鼻咽癌在西方相當罕見，然而在南中國卻是第三普遍的惡性腫瘤疾病，雖然已發現多種與鼻咽癌有密切關係的危險因子，如：遺傳、生活環境、EB病毒等，且隨著生活環境改善、治療方法不斷地進步，鼻咽癌患者的死亡率已逐漸降低。然而，直到目前，鼻咽癌形成的分子機制仍不是很清楚。
p57是調控細胞週期相關蛋白的Cip/Kip家族成員。p57基因在人類腫瘤中很少發生突變，顯示其功能的缺失主要起因於轉錄上或轉譯後表現受到了抑制。目前研究發現p57的功能缺失與多種發育缺陷和癌症的形成有關，如貝克維斯魏德曼症候群(BWS)、胃癌、膀胱癌等。由於訊息調控的研究大多集中在p21或是p27上，p57於生理上的表現和與細胞生長、基因的調控機制等的詳細機制都需要更進一步的特性分析研究。
科學家們對荷爾蒙與癌症間關聯性的研究已進行多年，由於鼻咽癌與荷爾蒙的相關研究非常的少，我們檢測數種荷爾蒙對鼻咽癌細胞的影響。結果發現其中progesterone與一MEK抑制劑-U0126有類似的作用機制，抑制了MAPK訊息傳遞分子的活化，且引發了p57的表現，並使得細胞生長受到抑制，細胞也停留在細胞週期的G1 phase；此外，在progesterone處理的同時加入其拮抗劑mifepristone (RU-486)後，抑制作用也受到了中和。因此，推論progesterone可能藉progesterone的receptor和MAPK訊息傳遞途徑調控p57的表現，來影響鼻咽癌細胞的細胞週期進行與生長。

Nasopharyngeal carcinoma (NPC) occurs occasionally in the west but is popular in south-eastern China and Hong Kong where it is the third most common form of malignancy among ethnic Chinese people. The possibly relevant factors associated with NPC have been found, for example, environment, genetics and EB virus, etc. Recently, as a result of improved environment of living and therapies, the death rate of NPC is decreasing year by year. However, the molecules mechanism underlying its tumorigenicity is still unclear.
The p57 protein is a maternally expressed, paternally imprinted cyclin-dependent kinases inhibitor (CDI). p57 mutations are rare in the human cancers, suggesting that other mechanisms of transcriptional or post translational silencing are involved in the loss of p57 function. Decreased expression of p57 has been found in Beckwith-Wiedemann syndrome (BWS), gastric cancer and bladder carcinoma. So far, many studies in signal transduction have been focused on p21 or p27. The relationship between p57 expression and signal transduction and cell proliferation under physiological circumstances requires further exploration.
Studies of the relationship between hormones and cancers have been proceeded for years, nevertheless, there is few about NPC. We examined several hormones’ effect on the NPC cell lines. We found that in response to progesterone treatment were the marked inhibition of pMAPK and up-regulation of p57, just like the influence of MEK inhibitor, U0126. Progesterone also induced growth inhibition and a slight accumulation of cells in the G1 phase of the cell cycle. On the other hand, the effects of progesterone were diminished in the presence of its antagonist mifepristone (RU-486). Taken together, these results suggest that progesterone treatment may induce the expression of the p57 on the mRNA and protein level by the progesterone receptor and that the MAPK signaling pathway may be involved in the progesterone-induced antiproliferative effect.

摘要-----------------------------------------------------------------------------1
Abstract--------------------------------------------------------------------------2
緒論-----------------------------------------------------------------------------4
一、前言-----------------------------------------------------------------4
二、細胞週期(Cell Cycle)--------------------------------------------5
三、細胞週期調節基因p57------------------------------------------6
四、MAPK訊息傳遞途徑--------------------------------------------8
五、Progesterone--------------------------------------------------------9
六、鼻咽癌(Nasopharyngeal carcinoma)---------------------------11
實驗材料與方法--------------------------------------------------------------14
結果與討論--------------------------------------------------------------------24
結論-----------------------------------------------------------------------------29
參考文獻-----------------------------------------------------------------------30
附圖-----------------------------------------------------------------------------42

Caspary T., Cleary M. A., Perlman E. J., Zhang P., Elledge S. J. and Tilghman S. M. (1999) Oppositely imprinted genes p57(Kip2) and igf2 interact in a mouse model for Beckwith-Wiedemann syndrome. Genes Dev. 13: 3115–3124.
