Transcriptional CDK inhibitors, CYC065 and THZ1 promote Bim-dependent apoptosis in primary and recurrent GBM through cell cycle arrest and Mcl-1 downregulation

Viktorija Juric,Hanne Jahns,Martine L M Lamfers,Joanna Fay,Cathy E Richards,Maïté Verreault,Ahmed Idbaih,Ann M Hopkins,Lance Hudson,Franck Bielle,Markus Rehm,Brona M Murphy

doi:10.1038/s41419-021-04050-7

Abstract

Activation of cyclin-dependent kinases (CDKs) contributes to the uncontrolled proliferation of tumour cells. Genomic alterations that lead to the constitutive activation or overexpression of CDKs can support tumourigenesis including glioblastoma (GBM), the most common and aggressive primary brain tumour in adults. The incurability of GBM highlights the need to discover novel and more effective treatment options. Since CDKs 2, 7 and 9 were found to be overexpressed in GBM, we tested the therapeutic efficacy of two CDK inhibitors (CKIs) (CYC065 and THZ1) in a heterogeneous panel of GBM patient-derived cell lines (PDCLs) cultured as gliomaspheres, as preclinically relevant models. CYC065 and THZ1 treatments suppressed invasion and induced viability loss in the majority of gliomaspheres, irrespective of the mutational background of the GBM cases, but spared primary cortical neurons. Viability loss arose from G2/M cell cycle arrest following treatment and subsequent induction of apoptotic cell death. Treatment efficacies and treatment durations required to induce cell death were associated with proliferation velocities, and apoptosis induction correlated with complete abolishment of Mcl-1 expression, a cell cycle-regulated antiapoptotic Bcl-2 family member. GBM models generally appeared highly dependent on Mcl-1 expression for cell survival, as demonstrated by pharmacological Mcl-1 inhibition or depletion of Mcl-1 expression. Further analyses identified CKI-induced Mcl-1 loss as a prerequisite to establish conditions at which the BH3-only protein Bim can efficiently induce apoptosis, with cellular Bim amounts strongly correlating with treatment efficacy. CKIs reduced proliferation and promoted apoptosis also in chick embryo xenograft models of primary and recurrent GBM. Collectively, these studies highlight the potential of these novel CKIs to suppress growth and induce cell death of patient-derived GBM cultures in vitro and in vivo, warranting further clinical investigation.

Highlights

Glioblastoma (GBM) is the most common primary brain tumour in adults
Since it was previously shown that degradation of Mcl-1 during conditions of mitotic arrest paves the way for Bim-induced activation of mitochondrial apoptosis [40], we studied the expression of this proapoptotic Bcl-2 family member
We provide an in-depth analysis of the effectiveness of the transcriptional Cyclin-dependent kinases (CDKs) inhibitors, CYC065 and THZ1, targeting CDK2/ 9 or 7 respectively, to treat primary and recurrent GBM

Summary

INTRODUCTION

Glioblastoma (GBM) is the most common primary brain tumour in adults. Despite efforts to combat this disease with an aggressive standard of care (SOC) protocol, including surgery, radiotherapy and temozolomide (TMZ) chemotherapy [1, 2] patient survival remains low, with less than 5% of those diagnosed surviving longer than 5 years [3]. Its use in an orthotopic patient-derived xenograft (PDX) model of GBM showed moderate efficacy, the dosing regimen required to maintain peak levels of Roscovitine in the brain resulted in toxic side-effects which would preclude its clinical utilisation [15]. Patient-derived GBM cultures were plated in 96well plates as gliomaspheres (3000 cells/well) and treated with indicated concentrations of CYC065, THZ1 and/or TMZ Treatment efficacies were further confirmed in a semi in vivo chick embryo xenograft models These data demonstrate that CYC065 and THZ1 display high anti-cancer activity in primary and recurrent GBM and provide a scientific rationale for the further development of CDK inhibitors for potential clinical utilisation in the future.

MATERIALS AND METHODS

RESULTS

Juric et al 5

Findings

DISCUSSION