FoxM1 is up-regulated in gastric cancer and its inhibition leads to cellular senescence, partially d

Jiping Zeng, 1,2,3†   Lixiang Wang, 1,4†   Qiao Li, 1,2 Wenjuan Li, 1,2 Magnus Bjrkholm, 1 Jihui Jia2* and Dawei Xu1*

1 Department ofMedicine, Division of Haematology, Karolinska University Hospital, Solna, and Karolinska Institutet, Stockholm, Sweden

2 Department ofMicrobiology/Key Laboratory for Experimental Teratology of Chinese Ministry of Education, Shandong University, Jinan, People’s Republic of China

3 Department of Biochemistry, Shandong University, Jinan, People’s Republic of China

4 Department of Pharmacology, School ofMedicine, Shandong University, Jinan, People’s Republic of China

*Correspondence to:

Dawei Xu, Haematology          Laboratory, Centre for Molecular Medicine (CMM), L8 : 00,         Karolinska University Hospital,   Solna, SE- 171 76

Stockholm, Sweden.

E-mail: Dawei.Xu@ki.se

Jihui Jia, Department of             Microbiology, School ofMedicine, Shandong University, Jinan,          250012, PR China.

E-mail: jiajihui@sdu.edu.cn

† These authors contributed

equally to this work.

No conflicts of interest were

declared.

Abstract

The FoxM1 transcription factor, a master regulator of mitotic gene expression, promotes the pathogenesis of several malignancies. However, little is known about its expression and function in gastric cancer. In the present study we determined   whether FoxM1 is over- expressed in gastric cancer, and whether it is required to maintain an immortal phenotype of gastric cancer cells. The over-expression of FoxM1 was observed in 37/42 tumour specimens from patients with gastric cancer. When FoxM1 in gastric cancer cells was knocked-down, impaired clonogenicity and cellular senescence occurred independently ofp53 and p16 status. FoxM1   depletion led to the down-regulation of its target genes c-MYC   and Skp2, coupled with the accumulation of the CDK inhibitor p27kip1 . Importantly, the FoxM1   inhibition- mediated cellular senescence and clonogenic defect was attenuated by the abolition ofp27kip 1 induction. Telomerase reverse transcriptase, the key component of telomerase essential for cellular immortalization, was also inhibited in the FoxM1-depleted cells. Taken together, the FoxM1   gene is aberrantly activated in gastric cancer and its inhibition triggers p53- and p16-independent senescence of cancer cells by regulating the expression of p27kip1   and other targets. These findings provide mechanistic insights into the role of FoxM1 in the pathogenesis of gastric cancer, which may have diagnostic and therapeutic implications in gastric cancer.

Copyright    2009 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Keywords:    FoxM1; gastric cancer; p27; senescence

Introduction

Gastric cancer   is   one   of   the   most   common   malig- nancies and   the   second   leading   cause   of   cancer- related death   worldwide   [1 – 4].   The   intimate   rela- tionship   between   Helicobacter pylori   infection and gastric cancer   has   long   been   established   and   it   is proposed   that H. pylori   infection   is   involved   in an early stage of gastric cancer pathogenesis by causing chronic gastritis, the precursor to all the abnormalities characteristic   of gastric   carcinogenesis   [5 – 8].   How- ever, these   pre-malignant   lesions   only   develop   in   a proportion of infected subjects and do not necessar- ily progress   into   invasive   cancers.   Host   susceptibil- ity, the pathogen’s virulence, environmental and many other factors significantly affect the oncogenic process initiated by   H. pylori   infection,   but   the   underlying molecular mechanism is unclear   [1,5,7,9 – 12]. Addi- tionally, because of lack of reliable screening markers

and efficient intervention approaches, the prognosis of gastric cancer remains poor. Therefore, improving our understanding   of the pathogenesis   of gastric cancer, identifying new biomarkers, and exploring novel ther- apeutic targets for treatment are urgently demanding tasks [1 – 4].

A hallmark   of   cancers,   including   gastric   cancer, is unlimited cellular proliferation due to the aberrant expression of key factors regulating cell proliferation and survival   [13]. Recent studies have shed light on cellular senescence, an irreversible cell growth arrest status,   as   a   potent   tumour   suppression mechanism [14 – 17]. Evidence has also accumulated that induction of senescence contributes to the treatment efficacy of chemotherapeutic agents and other anti-cancer strate- gies   [18 – 20].   It   is   known   that   activation   of telom- erase through induction of its rate-limiting component telomerase reverse transcriptase (hTERT) is a critical

Copyright 2009 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

www.pathsoc.org.uk

426

activity [21,53]. We thus   suggest that p27kip1   coop- erates with   other   factors to   drive the   senescence   of

FoxM1-deficient GC cells.