Chan A. S., To K. F., Lo K. W., Mak K. F., Pak W., Chiu B., Tse G. M., Ding M., Li X., Lee J. C., Huang D. P. (2000) High frequency of chromosome 3p deletion in histologically normal nasopharyngeal epithelia from southern Chinese. Cancer Res. 60:5365-5370.
Chan A. T. C., Teo P. M. L. and Johnson P. J. (2002) Nasopharyngeal carcinoma. Ann. Oncol. 13:1007-1015.
Clarke C. L., Sutherland R. L. (1990) Progestin regulation of cellular proliferation. Endocr. Rev. 11:266-301.
Cobb M. H. (1999) MAP kinase pathways. Prog. Biophys. Mol. Biol. 71:479-500.
Delmas C., Manenti S., Boudjelal A., Peyssonnaux C., Eychenes A. and Darbon J. (2001) The p42/p44 mitogen-activated protein kinase activation triggers p27Kip1 degradation independently of CDK2/cyclin E in NIH 3T3 cells. J. Biol. Chem. 276:34958 -34965.
Gao J., Mazella J., Tang M., Tseng L. (2000) Ligand-activated progesterone receptor isoform hPR-A is a stronger transactivator than
hPR-B for the expression of IGFBP-1 (insulin-like growth factor binding protein-1) in human endometrial stroma cells. Mol. Endocrinol. 14:1954-1961.
Going J. J., Anderson T. J., Battersby S., Maclntyre C. C. (1988) Proliferative and secretory activity in human breast during natural and artificial menstrual cycles. Am. J. Pathol. 130:193-204.
Graham J. D., Clarke C. L. (1997). Physiological action of progesterone in target tissues. Endocr. Rev. 18:502-519.
Grandjean V., Smith J., Schofield P. N. and Ferguson-Smith A. C. (2000) Increased IGF-II protein affects p57kip2 expression in vivo and in vitro: implications for Beckwith-Wiedemann syndrome. Proc. Natl. Acad. Sci. USA 97:5279-5284.
Groshong S. D., Owen G. I., Grimison B., Schauer I. E., Todd M. C., Langan T. A., Sclafani R. A., Lange A., Horwitz K. B. (1997) Biphasic regulation of breast cancer cell growth by progesterone: role of the cyclin-dependent kinase inhibitors, p21 and p27Kip1 Mol. Endocrinol. 11:1593-1607.
Gulley M. L., Nicholls J. M., Schneider B. G., Amin M. B., Ro J. Y. and Geradts J. (1998) Nasopharyngeal carcinoma frequently lack the p16/ MTS1 tumor suppressor protein but consistently express the retinoblastoma gene product. Am. J. Pathol. 152:865-869.
Hatada I., Ohashi H., Fukushima Y., Kaneko Y., Inoue M., Komoto Y.,
Okada A., Ohishi S., Nabetani A., Morisaki H., Nakayama M., Niikawa N., Mukai T. (1996) An imprinted gene p57KIP2 is mutated in Beckwith-Wiedemann syndrome. Nat. Genet. 14:171-173.
Ho J. H. C. (1978) An epidemiologic and clinical study of nasopharyngeal carcinoma. Int. J. Radiat. Oncol. Biol. Phys. 4:183-205.
Huang D. P., Lo K. W., van Hasselt C. A., Woo J. K., Choi P. H., Leung S. F., Cheung S. T., Cairns P., Sidransky D., Lee J. C. (1994) A region of homozygous deletion on chromosome 9p21-22 in primary nasopharyngeal carcinoma. Cancer Res. 54:4003-4006.
Hui A. B., Lo K. W., Leung S. F., Teo P., Fung M. K., To K. F., Wong N., Choi P. H., Lee J. C., Huang D. P. (1999) Detection of recurrent chromosomal and losses in primary nasopharyngeal carcinoma by comparative genomic hybridization. Int. J. Cancer 82:498-503.
Jeng M. H., Parker C. J., Jordan V. C. (1992) Estrogenic potential of progestins in oral contraceptives to stimulate human breast cancer cell proliferation. Cancer Res. 52:6539-6546.
Kalkhoven E., Kwakkenbos-Isbrucker L., de Laat S. W., Van der Saag P. T., Van der Burg B. (1994) Synthetic progestins induce proliferation of breast tumor cell lines via progesterone or estrogen receptor. Mol. Cell Endocrinol. 102:45-52.
