BACKGROUND: Breast carcinomas represent a heterogeneous group of tumors diverse in behavior, outcome, and response to therapy. Identification of proteins resembling the tumor biology can improve the diagnosis, prediction, treatment selection, and targeting of therapy. Since the beginning of the post-genomic era, the focus of molecular biology gradually moved from genomes to proteins and proteomes and to their functionality. Proteomics can potentially capture dynamic changes in protein expression integrating both genetic and epigenetic influences. METHODS: We prepared primary cultures of epithelial cells from 23 breast cancer tissue samples and performed comparative proteomic analysis. Seven patients developed distant metastases within three-year follow-up. These samples were included into a metastase-positive group, the others formed a metastase-negative group. Two-dimensional electrophoretical (2-DE) gels in pH range 4-7 were prepared. Spot densities in 2-DE protein maps were subjected to statistical analyses (R/maanova package) and data-mining analysis (GUHA). For identification of proteins in selected spots, liquid chromatography-tandem mass spectrometry (LC-MS/MS) was employed. RESULTS: Three protein spots were significantly altered between the metastatic and non-metastatic groups. The correlations were proven at the 0.05 significance level. Nucleophosmin was increased in the group with metastases. The levels of 2,3-trans-enoyl-CoA isomerase and glutathione peroxidase 1 were decreased. CONCLUSION: We have performed an extensive proteomic study of mammary epithelial cells from breast cancer patients. We have found differentially expressed proteins between the samples from metastase-positive and metastase-negative patient groups.

Department of Oncology 1st Faculty of Medicine Charles University Prague Czech Republic

Zobrazit více v PubMed

Li Y, Welm B, Podsypanina K, Huang SX, Chamorro M, Zhang XM, Rowlands T, Egeblad M, Cowin P, Werb Z, Tan LK, Rosen JM, Varmus HE. Evidence that transgenes encoding components of the Wnt signaling pathway preferentially induce mammary cancers from progenitor cells. Proceedings of the National Academy of Sciences of the United States of America. 2003;100:15853–15858. doi: 10.1073/pnas.2136825100. PubMed DOI PMC

Charafe-Jauffret E, Ginestier C, Monville F, Fekairi S, Jacquemier J, Birnbaum D, Bertucci F. How to best classify breast cancer: Conventional and novel classifications (Review) International Journal of Oncology. 2005;27:1307–1313. PubMed

Sorlie T. Molecular portraits of breast cancer: tumour subtypes as distinct disease entities. European Journal of Cancer. 2004;40:2667–2675. doi: 10.1016/j.ejca.2004.08.021. PubMed DOI

Kapoor A, Vogel VG. Prognostic factors for breast cancer and their use in the clinical setting. Expert Review of Anticancer Therapy. 2005;5:269–281. doi: 10.1586/14737140.5.2.269. PubMed DOI

Hondermarck H, Vercoutter-Edouart AS, Revillion F, Lemoine J, El-Yazidi-Belkoura I, Nurcombe V, Peyrat JP. Proteomics of breast cancer for marker discovery and signal pathway profiling. Proteomics. 2001;1:1216–1232. doi: 10.1002/1615-9861(200110)1:10<1216::AID-PROT1216>3.0.CO;2-P. PubMed DOI

Bini L, Magi B, Marzocchi B, Arcuri F, Tripodi S, Cintorino M, Sanchez JC, Frutiger S, Hughes G, Pallini V, Hochstrasser DF, Tosi P. Protein expression profiles in human breast ductal carcinoma and histologically normal tissue. Electrophoresis. 1997;18:2832–2841. doi: 10.1002/elps.1150181519. PubMed DOI

Selicharova I, Sanda M, Mladkova J, Ohri SS, Vashishta A, Fusek M, Jiracek J, Vetvicka V. 2-DE Analysis of Breast Cancer Cell Lines 1833 and 4175 with Distinct Metastatic Organ-specific Potentials and Comparison with Parental Cell Line MDA-MB-231. Oncology Reports. 2008;19:1237–1244. PubMed

