Effects of DSPP and MMP20 Silencing on Key Signaling Pathways in Oral Squamous Cell Carcinoma Cells

Abstract

Introduction: Oral carcinogenesis is a multistage process, featuring genetic and molecular alterations leading to rapid cell division, invasion, metastasis, and increased cell survival. Many of these alterations are due to perturbations in the cell signaling networks, which in turn lead to constitutive deregulation of the proteins involved in the regulatory pathways. Our recent reports show that the silencing of dentin sialophosphoprotein (DSPP) and its cognate matrix metalloproteinases 20 (MMP20) alters key tumorigenic hallmarks of oral squamous cell carcinoma (OSCC). Objective: This study, intended to advance our recent findings, focuses on determining the effects of silencing DSPP and its cognate MMP20 on the signaling pathways that control cell proliferation, differentiation, invasion and metastasis. Materials and Methods: DSPP and MMP20 were silenced individually and in combination, using adenovirus-mediated short hairpin RNA (shRNA) in OSCC cell line, OSC2, and the effects of silencing on the following pathways: EFGR; RAS-RAF; MEK; MAPK; ERK; JNK; NF-kB; TGFβ; and GSK3β, were analysed by western blot. Results: DSPP and MMP20 silencing decreased EGFR, KRAS, MEK1/2, MAPK, ERK, MEEK1, JNK, CREBP, p300, NF-kB,TGF β, SMAD7, GSK3 β, and β-catenin expressions. In contrast, the expression of IKKα and SMAD4 were increased in DSPP/MMP20-silenced group, compared with control group. Furthermore, DSPP-silencing alone was more effective than MMP20, or combined DSPP-MM20 silencing, in altering the levels of key proteins of each signaling pathway investigated. Conclusion: Our findings provide the basis for further studies aimed at verifying the effects of these alterations in the profiles of these proteins on the various hallmarks of oral carcinogenesis, and for understanding the molecular role of DSPP and MMP20 in OSCC. This is with a view to evaluating their diagnostic and prognostic utility as well as the values of DSPP/MMP20 as potential targets for design of chemotherapeutic agents for the treatment of OSCC patients.