Detection of circulating breast cancer cells with multiple-marker RT-PCR assay

RT-PCR assay for multiple markers has been shown to increase the detection rate of circulating tumor cells. This assay targeted against cytokeratin 19 (CK19), cytokeratin 20 (CK20) and beta-subunit of human chorionic gonadotropin (beta-hCG) was used to detect circulating breast cancer cells. 5 ml of peripheral blood was drawn before any surgical procedures from 72 breast cancer patients and 30 cases with benign breast disease. Total RNA was extracted from peripheral blood mononuclear cells, reverse-transcripted and amplified. For the benign cases, 10% (3/30) were positive for CK19 and all were negative for CK20 and beta-hCG, whereas 9.72% (7/72), 2.78% (2/72) and 12.5% (9/72) of the malignant cases were positive for CK19, CK20 and beta-hCG respectively. The detection rate of circulating breast cancer cells was unchanged when CK19 was combined with CK20, but it increased to 18.1% (13/72) when the marker was combined with beta-hCG. A significant difference was observed for beta-hCG between benign cases and affected patients with stage II, III and IV disease (p = 0.026). In conclusion, positive RT-PCR signals in blood samples of affected patients correlated with stage, in particular for beta-hCG. CK19/beta-hCG was a promising marker combination.