Skip to main content

Advertisement

SpringerLink
Log in

Urinary oncofetal ED-A fibronectin correlates with poor prognosis in patients with bladder cancer

  • Research Paper
  • Published:
Clinical & Experimental Metastasis Aims and scope Submit manuscript

Abstract

The extracellular matrix protein fibronectin (FN) contributes to the structural integrity of tissues as well as the adhesive and migratory functions of cells. While FN is abundantly expressed in adult tissues, the expression of several alternatively spliced FN isoforms is restricted to embryonic development, tissue remodeling and cancer. These FN isoforms, designated ED-A and ED-B, are frequently expressed by cancer cells, tumor-associated fibroblasts and newly forming blood vessels. Using a highly sensitive collagen-based indirect ELISA, we evaluated the correlation of urinary ED-A and ED-B at time of cystectomy with overall survival in patients with high-grade bladder cancer (BCa). Detectable levels of total FN as well as ED-A and ED-B were found in urine from 85, 73 and 51 % of BCa patients, respectively. The presence of urinary ED-A was a significant independent predictor of 2-year overall survival (OS) after adjusting for age, tumor stage, lymph node stage, and urinary creatinine by multivariable Logistic Regression (p = 0.029, OR = 4.26, 95 % CI 1.16–15.71) and improved accuracy by 3.6 %. Furthermore, detection of ED-A in the urine was a significant discriminator of survival specifically in BCa patients with negative lymph node status (Log-Rank, p = 0.006; HR = 5.78, 95 % CI 1.39–24.13). Lastly, multivariable Cox proportional hazards analysis revealed that urinary ED-A was an independent prognostic indicator of 5-year OS rate for patients with BCa (p = 0.04, HR = 2.20, 95 % CI 1.04–4.69). Together, these data suggest that cancer-derived, alternatively spliced FN isoforms can act as prognostic indicators and that additional studies are warranted to assess the clinical utility of ED-A in BCa.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

Similar content being viewed by others

References

  1. Potts JR, Campbell ID (1996) Structure and function of fibronectin modules. Matrix Biol 15:313–320

    Article  CAS  PubMed  Google Scholar 

  2. To WS, Midwood KS (2011) Plasma and cellular fibronectin: distinct and independent functions during tissue repair. Fibrogenesis Tissue Repair 4:21

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  3. Tamkun JW, Hynes RO (1983) Plasma fibronectin is synthesized and secreted by hepatocytes. J Biol Chem 258(7):4641–4647

    CAS  PubMed  Google Scholar 

  4. Mosher DF (2006) Plasma fibronectin concentration: a risk factor for arterial thrombosis? Arterioscler Thromb Vasc Biol 26(6):1193–1195

    Article  CAS  PubMed  Google Scholar 

  5. Ting KM et al (2000) Overexpression of the oncofetal Fn variant containing the EDA splice-in segment in the dermal-epidermal junction of psoriatic uninvolved skin. J Investig Dermatol 114:706–711

    Article  CAS  PubMed  Google Scholar 

  6. Matsuura H, Hakomori S (1985) The oncofetal domain of fibronectin defined by monoclonal antibody FDC-6: its presence in fibronectins from fetal and tumor tissues and its absence in those from normal adult tissues and plasma. PNAS 82(19):6517–6521

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  7. Borsi L et al (1987) Monoclonal antibodies in the analysis of fibronectin isoforms generated by alternative splicing of mRNA precursors in normal and transformed human cells. J Cell Biol 104(3):595–600

    Article  CAS  PubMed  Google Scholar 

  8. Carnemolla B et al (1989) A tumor-associated fibronectin isoform generated by alternative splicing of messenger RNA precursors. J Cell Biol 108(3):1139–1148

    Article  CAS  PubMed  Google Scholar 

  9. Glukhova MA et al (1990) Expression of fibronectin variants in vascular and visceral smooth muscle cells in development. Dev Biol 141(1):193–202

