Metalloproteases involved in the Temozolomide (TMZ) resistance of U87-MG glioma cells

Glioblastoma multiforme (GBM) is the most malignant type of brain tumours bearing a grim prognosis with a median survival of 14.8 months. Despite therapeutic advances over the past decades, GBM treatment remains ineffective with temozolomide (TMZ) as a standard chemotherapeutic agent. Even using TMZ combined with surgical resection and radiotherapy, the 2-year survival for patients of high-grade gliomas remains very low. Understanding potential molecular mechanisms contributing to the resistance of glioma cells to TMZ is important for optimizing the existing and developing novel therapeutic strategies. Metalloproteases (MPs) have been found to be increased in GBM and are associated with the malignant phenotype. In our study, we have systematically examined the changes in the expression of MPs in U87-MG cells after TMZ treatment and found that treatment of U87-MG glioblastoma cells with TMZ for at least 5 days induces expression of metalloproteases ADAM8, MMP-1, MMP-9 and MMP-14 in surviving U87-MG cells. To analyse the function of these MPs, a broad-spectrum metalloprotease inhibitor batimastat (BB-94) was applied to U87-MG cells in conjunction with TMZ. BB-94 causes increased susceptibility of U87-MG cells to TMZ induced cell death and reduces the invasive potency of U87-MG cells, indicating that the induction of MPs by TMZ contributes to chemoresistance and recurrence of glioblastoma. As a potential mechanism of MP induction by TMZ we found that ERK1/2 phosphorylation in U87-MG cells is enhanced after TMZ treatment. Application of UO126, a specific ERK1/2 inhibitor, abrogates TMZ-induced expression of ADAM8, MMP-1 and MMP-9, but not of MMP-14, implicating that there are alternative pathways for the induction of MMP-14 by TMZ. PS/γ-secretase combined with MPs participates in trans-membrane processing of numerous proteins thereby regulating various intracellular pathways. Inhibition of γ-secretase using DAPT had a similar effect as BB-94 on sensitization of U87-MG cells to TMZ, suggesting that γ-secretase could function as an adjuvant peptidase to MPs to be involved in the chemoresistance of glioblastoma cells. Taken together, we conclude that the induction of particular MPs MMP-1, MMP-9, MMP-14, and ADAM8 by TMZ causes chemoresistance of glioblastoma cells, and these MPs confer chemoresistance via cleaving membrane-bound proteins, for example CD44, Met or integrins ß1 and/or alphav, which might be involved in DNA damage response (DDR) or in proliferation and invasiveness of tumour cells. Therefore, TMZ treatment combined with MP inhibitor appears as a feasible therapy option to optimize TMZ therapies and to prevent recurrent glioma formation.