- Original Article
- Open Access
Proteomic analysis of urinary fibrinogen degradation products in patients with urothelial carcinomas
Clinical Proteomics volume 2, pages 45–65 (2006)
Despite many years of research efforts and continued progress in the identification of urine markers for detection of bladder cancer, none of the markers described to date has been able to replace cystoscopy and urine cytology, the current gold standards for diagnosis. Here, we present a comprehensive gel-based proteomic study in which we have analyzed the presence and origin of fibrinogen (FG) and its degradation products (FDPs) in the urine of patients with and without urothelial carcinoma (UCs), with the aim of evaluating the potential of two-dimensional (2D) gel FDP profiling for detecting bladder cancer. A total of 151 urine samples collected from patients with Ucs of varying degrees of atypia and stages of invasion were compared with a control group consisting of 34 healthy volunteers with no clinical history of bladder cancer. The level and degree of degradation of FG in the urine were determined by 2D gel Western blotting in combination with enhanced chemilumenscence detection. Elevated levels of urine FG/FDPs were found in 99% of patients bearing superficial tumors, in 97% of the cases with early invasive disease, and in 96% of patients with highly invasive tumors. 2D gel profiling of urine FG/FDPs showed that the FG chains exhibited differential susceptibility to in situ proteolysis, with the α-chain being the most susceptible and the γ-chain the most resistant. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry identified peptide sequence regions in several of the most representative and common FDPs, which can be of value for producing novel specific antibodies to detect FG/FDPs in the urine. In addition, we present evidence indicating that FG is not produced by normal or malignant urothelium, although it is present both in the stroma of malignant tissue and tumor lesions. Taken together, the data indicate that increased levels of FG/FDPs amounts in the urine are a characteristic feature of bladder cancer, and emphasize the value of 2D gel profiling of urine FG/FDPs for detecting low-grade, noninvasive UCs.
Freidell, G. H., Nagy, G. K., and Cohen, S. M. (1983) Pathology of human bladder cancer and related lesions, in The Pathology of Bladder Cancer, vol. 1 (Bryan, G. T. and Cohen, S. M., eds.), CRC Press, Boca Raton, FL, pp. 11–42.
Pauli, B. U., Alroy, J., and Weinstein, R. S. (1983) The ultrastructure and pathobiology of urinary bladder cancer, in The Pathology of Bladder cancer, vol. 2 (Bryan, G. T. and Cohen, S. M., eds.), CRC Press, Boca Raton, FL, pp. 41–140.
Shokeir, A. A. (2004) Squamous cell carcinoma of the bladder: pathology, diagnosis and treatment. BJU Int. 93, 216–220.
Mostofi, F. K., Davis, C. J., Jr., and Sesterhenn, I. A. (1990) Current understanding of pathology of bladder cancer and attendant problems. J. Occup. Med. 32, 793–796.
Stein, J. P., Grossfeld, G. D., Ginsberg, D. A., et al. (1998) Prognostic markers in bladder cancer: a contemporary review of the literature. J. Urol. 160, 645–659.
Mostofi, F. K. (1973) Proceedings: testicular tumors. Epidemiologic, etiologic, and pathologic features. Cancer 32, 1186–1201.
Donat, S. M. (2003) Evaluation and follow-up strategies for superficial bladder cancer. Urol. Clin. North Am. 30, 765–776.
Messing, E. M., Young, T. B., Hunt, V. B., et al. (1995) Comparison of bladder cancer outcome in men undergoing hematuria home screening versus those with standard clinical presentations. Urology 45, 387–396.
Chopin, D. K. and Gattegno, B. (2002) Superficial bladder tumors. Eur. Urol. 42, 533–541.
Brown, F. M. (2000) Urine cytology. It is still the gold standard for screening?. Urol. Clin. North Am. 27, 25–37.
Rife, C. C., Farrow, G. M., and Utz, D. C. (1979) Urine cytology of transitional cell neoplasms. Urol. Clin. North Am. 6, 599–612
Cajulis, R. S., Haines, G. K., 3rd., Frias-Hidvegi, D., McVary, K., and Bacus, J. W. (1995) Cytology, flow cytometry, image analysis, and interphase cytogenetics by fluoresce in situ hybridization in the diagnosis of transitional cell carcinoma in bladder washes: a comparative study. Diagn. Cytopathol. 13, 214–223.
Pode, D., Golijanin, D., Sherman, Y., Lebensart, P., and Shapiro, A. (1998) Immunostaining of Lewis X in cells from voided urine, cytopathology and ultrasound for noninvasive detection of bladder tumors. J. Urol. 159, 389–392.