    Article  CAS  PubMed  Google Scholar 

  10. Castellani P et al (1986) Transformed human cells release different fibronectin variants than do normal cells. J Cell Biol 103(5):1671–1677

    Article  CAS  PubMed  Google Scholar 

  11. Zardi L et al (1987) Transformed human cells produce a new fibronectin isoform by preferential alternative splicing of a previously unobserved exon. EMBO J 6(8):2337–2342

    PubMed Central  CAS  PubMed  Google Scholar 

  12. D’Ovidio MC et al (1998) Intratumoral microvessel density and expression of ED-A/ED-B sequences of fibronectin in breast carcinoma. Eur J Cancer 34(7):1081–1085

    Article  PubMed  Google Scholar 

  13. Rybak J-N et al (2007) The extra-domain a of fibronectin is a vascular marker of solid tumors and metastases. Cancer Res 67(22):10948–10957

    Article  CAS  PubMed  Google Scholar 

  14. Manabe R-I et al (1997) Modulation of cell-adhesive activity of fibronectin by the alternatively spliced EDA segment. J Cell Biol 139(1):295–307

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  15. Nicolo G et al (1990) PII: 0922–3371(90), 90056–3. Cell Differ Dev 32:401–408

    Article  CAS  PubMed  Google Scholar 

  16. Pujuguet P et al (1996) Expression of fibronectin ED-A+ and ED-B+ isoforms by human and experimental colorectal cancer contribution of cancer cells and tumor-associated myofibroblasts. Am J Pathol 148(2):579–592

    PubMed Central  CAS  PubMed  Google Scholar 

  17. Kohan M et al (2010) EDA-containing cellular fibronectin induces fibroblast differentiation through binding to α4β7 integrin receptor and MAPK/Erk 1/2-dependent signaling. FASEB J 24(11):4503–4512

    Article  CAS  PubMed  Google Scholar 

  18. Manabe R, Oh-e N, Sekiguchi K (1999) Alternatively spliced EDA segment regulates fibronectin-dependent cell cycle progression and mitogenic signal transduction. J Biol Chem 274(9):5919–5924

    Article  CAS  PubMed  Google Scholar 

  19. Booth AJ et al (2012) Recipient-derived EDA fibronectin promotes cardiac allograft fibrosis. J Pathol 226(4):609–618

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  20. Khan MM et al (2012) Alternatively-spliced extra domain A of fibronectin promotes acute inflammation and brain injury after cerebral ischemia in mice. Stroke 43(5):1376–1382

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  21. McFadden JP et al (2010) Psoriasis and extra domain A fibronectin loops. Br J Dermatol 163(1):5–11

    CAS  PubMed  Google Scholar 

  22. Castellani P et al (1994) The fibronectin isoform containing the ED-B oncofetal domain: a marker of angiogenesis. Int J Cancer J Int Du Cancer 59(5):612–618

    Article  CAS  Google Scholar 

  23. Khan ZA et al (2005) EDB fibronectin and angiogenesis—a novel mechanistic pathway. Angiogenesis 8(3):183–196

    Article  CAS  PubMed  Google Scholar 

  24. Astrof S et al (2004) Direct test of potential roles of EIIIA and EIIIB alternatively spliced segments of fibronectin in physiological and tumor angiogenesis. Mol Cell Biol 24(19):8662–8670

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  25. Chen W, Culp LA (1996) Adhesion mediated by fibronectin’s alternatively spliced EDb (EIIIB) and its neighboring type III repeats. Exp Cell Res 223(1):9–19

    Article  CAS  PubMed  Google Scholar 

  26. Carnemolla B et al (1992) The inclusion of the type III repeat ED-B in the fibronectin molecule generates conformational modifications that unmask a cryptic sequence. J Biol Chem 267(34):24689–24692

