An evaluation of molecular markers for improved detection of breast cancer metastases in sentinel nodes

Abstract

Background and objectives:In patients with breast cancer (BC), the sentinel node (SN) is the first node in the axillary basin that receives the primary lymphatic flow and can be used to accurately assess the axillary nodal status without removal of the axillary contents. Currently, histology and/or immunohistochemistry are the routine methods of SN analysis. The primary objective of this study was to develop a reproducible reverse transcription (RT) PCR assay, with emphasis on achieving high specificity for accurate detection of BC micrometastases in the SN. To correct for the heterogeneity of BC cells, a multimarker approach was followed, with the further aim of improving the detection rate of the assay. Methods: In total, 73 markers were evaluated, of which 7 were breast epithelial markers and 66 were either cancer testis or tumour associated antigens. Twelve BC cell lines and 30 SNs (from 30 patients) were analysed using RT-PCR to determine the in vitro and in vivo detection rates for each of the markers. In addition, 20 axillary nodes obtained from a patient with brain death were used as controls to optimise the PCR cycle numbers for all the markers. Results: Of the 30 SNs, 37% (11/30) were positive on haematoxylin and eosin analysis. Extensive immunohistochemical (IHC) analyses of the haematoxylin and eosin negative nodes confirmed the presence of very small numbers of BC cells in an additional 40% (12/30) of SNs. Molecular analysis with the hMAM-A alone identified metastases in 70% (21/30) of SNs. Using MAGE-A3 in combination with hMAM-A identified metastases in 90% (27/30) of patients. Seven SNs (23%) were negative for micrometastases (with haematoxylin and eosin and IHC) but RT-PCR positive for either hMAM-A or MAGE-A3. Conclusions: As IHC analysis resulted in a 77% detection rate compared with 37% for haematoxylin and eosin analysis, we consider that IHC is essential in order not to miss SN micrometastases. Molecular analysis with hMAM-A and MAGE-A3 allows detection of BC micrometastases with a 90% detection rate. However, the clinical value of histologically negative but RT-PCR positive SNs can only be determined with long term follow up.

Patterns of DNA copy number changes in sentinel lymph node breast cancer metastases

“…Genetic markers such as TP53, VEGF, MAM, PTGS2, NME1, and other cancer genes (Abdul-Rasool et al, 2006;Nissan et al, 2006;Zhu et al, 2006) have been shown to have significant prognostic potential in breast cancer with respect to the axillary and systemic metastasis. New information is now emerging from the data obtained from genome-wide studies using high-throughput methodologies where many genes show different characteristics in expression profiles, which can affect the clinical outcome including the development of lymph node metastases (Bertucci et al, 2000;Ahr et al, 2001;van 't Veer et al, 2002).…”

Molecular detection of breast cancer metastasis in sentinel lymph nodes by reverse transcriptase polymerase chain reaction (RT-PCR): identifying, evaluating and establishing multi-marker panels

“…Various studies have suggested that the combination of pathological evaluation and molecular analysis of SLNs yields the highest sensitivity for prediction of the pathological status of ALNs [18] as compared with IHC analysis [28], which in turn shows a higher metastasis detection rate than HE staining alone [11,29]. Expression of transcripts of several genes, including CK19, CEA, MGB1 and MGB2 and NY-BR-1, in SLN have been shown to increase the number of detected positive lymph nodes [28,[30][31][32][33][34].…”

The role of OCT4 immunohistochemistry in evaluation of retroperitoneal lymph node dissections: a pilot study

“…[33][34][35][36] Additionally, multiple molecular markers, including hMAM-A and MAGE-A3, have been successfully used for detection of micrometastasis in breast sentinel lymph nodes. 37 However, the impact of these findings on clinical outcome is debatable, and it has been suggested that the clinical benefit of such additional evaluation, 18 including immunohistochemistry, in some cases may be limited. 23,24 Similar to these other organs, immunohistochemical evaluation of lymph nodes for occult metastases may be of value in directing prognosis and therapy for germ cell tumor patients who undergo retroperitoneal lymph node dissection, particularly if cases are categorized according to clinical suspicion.…”