Serum Metastasin mRNA is an Important Survival Predictor in Breast

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Lymph node metastasis
A significant difference (P=0.05) was observed between serummetastasin mRNA and the number of involved lymph nodes. Expressionof serum metastasin mRNA was detected in 100% (12/12) and 33.3% (1/3) of cases with positive LNM, ranging from 1-9 and >/=10,respectively.
Tumour size and vascular density
Although high expression of serum metastasin was associated withtumour size >/=2 cm and microvessels >/=100/mm^sup 2^, thiscorrelation was not statistically significant.
Prognosis
Survival and metastasis rates were 60% (12/20) and 30% (6/20),respectively. Based on the prognostic index, which includes tumoursize, grade and LNM, 65% (13/20) and 35% (7/20) of the patientswere expected to have a five-year survival rate >60% (group withbetter survival) and <60% (group with poor survival),respectively.
The actual survival rate for the first group (PI < 4.5) was84.6% (11/13), while that for the second group (PI>4.5) was14.3% (1/7) over the 30-month follow-up period. The rate ofmetastasis was 15.4% (2/13) and 57% (4/7) in the two groups,respectively.
The discrepancy in survival and metastatic rates between the twogroups was partially explained by the 1.8- and 2.2-fold increase inrelative expression of metastasin (>/=1, Fig. 3) andangiogenesis (>/ =100 microvesseles/mm^sup 2^), respectively.
Discussion
Metastasin protein is a well-established marker of tumourprogression, invasion and metastasis formation, as well as anindicator of poor prognosis.10-12 Strong overexpression ofmetastasin protein has been found in different types of tumours,including breast,17 oesophageal squamous18 and colon carcinomas,19invasive pancreatic carcinomas,20 nonsmall cell lung cancers,21primary gastric cancers22 and bladder cancer.23
To the authors' knowledge, no previous investigations have beenperformed to examine the presence of metastasin mRNA in the serumof breast cancer patients. The present study aimed to provideopportunities to establish a noninvasive test for the earlydetection of breast cancer using serum metastasin mRNA. In thepresent study, a 7.2-fold increase (P<0.001) was observedbetween serum metastasin levels in benign cases and malignantcases. As the positive benign cases were associated withhyperplasia, metastasin might be useful as a marker to monitorcancer development and/or progression.
These results were consistent with work conducted on cell lines andhuman specimens.24 The mRNA of metastasin (S100A4) was expressed atthree- to 25-fold higher levels in cultured infiltrating ductalcarcinoma (IDC) cell lines than in normal or benign human breastcell lines. A statistically significant higher mean level of S100A4mRNA was detected in invasive carcinoma (124 cases: 113IDC and 11carcinoma in situ) than in the benign lesions (20 cases: fivefibrocystic disease and 15 fibroadenomas).
Although other investigators6 were able to detect several genes inhealthy subjects, no metastasin mRNA was detected in our controls.Nikitenko et al.25 demonstrated the presence of S100A4 mRNA in thestromal regions surrounding the epithelial ducts in normal breasttissue specimens. Analysis of 68 tumour biopsies showed that S100A4protein is expressed preferentially by macrophages, fibroblasts andactivated lymphocytes present in the tumour microenvironment,rather than by the tumour cells.26 Moreover, it has been shown thatit is externalised by the stromal cells to the fluid of the tumourmicroenvironment. A significantly higher concentration of S100A4protein was detected in the tumour interstitial fluid (TIF)compared to that in the normal counterparts. The absence ofmetastasin mRNA in control subjects reflects its specialdifferential role in health and disease.
Serum beta-actin mRNA was detected in all tested subjects,confirming the presence and integrity of serum RNA, and of equalloading. The same results were achieved using GAPDH RNA as areference gene.27 Serum GAPDH mRNA was detected in all cancerpatients, those with benign disease, and in the controls.
Nine breast rumour specimens from the group of patients tested forserum metastasin mRNA were selected randomly to investigate theexpression of tissue metastasin mRNA. Serum metastasin wasrepresentative of tissue metastasin in all nine tested samples.Therefore, serum metastasin mRNA may offer a new non-invasiveapproach for cancer screening and/or monitoring of hyperplasticactivity.