Mischak H, Apweiler R, Banks RE, Conaway M, Coon J, Dominiczak A, Ehrich JHH, Fliser D, Girolami M, Hermjakob H, Hochstrasser D, Jankowski J, Julian BA, Kolch W, Massy ZA, Neusuess C, Novak J, Peter K, Rossing K, Schanstra J, Semmes OJ, Theodorescu D, Thongboonkerd V, Weissinger EM, Van Eyk JE, Yamamoto T. Clinical proteomics: A need to define the field and to begin to set adequate standards. Proteomics Clinical Applications. 2007;1:148–156. doi: 10.1002/prca.200600771. PubMed DOI

Challapalli KK, Zabel C, Schuchhardt J, Kaindl AM, Klose J, Herzel H. High reproducibility of large-gel two-dimensional electrophoresis. Electrophoresis. 2004;25:3040–3047. doi: 10.1002/elps.200405979. PubMed DOI

Biron DG, Brun C, Lefevre T, Lebarbenchon C, Loxdale HD, Chevenet F, Brizard JP, Thomas F. The pitfalls of proteomics experiments without the correct use of bioinformatics tools. Proteomics. 2006;6:5577–5596. doi: 10.1002/pmic.200600223. PubMed DOI

Hondermarck H. Breast cancer: when proteomics challenges biological complexity. Molecular & Cellular Proteomics. 2003;2:281–291. PubMed

Matouskova E, Dudorkinova D, Krasna L, Vesely P. Temporal in vitro expansion of the luminal lineage of human mammary epithelial cells achieved with the 3T3 feeder layer technique. Breast Cancer Research and Treatment. 2000;60:241–249. doi: 10.1023/A:1006409605067. PubMed DOI

Villadsen R. In search of a stem cell hierarchy in the human breast and its relevance to breast cancer evolution. Apmis. 2005;113:903–921. doi: 10.1111/j.1600-0463.2005.apm_344.x. PubMed DOI

Polyak K. Breast cancer: origins and evolution. Journal of Clinical Investigation. 2007;117:3155–3163. doi: 10.1172/JCI33295. PubMed DOI PMC

Selicharova I, Smutna K, Sanda M, Ubik K, Matouskova E, Bursikova E, Brozova M, Vydra J, Jiracek J. 2-DE analysis of a new human cell line EM-G3 derived from breast cancer progenitor cells and comparison with normal mammary epithelial cells. Proteomics. 2007;7:1549–1559. doi: 10.1002/pmic.200600907. PubMed DOI

Brozova M, Kleibl Z, Netikova I, Sevcik J, Scholzova E, Brezinova J, Chaloupkova A, Vesely P, Dundr P, Zadinova M, Krasna L, Matouskova E. Establishment, growth and in vivo differentiation of a new clonal human cell line, EM-G3, derived from breast cancer progenitors. Breast Cancer Res Tr. 2006;103:247–257. doi: 10.1007/s10549-006-9358-x. PubMed DOI

Krasna L, Dudorkinova D, Vedralova J, Vesely P, Pokorna E, Kudlackova I, Chaloupkova A, Petruzelka L, Danes J, Matouskova E. Large expansion of morphologically heterogeneous mammary epithelial cells, including the luminal phenotype, from human breast tumours. Breast Cancer Research and Treatment. 2002;71:219–235. doi: 10.1023/A:1014457731494. PubMed DOI

The R Project for Statistical computing . 2007. PubMed

maanova . 2007.

Coufal D, Vydra J, Selicharova I. GUHA analysis of proteomic oncological data. Neural Network World. 2007;17:447–457.

Storey JD, Taylor JE, Siegmund D. Strong control, conservative point estimation and simultaneous conservative consistency of false discovery rates: a unified approach. Journal of the Royal Statistical Society Series B-Statistical Methodology. 2004;66:187–205. doi: 10.1111/j.1467-9868.2004.00439.x. DOI

Breiman L, Friedman J, Olshen R, Stone C. Classification and regression trees. Boca Raton, FL: Chapman & Hall/CRC; 1984.

