Association Between Thrombin Activatable Fibrrinolysis Inhibitor And Breast Cancer

Part I Clinical investigationObjective The aim of this study was to investigate if thrombin activatable fibrinolysis inhibitor antigen and activity (TAFI Ag and TAFI Act) levels and TAFI Thr325lle and Thr147Ala polymorphism could be a risk marker of breast cancer in Chinese Han patients.Methods:The plasma TAFI Ag and Act was determined using ELISA and chromogenic assay in256patients with breast cancer and192healthy controls. Besides fibrinogen and D-dimer levels were routinely measured, TAFI Thr325lle (rs1926447) and Thr147Ala (rs3742264) polymorphism was genotyped in both patient and control groups using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis.Results:TAFI Ag and Act levels were significantly higher in breast cancer patients than in controls (100.6±15.2%vs82.7±11.2%and30.5±2.8μg/ml vs23.5±1.6μg/ml, P<0.001respectively). The plasma fibrinogen and D-dimer levels were also significant increased in comparison with those in controls (3.7±1.2g/L vs.2.6±1.4g/L and598.5±115.3ng/mL vs.125.2±55.4ng/mL, P<0.001, respectively). TAFI Ag levels are correlated with metastasis of breast cancer (P<0.001). The Thr/lle (CT) and lle/lle (TT) genotypes were more frequently in patients group compared with the control group [OR:2.106;(95%CI:1.379-3.217); P<0.001]. The high-risk T alleles frequency was also higher in patients compared with healthy controls [OR:1.718;(95%CI:1.316-2.243); P<0.001]. The polymorphism was significantly correlated with TAFI Ag levels in either group (P<0.001). The lle/lle (TT) genotype had the lowest TAFI Ag level, while the Thr/Thr (CC) had the highest one. Thr147Ala gene polymorphism in the case group and control group in the frequency distribution, as well as TAFI antigen level were no significant difference.Conclusions:The plasma TAFI levels and TAFI Thr325lle genotypes were associated with breast cancer patients in Chinese Han populations and can be considered as the risk markers of breast cancer. Patients with breast cancer are at high risk of high coagulation state, which pathogenesis is complex involves both coagulation activation and fibrinolytic inhibition. Part Ⅱ Experiment researchObjective To investigate the effects of CPB2gene silencing on thrombin activated fibrinolysis inhibitor (TAFI) expression and metastatic in the breast cancer cell line MDA-MB-231.Methods Based on the sequence of CPB2, three siRNA were designed and chemically synthesized. They were transfected into the breast cancer cell line MDA-MB-231with LipofectamineTM2000. The TAFI mRNA and protein levels were determined by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot. Cell invasive and metastasis ability and growth inhibition were determined by transwell assay and MTT assay.Results siRNAs targeting CPB2significantly down-regulated the mRNA and the protein level of TAFI (P<0.05), and the inhibition ratio range from55.4%to42.6%. The growth of breast cell line MDA-MB-231were inhibited (P<0.05) and the invasive and metastasis ability were also significantly inhibited compared to the control group (P<0.05).Conclusions CPB2-siRNA can significantly down-regulate TAFI expression in transcriptional and translational levels, inhibit cell growth, invasive and metastasis ability in the breast cell line MDA-MB-231. It can be used as a new target for the breast cancer in therapy of metastasis.