    CAS  PubMed  Google Scholar 

  27. Schiefner A, Gebauer M, Skerra A (2012) Extra-domain B in oncofetal fibronectin structurally promotes fibrillar head-to-tail dimerization of extracellular matrix protein. J Biol Chem 287(21):17578–17588

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  28. Ioachim E et al (2005) A clinicopathological study of the expression of extracellular matrix components in urothelial carcinoma. BJU Int 95(4):655–659

    Article  PubMed  Google Scholar 

  29. Yang X et al (2013) Diagnostic value of bladder tumor fibronectin in patients with bladder tumor: a systematic review with meta-analysis. Clin Biochem 46(15):1377–1382

    Article  CAS  PubMed  Google Scholar 

  30. Fernandez-Garcia B et al (2014) Expression and prognostic significance of fibronectin and matrix metalloproteases in breast cancer metastasis. Histopathology 64(4):512–522

    Article  PubMed  Google Scholar 

  31. Ma LJ et al (2014) Fibronectin overexpression is associated with latent membrane protein 1 expression and has independent prognostic value for nasopharyngeal carcinoma. Tumour Biol 35(2):1703–1712

    Article  CAS  PubMed  Google Scholar 

  32. Inufusa H et al (1995) Localization of oncofetal and normal fibronectin in colorectal cancer correlation with histologic grade, liver metastasis, and prognosis. Cancer 75(12):2802–2808

    Article  CAS  PubMed  Google Scholar 

  33. Ylätupa S et al (1995) An improved method for quantification of extra domain A-containing cellular fibronectin (EDAcFN) in different body fluids. Clin Chim Acta 234:79–90

    Article  PubMed  Google Scholar 

  34. Hegele A et al (2003) Cellular fibronectin in patients with transitional cell carcinoma of the bladder. Urol Res 30(6):363–366

    CAS  PubMed  Google Scholar 

  35. Richter P et al (2008) IIICS de novo glycosylated fibronectin as a marker for invasiveness in urothelial carcinoma of the urinary bladder (UBC). J Cancer Res Clin Oncol 134(10):1059–1065

    Article  CAS  PubMed  Google Scholar 

  36. Blick CGT et al (2011) Evaluation of diagnostic strategies for bladder cancer using computed tomography (CT) urography, flexible cystoscopy and voided urine cytology: results for 778 patients from a hospital haematuria clinic. BJU Int 110(1):84–94

    Article  PubMed  Google Scholar 

  37. Planz B et al (2005) The role of urinary cytology for detection of bladder cancer. Eur J Surg Oncol 31(3):304–308

    Article  CAS  PubMed  Google Scholar 

  38. Alias-Melgar A et al (2013) Association of urine oncofetal fibronectin levels with urology's most common disorders. Ann Clin Lab Sci 43(4):420–423

    CAS  PubMed  Google Scholar 

  39. Eissa SS et al (2002) Comparative evaluation of the nuclear matrix protein, fibronectin, urinary bladder cancer antigen and voided urine cytology in the detection of bladder tumors. J Urol 168(2):465–469

    Article  CAS  PubMed  Google Scholar 

  40. Katayama M et al (1991) Urinary fibronectin fragments (a potential tumor marker) measured by immunoenzymometric assay with domain-specific monoclonal antibodies. Clin Chem 37(3):466–471

    CAS  PubMed  Google Scholar 

  41. Malmström PU et al (1993) Increasing survival of patients with urinary bladder cancer. A nationwide study in Sweden 1960–1986. Eur J Cancer 29A(13):1868–1872

    Article  PubMed  Google Scholar 

  42. Saito M et al (2005) Proteome analysis of gelatin-bound urinary proteins from patients with bladder cancers. Eur Urol 48(5):865–871

    Article  CAS  PubMed  Google Scholar 

  43. Eissa SS et al (2011) The clinical relevance of urine-based markers for diagnosis of bladder cancer. Med Oncol 28(2):513–518

