Expression Of CK19,CEAmRNA In Peripheral Blood From Patients With Breast Cancer Detected By Real-time Quantitative PCR And The Relationship With Metastasis Related Factors

Objective To explore the clinical significance of detection of cytokeratin 19(CK19)mRNA and carcinoembryonic antigen(CEA) mRNA expression for the diagnosis ofmicrometastasis in the peripheral blood of patients with breast cancer,and the relationshipwith the metastasis related factors including variant exon 6-containing isoforms of CD44(CD44v6),matrix metalloproteinase-9(MMP9),vascular endothelial growth factor(VEGF)and other clinicopathological characteristics .Methods β-actin was used as internal control. Total RNA extracted from breast carcinomacell line(MCF7) and 19 cases of fresh breast carcinoma tissue were detected of CK19,CEAmRNA by real-time quantitative polymerase chain reaction(RQ-PCR). Theexpression of CK19,CEAmRNA by RQ-PCR in peripheral blood samples from 28 normalhealthy volunteers,20 patients with benign breast disease and 108 patients with breastcancer(88 patients with operable breast cancer and 20 patients with metastatic breast cancer)were also tested. The expression of CD44v6, MMP9 and VEGF were detected byimmunohistochemical techniques(Elivision) in 88 specimens of operable breast carcinoma .Results The expression of CK19,CEAmRNA of breast carcinoma cell line(MCF7) was allpositive in 15 tests,and the positive rate was 100%. In 19 fresh breast carcinoma tissues, theCK19mRNA positive rate was 100%(19/19), while the CEAmRNA was 78.9%(15/19). Itwas considered positive with micrometastasis if either CK19mRNA or CEAmRNA wasdetected in peripheral blood samples.Of 28 samples from normal volunteers,none waspositive for both CK19mRNA and CEAmRNA. None of the blood samples from patientswith benign breast disease was positive for CEAmRNA,but one was positive forCK19mRNA (5%). The positive rates of CK19mRNA and CEAmRNA from the blood ofbreast cancer patients were 24.1%(26/108) and 21.3%(23/108) respectively, the positiverate of both CK19mRNA and CEAmRNA was 13.9%(15/108). The positive rates ofCK19mRNA and(or) CEAmRNA for metastatic breast cancer patients were higher thanthat for operable breast cancer patients. Among 22 CK19mRNA and(or) CEAmRNApositive operable breast cancer patients, 2(9.1%) relapsed within 1 year after surgery.In 66other patients whose both mRNA were negative, none replased within 1 year aftersurgery.In the breast carcinoma tissues of 88 patients with operable breast cancer, 81(92%)got positive expression of CD44v6, 76(86.4%) had positive MMP9 expression, 71(80.7%)got positive expression of VEGF.In the group of operable breast cancer with lymph nodemetastasis, the expression of VEGF increased significantly, compared to the group ofnon-lymph node metastasis (p<0.05), but there wasn,t statistically significant differencebetween the two group in CD44v6 and MMP9 (p>0.05). Statistically significant differenceswere found between operable breast cancer patients with micrometastases in peripheral bloodand patients without micrometastases among the tumor size, clinical stage,expressions ofki67,CD44v6, VEGF status (p<0.05).There was no statistically significant differencebetween the two groups in age, tumor location, pathological histology type, nodalmetastasis, expressions of estrogen receptor(ER), pregnant receptor(PR),CerbB2,MMP9status (p>0.05). Multivariate analysis showed that tumor size, clinical stage, expressionsof ki67,CD44v6,VEGF status were associated with micrometastases in peripheral blood.Conclusion CK19 ,CEAmRNA would be good markers for detecting themicrometastases in the peripheral blood of breast cancer patients. The combined detectionof CK19 and CEAmRNA could improve the detection rates of micrometastases in theperipheral blood. Detection of micrometastases in peripheral blood of breast cancer patientsmight be helpful to predict the prognosis of patients. Tumor size, clinical stage, expressionsof ki67, CD44v6, VEGF status were effective predictors of micrometastases in theperipheral blood of breast cancer patients.