The cases in the present study were mainly positive for LNM (15/20), but serum metastasin mRNA was expressed in both LNM-negativeand LNM-positive cases. However, expression showed significantvariation (P=0.05) between the number of involved lymph nodes,which ranged from 1-9 and >/=10. This might further confirminvolvement of metastasin in early metastasis. While otherinvestigators28- 29 have shown an association between metastasinprotein and tumour spread to the regional lymph nodes, specificallyin IDC, others have not detected a significant correlation betweenS100A4 mRNA and the number of nodes affected.24 As the number ofaffected lymph nodes is an important prognostic indicator in breastcancer, these conflicting results require further study.
Vascular density was significantly higher in tumour tissue than innormal tissue (P<0.001). In the tumour specimens, vasculardensity varied considerably and showed no significant relationshipto any histopathological parameter or to serum metastasin mRNAlevel. In contrast, extracellular metastasin protein has been shownto have an angiogenic and motility stimulating effect.30 The bloodvessel network of S100-positive tumours is more pronounced than inS100-negative tumours. This discrepancy could be due simply tofunctional difference between serum metastasin mRNA and theextracellular metastasin oligomeric form of the protein, whichmodulates angiogenesis via inhibition of the thrombospodin gene andinteraction with the annexin II receptor.31,32
Non-significant correlation was observed between hormone receptorsand serum metastasin in the present study, while a non- significantor only weakly significant correlation has been reported by otherworkers;33,34 however, an inverse relationship was detected betweenS100A4 mRNA and oestrogen receptor level.35
Similar non-significant correlation was observed between tumourgrade and serum metastasin. In contrast, a significant correlationbetween S100A4 protein and histological grade was detected in astudy of 62 breast carcinomas.36 This discrepancy between theresults of the present study and those of other studies might beexplained by differences in sample size.
Extracellular S100A4 protein level has been shown to increase inaged mice;37 however, no comments on age were reported in patientswith colorectal cancer.38 No significant correlation between ageand serum metastasin mRNA level was detected in the present study,perhaps because only four patients were aged over 60 years. Theremainder of the patients (n=16) were in the 35-58 age range.
A significant relationship with prognosis was detected in thepresent study. Therefore, high serum metastasin mRNA levels mayprove to be an indicator of poor prognosis in breast cancerpatients. The same poor prognosis was demonstrated for survival ofS100A4 patients in other studies.17,39
Patients who were serum metastasin mRNA-positive developed distantmetastases in the lung, bone and liver, while patients who wereserum metastasin mRMA-negative developed local recurrence. Similarresults have been reported previously,24 where the level of S100A4mRNA proved to be predictive of the time of distant metastasis butnot local metastasis. Biochemical and cell-based studies havedemonstrated the involvement of metastasin in remodelling of theextracellular matrix40 and cellular motility,41,42 which are twosteps in the metastatic cascade.
In conclusion, the present study shows that analysis of metastasinin serum serves as an important prognostic marker in breastcarcinoma and may identify patients who are at high risk ofmetastasis and/or death.
References
1 Leon SA, Shapiro B, Sklaroff DM, Yaros MJ. Free DNA in the serumof cancer patients and the effect of therapy. Cancer Res 1977; 37:646-50.
2 Stroun M, Anker P, Maurice P, Lyautey J, Lederrey C, Beljanski M.Neoplastic characteristics of the DNA found in the plasma of cancerpatients. Oncology 1989; 46: 318-22.
3 Anker P, Mulcahy H, Chen XQ, Stroun M. Detection of circulatingtumor DNA in the blood (plasma/serum) of cancer patients. CancerMetast Rev 1999; 18 (Suppl): 65-73.
4 Kopreski MS, Benko FA, Kwak LW, Gocke CD. Detection of tumormessenger RNA in the serum of patients with malignant melanoma.Clin Cancer Res 1999; 5:1961-5.
5 Chan KCA, Lo YMD. Circulating tumor-derived nucleic acids incancer patients: potential applications as tumor markers. Br JCancer 2007; 96: 681-5.
6 Fleischhacker M, Schmidt B. Circulating nucleic acids (CNAs) andcancer: a survey. Biochim Biophys Acta 2007; 1775:181-232.
7 Reddi KK, Holland JF. Elevated serum ribonuclease in patientswith pancreatic cancer. Proc Natl Acad Sd USA 1976; 73: 2308-10.
