Expression Of KIF3A In Triple Negative Breast Cancer And Its Effect On Proliferation And Metastasis In Vitro

Objective To explore the role and significance of KIF3A on the tumorigenesis and development of triple negative breast cancer,by detecting the expression of KIF3A mRNA and protein in triple negative breast cancer,analyzing its relationship with clinicopathological features and investigating the effects of KIF3A on proliferation,migration and invasion in triple negative breast cancer cell lines.Methods Quantitative real-time polymerase chain reaction(qRT-PCR)and Western blot were used to respectively explore the mRNA and protein levels of KIF3A in 9 pairs of triple negative breast cancer fresh tissue and the corresponding adjacent tissues.The expression of KIF3A was examined immunohistochemically in 98 cases of primary triple negative breast cancer and corresponding adjacent tissues.The relationship was then analyzed between the expression of KIF3A in triple negative breast cancer and clinicopathological characters.With lentivirus-mediated interference technology made KIF3A gene silence in triple negative breast cancer MDA-MB-231 and BT549 cells.And KIF3A gene was transfected into triple negative breast cancer MDA-MB-468 cells bv liposome method.MTT assay and colony formation assay were applied to examine proliferation ability of triple negative breast cancer cells.The effect of KIF3A on triple negative breast cancer cell cycle was detected by flow cytometry.Wound healing assay and transwell assay were applied to estimate migration and invasion ability of triple negative breast cancer cells.Western blot was performed to detect the effect of KIF3A on Rb-E2F signaling pathway and epithelial-mesenchymal transition related proteins in triple negative breast cancer.Results The qRT-PCR and Western blot results showed that KIF3A mRNA and protein levels both upregulated in 9 pairs fresh triple negative breast cancer tissues.IHC staining showed that 70 out of 98 triple negative breast cancer cases(71.4%)expressed higher levels of KIF3A than the adjacent tissues.We also found that 25 of 32 cases(78.1%)showed stronger KIF3A expression in the metastatic cancer cells in the lymph node than in the primary cancer tissues.In addition,the expression of KIF3A in triple negative breast cancer was related to lymph node metastasis and recurrence(P < 0.05).There were no significant differences between groups for age,tumor size or grade(P > 0.05).MTT assay and colony formation assay results suggested that silencing of KIF3A inhibited proliferation of triple negative breast cancer cells(P < 0.01)and KIF3A overexpression promoted proliferation of triple negative breast cancer cells(P < 0.05).The proportion of cells in the G1 phase was significantly increased with a decrease in the proportion of cells in the S phase after KIF3A knock-down of triple negative breast cancer cells(P < 0.01).The proportion of cells in the G1 phase was significantly decreased with an increase in the proportion of cells in the S phase after KIF3A over-expression of triple negative breast cancer cells(P < 0.05).It was confirmed via the wound healing assay and transwell assay results that KIF3A knock-down suppressed triple negative breast cancer cells migration and invasion and KIF3A overexpression promoted triple negative breast cancer cells migration and invasion.What’s more,Rb-E2F signaling pathway related proteins Phospho-Rb,E2F1,CyclinE1,CyclinD1 were apparently decreased and P21 was apparently increased after KIF3A knock-down of triple negative breast cancer cells(P < 0.05).And phospho-Rb,E2F1,CyclinE1,CyclinD1 were apparently increased and P21 was apparently decreased after KIF3A over-expression of triple negative breast cancer cells(P < 0.05).The expression level of epithelial-mesenchymal transition related proteins ZEB1,Vimentin,MMP-9 and MMP-2 was decreased,whereas the expression level of E-cadherin was increased after KIF3A knock-down of triple negative breast cancer cells.And this correlation was further confirmed by KIF3A over-expression in triple negative breast cancer cells(P < 0.05),where increased ZEB1,Vimentin,MMP-9 and MMP-2 expressions and a decreased E-cadherin expression were observed(P < 0.05).Conclusion KIF3A may promote tumorigenesis and progression of triple negative breast cancer.Silencing of KIF3A in triple negative breast cancer cells can inhibit the G1/S transition by suppressing the Rb-E2F signaling pathway,which increases cell cycle progression and finally promotes triple negative breast cancer cells proliferation in vitro.Silencing of KIF3A in triple negative breast cancer cells can inhibit cells migration and invasion by repressing EMT.KIF3A might holds potential as a therapeutic drug target for human triple negative breast cancer.