Katzenellenbogen B. S. (2000) Mechanisms of action and cross-talk between estrogen receptor and progesterone receptor pathways. J. Soc.
Gynecol. Invest. 7:S33-37.
Kortylewski M., Heinrich P. C., Kauffmann M. E., Bohm M., Mackiewicz A. and Behrmann I. (2001) Mitogen-activated protein kinase control p27/Kip1 expression and growth of human melanoma cells. J. Biochem. 357:297-303.
Kretzschmar M., Doody J., Timokhina I. and Massague J. (1999) A mechanism of repression of TGFβ/Smad signaling by oncogenic Ras. Genes Dev. 13:804-816.
Lee M. H., Reynisdottir I. and Massague J. (1995) Cloning of p57KIP2, a cyclin-dependent kinase inhibitor with unique domain structure and tissue distribution. Genes Dev. 9:639 -649.
Lee M. H. and Yang H. Y. (2001) Negative regulators of cyclin-dependent kinases and their roles in cancers. Cell. Mol. Life Sci. 58:1907-1922.
Lin C. T., Chan W. Y., Chen W., Huang H. M., Wu H. C., Hsu M. M., Chuang S. M. and Wang C. C. (1993) Characterization of seven newly established nasopharyngeal carcinoma cell lines. Lab. Invest. 68:716-727.
Lin V. C., Ng E. H., Aw S. E., Tan M. G., Ng E. H., Chan V. S., Ho G. H. (1999) progestins inhibit the growth of MDA-MB-231 cells transfected with progesterone receptor cDNA. Clin. Cancer Res. 5:395-404.
Lin V. C., Ng E. H., Aw S. E., Tan M. G., Ng E. H., Bay B. H. (2000)
Progesterone induces cell spreading and focal adhesion in breast cancer cells MDA-MB-231 transfected with progesterone receptor cDNA. Mol. Endocrinol. 14:348-358.
Lin V. C., Aw S. E., Ng E. H., Ng E. H., Chowdhury S. H. (2001) Effect of progesterone on the invasive properties and tumor growth of progesterone receptor (PR)-transfected breast cancer cells MDA-MB-231. Clin. Cancer Res. 7:2880-2886.
Lin V. C., Woon C. T., Aw S. E., Guo C. H. (2003) Distinct molecular pathways mediate progesterone-induced growth inhibition and focal adhesion. Endocrinology 144 (12):5650 -5657.
Lo K. W., Cheung S. T., Leung S. F., van Hasselt A., Tsang Y. S., Mak K. F., Chung Y. F., Woo J. K., Lee J. C., Huang D. P. (1996) Hypermethylation of the p16 gene in nasopharyngeal carcinoma. Cancer Res. 56:2721-2725.
Lo K. W., Teo P. M., Hui A. B. To K. F., Tsang Y. S., Chan S. Y., Mak K. F., Lee J. C., Huang D. P. (2000) High resolution allelotype of microdissected primary nasopharyngeal carcinoma. Cancer Res. 60:3348-3353.
Lo K. W., Kwong J., Hui A. B., Chan S. Y., To K. F., Chan A. S., Chow L. S., Teo P. M., Johnson P. J., Huang D. P. (2001) High frequency of promoter hypermethylation of RASSF1A in nasopharyngeal carcinoma. Cancer Res. 61:3877-3881.
Luo J., Xiao J., Tao Z. and Li X. (1997) Detection of c-Myc gene expression in nasopharyngeal carcinoma by nonradioactive in situ hybirdization and immunohistochemistry. Chin. Med. J. (England) 110:229-232.
Lydon J. P., Demayo F. J., Funk C. R., Mani S. K., Hughes A. R., Montgomery Jr C. A., Shyamala G., Conneely O. M., O’Malley B. W. (1995) Mice lacking progesterone receptor exhibit pleiotropic reproductive abnormalities. Genes Dev. 9:2266-2278.
Marks J. E., Phillips J. L., Menck H. R. (1998) The National Cancer Data Base report on the relationship of race and national origin to the histology of nasopharyngeal carcinoma. Cancer 83:582-588.
Matsuoka S., Edwards M. C., Bai C., Parker S., Zhang P., Baldini A., Harper J. W., Elledge S. J. (1995) p57KIP2, a structurally distinct member of the p21CIP1 Cdk inhibitor family, is a candidate tumor suppressor gene. Genes Dev. 9:650-662.
