Detection And Clinical Significance Of Circulating Tumor Cells And Tumor Stem Cells In Patients With Primary Breast Cancer

Objective: To detect circulating tumor cells(CTC) and tumor stem cells(TSC) in patients with primary breast cancer, and evaluate the relation between CTC and clinical pathology.Methods:1 Peripheral blood samples(10 ml in EDTA) from 44 patients with primary breast cancer and from 15 healthy volunteers were prepared for Peripheral blood mononuclear cells(PBMC).2 PBMC were incubated with cytokeratins 19(CK19) mouse monoclonal antibodies, as the marker of circulating tumor cells (CTC).3 The cells were selected with Magnetic-activated cell sorting(MACS) for positive selection of cytokeratin-19-positive (CK¹⁹⁺)cells.4 A double immunohistochemical staining was applied on the CK¹⁹⁺ cells using the anti-CD44 mouse monoclonal antibodies and anti-CD24 mouse monoclonal antibodies, detection of CD44⁺/CD24^—/low cells as the marker of tumor stem cells(TSC).5 Selected the CD44⁺/CD24^—/low cells using fluorescence microscope.6 The data was analyzed with SPSS13.0 statistical software.Results:1 There was no CK19 positive cells in peripheral blood of 15 healthy volunteers. Detection of 44 patients with primary breast cancer showed that 33 of 44(75.00%) patients had CK19 positive cells in PBMC. It showed statistical difference between tow groups(P< 0.05).2 44 patients were divided into three groups based on the TNM system, 5 of 12(41.67%) samples in stageâ… , 15 of 18(83.33%) samples in stageâ…¡and 13 of 14(92.86%) samples in stageâ…¢had CK19 positive cells. It showed statistical difference between different clinical stage(P< 0.05). 3 44 samples were divided into two groups based on lymph node metastasis, 15 of 24(62.50%) in N0 and 18 of 20(90.00%) in N_1-3 had CK19 positive cells. It showed statistical difference between different clinical stage(P< 0.05).4 CD44⁺/CD24^—/low cells were detected in 21 of 33 (63.63%) patients who had CK19 positive cells in peripheral blood. Three groups were divided based on the TNM system, 2 of 5(40.00%) samples in stageâ… , 10 of 15(66.67%) samples in stageâ…¡and 9 of 13(69.23%)samples in stageâ…¢had CD44+/CD24—/low cells.5 24 of 33(72.73%) found CK19 positive cells in ER-positive samples, 15 of 24(62.50%) samples found CD44⁺/CD24^—/low cells; 9 of 11(81.82%) found CK19 positive cells in ER-negative samples, 6 of 9(66.67%) samples found CD44⁺/CD24^—/low cells.6 4 of 6(66.67%) found CK19 positive cells in C-erbB2 positive samples, 2 of 4(50.00%) samples found CD44⁺/CD24^—/low cells; 16 of 24(66.67%) found CK19 positive cells in C-erbB2 negative samples, 11 of 16(68.75%) samples found CD44⁺/CD24^—/low cells.7 9 of 13(69.23%) found CK19 positive cells in angiolymphatic invasion samples, 8 of 9(88.89%) samples found CD44⁺/CD24^—/low cells; 24 of 31(77.42%) found CK19 positive cells in non-angiolymphatic-invasion samples, 13 of 24(54.17%) samples found CD44⁺/CD24^—/low cells.8 All cases had been classified into 3 subgroups (triple-negative, ER- positive, and C-erbB2 positive) based on gene expression profiles. 5 of 5(100.00%) found CK19 positive cells in triple-negative samples, 4 of 5(80.00%) samples found CD44⁺/CD24^—/low cells; 11 of 19(57.89%) found CK19 positive cells in ER-positive samples, 7 of 11(63.64%) samples found CD44⁺/CD24^—/low cells; 3 of 4(75.00%) found CK19 positive cells in C-erbB2 positive samples, 2 of 3(66.67%) samples found CD44⁺/CD24^—/low cells.Conclusion:1 CTC and TSC can be detected in peripheral blood from patient with primary breast cancer.2 The ratio of CTC is related to clinical stage and metastastc lymph node. 3 TSC is related to clinical stage, and there are more TSC in later stage patients than eraly stage patients.4 The ratio of CTC and TSC is related tuo ER, PR, C-erbB2, angiolymphatic invasion and gene expression profiles.