8 Tsui NB, Ng EK, Lo YMD. Stability of endogenous and added RNA inblood specimens, serum and plasma. Clin Chem 2002; 48: 1647-53.
9 Ng EK, Tsui NB, Lam NY et al. Presence of filterable andnonfilterable mRNA in the plasma of cancer patients and healthyindividuals. Clin Chem 2002; 48:1212-7.
10 Yao R, Davidson DD, Lopez-Beltran A, MacLennan GT, Montironi R,Cheng L. The SlOO proteins for screening and prognostic grading ofbladder cancer. Histol Histopathol 2007; 22: 1025-32.
11 Tarabykina S, Griffiths TR, Tulchinsky E, Mellon JK, BronsteinIB, Kriajeyska M. Metastsis-associated protein S100A4: spotlight onits role in cell migration. Curr Cancer Drug Targets 2007; 7: 217-28.
12 Garrett SC, Varney KM, Weber DJ, Bresnick AR. S100A4, a mediatorof metastasis. ] Biol Chem 2006; 281: 677-80.
13 El-Abd E, Morsi M, Zaki S, Sobhi A. Relationship between serumhormones and angiogenesis with clinicopathological behaviour inEgyptian breast cancer. Bull Alex Foc Med 2003; 39: 257-64.
14 Haybittle Jl, Blarney RW, Elstonc W et al. A prognostic index inprimary breast cancer. Br J Cancer 1982; 45: 361-6.
15 Sambrook J, Fritsch EF, Maniatis T. Molecular cloning: alaboratory manual 2nd edn. Cold Spring Harbor University Press,1989.
16 Saad A, Sheta M, Sobhi A, Aly E, El-Abd E. Clinical andpathological assessment of sentinel lymph node micrometastasis inbreast cancer using RT-PCR. J Egypt Soc Path 2005; 25: 81-7.
17 Rudland PS, Platt-Higgins A, Renshaw C et al. Prognosticsignificance of the metastasis-inducing protein S100A4 (p9Ka) inhuman breast cancer. Cancer Res 2000; 60: 1595-603.
18 Ninomiya I, Ohta T, Fushida S et al. Increased expression ofS100A4 and its prognostic significance in esophageal squamous cellcarcinoma. Int J Oncol 2001; 18: 715-20.
19 Taylor S, Herrington S, Prime W, Rudland PS, Barraclough R.S100A4 (p9Ka) protein in colon carcinoma and liver metastases;association with carcinoma cells and T-lymphocytes. Br J Cancer2002; 86: 409-16.
20 Rosty C, Ueki T, Argani P et al. Overexpression of S100A4 inpancreatic ductal adenocarcinomas is associated with poordifferentiation and DNA hypomethylation. Am J Pathol 2002; 160: 45-50.
21 Kimura K, Endo Y, Yonemura Y et al. Clinical significance ofS100A4 and E-cadherin-related adhesion molecules in nonsmall celllung cancer. Int J Oncol 2000; 16:1125-31.
22 Yonemura Y, Endou Y, Kimura K et al. Inverse expression ofS100A4 and E-cadherin is associated with metastatic potential ingastric cancer. Clin Cancer Res 2000; 6: 4234-42.
23 Davies BR, O'Donnell M, Durkan GC et al. Expression of S100A4protein is associated with metastasis and reduced survival in humanbladder cancer. J Pathol 2002; 196: 292-9.
24 Barraclough R, Chen H, Davies BR et al. Use of DNA transfer inthe induction of metastasis in experimental mammary systems.Biochem Soc Symp 1998; 63: 273-94. 25 Nikitenko LL, Lloyd BH,Rudland PS, Fear S, Barraclough R. Localisation by in situhybridisation of S100A4 (p9Ka) mRNA in primary human breast tumourspecimens. Int J Cancer 2000; 86: 219-28.
26 Cabezon T, Celis JE, Skibshaj I et al. Expression of S100A4 by avariety of cell types present in the tumor microenvironment ofhuman breast cancer. Int ] Cancer 2007; 121:1433-44.
27 Chen XQ, Bonnefoi H, Pelte M et al. Telomerase RNA as adetection marker in the serum of breast cancer patients. ClinCancer Res 2000; 6: 3823-6.