Migliaccio A., Piccolo D., Castoria G., Di Domenico M., Bilancio A., Lombardi M., Gong W., Beata M., Auricchio F. (1998) Activation of the Src/p21ras/Erk pathway by progesterone receptor via cross-talk with estrogen receptor. EMBO J. 17:2008-2018.
Mulac-Jericevic B., Mullinax R. A., DeMayo F. J., Lydon J. P., Conneely O. M. (2001) Subgroup of reproductive functions of progesterone mediated by progesterone receptor B isoform. Science 289:1751-1754.
Murakami Y. and Sekiya T. (1998) Accumulation of genetic alterations and their significance in each primary human cancer and cell lines. Mut. Res. 400:421-437.
Musgrove E. A., Lee C. S., Sutherland R. L. (1991) Progestins both stimulate and inhibit breast cancer cell cycle progression while increasing expression of transforming growth factor α, epidermal growth factor receptor, c-fos, and c-myc genes. Mol. Cell Biol. 11:5032-5043.
Musgrove E. A., Swarbrick A., Lee C. S., Cornish A. L., Sutherland R. L. (1998) Mechanisms of cyclin-dependent kinase inactivation by progestins. Mol. Cell Bio. 18:1812-1825.
Nijjar T., Wigington D., Garbe J. C., Waha A., Stampfer M. R. and Yaswen P. (1999) p57KIP2 expression and loss of heterozygosity during immortal conversion of cultured human mammary epithelial cells. Cancer Res. 59:5112-5118.
Nishimori S., Tanaka Y., Chiba T., Fujii M., Imamura T., Miyazono K. Ogasawara T, Kawaguchi H, Igarashi T, Fujita T, Tanaka K, Toyoshima H. (2001) Smad-mediated transcription is required for transforming growth factor-beta 1-induced p57(Kip2) proteolysis in osteoblastic cells. J. Biol. Chem. 276:10700-10705.
Nojima H. (2004) G1 and S-phase checkpoints, chromosome instability, and cancer. Methods Mol. Biol. 280:3-50.
Orlow I., Iavarone A., Crider-Miller S. J., Bonilla F., Latres E., Lee M. H.,
Gerald W. L., Massague J., Weissman B. E., Cordon-Cardo C. (1996) Cyclin-dependent kinase inhibitor p57KIP2 in soft tissue sarcomas and Wilms tumors. Cancer Res. 56:1219-1221.
O’Shea J. J., Gadina M., Schreiber R. D. (2002) Cytokine signaling in 2002: new surprises in the Jak/Stat pathway. Cell 109 Suppl:S121-131.
Oya M. and Schulz W. A. (2000) Decreased expression of p57(KIP2) mRNA in human bladder cancer. Br. J. Cancer 83:626-631.
Porter A. C. and Vaillancourt R. R. (1999) Tyrosine kinase receptor-activated signal transduction pathways which lead to oncogenesis. Oncogene 17:1343-1352.
Rane S. G. (1999) Ion channels as physiological effectors for growth factor receptor and Ras/ERK signaling pathways. Adv. Second Messenger Phosphoprotein Res. 33:107-127.
Reddy S. P., Raslan W.F., Gooneratne S., Kathuria S., Marks J. E. (1995) Prognostic significance of keratinization in nasopharyngeal carcinoma. Am. J. Otolaryngol. 16:103-108.
Reid L. H., Crider-Miller S. J., West A., Lee M. H., MassagueJ. and Weissman B. E. (1996) Genomic organization of the human p57KIP2 gene and its analysis in the G401 Wilms’ tumor assay. Cancer Res. 56:1214-1218.
Samuelsson M. K., Pazirandeh A., Davani B. and Okret S. (1999)
p57Kip2, a glucocorticoid-induced inhibitor of cell cycle progression in HeLa cells. Mol. Endocrinol. 13:1811-1822.
Schoonen W. G., Joosten J. W., Kloosterboer H. J. (1995) Effects of two classes of progestagens, pregnane and 19-nortestosterone derivatives, on cell growth of human breast tumor cells: I. MCF-7 cell lines. J. Steroid Biochem. Mol. Biol. 55:423-437.
Shanmugaratnam K., Sobin L. H. (1993) The World Health Organization histological classification of tumours of the upper respiratory tract and ear. A commentary on the second edition. Cancer 71:2689-2697.
