CINALLI R M,et al. The serine/threonine kinase Pim-2 is a transcriptionally regulated apoptotic inhibitor[J]. Genes Dev,2003,17(15):1841-1854. DOI:10. 1101/gad. 1105003.
[7] KEANE N A,REIDY M,NATONI A,et al. Targeting the Pim kinases in multiple myeloma[J]. Blood Cancer J,2015:e325. DOI:10. 1038/bcj. 2015. 46.
[8] DAI H,LI R,WHEELER T,et al. Pim-2 upregulation:biological implications associated with disease progression and perinueral invasion in prostate cancer[J]. Prostate,2005,65(3):276-286. DOI:10. 1002/pros. 20294.
[9] DAI J M,ZHANG S Q,ZHANG W,et al. Antisense oligodeoxynucleotides targeting the serine/threonine kinase Pim-2 inhibited proliferation of DU-145 cells[J]. Acta Pharmacol Sin,2005,26(3):364-368. DOI:10. 1111/j. 1745-7254. 2005. 00050. x.
[10] WHITE E. The pims and outs of survival signaling:role for the Pim-2 protein kinase in the suppression of apoptosis by cytokines[J]. Genes Dev,2003,17(15):1813-1816.DOI:10. 1101/gad. 1123103.
[11] PENG C,KNEBEL A,MORRICE N A,et al. Pim kinase substrate identification and specificity[J]. J Biochem,2007,141(3):353-362. DOI:10. 1093/jb/mvm040.
[12] REN K,GOU X,XIAO M,et al. The over-expression of Pim-2 promote the tumorigenesis of prostatic carcinoma through phosphorylating e IF4B[J]. Prostate,2013,73(13):1462-1469. DOI:10. 1002/pros. 22693.
[13] OH H J,LEE J S,SONG D K,et al. D-glucosamine inhibits proliferation of human cancer cells through inhibition of p70S6K[J]. Biochem Biophys Res Commun,2007,360(4):840-845. DOI:10. 1016/j. bbrc. 2007. 06. 137.
[14] SHAHBAZIAN D,PARSYAN A,PETROULAKIS E,et al.Control of cell survival and proliferation by mammalian eukaryotic initiation factor 4B[J]. Mol Cell Biol,2010,30(6):1478-1485. DOI:10. 1128/MCB. 01218-09.
[15] YE C,ZHANG C,HUANG H,et al. The natural compound myricetin effectively represses the malignant progression of prostate cancer by inhibiting PIM1 and disrupting the PIM1/CXCR4 interaction[J]. Cell Physiol Biochem,2018,48(3):1230-1244. DOI:10. 1159/000492009.
[16] MOLOGNI L,MAGISTRONI V,CASUSCELLI F,et al.The novel PIM1 inhibitor NMS-P645 reverses PIM1-dependent effects on TMPRSS2/ERG positive prostate cancer cells and shows anti-proliferative activity in combination with PI3K inhibition[J]. J Cancer,2017,8(1):140-145.
[17] QU Y,ZHANG C,DU E,et al. Pim-3 is a critical risk factor in development and prognosis of prostate cancer[J]. Med Sci Monit,2016,22:4254-4260. DOI:10. 12659/msm.898223.
[18] SHAHBAZIAN D,ROUX P P,MIEULET V,et al. The m TOR/PI3K and MAPK pathways converge on e IF4B to control its phosphorylation and activity[J]. EMBO J,2006,25(12):2781-2791. DOI:10. 1038/sj. emboj. 7601166.
[19] KROCZYNSKA B,KAUR S,KATSOULIDIS E,et al.Interferon-dependent engagement of eukaryotic initiation factor 4B via S6 kinase(S6K)-and ribosomal protein S6Kmediated signals[J]. Mol Cell Biol,2009,29(10):2865-2875. DOI:10. 1128/MCB. 01537-08.
[20] CHEN L S,REDKAR S,TAVERNA P,et al. Mechanisms of cytotoxicity to Pim kinase inhibitor,SGI-1776,in acute myeloid leukemia[J]. Blood,2011,118(3):693-702.DOI:10. 1182/blood-2010-12-323022.
[21] ZEMSKOVA M,SAHAKIAN E,BASHKIROVA S,et al.The PIM1 kinase is a critical component of a survival pathway activated by docetaxel and promotes survival of docetaxel-treated prostate cancer cells[J]. J Biol Chem,2008,283(30):20635-20644. DOI:10. 1074/jbc. M709479200.
[22] KAPELKO-SOWIK K,URBANIAK-KUJDA D, WOOWIEC D,et al. Expression of Pim-2 and NF-κB genes is increased in patients with acute myeloid leukemia(AML)and acute lymphoblastic leukemia(ALL)and is associated with complete remission rate and overall survival[J].Postepy Hig Med Dosw(Online),2013,67:553-559.
[23] ASANO J,NAKANO A,ODA A,et al. The serine/threonine kinase Pim-2 is a novel anti-apoptotic mediator in myeloma cells[J]. Leukemia,2011,25(7):1182-1188.DOI:10. 1038/leu. 2011. 60.
[24] PARK Y H,SEO S Y,LEE E,et al. Simvastatin induces apoptosis in castrate resistant prostate cancer cells by deregulating nuclear factor-κB pathway[J]. J Urol,2013,189(4):1547-1552. DOI:10. 1016/j. juro. 2012. 10. 030.
[25] KIM H R,PARK C G,JUNG J Y. Acacetin(5,7-dihydroxy-4’-methoxyflavone)exhibits in vitro and in vivo anticancer activity through the suppression of NF-κB/Akt signaling in prostate cancer cells[J]. Int J Mol Med,2014,33(2):317-324. DOI:10. 3892/ijmm. 2013. 1571.
[26] RODRíGUEZ-BERRIGUETE G,TORREALBA N,ORTEGA M A,et al. Prognostic value of inhibitors of apoptosis proteins(IAPs)and caspases in prostate cancer:caspase-3forms and XIAP predict biochemical progression after radical prostatectomy[J]. BMC Cancer,2015,15:809. DOI:10.1186/s12885-015-1839-z.
[27] SELIGSON D B,HONGO F,HUERTA-YEPEZ S,et al.Expression of X-linked inhibitor of apoptosis protein is a strong predictor of human prostate cancer recurrence[J].Clin Cancer Res,2007,13(20):6056-6063. DOI:10.1158/1078-0432. CCR-07-0960.
[28] HWANG C,OETJEN K A,KOSOFF D,et al. X-linked inhibitor of apoptosis deficiency in the TRAMP mouse prostate cancer model[J]. Cell Death Differ,2008,15(5):831-840. DOI:10. 1038/cdd. 2008. 15.
[29] OWENS T W,FOSTER F M,VALENTIJN A,et al. Role for X-linked inhibitor of apoptosis protein upstream of mitochondrial permeabilization[J]. J Biol Chem,2010,285(2):1081-1088. DOI:10. 1074/jbc. M109. 072322.
[30] GOGADA R,PRABHU V,AMADORI M,et al. Resveratrol induces p53-independent,X-linked inhibitor of apoptosis protein(XIAP)-mediated bax protein oligomerization on mitochondria to initiate cytochrome crelease and caspase activation[J]. J Biol Chem,2011,286(33):28749-28760.DOI:10. 1074/jbc. M110. 202440.
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