Abstract 2227: Effects of cathepsin S in differentiated and stem-like glioblastoma cells

Abstract

Abstract Background: Glioblastoma (GBM) is the most common and aggressive brain tumour subtype with a poor prognosis of ∼15 months. A GBM stem-like population within the tumour is hypothesised to promote resistance to current treatment and invasion through normal tissue. Cathepsin S (Cat S) is a lysosomal cysteine protease that remains catalytically active at neutral pH and has been shown to play a role in radiation resistance and invasion of cancer cells. It is also an independent predictor of GBM patient survival. Materials and methods: GBM cell lines (U87MG and LN229) with lentiviral shRNA knockdown of Cat S are grown in serum-free medium with growth factors that allow de-differentiation of the cells into a stem-like state. A transwell system is used for quantifying monolayer invasion. Under stem conditions, spheroid structures are formed and their invasion in 3D is analysed with GFP expression. Protein expression determined via Western blot. Radiation response determined via clonogenic survival assay. Proliferation measured using MTT assay. Cat S-like activity measured using fluorogenic peptide substrate cleavage. Results: Radiation induces Cat S protein expression and protease activity in both GBM cell lines at 24 hours. Knockdown of Cat S induces radiation sensitivity in both cell lines when differentiated. In stem-like conditions, this is observed for U87MG but not for LN229 cells. Similarly, Cat S knockdown has no effect on proliferation of cell lines, except LN229 stem-like cells where it is increased. Finally, Cat S knockdown reduces the invasive capacity of both differentiated cell lines and U87MG stem-like cells 3D invasion through both collagen and matrigel, but has no effect on LN229 stem-like cells. Cat S-like activity is reduced in all but stem LN229, despite complete knockdown. Further investigation into differential effects in stem-like cells will examine possible role of autophagy, which is linked to radiation resistance and Cat S inhibition. Conclusion Cat S is a potential drug target in GBM as it reduces invasion and radio-resistance of glioma cell lines. The differential effect seen in stem-like cells suggests that the genetic background of tumours may be an important factor in determining utility of Cat S therapy in targeting the stem-like GBM population thought to be responsible for recurrence. Citation Format: Robyn A. Foster, Shahnaz T. Al Rashid, Roberta E. Burden, Christopher J. Scott, Kevin M. Prise, Thomas J. Flannery. Effects of cathepsin S in differentiated and stem-like glioblastoma cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2227. doi:10.1158/1538-7445.AM2015-2227