Limitations of specific reverse-transcriptase polymerase chain reaction markers in the detection of metastases in the lymph nodes and blood of breast cancer patients.
OBJECTIVE
This study was performed to evaluate the potential of specific mRNA markers to detect micrometastases by reverse-transcriptase polymerase chain reaction (RT-PCR) and Southern blot analysis of sentinel lymph nodes (SNs) and blood from patients with breast cancer.


METHODS
We assessed the specificity of carcinoembryonic antigen (CEA), cytokeratin-19 (CK-19), CK-20, gastrointestinal tumor-associated antigen-733.2 (GA733.2), and mucin-1 (MUC-1) in the blood of healthy donors (n = 13) and lymph nodes from patients without cancer (n = 3) by RT-PCR assay. The sensitivity of the RT-PCR assay for the target mRNA markers was assessed in breast cancer cell lines (n = 4), primary breast tumors (n = 8), and the frozen sections of SNs (n = 22) from 22 patients with American Joint Committee on Cancer (AJCC) stages I to IIIA breast cancer.


RESULTS
CK-20 was the only mRNA marker not detected in lymph nodes or blood from patients without cancer. Both the blood and lymph nodes from patients without cancer expressed CEA, CK-19, GA733.2, and MUC-1 mRNA. All four breast cancer cell lines and six of eight primary breast tumors expressed all five mRNA markers. Expression of mRNA by the RT-PCR assay in the frozen-section SNs (n = 12) without metastases by conventional histopathology ranged from 8% (CK-20) to 92% (GA733.2). Detection of RT-PCR cDNA products in frozen-section SNs was increased with Southern blot analysis compared with ethidium bromide gel electrophoresis (EtBr) for all mRNA markers except CK-19.


CONCLUSIONS
CEA, CK-19, GA733.2, and MUC-1 show no diagnostic value as mRNA markers for the detection of micrometastases by the RT-PCR assay because they are expressed in the blood and lymph nodes of patients without cancer. Further studies are needed to assess the sensitivity of CK-20 to detect micrometastases by the RT-PCR assay in the blood and frozen-section SNs of patients with breast cancer.
Department of Molecular Oncology, John Wayne Cancer Institute at Saint John's Health Center, Santa Monica, CA 90404, USA.
P J
PJ
Bostick
S
S
Chatterjee
D D
DD
Chi
K T
KT
Huynh
A E
AE
Giuliano
R
R
Cote
D S
DS
Hoon
D000951:Antigens, Neoplasm / Q000032:analysis; D014408:Biomarkers, Tumor / Q000097:blood; D015139:Blotting, Southern; D001943:Breast Neoplasms / Q000737:chemistry* / Q000473:pathology*; D002272:Carcinoembryonic Antigen / Q000032:analysis; D015815:Cell Adhesion Molecules / Q000032:analysis; D000071858:Epithelial Cell Adhesion Molecule; D005260:Female; D005629:Frozen Sections; D006801:Humans; D007381:Intermediate Filament Proteins / Q000032:analysis; D053554:Keratin-20; D007633:Keratins / Q000032:analysis; D008198:Lymph Nodes / Q000737:chemistry; D008207:Lymphatic Metastasis; D018396:Mucin-1 / Q000032:analysis; D009360:Neoplastic Cells, Circulating; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000032:analysis; D012194:RNA-Directed DNA Polymerase; D012680:Sensitivity and Specificity; D014407:Tumor Cells, Cultured
D000951:Antigens, Neoplasm; D014408:Biomarkers, Tumor; D002272:Carcinoembryonic Antigen; D015815:Cell Adhesion Molecules; D000071858:Epithelial Cell Adhesion Molecule; D007381:Intermediate Filament Proteins; C507499:KRT20 protein, human; D053554:Keratin-20; D018396:Mucin-1; D012333:RNA, Messenger; D007633:Keratins; D012194:RNA-Directed DNA Polymerase