Ramakumar, S., Bhuiyan, J., Besse, J. A., et al. (1999) Comparison of screening methods in the detection of bladder cancer. J. Urol. 161, 388–394.
Konety, B. R. and Getzenberg, R. H. (2001) Urine based markers of urological malignancy. J. Urol. 165, 600–611.
Han, K. R., Pantuck, A. J., Belldegrun, A. S., and Rao, J. Y. (2002) Tumor markers for the early detection of bladder cancer. Front Biosci. 7, 19–26.
Eissa, S., Kassim, S., and El-Ahmady, O. (2003) Detection of bladder tumours: role of cytology, morphology-based assays, biochemical and molecular markers. Curr. Opin. Obstet. Gynecol. 15, 395–403.
Glas, A. S., Roos, D., Deutekom, M., Zwinderman, A. H., Bossuyt, P. M., and Kurth, K. H. (2003) Tumor markers in the diagnosis of primary bladder cancer. A systematic review. J. Urol. 169, 1975–1982.
Lokeshwar, V. B. and Soloway, M. S. (2002) Urine based markers of urological malignancy J. Urol. 167, 1406–1407.
Simon, M. A., Lokeshwar, V. B., and Soloway, M. S. (2003) Current bladder cancer tests: unnecessary or beneficial? Crit. Rev. Oncol. Hematol. 47, 91–107.
Ozen, H. and Hall, M. C. (2000) Bladder cancer. Curr. Opin. Oncol. 12, 255–259.
Dey, P. (2004) Urinary markers of bladder carcinoma. Clin. Chim. Acta. 340, 57–65.
Halachmi, S., Linn, J. F., Amiel, G. E., Moskovitz, B., and Nativ, O. (1998) Urine cytology, tumour markers and bladder cancer. Br. J. Urol. 82, 647–654.
Topsakal, M., Karadeniz, T., Anac, M., Donmezer, S., and Besisik, A. (2001) Assessment of fibrin-fibrinogen degradation products (Accu-Dx) test in bladder cancer patients. Eur. Urol. 39, 287–291.
Schmetter, B. S., Habicht, K. K., Lamm, D. L., et al. (1997) A multicenter trial evaluation of the fibrin/fibrinogen degradation products test for detection and monitoring of bladder cancer. J. Urol. 158, 801–805.
Wajsman, Z., Williams, P. D., Greco, J., and Murphy, G. P. (1978) Further study of fibrinogen degradation products in bladder cancer detection. Urology 12, 659–661.
McCabe, R. P., Lamm, D. L., Haspel, M. V., et al. (1984) A diagnostic-prognostic test for bladder cancer using a monoclonal antibody-based enzyme-linked immunoassay for detection of urinary fibrin(ogen) degradation products. Cancer Res. 44, 5886–5893.
Misra, K., Chowhan, J. S., Gupta, R. L., and Sagreiya, K. (1985) Diagnostic role of urine cytology and fibrinogen degradation products in carcinoma of bladder. Indian J. Cancer 22, 145–151.
Ewing, R., Tate, G. M., and Hetherington, J. W. (1987) Urinary fibrin/fibrinogen degradation products in transitional cell carcinoma of the bladder. Br. J. Urol. 59, 53–58.
Tsihlias, J. and Grossman, H. B. (2000) The utility of fibrin/fibrinogen degradation products in superficial bladder cancer. Urol. Clin. North Am. 27, 39–46.
Johnston, B., Morales, A., Emerson, L., and Lundie, M. (1997) Rapid detection of bladder cancer: a comparative study of point of care tests. J. Urol. 158, 2098–2101.
Siemens, D. R., Morales, A., Johnston, B., and Emerson, L. (2003) A comparative analysis of rapid urine tests for the diagnosis of upper urinary tract malignancy. Can. J. Urol. 10, 1754–1758.
Sobin, L. H. (1978) The WHO histological classification of urinary bladder tumours. Urol. Res. 6, 193–195.
Celis, J. E., Gromov, P., Cabezon, T., et al. (2004) Proteomic characterization of the interstitial fluid perfusing the breast tumor microenvironment: a novel resource for biomarked and therapeutic target discovery. Mol. Cell Proteomics 3, 327–344.
Celis, J. E., Trentemølle, S., and Gromov, P. (2006) Gel-based proteomics: high-resolution two-dimensional gel electrophoresis of proteins isoelectric focusing (IEF) and nonequilibrium pH gradient electrophoresis (NEPHGE), in Cell Biology. A Laboratory Handbook, vol. 4, (Celis, J. E., Carter, N., Hunter, T., Shotton, D., Simons, K., and Small, J. V., eds.), Academic Press, San Diego, CA, pp. 165–174.
