TY - JOUR
T1 - Establishment and validation of real-time polymerase chain reaction method for CDH1 promoter methylation
AU - Toyooka, Kiyomi O.
AU - Toyooka, Shinichi
AU - Maitra, Anirban
AU - Feng, Qinghua
AU - Kiviat, Nancy C.
AU - Smith, Alice
AU - Minna, John D.
AU - Ashfaq, Raheela
AU - Gazdar, Adi F.
PY - 2002
Y1 - 2002
N2 - Aberrant methylation of the promoter region has emerged as the major mechanism for silencing tumor suppressor genes. However, for some genes, such as E-cadherin (CDH1), methylation and protein expression demonstrate considerable heterogeneity, making correlations difficult. We compared methylation and protein expression status of CDH1 in 56 primary breast carcinomas using semiquantitative assays. Aberrant CDH1 methylation was studied by methylation-specific polymerase chain reaction (MSP) and semiquantitative real-time MSP assays. The Cdh1 expression was investigated by immunostaining on archival formalin-fixed sections from 34 primary carcinomas and their accompanying normal epithelium and preinvasive and metastatic lesions. Membrane-specific Cdh1 expression in the neoplastic cells was quantified by image analysis using an automated cellular imaging system and a continuous score. Aberrant promoter methylation of the CDH1 was present in 24 of 56 (43%) breast carcinomas by MSP assay. There was excellent concordance between the standard MSP assay and the real-time assay (91%, P < 0.0001). The concordance between loss of Cdh1 expression and CDH1 methylation by standard MSP was 71% (P = 0.02). Furthermore, there was a strong correlation between the semiquantitative assays for methylation and protein expression (r = 0.47, P = 0.005). We conclude that promoter methylation of CDH1 significantly correlated with the Cdh1 expression level, demonstrating that epigenetic silencing is a valid pathway for silencing of tumor suppressor genes in primary breast carcinomas.
AB - Aberrant methylation of the promoter region has emerged as the major mechanism for silencing tumor suppressor genes. However, for some genes, such as E-cadherin (CDH1), methylation and protein expression demonstrate considerable heterogeneity, making correlations difficult. We compared methylation and protein expression status of CDH1 in 56 primary breast carcinomas using semiquantitative assays. Aberrant CDH1 methylation was studied by methylation-specific polymerase chain reaction (MSP) and semiquantitative real-time MSP assays. The Cdh1 expression was investigated by immunostaining on archival formalin-fixed sections from 34 primary carcinomas and their accompanying normal epithelium and preinvasive and metastatic lesions. Membrane-specific Cdh1 expression in the neoplastic cells was quantified by image analysis using an automated cellular imaging system and a continuous score. Aberrant promoter methylation of the CDH1 was present in 24 of 56 (43%) breast carcinomas by MSP assay. There was excellent concordance between the standard MSP assay and the real-time assay (91%, P < 0.0001). The concordance between loss of Cdh1 expression and CDH1 methylation by standard MSP was 71% (P = 0.02). Furthermore, there was a strong correlation between the semiquantitative assays for methylation and protein expression (r = 0.47, P = 0.005). We conclude that promoter methylation of CDH1 significantly correlated with the Cdh1 expression level, demonstrating that epigenetic silencing is a valid pathway for silencing of tumor suppressor genes in primary breast carcinomas.
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U2 - 10.1016/S0002-9440(10)64218-6
DO - 10.1016/S0002-9440(10)64218-6
M3 - Article
C2 - 12163387
AN - SCOPUS:0036965997
SN - 0002-9440
VL - 161
SP - 629
EP - 634
JO - American Journal of Pathology
JF - American Journal of Pathology
IS - 2
ER -