Gene promoter hypermethylation is found in sentinel lymph nodes of breast cancer patients, in samples identified as positive by one-step nucleic acid amplification of cytokeratin 19 mRNA.
We analysed the promoter methylation status of five genes, involved in adhesion (EPB41L3, TSLC-1), apoptosis (RASSF1, RASSF2) or angiogenesis (TSP-1), in intraoperative sentinel lymph node (SLN) biopsy samples from patients with breast cancer, that had been processed by the one-step nucleic acid amplification (OSNA) technique. SLN resection is performed to estimate the risk of tumour cells in the clinically negative axilla, to avoid unnecessary axillary lymph node dissection. OSNA is currently one of the eligible molecular methods for detecting tumour cells in SLNs. It is based on the quantitative evaluation of cytokeratin 19 mRNA which allows distinguishing between macrometastasis, micrometastasis and isolated tumour cells, on the basis of the quantity of tumour cells present. There have been no prior studies on the question whether or not samples processed by OSNA can be used for further molecular studies, including epigenetic abnormalities which are some of the most important molecular alterations in breast cancer. Genomic DNA was extracted from samples obtained from 50 patients diagnosed with primary breast cancer. The content of tumour cells in SLNs was evaluated by OSNA, and the promoter methylation status of the selected genes was analysed by methylation-specific PCR. All were found to be hypermethylated to a variable degree, and RASSF1 hypermethylation was significantly associated with macrometastasis, micrometastasis and isolated tumour cells (p = 0.002). We show that samples used for OSNA are suitable for molecular studies, including gene promoter methylation. These samples provide a new source of material for the identification of additional biomarkers.
Cancer Epigenetics Group, Navarrabiomed-Biomedical Research Center, Instituto de Investigaciones Sanitarias de Navarra-IdiSNA, Pamplona, Navarra, Spain.
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Martín-Sánchez
Cancer Epigenetics Group, Navarrabiomed-Biomedical Research Center, Instituto de Investigaciones Sanitarias de Navarra-IdiSNA, Pamplona, Navarra, Spain.
E
E
Pernaut-Leza
Cancer Epigenetics Group, Navarrabiomed-Biomedical Research Center, Instituto de Investigaciones Sanitarias de Navarra-IdiSNA, Pamplona, Navarra, Spain.
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S
Mendaza
Cancer Epigenetics Group, Navarrabiomed-Biomedical Research Center, Instituto de Investigaciones Sanitarias de Navarra-IdiSNA, Pamplona, Navarra, Spain.|Department of Pathology, Navarra Health Service, Complejo Hospitalario de Navarra, Pamplona, Navarra, Spain.
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Cordoba
Cancer Epigenetics Group, Navarrabiomed-Biomedical Research Center, Instituto de Investigaciones Sanitarias de Navarra-IdiSNA, Pamplona, Navarra, Spain.|Department of Surgery, Navarra Health Service, Complejo Hospitalario de Navarra, Pamplona, Navarra, Spain.
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Vicente-Garcia
Cancer Epigenetics Group, Navarrabiomed-Biomedical Research Center, Instituto de Investigaciones Sanitarias de Navarra-IdiSNA, Pamplona, Navarra, Spain.
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I
Monreal-Santesteban
Department of Hematology, Navarra Health Service, Complejo Hospitalario de Navarra, Pamplona, Navarra, Spain.
J Pérez
JP
Vizcaino
Department of Pathology, Navarra Health Service, Complejo Hospitalario de Navarra, Pamplona, Navarra, Spain.
M J Díaz
MJ
De Cerio
Cancer Epigenetics Group, Navarrabiomed-Biomedical Research Center, Instituto de Investigaciones Sanitarias de Navarra-IdiSNA, Pamplona, Navarra, Spain.
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Perez-Janices
Cancer Immunomodulation Group, Navarrabiomed-Biomedical Research Center, Instituto de Investigaciones Sanitarias de Navarra-IdiSNA, Pamplona, Navarra, Spain.
I
I
Blanco-Luquin
Cancer Immunomodulation Group, Navarrabiomed-Biomedical Research Center, Instituto de Investigaciones Sanitarias de Navarra-IdiSNA, Pamplona, Navarra, Spain.
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Escors
Cancer Epigenetics Group, Navarrabiomed-Biomedical Research Center, Instituto de Investigaciones Sanitarias de Navarra-IdiSNA, Pamplona, Navarra, Spain.
A
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Ulazia-Garmendia
Cancer Epigenetics Group, Navarrabiomed-Biomedical Research Center, Instituto de Investigaciones Sanitarias de Navarra-IdiSNA, Pamplona, Navarra, Spain. dguerres@navarra.es.|Department of Pathology, Navarra Health Service, Complejo Hospitalario de Navarra, Pamplona, Navarra, Spain. dguerres@navarra.es.
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Guerrero-Setas
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D001943:Breast Neoplasms / Q000235:genetics* / Q000473:pathology*; D019175:DNA Methylation; D005260:Female; D016147:Genes, Tumor Suppressor / Q000502:physiology; D006801:Humans; D053539:Keratin-19 / Q000235:genetics* / Q000378:metabolism; D008198:Lymph Nodes / Q000473:pathology; D008207:Lymphatic Metastasis; D008875:Middle Aged; D061206:Neoplasm Micrometastasis / Q000235:genetics / Q000473:pathology*; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics; D000071036:Sentinel Lymph Node / Q000378:metabolism*; D021701:Sentinel Lymph Node Biopsy / Q000379:methods
D053539:Keratin-19; D012333:RNA, Messenger
Breast cancer; DNA methylation; EPB41L3; OSNA; RASSF1; RASSF2; SLN; TSLC-1; TSP-1