Shin J. Y., Kim H. S., Park J., Park J. B. and Lee J. Y. (2000) Mechanism for inactivation of the KIP family cyclin-dependent kinase inhibitor genes in gastric cancer cells. Cancer Res. 60:262-265.
Shin J. Y., Kim H. S., Lee K. S., Kim J., Park J. B., Won M. H., Chae S. W., Choi Y. H., Choi K. C., Park Y. E., Lee J. Y. (2000) Mutation and expression of the p27KIP1 and p57KIP2 genes in human gastric cancer. Exp. Mol. Med. 32:79-83.
Spitz I. M. (2003) Progesterone antagonists and progesterone receptor modulators. Expert Opin. Investig. Drugs 12(10):1693-707.
Sutherland R. L., Hall R. E., Pang G. Y. N., Musgrove E. A., Clarke C. L. (1988) Effect of medroxyprogesterone acetate on proliferation and cell cycle kinetics in human mammary carcinoma cells. Cancer Res. 48:5084-5091.
Sutherland R. L., Prall O. W. J., Watts C. K. W., Musgrove E. A. (1998) Estrogen and progestin regulation of cell cycle progression. J. Mammary Gland Biol. Neoplasia 3:63-72.
Tsugu A., Sakai K., Dirks P. B., Jung S., Weksberg R., Fei Y. L., Mondal S., Ivanchuk S., Ackerley C., Hamel P, A., Rutka J, T. (2000) Expression of p57(KIP2) potently blocks the growth of human astrocytomas and induces cell senescence. Am. J. Pathol. 157:919-932.
Tung L., Mohamed K. M., Hoeffler J. P., Takimoto G. S., Horwitz K. B. (1993) Antagonist occupied human progesterone B- receptors activate transcription without binding to progesterone response elements and are dominantly inhibited by A-receptors. Mol. Endocrinol. 7:1256-1265.
Urano T., Yashiroda H., Muraoka M., Tanaka K., Hosoi T., Inoue S., Ouchi Y., Toyoshima H. (1999) p57(Kip2) is degraded through the proteasome in osteoblasts stimulated to proliferation by transforming growth factor beta1. J Biol. Chem. 274(18):12197-12200.
US Mortality Pubilc Use Data Tape (2001)
Van der Burg B., Kalhoven E., Isbrucker L., de Laat S. W. (1992) Effects of progestins on the proliferation of estrogen- dependent human breast cancer cells under growth factor-defined conditions. J. Steroid Biochem. Mol. Biol. 42:457-465.
Veronesi U., Luini A., Mariani L., Del Vecchio M., Alvez D., Andreoli C., Giacobone A., Merson M., Pacetti G., Raselli R., Saccozzi R. (1994)
Effect of menstrual phase on surgical treatment of breast cancer. Lancet 343:1545-1547.
Wang G. L., Lo K. W., Tsang K. S., Chung N. Y., Tsang Y. S., Cheung S. T., Lee J. C. and Huang D. P. (1999) Inhibiting tumorigenic potential by restoration of p16 in nasopharyngeal carcinoma. Br. J. Cancer 81:1122-1126.
Weinberg R. A., How cancer arises? (1996) Sci. Am. 275:62-67.
Wen D. X., Xu Y. F., Mais D. E., Goldman M. E., McDonnelle D. P. (1994) The A and B isoforms of human progesterone receptor operate through distinct signaling pathways within target cells. Mol. Cell Biol. 14:8356-8364.
Xu B., Hu P., Wu Q., Hou J., Zhang B., Chen X., Huang G. (1999) Relationship between expression of estrogen receptor progestrone receptor and the biological characteristics of nasopharyngeal carcinoma. Lin Chuang Er Bi Yan Hou Ke Za Zhi 13(8):347-349.
Yan Y., Frisen J., Lee M. H., Massague J. and Barbacid M. (1997) Ablation of the CDK inhibitor p57Kip2 results in increased apoptosis and delayed differentiation during mouse development. Genes Dev. 11:973-983.
Zhang P., Liegeois N. J., Wong C., Finegold M., Hou H.,Thompson J. C. Silverman A, Harper J. W., DePinho R. A., Elledge S. J. (1997) Altered cell differentiation and proliferation in mice lacking p57KIP2 indicates a
role in Beckwith-Wiedemann syndrome. Nature 387:151-158.
行政院衛生署衛生統計資訊網台灣地區主要死因趨勢圖(2004)。