title: Limitations of reverse-transcriptase polymerase chain reaction analyses for detection of micrometastatic epithelial cancer cells in bone marrow.
abstract: OBJECTIVE
  This study was designed to evaluate the potential of reverse-transcriptase polymerase chain reaction (RT-PCR) analyses for the detection of micrometastatic carcinoma cells in bone marrow (BM).
  
  
  METHODS
  The specificity of RT-PCR assays with primers specific for various tumor-associated and organ-specific mRNA species was examined by analysis of 53 BM aspirates from control patients with no epithelial malignancy. In addition, BM samples from 63 patients with prostate cancer (n = 53) or breast cancer (n = 10) were analyzed by RT-PCR with primers specific for prostate-specific antigen (PSA) mRNA. As a reference method, all samples were analyzed simultaneously by an established immunocytochemical assay, using monoclonal antibodies (mAbs) against cytokeratins (CK) for tumor-cell detection.
  
  
  RESULTS
  Seven of eight marker species could be detected in a considerable number of BM samples from control patients: epithelial glycoprotein-40 (EGP-40; 53 of 53 samples), desmoplakin I (DPI I; five of five), carcinoembryonic antigen (CEA; five of 19), erb-B2 (five of seven), erb-B3 (six of seven), prostate-specific membrane antigen (PSM; four of nine), and CK18 (five of seven). Only PSA mRNA was not detected in any of the 53 control BM samples. In serial dilution experiments, the PSA RT-PCR assay was able to detect five LNCaP prostate carcinoma cells in 4 x 10(6) BM cells. CK-positive cells were found in 20 patients (37.7%) with prostate cancer, while PSA mRNA was found in only 15 (28.3%; P = .04). Moreover, despite the recent observation that PSA is also expressed in mammary carcinomas, none of the 10 CK-positive BM samples were PSA mRNA-positive.
  
  
  CONCLUSIONS
  Limiting factors in the detection of micrometastatic tumor cells by RT-PCR are (1) the illegitimate transcription of tumor-associated or epithelial-specific genes in hematopoietic cells, and (2) the deficient expression of the marker gene in micrometastatic tumor cells.
authors.affiliation: Institut für Immunologie der Universität Munchen, Germany.
authors.forename: A
authors.initials: A
authors.lastname: Zippelius
authors.forename: P
authors.initials: P
authors.lastname: Kufer
authors.forename: G
authors.initials: G
authors.lastname: Honold
authors.forename: M W
authors.initials: MW
authors.lastname: Köllermann
authors.forename: R
authors.initials: R
authors.lastname: Oberneder
authors.forename: G
authors.initials: G
authors.lastname: Schlimok
authors.forename: G
authors.initials: G
authors.lastname: Riethmüller
authors.forename: K
authors.initials: K
authors.lastname: Pantel
mesh_terms: D014408:Biomarkers, Tumor / Q000032:analysis*; D019046:Bone Marrow Neoplasms / Q000175:diagnosis* / Q000201:enzymology / Q000235:genetics / Q000556:secondary*; D002277:Carcinoma / Q000175:diagnosis* / Q000201:enzymology / Q000235:genetics / Q000556:secondary*; D017931:DNA Primers; D005260:Female; D006801:Humans; D008297:Male; D016133:Polymerase Chain Reaction / Q000379:methods*; D011237:Predictive Value of Tests; D012333:RNA, Messenger / Q000032:analysis; D012334:RNA, Neoplasm / Q000032:analysis; D012194:RNA-Directed DNA Polymerase; D012680:Sensitivity and Specificity
chemical_list: D014408:Biomarkers, Tumor; D017931:DNA Primers; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D012194:RNA-Directed DNA Polymerase