Cyclin-dependent kinase-associated protein phosphatase is overexpressed in alcohol-related hepatocellular carcinoma 
and influences xenograft tumor growth

Lin,Wey-Ran; Lai,Ming-Wei; Yeh,Chau-Ting

doi:10.3892/or.2012.2208

Cyclin-dependent kinase-associated protein phosphatase is overexpressed in alcohol-related hepatocellular carcinoma and influences xenograft tumor growth

Authors:
- Wey-Ran Lin
- Ming-Wei Lai
- Chau-Ting Yeh
View Affiliations

Affiliations: Department of Gastroenterology and Hepatology, Linkou Chang Gung Memorial Hospital, Taoyuan, Taiwan, R.O.C., Liver Research Center, Taipei Chang Gung Memorial Hospital, Taipei, Taiwan, R.O.C.
Published online on: December 24, 2012 https://doi.org/10.3892/or.2012.2208
Pages: 903-910
Copyright: © Lin et al. This is an open access article distributed under the terms of Creative Commons Attribution License [CC BY_NC 3.0].

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

The cyclin-dependent kinase (Cdk)-associated protein phosphatase (KAP) is a dual-specificity phosphatase that dephosphorylates Cdk2 and inhibits cell cycle progression. The overexpression of KAP has been found in breast, prostate and renal cell carcinomas. However, the role of KAP in hepatocellular carcinoma (HCC) remains unclear. Therefore, the aim of this study was to investigate the expression of KAP in HCC and elucidate its role in tumorigenesis. HCC tissues from 117 patients undergoing surgical resection were collected for western blot analysis and immunohistochemichal analysis to establish clinical correlation. The antisense-mediated inhibition of KAP expression was performed in Huh-7 cell lines for tumorigenicity and growth regulation experiments. Clinicopathological analysis indicated that KAP was overexpressed in HCC tissue from alcoholic patients (P<0.001). It was significantly overexpressed in patients with a tumor number of <3 (P=0.0271), suggesting the potential role of KAP in tumorigenesis during early‑stage alcohol-related HCC. Additionally, the antisense-mediated inhibition of KAP in Huh-7 HCC cells interfered with cell cycle progression, decreased cell proliferation, reduced the colony‑forming ability of the cells and increased apoptosis. Tumorigenicity experiments showed that the KAP knockdown in Huh-7 cells generated smaller tumors in nude mice compared with the mock controls (P=0.018). In the cells in which KAP had been knocked down, the physical interaction between KAP and Cdk2 significantly increased, despite the reduced expression levels of KAP. The phosphorylation of cell proliferation and apoptosis-associated proteins, including phosphatase and tensin homolog (PTEN), glycogen synthase kinase (GSK), p44/42 and Akt, was decreased. Therefore, it can be concluded that KAP is overexpressed in alcohol-related HCC. The antisense-mediated knockdown of KAP in Huh-7 cells decreased cell proliferation, reduced the colony‑forming ability of the cells, interfered with cell cycle progression and suppressed xenograft tumor formation, partly through enhanced KAP and Cdk2 interaction.