The MathWorks . 2007.

Candiano G, Bruschi M, Musante L, Santucci L, Ghiggeri GM, Carnemolla B, Orecchia P, Zardi L, Righetti PG. Blue silver: A very sensitive colloidal Coomassie G-250 staining for proteome analysis. Electrophoresis. 2004;25:1327–1333. doi: 10.1002/elps.200305844. PubMed DOI

Garcia M, Platet N, Liaudet E, Laurent V, Derocq D, Brouillet JP, Rochefort H. Biological and clinical significance of cathepsin D in breast cancer metastasis. Stem Cells. 1996;14:642–650. PubMed

Mai J, Waisman DM, Sloane BF. Cell surface complex of cathepsin B/annexin II tetramer in malignant progression. Biochim Biophys Acta. 2000;1477:215–230. PubMed

Torre GC. SCC antigen in malignant and nonmalignant squamous lesions. Tumor Biology. 1998;19:517–526. doi: 10.1159/000030045. PubMed DOI

Guo J, Shou C, Meng L, Jiang B, Dong B, Yao L, Xie Y, Zhang J, Chen Y, Budman DR, Shi YE. Neuronal protein synuclein gamma predicts poor clinical outcome in breast cancer. International Journal of Cancer. 2007;121:1296–1305. doi: 10.1002/ijc.22763. PubMed DOI

Gusterson BA, Ross DT, Heath VJ, Stein T. Basal cytokeratins and their relationship to the cellular origin and functional classification of breast cancer. Breast Cancer Research. 2005;7:143–148. doi: 10.1186/bcr1041. PubMed DOI PMC

Marahatta SB, Punyarit P, Bhudisawasdi V, Paupairoj A, Wongkham S, Petmitr S. Polymorphism of glutathione S-transferase omega gene and risk of cancer. Cancer Letters. 2006;236:276–281. doi: 10.1016/j.canlet.2005.05.020. PubMed DOI

Thornalley PJ. Glyoxalase I - structure, function and a critical role in the enzymatic defence against glycation. Biochemical Society Transactions. 2003;31:1343–1348. PubMed

Sarto C, Binz PA, Mocarelli P. Heat shock proteins in human cancer. Electrophoresis. 2000;21:1218–1226. doi: 10.1002/(SICI)1522-2683(20000401)21:6<1218::AID-ELPS1218>3.0.CO;2-H. PubMed DOI

Janssen U, Fink T, Lichter P, Stoffel W. Human Mitochondrial 3,2-Trans-Enoyl-Coa Isomerase (Dci) - Gene Structure and Localization to Chromosome 16P13.3. Genomics. 1994;23:223–228. doi: 10.1006/geno.1994.1480. PubMed DOI

He Q, Shkarin P, Hooley RJ, Lannin DR, Weinreb JC, Bossuyt VI. In vivo MR spectroscopic imaging of polyunsaturated fatty acids (PUFA) in healthy and cancerous breast tissues by selective multiple-quantum coherence transfer (Sel-MQC): A preliminary study. Magn Reson Med. 2007;58:1079–1085. doi: 10.1002/mrm.21335. PubMed DOI

Lei XG, Cheng WH, McClung JP. Metabolic regulation and function of glutathione peroxidase-1. Annu Rev Nutr. 2007;27:41–61. doi: 10.1146/annurev.nutr.27.061406.093716. PubMed DOI

Lim MJ, Wang XW. Nucleophosmin and human cancer. Cancer Detection and Prevention. 2006;30:481–490. doi: 10.1016/j.cdp.2006.10.008. PubMed DOI PMC

Zhang Y. The ARF-B23 connection: implications for growth control and cancer treatment. Cell Cycle. 2004;3:259–262. PubMed

Phenotyping breast cancer cell lines EM-G3, HCC1937, MCF7 and MDA-MB-231 using 2-D electrophoresis and affinity chromatography for glutathione-binding proteins