In vitro   experimental   and   animal   studies   suggest that FoxM1 is a novel anti-cancer target, and FoxM1- specific inhibitors have been developed for this pur- pose [40,41,54].   The   present   findings   indicate   that the same   strategy may be applied to gastric cancer. Importantly, the recognition of p27kip1 -dependent and p53/p16-pRb-independent   senescence   resulting   from FoxM1 depletion in GC cells may provide a rationale to manipulate cancer cell senescence with an ultimate therapeutic goal.

Acknowledgements

We thank Dr Juan SH (Taipei Medical University, Taiwan) for providing the p27 reporter plasmid. The study was supported by the Swedish Cancer Society, the Swedish Research Council, the Cancer   Society   in   Stockholm,   the   Swedish   Society   of Medicine, the   Karolinska Institutet Foundation, the National Natural Science   Foundation   of China   (Grant   Nos   30770118 and 30772545), the National Basic Research Program of China (Grant   No.   973   Program   2007CB512001)   and   the   Science Foundation of Shandong Province (Grant Nos 2005GG3202087 and Y2004C03).

Supporting information

Supporting information   may be found   in the   online version of this article.

References

1.   Vogiatzi P,   Vindigni C,   Roviello F,   Renieri A,   Giordano A. Deciphering the underlying genetic and epigenetic events leading to gastric carcinogenesis. J Cell Physiol 2007;211:287 – 295.

2. Di    Mario F,    Cavallaro LG,    Cavestro GM,    Franze A.    Are there   useful   biomarkers   for   gastric   cancer?   Dig   Liver Dis 2006;38:308 – 309.

3.   Zheng L, Wang L, Ajani J, Xie K. Molecular basis of gastric can- cer development and progression. Gastric Cancer 2004;7:61 – 77.

4.   Smith MG,   Hold GL,   Tahara E,   El-Omar EM.   Cellular   and molecular    aspects   of   gastric   cancer.   World   J   Gastroenterol 2006;12:2979 – 2990.

5.   Correa P,   Houghton J.   Carcinogenesis   of   Helicobacter pylori . Gastroenterology 2007;133:659 – 672.

6.   Knekt P,   Teppo L,   Aromaa A,   Rissanen H,   Kosunen TU.   Heli- cobacter pylori   IgA and IgG antibodies, serum pepsinogen I and the risk of gastric cancer: changes in the risk with extended follow- up period. Int J Cancer 2006;119:702 – 705.

7.   Peek RM   Jr,   Crabtree JE.   Helicobacter   infection   and gastric neoplasia. J Pathol 2006;208:233 – 248.

8.   Du Y,   Danjo K,   Robinson PA,   Crabtree JE.   In-cell   Western analysis    of   Helicobacter   pylori-induced   phosphorylation   of extracellular-signal related kinase via the transactivation of the epi- dermal growth factor receptor. Microbes Infect 2007;9:838 – 846.

9.   Buffart TE, Carvalho B, Hopmans E, Brehm V, Kranenbarg EK, Schaaij-Visser TB,   et al.   Gastric   cancers   in   young   and   elderly patients       show      different     genomic     profiles.     J     Pathol 2007;211:45 – 51.

10.   Tsukamoto Y,   Uchida T,   Karnan S,   Noguchi T,   Nguyen LT, Tanigawa M, et al. Genome-wide analysis of DNA copy number

J Zeng et al

alterations    and   gene   expression   in   gastric   cancer.   J   Pathol 2008;216:471 – 482.

11.   El-Omar EM. Role of host genes in sporadic gastric cancer. Best Pract Res Clin Gastroenterol 2006;20:675 – 686.

12.   Chochi K,   Ichikura T,   Kinoshita M,   Majima T,   Shinomiya N, Tsujimoto H, et al. Helicobacter pylori augments growth of gastric cancers   via   the   lipopolysaccharide-toll-like   receptor   4   pathway whereas its lipopolysaccharide   attenuates   antitumor   activities   of human mononuclear cells. Clin Cancer Res 2008;14:2909 – 2917.