    Article  CAS  PubMed  Google Scholar 

  44. Shiozawa K, Hino K, Shiozawa S (2001) Alternatively spliced EDA-containing fibronectin in synovial fluid as a predictor of rheumatoid joint destruction. Rheumatology 40(7):739–742

    Article  CAS  PubMed  Google Scholar 

  45. Engvall E, Ruoslahti E (1977) Binding of soluble form of fibroblsat surface protein, fibronectin, to collagen. Int J Cancer 20:1–5

    Article  CAS  PubMed  Google Scholar 

  46. Engvall E, Ruoslahti E, Miller EJ (1978) Affinity of fibronectin to collagens of different genetic types and to fibrinogen. J Exp Med 8:1–12

    Google Scholar 

  47. Menzin AW et al (1998) Identification of oncofetal fibronectin in patients with advanced epithelial ovarian cancer: detection in ascitic fluid and localization to primary sites and metastatic implants. Cancer 82:152–158

    Article  CAS  PubMed  Google Scholar 

  48. Menéndez V et al (2005) Diagnosis of bladder cancer by analysis of urinary fibronectin. Urology 65(2):284–289

    Article  PubMed  Google Scholar 

  49. Witjes JA, Moonen PMJ, van der Heijden AG (2005) Comparison of hexaminolevulinate based flexible and rigid fluorescence cystoscopy with rigid white light cystoscopy in bladder cancer: results of a prospective phase II study. Eur Urol 47(3):319–322

    Article  PubMed  Google Scholar 

  50. Otto W et al (2013) Introduction and first clinical application of a simplified immunohistochemical validation system confirms prognostic impact of Ki-67 and CK20 for stage T1 urothelial bladder carcinoma: single-center analysis of eight biomarkers in a series of three hundred six patients. Clin Genitourin Cancer 11(4):537–544

    Article  PubMed  Google Scholar 

  51. Riester M et al (2012) Combination of a novel gene expression signature with a clinical nomogram improves the prediction of survival in high-risk bladder cancer. Clin Cancer Res 18(5):1323–1333

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  52. Tarin TV et al (2012) Lymph node-positive bladder cancer treated with radical cystectomy and lymphadenectomy: effect of the level of node positivity. Eur Urol 61(5):1025–1030

    Article  PubMed Central  PubMed  Google Scholar 

  53. Eissa S et al (2010) Diagnostic value of fibronectin and mutant p53 in the urine of patients with bladder cancer: impact on clinicopathological features and disease recurrence. Med Oncol 27(4):1286–1294

    Article  CAS  PubMed  Google Scholar 

  54. Bellmunt J et al (2006) Gene expression of ERCC1 as a novel prognostic marker in advanced bladder cancer patients receiving cisplatin-based chemotherapy. Ann Oncol 18(3):522–528

    Article  Google Scholar 

  55. Borsi L, Balza E, Allemanni G, Zardi L (1992) Differential expression of the fibronectin isoform containing the ED-B oncofetal domain in normal human fibroblast cell lines originating from different tissues. Exp Cell Res 199:98–105

    Article  CAS  PubMed  Google Scholar 

  56. Barone MV, Henchcliffe C, Baralle FE, Paolella G (1989) Cell type specific trans-acting factors are involved in alternative splicing of human fibronectin pre-mRNA. EMBO J Eur Mol Biol Organ 8:1079–1085

    CAS  Google Scholar 

  57. Zijlstra A, McCabe NR, Schelling ME (1999) Expression and assembly of the angiogenic marker B-fibronectin by endothelial cells in vitro: regulation by confluency. Angiogenesis 3:77–87

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

We gratefully acknowledge the laboratory of Bob Matusik including the technician responsible for long-term sample collection, Manik Paul. Many thanks to Paul Knoll and Chaochen You for development and maintenance of the urology database. We would also like to thank Alison Woodworth and Sherrie Stafford in the Vanderbilt Clinical Chemistry laboratory for collection of normal controls.

