Sensitive detection of micrometastases in bone marrow from patients with breast cancer using immunomagnetic isolation of tumor cells in combination with reverse transcriptase/polymerase chain reaction for cytokeratin-19

2000 ◽  
Vol 126 (4) ◽  
pp. 212-218 ◽  
Author(s):  
Xiao Yan Zhong ◽  
Sepp Kaul ◽  
Yung Sheng Lin ◽  
Astrid Eichler ◽  
Gunther Bastert
Keyword(s):  
Breast Cancer ◽  
Bone Marrow ◽  
Polymerase Chain Reaction ◽  
Reverse Transcriptase ◽  
Tumor Cells ◽  
Sensitive Detection ◽  
Cytokeratin 19 ◽  
Chain Reaction ◽  
Immunomagnetic Isolation ◽  
Polymerase Chain

Reverse transcriptase/polymerase chain reaction detection of cytokeratin-19 mRNA in bone marrow and blood of breast cancer patients

1996 ◽  
Vol 122 (11) ◽  
pp. 679-686 ◽  
Author(s):  
William Kr�ger ◽  
Cordula Krzizanowski ◽  
Michael Holweg ◽  
Marcus Stockschl�der ◽  
Nicolaus Kr�ger ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Bone Marrow ◽  
Polymerase Chain Reaction ◽  
Reverse Transcriptase ◽  
Cancer Patients ◽  
Cytokeratin 19 ◽  
Breast Cancer Patients ◽  
Polymerase Chain Reaction Detection ◽  
Chain Reaction ◽  
Polymerase Chain

Specific detection of carcinoembryonic antigen-expressing tumor cells in bone marrow aspirates by polymerase chain reaction.

1994 ◽  
Vol 12 (4) ◽  
pp. 725-729 ◽  
Author(s):  
M Gerhard ◽  
H Juhl ◽  
H Kalthoff ◽  
H W Schreiber ◽  
C Wagener ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Bone Marrow ◽  
Polymerase Chain Reaction ◽  
Tumor Cells ◽  
Normal Bone Marrow ◽  
Specific Detection ◽  
Chain Reaction ◽  
Pcr Assay ◽  
Normal Bone ◽  
Polymerase Chain

PURPOSE To establish a sensitive assay for the specific detection of carcinoembryonic antigen (CEA)-expressing tumor cells in the bone marrow of patients with colorectal cancer and other CEA-positive carcinomas. PATIENTS AND METHODS A CEA-specific nested reverse transcriptase (RT) polymerase chain reaction (PCR) assay was developed and optimized using limiting dilutions of a CEA-positive cancer cell line mixed with normal bone marrow cell specimens. The optimized test was then used to examine bone marrow samples obtained from 15 patients with abdominal carcinomas (colorectal, n = 10; pancreatic, n = 3; gastric, n = 2) and six patients with breast cancer. Specificity was assessed by examination of 56 negative controls (malignant hematologic disease, n = 28; nonmalignant disease conditions, n = 5; healthy bone marrow donors, n = 8; normal peripheral-blood samples, n = 15). For 11 patients with abdominal carcinomas, immunostaining evaluations were performed using an anti-CEA and an anticytokeratin antibody, and the results compared with the nested PCR assay. RESULTS In the sensitivity calibration system, single CEA-expressing tumor cells were detected in 2 to 5 x 10(7) normal bone marrow cells. All 56 control samples failed to amplify. This demonstrates that mRNAs coding for highly homologous CEA-related antigens expressed by various lineages of blood cells do not interfere. Bone marrow samples from 10 of 15 patients with abdominal cancers and four of six breast cancer patients scored positive, indicating micrometastatic bone disease. Four of 11 samples from the gastrointestinal cancer patients were found to be positive by the PCR method, but were negative with the immunocytology method. CONCLUSION Since approximately 30% of the colorectal carcinoma patients that score negative in immunocytology staining of bone marrow samples have been reported to relapse, earlier diagnosis of the presence of malignant cells is needed. Our result that samples scoring positive in the described CEA-specific PCR test remained negative by two immunostaining methods suggests a higher sensitivity. We conclude that PCR amplification of CEA mRNA may lead to an earlier diagnosis of micrometastatic bone disease in patients with CEA-expressing carcinomas.

Limitations of reverse-transcriptase polymerase chain reaction analyses for detection of micrometastatic epithelial cancer cells in bone marrow.

1997 ◽  
Vol 15 (7) ◽  
pp. 2701-2708 ◽  
Author(s):  
A Zippelius ◽  
P Kufer ◽  
G Honold ◽  
M W Köllermann ◽  
R Oberneder ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Bone Marrow ◽  
Polymerase Chain Reaction ◽  
Reverse Transcriptase ◽  
Tumor Cells ◽  
Carcinoma Cells ◽  
Cell Detection ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Polymerase Chain

PURPOSE 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). PATIENTS AND 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. CONCLUSION 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.

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