13.   Hanahan D,   Weinberg RA.   The   hallmarks   of   cancer.   Cell 2000;100:57 – 70.

14.   Shay JW, Wright WE. Hallmarks of telomeres in ageing research. J Pathol 2007;211:114 – 123.

15.   Beausejour CM,   Campisi J.   Ageing:   balancing   regeneration   and cancer. Nature 2006;443:404 – 405.

16.   Braig M,    Lee S,    Loddenkemper C,    Rudolph C,    Peters AH, Schlegelberger B, et al. Oncogene-induced senescence as an initial barrier in lymphoma development. Nature 2005;436:660 – 665.

17.   Chen Z,   Trotman LC,   Shaffer D,   Lin HK,   Dotan ZA,   Niki M, et al. Crucial role of p53-dependent cellular senescence in suppres- sion of Pten-deficient tumorigenesis. Nature 2005;436:725 – 730.

18.   Schmitt CA,   Fridman JS,   Yang M,   Lee S,   Baranov E,   Hoff- man RM,   et al.   A   senescence   program   controlled   by   p53   and p16INK4a   contributes   to   the   outcome   of   cancer   therapy.   Cell 2002;109:335 – 346.

19.   Xue   W,   Zender   L,   Miething   C,   Dickins   RA,   Hernando   E, Krizhanovsky V,   et al.   Senescence   and   tumour   clearance   is triggered by p53 restoration in murine liver carcinomas. Nature 2007;445:656 – 660.

20.   Wu CH,    van    Riggelen J,    Yetil A,    Fan AC,    Bachireddy P, Felsher DW. Cellular   senescence is an important mechanism of tumor regression upon c-Myc inactivation. Proc Natl Acad Sci USA 2007;104:13028 – 13033.

21.   Cong YS,   Wright WE,   Shay JW.   Human   telomerase   and   its regulation. Microbiol Mol Biol Rev 2002;66:407 – 425.

22.   Liu L, Lai S, Andrews LG, Tollefsbol TO. Genetic and epigenetic modulation of telomerase   activity   in   development   and   disease. Gene 2004;340:1 – 10.

23.   Papazoglu C, Mills AA. p53: at the crossroad between cancer and ageing. J Pathol 2007;211:124 – 133.

24.   Kim WY, Sharpless NE. The regulation of INK4/ARF in cancer and aging. Cell 2006;127:265 – 275.

25.   Wierstra I,   Alves J.   FOXM1,   a   typical   proliferation-associated transcription factor. Biol Chem 2007;388:1257 – 1274.

26.   Laoukili J,   Stahl M, Medema RH. FoxM1:   at the   crossroads   of ageing and cancer. Biochim Biophys Acta 2007;1775:92 – 102.

27.   Wonsey DR,   Follettie MT.   Loss   of   the   forkhead transcription factor    FoxM1   causes   centrosome   amplification   and   mitotic catastrophe. Cancer Res 2005;65:5181 – 5189.

28.   Wang   IC,   Chen   YJ,   Hughes   DE,   Ackerson   T,   Major   ML, Kalinichenko VV, et al. FoxM1 regulates transcription of JNK1 to promote the G1/S transition and tumor cell invasiveness. J Biol Chem 2008;283:20770 – 20778.

29.   Wang IC, Meliton L, Tretiakova M, Costa RH, Kalinichenko VV, Kalin TV.   Transgenic   expression   of   the   forkhead   box   M1 transcription factor induces formation of lung tumors. Oncogene 2008;27:4137 – 4149.

30.   Teh MT, Wong ST, Neill GW, Ghali LR, Philpott MP, Quinn AG. FOXM1 is a downstream target of Gli1 in basal cell carcinomas. Cancer Res 2002;62:4773 – 4780.

31.   Pilarsky C,   Wenzig M,   Specht T,   Saeger HD,   Grutzmann R. Identification   and   validation   of commonly   overexpressed   genes in solid   tumors   by   comparison   of   microarray   data.   Neoplasia 2004;6:744 – 750.

32.   Madureira   PA,   Varshochi   R,   Constantinidou   D,   Francis   RE, Coombes RC,   Yao   KM,   et al.   The   Forkhead   box   M1   protein regulates the transcription of the estrogen receptor alpha in breast cancer cells. J Biol Chem 2006;281:25167 – 25176.

33.   Liu M,   Dai B,   Kang SH,   Ban K,   Huang FJ,   Lang FF,   et al. FoxM1B is overexpressed in human glioblastomas and critically regulates    the   tumorigenicity   of   glioma   cells.   Cancer   Res 2006;66:3593 – 3602.