Author contributions

SA, HL, CA and AZ were responsible for the study concept and design as well as manuscript preparation. SA performed specimen processing and analysis, database construction and maintenance, and all data/statistical analyses. HL performed all ELISA assays and contributed to the data analysis. PC contributed to the VUMC cohort selection and collection, as well as manuscript editing. TK optimized and performed IF.

Funding

This work was supported by the National Institutes of Health for SA (VCORCDP K12-CA9060625 and CTSA UL1-TR000445), HL (T32-CA009593), CA (DK094900 and DK091491), and AZ (R01-CA143081 and P01-CA040035). SA was also supported by the Department of Veterans Affairs (IK2BX002498).

Author information

Author notes

    Authors and Affiliations

    1. Department of Veterans Affairs, Nashville, TN, 37212, USA

      Shanna A. Arnold

    2. Department of Pathology, Microbiology, and Immunology, Vanderbilt University, Nashville, TN, 37232, USA

      Shanna A. Arnold, Tatiana Ketova & Andries Zijlstra

    3. Department of Cancer Biology, Vanderbilt University, Nashville, TN, 37232, USA

      Holli A. Loomans, Claudia D. Andl & Andries Zijlstra

    4. Department of Surgery, Vanderbilt University, Nashville, TN, 37232, USA

      Claudia D. Andl & Peter E. Clark

    5. Vanderbilt-Ingram Cancer Center, Vanderbilt University Medical Center, 1161 21st Ave S., C-2102B Medical Center North, Nashville, TN, 37232, USA

      Claudia D. Andl, Peter E. Clark & Andries Zijlstra

    6. Epithelial Biology Center, Vanderbilt University, Nashville, TN, 37232, USA

      Claudia D. Andl

    Authors
    1. Shanna A. Arnold
      View author publications

      You can also search for this author in PubMed Google Scholar

    2. Holli A. Loomans
      View author publications

      You can also search for this author in PubMed Google Scholar

    3. Tatiana Ketova
      View author publications

      You can also search for this author in PubMed Google Scholar

    4. Claudia D. Andl
      View author publications

      You can also search for this author in PubMed Google Scholar

    5. Peter E. Clark
      View author publications

      You can also search for this author in PubMed Google Scholar

    6. Andries Zijlstra
      View author publications

      You can also search for this author in PubMed Google Scholar

    Corresponding author

    Correspondence to Andries Zijlstra.

    Ethics declarations

    Animal welfare

    This article does not contain any studies with animals performed by any of the authors.

    Conflict of interest

    The authors have no competing interests to disclose.

    Impact statement

    Despite improvements in the detection and treatment of bladder cancer (BCa), few advances have been made in predicting outcome and non-invasively monitoring disease progression. We show that the ED-A isoform of fibronectin and not total fibronectin is a prognostic urinary biomarker in BCa. Furthermore, the greatest discrimination with urinary ED-A occurs in lymph node negative patients where those with undetectable levels of urinary ED-A are 10 times more likely to survive 2 years following cystectomy compared to those with any detectable level of urinary ED-A.

    Additional information

    Shanna A. Arnold and Holli A. Loomans have contributed equally to this work.

    Electronic supplementary material

    Below is the link to the electronic supplementary material.

    Supplementary material 1 (DOCX 519 kb)

    Rights and permissions

    Reprints and permissions

    About this article

    Check for updates. Verify currency and authenticity via CrossMark

    Cite this article

    Arnold, S.A., Loomans, H.A., Ketova, T. et al. Urinary oncofetal ED-A fibronectin correlates with poor prognosis in patients with bladder cancer. Clin Exp Metastasis 33, 29–44 (2016). https://doi.org/10.1007/s10585-015-9754-x

    Download citation

    • Received:

    • Accepted:

    • Published:

    • Issue Date:

    • DOI: https://doi.org/10.1007/s10585-015-9754-x

    Keywords

    Search

    Navigation