title: High-sensitivity detection of minimal residual breast carcinoma using the polymerase chain reaction and primers for cytokeratin 19.
abstract: We have developed a reverse transcriptase-polymerase chain reaction (RT-PCR) assay to identify breast carcinoma cells in bone marrow aspirates with high sensitivity and specificity. This assay relies on the detection of cytokeratin 19 (K19) RNA by nested primer PCR followed by annealing to a (32P)-labeled internal sequence probe and autoradiography. In reconstitution experiments, this assay is capable of detecting 10 fg of admixed mammary tumor RNA in 1 microgram of normal marrow RNA (a dilution of 1:10(7)). Thirty of 30 primary breast tumor specimens, 19 of 19 cytologically positive bone marrow aspirate specimens, and three of 11 aspirate negative/biopsy positive specimens showed detectable K19 transcript. This assay shows high specificity, with 50 of 52 negative control aspirates showing no detectable amplification product. False-positive amplification was noted in two of 18 aspirates obtained from patients with active chronic myelogenous leukemia. Of stage II and III postsurgical breast carcinoma patients with histologically negative bone marrows and no radiographic bone disease, 14 of 30 were K19 positive by PCR. RT-PCR analysis of K19 transcript is a highly sensitive and specific method of detecting and monitoring low-level metastatic disease in patients with primary carcinoma of the breast. The presence of K19 RNA in histologically negative bone marrows suggests that this assay may prove a powerful monitor for patients undergoing curative therapy as well as a novel prognostic indicator.
authors.affiliation: Department of Pathology, H. Lee Moffitt Cancer Center and Research Institute, University of South Florida, Tampa 33612, USA.
authors.forename: L C
authors.initials: LC
authors.lastname: Moscinski
authors.forename: W L
authors.initials: WL
authors.lastname: Trudeau
authors.forename: K K
authors.initials: KK
authors.lastname: Fields
authors.forename: G J
authors.initials: GJ
authors.lastname: Elfenbein
mesh_terms: D001943:Breast Neoplasms / Q000175:diagnosis* / Q000235:genetics*; D002277:Carcinoma / Q000175:diagnosis* / Q000235:genetics*; D017931:DNA Primers; D005260:Female; D006801:Humans; D007633:Keratins / Q000032:analysis*; D018365:Neoplasm, Residual / Q000175:diagnosis* / Q000235:genetics*; D016133:Polymerase Chain Reaction; D012194:RNA-Directed DNA Polymerase; D012680:Sensitivity and Specificity; D014407:Tumor Cells, Cultured
chemical_list: D017931:DNA Primers; D007633:Keratins; D012194:RNA-Directed DNA Polymerase