Prognostic relevance of Wnt-inhibitory factor-1 (WIF1) and Dickkopf-3 (DKK3) promoter methylation in human breast cancer.
BACKGROUND
Secreted Wnt signaling antagonists have recently been described as frequent targets of epigenetic inactivation in human tumor entities. Since gene silencing of certain Wnt antagonists was found to be correlated with adverse patient survival in cancer, we aimed at investigating a potential prognostic impact of the two Wnt antagonizing molecules WIF1 and DKK3 in breast cancer, which are frequently silenced by promoter methylation in this disease.


METHODS
WIF1 and DKK3 promoter methylation were assessed by methylation-specific PCR with bisulfite-converted DNA from 19 normal breast tissues and 150 primary breast carcinomas. Promoter methylation was interpreted in a qualitative, binary fashion. Statistical evaluations included two-sided Fisher's exact tests, univariate log-rank tests of Kaplan-Meier curves as well as multivariate Cox regression analyses.


RESULTS
WIF1 and DKK3 promoter methylation were detected in 63.3% (95/150) and 61.3% (92/150) of breast carcinoma samples, respectively. In normal breast tissues, WIF1 methylation was present in 0% (0/19) and DKK3 methylation in 5.3% (1/19) of samples. In breast carcinomas, WIF1 methylation was significantly associated with methylation of DKK3 (p = 0.009). Methylation of either gene was not associated with clinicopathological parameters, except for DKK3 methylation being associated with patient age (p = 0.007). In univariate analysis, WIF1 methylation was not associated with clinical patient outcome. In contrast, DKK3 methylation was a prognostic factor in patient overall survival (OS) and disease-free survival (DFS). Estimated OS rates after 10 years were 54% for patients with DKK3-methylated tumors, in contrast to patients without DKK3 methylation in the tumor, who had a favorable 97% OS after 10 years (p < 0.001). Likewise, DFS at 10 years for patients harboring DKK3 methylation in the tumor was 58%, compared with 78% for patients with unmethylated DKK3 (p = 0.037). Multivariate analyses revealed that DKK3 methylation was an independent prognostic factor predicting poor OS (hazard ratio (HR): 14.4; 95% confidence interval (CI): 1.9-111.6; p = 0.011), and short DFS (HR: 2.5; 95% CI: 1.0-6.0; p = 0.047) in breast cancer.


CONCLUSIONS
Although the Wnt antagonist genes WIF1 and DKK3 show a very similar frequency of promoter methylation in human breast cancer, only DKK3 methylation proves as a novel prognostic marker potentially useful in the clinical management of this disease.
Veeck
Jürgen
J
Molecular Oncology Group, Institute of Pathology, University Hospital of the RWTH Aachen, Pauwelsstrasse 30, D-52074 Aachen, Germany. juergen.veeck@rwth-aachen.de
Wild
Peter J
PJ
Fuchs
Thomas
T
Schüffler
Peter J
PJ
Hartmann
Arndt
A
Knüchel
Ruth
R
Dahl
Edgar
E
D048868:Adaptor Proteins, Signal Transducing; D001943:Breast Neoplasms / Q000175:diagnosis / Q000235:genetics*; D018925:Chemokines; D019175:DNA Methylation; D044127:Epigenesis, Genetic; D020868:Gene Silencing; D005819:Genetic Markers; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics*; D015999:Multivariate Analysis; D016133:Polymerase Chain Reaction; D011379:Prognosis; D011401:Promoter Regions, Genetic; D016016:Proportional Hazards Models; D012307:Risk Factors; D015398:Signal Transduction; D051155:Wnt1 Protein / Q000235:genetics*
D048868:Adaptor Proteins, Signal Transducing; D018925:Chemokines; C498624:DKK3 protein, human; D005819:Genetic Markers; D036341:Intercellular Signaling Peptides and Proteins; C495247:WNT1 protein, human; D051155:Wnt1 Protein