Kinesin family member 11 contributes to the progression and prognosis of human breast cancer

Abstract

The present study aimed to clarify the association between kinesin family member 11 (KIF11) and human breast cancer, and the effect of KIF11 on breast cancer cell progression. Western blot analysis, reverse transcription-quantitative polymerase chain reaction (RT-qPCR) analysis, retroviral infection, immunohistochemistry staining, MTT assay, anchorage-independent growth ability assay and tumorigenicity assay were all used in the present study. Western blot and RT-qPCR analysis revealed that the expression of KIF11 was markedly increased in malignant cells compared with that in non-tumorous cells at the mRNA and protein level. Immunohistochemical analysis revealed that KIF11 expression was upregulated in 256/268 (95.8%) paraffin-embedded archival breast cancer biopsies. Statistical analysis demonstrated a significant association between the upregulation of KIF11 expression and the progression of breast cancer. Multivariate analysis revealed that KIF11 upregulation represents an independent prognostic indicator for the survival of patients with breast cancer. Tumorigenicity experiments were further used to evaluate the effect of KIF11 in non-obese diabetic/severe combined immunodeficient mice. Silencing endogenous KIF11 by short hairpin RNAs inhibited the proliferation of breast cancer cells in vitro and in vivo. The present results suggest that KIF11 may serve an important function in the proliferation of breast cancer and may represent a novel and useful prognostic marker for breast cancer.

Keywords: breast cancer; kinesin family member 11; prognosis; progression.

Figure 1.

Expression analysis of KIF11 protein and mRNA in breast cancer cells. (A) Western blot analysis of KIF11 protein expression in two NBEC and breast cancer cell lines. NBECs were used as the negative control. (B) Expression of KIF11 mRNA in NBECs and breast cancer cell lines. KIF11, kinesin family member 11; NBEC, normal breast epithelial cells.

Figure 2.

Expression analysis of KIF11 protein and mRNA in breast cancer tissues paired with ANT. (A) Western blot investigation of expression of KIF11 protein in six paired breast cancer tissues. β-actin was used the loading control. (B) Reverse transcription-quantitative polymerase chain reaction analysis of KIF11 expression in the indicated fresh tissues. GAPDH was used as the loading control. (C) Immunohistochemical staining of KIF11 protein in the indicated primary tissues. P<0.05 vs. ANT. KIF11, kinesin family member 11; ANT, adjacent non-tumorous tissues; T, breast cancer tissues.

Figure 3.

Overexpression of KIF11 in archived breast cancer tissues. (A) Representative immunohistochemistry of KIF11 expression in normal breast tissue and breast cancer specimens of different clinical stages. (B) Statistical quantification of the average MOD of KIF11 staining between normal breast tissues and breast cancer specimens of different clinical stages. The average mean absorbance of KIF11 staining increases as breast cancer progresses to a higher clinical stage. (C) Kaplan-Meier curves with univariate analyses (log-rank test) for patients with low KIF11 expression vs. intensive KIF11 expression. KIF11, kinesin family member 11; MOD, mean optical density.

Figure 4.

Downregulation of endogenous KIF11 suppressed MCF-7 and MDA-MB-231 cell proliferation in vitro. (A) Western blot analysis of increased KIF11 expression. (B) MTT assay showing that KIF11-transfected cells grew slower than vector-transfected cells. (C) Representative micrographs and quantification of crystal violet-stained cell colonies. (D) Representative micrographs and colony numbers in the anchorage-independent growth assay. Each bar represents the mean of three independent experiments. *P<0.05 vs. vector. KIF11, kinesin family member 11.

Figure 5.

In vivo assays of the effect of KIF11 on cell proliferation. (A) Excised tumours 35 days subsequent to injection into non-obese diabetic/severe combined immunodeficient mice. (B) Tumour growth curve measured every 5 days. (C) The average weight of the excised tumours in the group injected with vector-transfected cells or KIF11-RNAi1-transfected cells. *P<0.05 vs. vector.