Higher levels of TIMP-1 expression are associated with a poor prognosis in triple-negative breast cancer.

Xiaoming Zhou,Guangcun Cheng,Xuemei Fan,Mingang Hao,Jinglong Wang,Xueqing Sun

doi:10.1186/s12943-016-0515-5

Abstract

BackgroundTissue inhibitor of metalloproteinases-1 (TIMP-1) is a multifunctional protein that can directly regulate apoptosis and metastasis. In this study, we investigated the functional and molecular mechanisms by which TIMP-1 influences triple-negative breast cancer (TNBC).MethodsThe expression level of TIMP-1 in breast cancer tissues was analyzed using the ONCOMINE microarray database. The overall survival of patients with distinct molecular subtypes of breast cancer stratified by TIMP-1 expression levels was evaluated using Kaplan–Meier analysis. Bisulfate sequencing PCR (BSP) was used to analyze the methylation status of the TIMP-1 promoter. Real-time-PCR (RT-PCR), Western blot and ELISA assays were used to evaluate gene and protein expression in cell lines and human tissue specimens. In addition, TIMP-1 function was analyzed using a series of in vitro and in vivo assays with cells in which TIMP-1 was inhibited using RNAi or neutralizing antibodies.ResultsWe found that serum TIMP-1 levels were strongly enhanced in patients with TNBC and that elevated TIMP-1 levels were associated with a poor prognosis in TNBC. However, TIMP-1 levels were not significantly associated with overall survival in other subtypes of breast cancer or in the overall population of breast cancer patients. We also report the first evidence that the TIMP-1 promoter is hypomethylated in TNBC cell lines compared with non-TNBC cell lines, suggesting that aberrant TIMP-1 expression in TNBC results from reduced DNA methylation. RNAi-mediated silencing of TIMP-1 in TNBC cells induced cell cycle arrest at the G1 phase and reduced cyclin D1 expression. In addition, mechanistic analyses revealed that the p-Akt and p-NF-κB signaling pathways, but not the GSK-3β and MAPK1/2 pathways, are associated with TIMP-1 overexpression in TNBC cells. Moreover, neutralizing antibodies against TIMP-1 significantly decreased the rate of tumor growth in vivo.ConclusionsOur findings suggest that TIMP-1 is a biomarker indicative of a poor prognosis in TNBC patients and that targeting TIMP-1 may provide an attractive therapeutic intervention specifically for triple-negative breast cancer patients.

Highlights

Tissue inhibitor of metalloproteinases-1 (TIMP-1) is a multifunctional protein that can directly regulate apoptosis and metastasis
We found that TIMP-1 expression was significantly increased in invasive breast carcinoma (Fig. 1a) and ductal breast carcinoma (Fig. 1b) compared with normal breast tissues
We evaluated the levels of TIMP-1 mRNA and protein in breast cancer cell lines and found that TIMP-1 expression was significantly elevated in the triple-negative breast cancer (TNBC) cell lines

Summary

Introduction

Tissue inhibitor of metalloproteinases-1 (TIMP-1) is a multifunctional protein that can directly regulate apoptosis and metastasis. We investigated the functional and molecular mechanisms by which TIMP-1 influences triple-negative breast cancer (TNBC). There has been a focus on further characterizing the various molecular markers and biomarkers associated with TNBC, including EGFR, VEGFR, c-Myc, C-kit, Poly (ADP-ribose) polymerase-1, HSP90, TOP-2A and spleen tyrosine kinase (SYK) [7, 8]. These biomarkers might be valuable prognostic indicators and might represent potential therapeutic targets of TNBC treatment. Identifying novel biomarkers of TNBC might further contribute to the development of effective TNBC treatment approaches

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Conclusion