Detection of metastases in sentinel lymph nodes of breast cancer patients by multiple-marker RT-PCR

Abstract

This study was undertaken to assess a multiple-marker RT-PCR and Southern blot assay for detection of metastases in frozen sections of sentinel lymph nodes from breast cancer patients. Sentinel lymphadenectomy was performed in 41 AJCC (American Joint Committee on Cancer) stage I-IIIA breast cancer patients and 57 sentinel nodes (SNs) were excised. The SN, which is the first node in the lymphatic basin to receive metastases from the primary tumor, was identified using isosulfan blue dye. Hematoxylin and eosin (H&E), immuno-histochemistry (IHC) and RT-PCR were performed on adjacent sections of the SN. Six consecutive 12-microm frozen sections of each SN were obtained for the RT-PCR assay to determine expression of mRNA tumor markers C-Met, beta1 --> 4GalNAc-T and P97. Metastatic breast cancer was detected by H&E in 10 of 57 (18%) SNs and by IHC in an additional 7 (12%). Only 1 of 17 (6%) SNs with metastases did not express any of the 3 tumor mRNA markers. C-Met, beta1 --> 4GalNAc-T and P97 tumor mRNA markers were expressed in 31 (78%), 21 (53%) and 25 (63%) of 40 SNs without metastases, respectively. At least 2 mRNA tumor markers were expressed in 25/40 (63%) histo-pathologically tumor-free SNs, whereas all 3 mRNA tumor markers were expressed in 17/40 (43%) SNs. Expression of all 3 mRNA tumor markers in a SN was significantly higher in patients with a family history of breast cancer (p = 0.05), prior history of breast cancer (p < 0.05), infiltrating lobular carcinoma (p = 0.06), estrogen receptor-negative (p = 0.04) tumor or a higher Bloom Richardson score (p = 0.04). The multiple-marker RT-PCR and Southern blot assay improves the detection of occult metastases in the SN when compared to conventional H&E and IHC analysis. Expression of all 3 tumor mRNA markers in the SN correlated with poor prognostic clinico-pathologic factors compared to expression of 0 to 2 markers.