Peripheral Circulating Tumor Cells Research Articles

Identification of Enhancer of Zeste Homolog 2 Expression in Peripheral Circulating Tumor Cells in Metastatic Prostate Cancer Patients: A Preliminary Study

PurposeEnhancer of zeste homolog 2 (EZH2), a kind of transcriptional repressor, is reportedly over-expressed in metastatic prostate cancer. In this study, we analyzed EZH2 mRNA in circulating tumor cells (CTCs) in peripheral blood as a biomarker in patients with metastatic prostate cancer.Patients and MethodsBer-EP4 coated immunomagnetic beads were used to harvest CTCs, and mRNA was isolated by oligo-dT conjugated immunomagnetic beads. Reverse transcriptase-polymerase chain reaction for EZH2 mRNA was performed and the expression density was measured. The sensitivity of this test for detection of EZH2 mRNA was determined by serial dilutions of a human prostate cancer cell line. Blood samples were collected from 20 patients each with metastatic or localized prostate cancer and 10 healthy volunteers.ResultsSensitivity experiments showed that the test was highly sensitive as it could detect 10 tumor cells per 5 mL. EZH2 mRNA expression was obtained from peripheral blood samples of patients and control subjects. EZH2 mRNA expression density in the metastatic prostate cancer group was significantly higher than in the control (p = 0.023) and localized prostate cancer groups (p = 0.019). There was no difference between the control and localized prostate cancer groups (p > 0.05).ConclusionEZH2 mRNA expression in circulating epithelial cells represents a promising marker for detecting early metastasis in prostate cancer. However, more specific and sensitive techniques for detection of CTCs are needed to avoid mononuclear cell contamination.

Detection of clonal excess in lymphoproliferative disease by kappa/lambda analysis: correlation with immunoglobulin gene DNA rearrangement

Previous studies have suggested that analysis of the distribution of surface immunoglobulin light chain isotypes by flow cytometry provides evidence for monoclonality of B cell tumors and may detect populations of circulating tumor cells in patients with lymphoproliferative disease. We have used simultaneous flow cytometry and DNA restriction enzyme analysis on 58 samples of tissue and blood to determine whether lymphocyte populations detected by "kappa/lambda" analysis are indeed monoclonal. In greater than 90% of cases, abnormalities detected by flow cytometry correlated with monoclonal rearrangements of immunoglobulin genes as detected by Southern blot analysis. By analyzing tissue and blood from the same patients, we have also demonstrated that monoclonal circulating cells detected by flow cytometry reflect peripheral circulating tumor cells, since DNA from these cells shows the same immunoglobulin rearrangement as DNA from the original tumors in these patients. Although mixing studies suggested that DNA rearrangement studies were more sensitive than was flow cytometry in detecting minor populations of monoclonal lymphocytes, we found only one case in which this affected the diagnostic accuracy of the kappa/lambda analysis, with one notable exception, that of detection of a monoclonal proliferation of B cells that did not express surface immunoglobulin. The kappa/lambda test thus offers a powerful diagnostic tool in the evaluation of lymphoproliferative disease.

Detection of Clonal Excess in Lymphoproliferative Disease by K/λ Analysis: Correlation With Immunoglobulin Gene DNA Rearrangement

Abstract Previous studies have suggested that analysis of the distribution of surface immunoglobulin light chain isotypes by flow cytometry provides evidence for monoclonality of B cell tumors and may detect populations of circulating tumor cells in patients with lymphoproliferative disease. We have used simultaneous flow cytometry and DNA restriction enzyme analysis on 58 samples of tissue and blood to determine whether lymphocyte populations detected by “kappa/lambda” analysis are indeed monoclonal. In greater than 90% of cases, abnormalities detected by flow cytometry correlated with monoclonal rearrangements of immunoglobulin genes as detected by Southern blot analysis. By analyzing tissue and blood from the same patients, we have also demonstrated that monoclonal circulating cells detected by flow cytometry reflect peripheral circulating tumor cells, since DNA from these cells shows the same immunoglobulin rearrangement as DNA from the original tumors in these patients. Although mixing studies suggested that DNA rearrangement studies were more sensitive than was flow cytometry in detecting minor populations of monoclonal lymphocytes, we found only one case in which this affected the diagnostic accuracy of the kappa/lambda analysis, with one notable exception, that of detection of a monoclonal proliferation of B cells that did not express surface immunoglobulin. The kappa/lambda test thus offers a powerful diagnostic tool in the evaluation of lymphoproliferative disease.