View Figures

View References

1	Brown NR, Noble ME, Lawrie AM, et al: Effects of phosphorylation of threonine 160 on cyclin-dependent kinase 2 structure and activity. J Biol Chem. 274:8746–8756. 1999. View Article : Google Scholar : PubMed/NCBI
2	Solomon MJ: Activation of the various cyclin/cdc2 protein kinases. Curr Opin Cell Biol. 5:180–186. 1993. View Article : Google Scholar : PubMed/NCBI
3	Atherton-Fessler S, Hannig G and Piwnica-Worms H: Reversible tyrosine phosphorylation and cell cycle control. Semin Cell Biol. 4:433–442. 1993. View Article : Google Scholar : PubMed/NCBI
4	Den Haese GJ, Walworth N, Carr AM and Gould KL: The Wee1 protein kinase regulates T14 phosphorylation of fission yeast Cdc2. Mol Biol Cell. 6:371–385. 1995.PubMed/NCBI
5	Coleman TR and Dunphy WG: Cdc2 regulatory factors. Curr Opin Cell Biol. 6:877–882. 1994. View Article : Google Scholar : PubMed/NCBI
6	Kaldis P, Sutton A and Solomon MJ: The Cdk-activating kinase (CAK) from budding yeast. Cell. 86:553–564. 1996. View Article : Google Scholar : PubMed/NCBI
7	Nigg EA: Cyclin-dependent kinase 7: at the cross-roads of transcription, DNA repair and cell cycle control? Curr Opin Cell Biol. 8:312–317. 1996. View Article : Google Scholar : PubMed/NCBI
8	Lew DJ and Kornbluth S: Regulatory roles of cyclin dependent kinase phosphorylation in cell cycle control. Curr Opin Cell Biol. 8:795–804. 1996. View Article : Google Scholar : PubMed/NCBI
9	Poon RY and Hunter T: Dephosphorylation of Cdk2 Thr160 by the cyclin-dependent kinase-interacting phosphatase KAP in the absence of cyclin. Science. 270:90–93. 1995. View Article : Google Scholar : PubMed/NCBI
10	Gyuris J, Golemis E, Chertkov H and Brent R: Cdi1, a human G1 and S phase protein phosphatase that associates with Cdk2. Cell. 75:791–803. 1993. View Article : Google Scholar : PubMed/NCBI
11	Hannon GJ, Casso D and Beach D: KAP: a dual specificity phosphatase that interacts with cyclin-dependent kinases. Proc Natl Acad Sci USA. 91:1731–1735. 1994. View Article : Google Scholar : PubMed/NCBI
12	Song H, Hanlon N, Brown NR, Noble ME, Johnson LN and Barford D: Phosphoprotein-protein interactions revealed by the crystal structure of kinase-associated phosphatase in complex with phosphoCDK2. Mol Cell. 7:615–626. 2001. View Article : Google Scholar : PubMed/NCBI
13	Yeh CT, Lu SC, Chen TC, Peng CY and Liaw YF: Aberrant transcripts of the cyclin-dependent kinase-associated protein phosphatase in hepatocellular carcinoma. Cancer Res. 60:4697–4700. 2000.PubMed/NCBI
14	Yeh CT, Lu SC, Chao CH and Chao ML: Abolishment of the interaction between cyclin-dependent kinase 2 and Cdk-associated protein phosphatase by a truncated KAP mutant. Biochem Biophys Res Commun. 305:311–314. 2003. View Article : Google Scholar : PubMed/NCBI
15	Yu Y, Jiang X, Schoch BS, Carroll RS, Black PM and Johnson MD: Aberrant splicing of cyclin-dependent kinase-associated protein phosphatase KAP increases proliferation and migration in glioblastoma. Cancer Res. 67:130–138. 2007. View Article : Google Scholar : PubMed/NCBI
16	Lee SW, Reimer CL, Fang L, Iruela-Arispe ML and Aaronson SA: Overexpression of kinase-associated phosphatase (KAP) in breast and prostate cancer and inhibition of the transformed phenotype by antisense KAP expression. Mol Cell Biol. 20:1723–1732. 2000. View Article : Google Scholar : PubMed/NCBI
17	Lai MW, Chen TC, Pang ST and Yeh CT: Overexpression of cyclin-dependent kinase-associated protein phosphatase enhances cell proliferation in renal cancer cells. Urol Oncol. 30:871–878. 2012. View Article : Google Scholar : PubMed/NCBI
18	Forner A, Llovet JM and Bruix J: Hepatocellular carcinoma. Lancet. 379:1245–1255. 2012. View Article : Google Scholar
19	El-Serag HB: Hepatocellular carcinoma. N Engl J Med. 365:1118–1127. 2011. View Article : Google Scholar : PubMed/NCBI
20	Farazi PA and DePinho RA: Hepatocellular carcinoma pathogenesis: from genes to environment. Nat Rev Cancer. 6:674–687. 2006. View Article : Google Scholar : PubMed/NCBI
21	Villanueva A, Newell P, Chiang DY, Friedman SL and Llovet JM: Genomics and signaling pathways in hepatocellular carcinoma. Semin Liver Dis. 27:55–76. 2007. View Article : Google Scholar : PubMed/NCBI
22	El-Serag HB and Rudolph KL: Hepatocellular carcinoma: epidemiology and molecular carcinogenesis. Gastroenterology. 132:2557–2576. 2007. View Article : Google Scholar : PubMed/NCBI
23	Morgan TR, Mandayam S and Jamal MM: Alcohol and hepatocellular carcinoma. Gastroenterology. 127(5 Suppl 1): S87–S96. 2004. View Article : Google Scholar : PubMed/NCBI
24	McKillop IH and Schrum LW: Role of alcohol in liver carcinogenesis. Semin Liver Dis. 29:222–232. 2009. View Article : Google Scholar : PubMed/NCBI
25	Brooks PJ and Theruvathu JA: DNA adducts from acetaldehyde: implications for alcohol-related carcinogenesis. Alcohol. 35:187–193. 2005. View Article : Google Scholar : PubMed/NCBI
26	Koop DR: Oxidative and reductive metabolism by cytochrome P450 2E1. FASEB J. 6:724–730. 1992.PubMed/NCBI
27	Lieber CS: Cytochrome P-4502E1: its physiological and pathological role. Physiol Rev. 77:517–544. 1997.PubMed/NCBI
28	Badger TM, Ronis MJ, Seitz HK, Albano E, Ingelman-Sundberg M and Lieber CS: Alcohol metabolism: role in toxicity and carcinogenesis. Alcohol Clin Exp Res. 27:336–347. 2003. View Article : Google Scholar : PubMed/NCBI
29	Tanaka E, Terada M and Misawa S: Cytochrome P450 2E1: its clinical and toxicological role. J Clin Pharm Ther. 25:165–175. 2000. View Article : Google Scholar : PubMed/NCBI
30	Yang CS, Yoo JS, Ishizaki H and Hong JY: Cytochrome P450IIE1: roles in nitrosamine metabolism and mechanisms of regulation. Drug Metab Rev. 22:147–159. 1990. View Article : Google Scholar : PubMed/NCBI
31	Carter EA and Wands JR: Ethanol inhibits hormone stimulated hepatocyte DNA synthesis. Biochem Biophys Res Commun. 128:767–774. 1985. View Article : Google Scholar : PubMed/NCBI
32	Lad PJ, Shier WT, Skelly H, de Hemptinne B and Leffert HL: Adult rat hepatocytes in primary culture. VII. Proliferative and functional properties of cells from ethanol-intoxicated animals: evidence for a reversible albumin ‘production defect’. Alcohol Clin Exp Res. 6:72–79. 1982.PubMed/NCBI
33	Duguay L, Coutu D, Hetu C and Joly JG: Inhibition of liver regeneration by chronic alcohol administration. Gut. 23:8–13. 1982. View Article : Google Scholar : PubMed/NCBI
34	Wands JR, Carter EA, Bucher NL and Isselbacher KJ: Inhibition of hepatic regeneration in rats by acute and chronic ethanol intoxication. Gastroenterology. 77:528–531. 1979.PubMed/NCBI
35	McKillop IH, Vyas N, Schmidt CM, Cahill PA and Sitzmann JV: Enhanced Gi-protein-mediated mitogenesis following chronic ethanol exposure in a rat model of experimental hepatocellular carcinoma. Hepatology. 29:412–420. 1999. View Article : Google Scholar