Journal Article

TIMP-2 regulates proliferation, invasion and STAT3-mediated cancer stem cell-dependent chemoresistance in ovarian cancer cells

Bilandzic, Maree, Kannourakis, George, Ahmed, Nuzhat, Western, Patrick, Findlay, Jock K., Kadife, Elif, Escalona, Ruth M.

2020

Overview

Background The metzincin family of metalloproteinases and the tissue inhibitors of metalloproteinases (TIMPs) are essential proteins required for biological processes during cancer progression. This study aimed to determine the role of TIMP-2 in ovarian cancer progression and chemoresistance by reducing TIMP-2 expression in vitro in Fallopian tube secretory epithelial (FT282) and ovarian cancer (JHOS2 and OVCAR4) cell lines. Methods FT282, JHOS2 and OVCAR4 cells were transiently transfected with either single or pooled TIMP-2 siRNAs. The expression of different genes after TIMP-2 knock down (T2-KD) or in response to chemotherapy was determined at the mRNA level by quantitative real time PCR (qRT-PCR) and at the protein level by immunofluorescence. Sensitivity of the cell lines in response to chemotherapy after TIMP-2 knock down was investigated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and 5-Ethynyl-2′-deoxyuridine (EdU) assays. Cell invasion in response to TIMP-2 knockdown was determined by xCELLigence. Results Sixty to 90 % knock down of TIMP-2 expression was confirmed in FT282, OVCAR4 and JHOS2 cell lines at the mRNA and protein levels. TIMP-2 knock down did not change the mRNA expression of TIMP-1 or TIMP-3. However, a significant downregulation of MMP-2 in T2-KD cells occurred at both the protein and activation levels, compared to Control (Cont; scrambled siRNA) and Parental cells (P, transfection reagent only). In contrast, membrane bound MT1-MMP protein levels were significantly upregulated in T2-KD compared to Cont and P cells. T2-KD cells exhibited enhanced proliferation and increased sensitivity to cisplatin and paclitaxel treatments. Enhanced invasion was observed in the T2-KD-JOSH2 and OVCAR4 cells but not in T2-KD-FT282 cells. Treatment with cisplatin or paclitaxel significantly elevated the expression of TIMP-2 in Cont cells but not in T2-KD cells, consistent with significantly elevated expression of chemoresistance and CSC markers and activation of STAT3. Furthermore, a potent inhibitor of STAT3 activation, Momelotinib, suppressed chemotherapy-induced activation of P-STAT3 in OVCAR4 cells with concomitant reductions in the expression of chemoresistance genes and CSC markers. Conclusions The above results suggest that TIMP-2 may have a novel role in ovarian cancer proliferation, invasion and chemoresistance.

• Publisher: BioMed Central,BioMed Central Ltd,Springer Nature B.V,BMC
• Subject: Antineoplastic Agents, Phytogenic - pharmacology / Benzamides - pharmacology / Biomedical and Life Sciences / Biomedicine / Bromine compounds / Cancer Research / Cancer stem cells / Cancer therapies / Cell adhesion & migration / Cell and molecular biology / Cell Line, Tumor / Cell proliferation / Cell Proliferation - physiology / Chemoresistance / Chemotherapy / Cisplatin / Cystadenocarcinoma, Serous - drug therapy / Cystadenocarcinoma, Serous - genetics / Cystadenocarcinoma, Serous - metabolism / Cystadenocarcinoma, Serous - pathology / Development and progression / Drug Resistance, Neoplasm / Fallopian tube / Fallopian tubes / Female / Gelatinase A / Gene expression / Health Promotion and Disease Prevention / Humans / Immunofluorescence / Laboratories / Medicine/Public Health / Membrane proteins / Metastasis / MMPs / Neoplasm Invasiveness / Neoplastic Stem Cells - drug effects / Neoplastic Stem Cells - metabolism / Neoplastic Stem Cells - pathology / Oncology / Ovarian cancer / Ovarian Neoplasms - drug therapy / Ovarian Neoplasms - genetics / Ovarian Neoplasms - metabolism / Ovarian Neoplasms - pathology / Paclitaxel / Paclitaxel - pharmacology / Pathology / Patients / Proliferation / Protein binding / Protein Kinase Inhibitors - pharmacology / Proteins / Pyrimidines - pharmacology / Research Article / RNA / siRNA / STAT3 / Stat3 protein / STAT3 Transcription Factor - antagonists & inhibitors / STAT3 Transcription Factor - metabolism / Stem cells / Surgical Oncology / TIMP-2 / Tissue inhibitor of metalloproteinase 1 / Tissue inhibitor of metalloproteinase 2 / Tissue inhibitor of metalloproteinase 3 / Tissue Inhibitor of Metalloproteinase-2 - metabolism / Transfection / Tumor cell lines / Tumors / Womens health