Comparative analysis of the antiglioblastoma activity of adenosine A2A receptor and CK1δ blockers

Abstract

Glioblastoma multiforme (GBM), a multifactorial deadliest cancer with constrained clinical efficacy due to heterogeneity, drug resistance, side effects of the chemotherapeutic drug, necessitating the development of novel cancer therapeutics. The adenosine A_2A receptor targeted binding of antagonist leads to regulation of downstream effectors, mediating the phosphorylation of Casein kinase 1δ kinase domain (CK1δ) in cancers. Here, we performed a comparative investigation of Food and Drug Administration (FDA) approved drugs, istradefylline and riluzole inhibiting adenosine A_2A receptor and CK1δ isoform in GBM cell growth. Molecular interaction of riluzole with CK1δ isoform and istradefylline with adenosine A_2A receptor was identified through molecular docking and dynamic simulations. The potential of these two FDA approved drugs in inhibiting GBM cell growth was investigated through various in-vitro analysis including dose-dependent dynamic assay, cell cycle assay, apoptosis assay by flow cytometry. Further the effect of these drugs on spheroid cell growth and cell size was measured. In silico analyses demonstrated that riluzole binds strongly to CK1δ isoform with a binding energy of -9.02 kcal/mol, whereas istradefylline binds to adenosine A_2A receptor with -9.88 kcal/mol. In vitro evaluation revealed that riluzole increased cell growth inhibition by 24 % in LN229 cells and 36 % in SNB19 cells than istradefylline. Riluzole arrested the cell cycle at S phase in both cell lines, whereas istradefylline arrest was cell line specific. Three-dimensional (3D) spheroid model of 1321N1 GBM cells further demonstrated that riluzole inhibits ∼50 % higher cell growth inhibition than istradefylline with effective reduction in spheroid volume and size. Overall, our analysis revealed that blocking of adenosine A_2A receptor downstream signaling pathway protein CK1δ with its inhibitor, riluzole, showed higher anti-GBM effect than its upstream signaling blocker, istradefylline. Thus, blocking adenosine A_2A receptor downstream effector signaling protein through its antagonist and blocking its effector protein CK1δ could provide an opportunity to develop targeted therapy for GBM.

Keywords: 3D spheroids; Adenosine A2A receptor; Casein kinase 1δ kinase domain; Cell death; In silico.