28 Sherbet GV, Lakshmi MS. S100A4 (MTS1) calcium binding protein incancer growth, invasion and metastasis. Anticancer Res 1998; 18:2415-22.
29 Albertazzi E, Cajone F, Leone BE, Naguib RN, Lakshmi MS, SherbetGV. Expression of metastasis-associated genes h-mtsl (S100A4) andnm23 in carcinoma of breast is related to disease progression. DNACell Biol 1998; 17: 335-42.
30 Helfman DM, Kirn EJ, Lukanidin E, Grigorian M. The metastasis-associated protein S100A4: role in tumor progression andmetastasis. Br J Cancer 2005; 92:1955-8.
31 Roberts DD. Regulation of tumor growth and metastasis bythrombospondinl. FASEB/1996; 10:1183-91.
32 Semov A, Moreno MJ, Onichtchenko A et al. Metastasisassociatedprotein S100A4 induces angiogenesis through interaction withannexin II and accelerated plasmin formation. J Biol Chem 2005;280: 20833- 41.
33 Pedrocchi M, Schafer BW, Muller H, Eppenberger U, Heizmann CW.Expression of Ca(2+)-binding proteins of the S100 family inmalignant human breast cancer cell lines and biopsy samples. Int JCancer 1994; 57: 684-90.
34 Platt-Higgins AM, Renshaw CA, West CR et al. Comparison of themetastasis-inducing protein S100A4 (p9Ka) with other prognosticmarkers in human breast cancer. Int J Cancer 2000; 89: 198-208.
35 Barraclough R. Calcium-binding protein S100A4 in health anddisease. Biochim Biophys Acta 1998; 1448:190-9.
36 Pedersen KB, Nesland JM, Fodstad O, Maelandsmo GM. Expression ofS100A4, E-cadherin, alpha- and beta-catenin in breast cancerbiopsies. Br J Cancer 2002; 87: 1281-6.
37 Ambartsumian N, Klingelhofer J, Grigorian M et al. Themetastasis-associated Mtsl (S100A4) protein could act as anangiogenic factor. Oncogene 2001; 20: 4685-95.
38 Flatmark K, Maelandsmo GM, Mikalsen SO et al. Immunofluorometricassay for the metastasis-related protein S100A4: release of S100A4from normal blood cells prohibits the use of S100A4 as a tumormarker in plasma and serum. Tumour Biol 2004; 1- 2: 31-10.
39 Lee WY, Su WC, Lin PW, Guo HR, Chang TW, Chen HH. Expression ofS100A4 and Met: potential predictors for metastasis and survival inearly-stage breast cancer. Oncology 2004; 66: 429-38.
40 Bjorland K, Winberg JO, Odegaard OT et al. S100A4 involvement inmetastasis: deregulation of matrix metalloproteinases and tissueinhibitors of matrix metalloproteinases in osteosarcoma cellstransfected with an anti-S100A4 ribozyme. Cancer Res 1999; 59:4702- 8.
41 Mandinova A, Atar D, Schafer BW, Spiess M, Aebi U, Heizmann CW.Distinct subcellular localization of calcium binding SlOO proteinsin human smooth muscle cells and their relocation in response torises in intracellular calcium. J Cell Sci 1998; 111: 2043-54.
42 Grigorian MS, Tulchinsky EM, Zain S et al. The mtsl gene andcontrol of tumour metastasis. Gene 1993; 135: 229-38.
E. EL-ABD*, R. EL-TAHAN[dagger], L FAHMY[dagger], S. ZAKI[doubledagger], W. FAID[section], A. SOBHIYen , K. KANDIL[section] and F.EL-KWISKY#
* Molecular Biology Department, Medical Technology Centre;
[dagger] Biochemistry Department, Oncology Unit, [section] SurgeryDepartment,
Yen Pathology Department and # Statistics Department, MedicalResearch Institute;
and [section] Biochemistry Department, Faculty of Science,Alexandria University, Egypt
Accepted: 8 April 2008
Correspondence to: Dr Eman El-Abd
Email: elabde@netscape.net
Copyright Step Communications Ltd. 2008
(c) 2008 British Journal of Biomedical Science. Provided byProQuest Information and Learning. All rights Reserved.

Source: British Journal of Biomedical Science
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