Gromova, I. and Celis, J. E. (2006) Protein detection in gels by silver staining: a procedure compatible with mass-spectrometry, in Cell Biology. A Laboratory Handbook, vol. 4 (Celis, J. E., Carter, N., Hunter, T., Shotton, D., Simons, K., and Small, J. V., eds.), Academic Press, San Diego, CA.
Celis, J. E. and Gromov, P. (2000) High-resolution two-dimensional gel electrophoresis and protein identification using western blotting and ECL detection. EXS 88, 55–67.
Hermanson, G. T., Krishna Mallia A., and Smith, P. K. (1992) Immobilized affinity ligand techniques. Academic Press, San Diego, CA pp. 224–226.
Shevchenko, A., Wilm, M., Vorm, O., and Mann, M. (1996) Mass spectrometric sequencing of proteins silver-stained polyacrylamide gels. Anal. Chem. 68, 850–858.
Pieper, R., Gatlin, C. L., McGrath, A. M. et al. (2004) Characterization of the human urinary proteome: a method for high-resolution display of urinary proteins on two-dimensional electrophoresis gels with a yield of nearly 1400 distinct protein spots. Proteomics 4, 1159–1174.
Celis, J. E., Gromova, I., Moreira, J. M., Cabezon, T., and Gromov, P. (2004) Impact of proteomics on bladder cancer research. Pharmacogenomics 5, 381–394.
Marshall, T. and Williams, K. M. (1998) Clinical analysis of human urinary proteins using high resolution electrophoretic methods. Electrophoresis 19, 1752–1770.
Waller, K. V., Ward, K. M., Mahan, J. D., and Wismatt, D. K. (1989) Current concepts in proteinuria. Clin. Chem. 35, 755–765.
Brown, L. F., Dvorak, A. M., and Dvorak, H. F. (1989) Leaky vessels, fibrin deposition, and fibrosis: a sequence of events common to solid tumors and to many other types of disease. Am. Rev. Respir. Dis. 140, 1104–1107.
Simpson-Haidaris, P. J. and Rybarczyk, B. (2001) Tumors and fibrinogen. The role of fibrinogen as an extracellular matrix protein. Ann. NY Acad. Sci. 936, 406–425.
Wajsman, Z., Merrin, C. E., Chu, T. M., Moore, R. H., and Murphy, G. P. (1975) Evaluation of biological markers in bladder cancer. J. Urol. 114, 879–893.
Brown, L. F., Van de Water, L., Harvey, V. S., and Dvorak, H. F. (1988) Fibrinogen influx and accumulation of cross-linked fibrin in healing wounds and in tumor stroma. Am. J. Pathol. 130, 455–465.
Dvorak, H. F., Nagy, J. A., Berse, B., et al. (1992) Vascular permeability factor, fibrin, and the pathogenesis of tumor stroma formation. Ann. NY Acad. Sci. 667, 101–111.
O'Brien, T., Cranston, D., Fuggle, S., Bicknell, R., and Harris, A. L. (1995) Different angiogenic pathways characterize superficial and invasive bladder cancer. Cancer Res. 55, 510–513.
Crew, J. P., O'Brien, T., Bicknell, R., Fuggle, S., Cranston, D., and Harris, A. L. (1999) Urinary vascular endothelial growth factor and its correlation with bladder cancer recurrence rates. J. Urol. 161, 799–804.
Brown, L. F., Berse, B., Jackman, R. W., et al. (1993) Increased expression of vascular permeability factor (vascular endothelial growth factor) and its receptors in kidney and bladder carcinomas. Am. J. Pathol. 143, 1255–1262.
Stohrer, M., Boucher, Y., Stangassinger, M., and Jain, R. K. (2000) Oncotic pressure in solid tumors is elevated. Cancer Res. 60, 4251–4255.
Boucher, Y., Leunig, M., and Jain, R. K. (1996) Tumor angiogenesis and interstitial hypertension. Cancer Res. 56, 4264–4266.
Gaffney, P. J. and Dobos, P. (1971) A structural aspect of human fibrinogen suggested by its plasmin degradation. FEBS Lett. 15, 13–16.
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Gromov, P., Moreira, J.M.A., Gromova, I. et al. Proteomic analysis of urinary fibrinogen degradation products in patients with urothelial carcinomas. Clin Proteom 2, 45–65 (2006). https://doi.org/10.1385/CP:2:1:45
- Bladder Cancer
- Urothelial Carcinoma
- Urine Cytology
- Fibrinogen Degradation Product