Cyclin-dependent kinase inhibitors exert distinct effects on patient-derived 2D and 3D glioblastoma cell culture models

Abstract

Current therapeutic approaches have met limited clinical success for glioblastoma multiforme (GBM). Since GBM harbors genomic alterations in cyclin-dependent kinases (CDKs), targeting these structures with specific inhibitors (CDKis) is promising. Here, we describe the antitumoral potential of selective CDKi on low-passage GBM 2D- and 3D models, cultured as neurospheres (NSCs) or glioma stem-like cells (GSCs). By applying selective CDK4/6i abemaciclib and palbociclib, and the more global CDK1/2/5/9-i dinaciclib, different effects were seen. Abemaciclib and dinaciclib significantly affected viability in 2D- and 3D models with clearly visible changes in morphology. Palbociclib had weaker and cell line-specific effects. Motility and invasion were highly affected. Abemaciclib and dinaciclib additionally induced senescence. Also, mitochondrial dysfunction and generation of mitochondrial reactive oxygen species (ROS) were seen. While autophagy was predominantly visible after abemaciclib treatment, dinaciclib evoked γ-H2AX-positive double-strand breaks that were boosted by radiation. Notably, dual administration of dinaciclib and abemaciclib yielded synergistic effects in most cases, but the simultaneous combination with standard chemotherapeutic agent temozolomide (TMZ) was antagonistic. RNA-based microarray analysis showed that gene expression was significantly altered by dinaciclib: genes involved in cell-cycle regulation (different CDKs and their cyclins, SMC3), mitosis (PLK1, TTK), transcription regulation (IRX3, MEN1), cell migration/division (BCAR1), and E3 ubiquitination ligases (RBBP6, FBXO32) were downregulated, whereas upregulation was seen in genes mediating chemotaxis (CXCL8, IL6, CCL2), and DNA-damage or stress (EGR1, ARC, GADD45A/B). In a long-term experiment, resistance development was seen in 1/5 cases treated with dinaciclib, but this could be prevented by abemaciclib. Vice versa, adding TMZ abrogated therapeutic effects of dinaciclib and growth was comparable to controls. With this comprehensive analysis, we confirm the therapeutic activity of selective CDKi in GBM. In addition to the careful selection of individual drugs, the timing of each combination partner needs to be considered to prevent resistance.

Implementation of a combined CDK inhibition and arginine-deprivation approach to target arginine-auxotrophic glioblastoma multiforme cells

“…Interference of CDKi/SpyADI with mitochondria (increased mitochondrial reactive oxygen species (ROS) production and mitochondrial membrane potential) and lysosomal formation were studied based on our previous observations under CDKi monotherapy [ 16 ]. Mitochondrial activity increased cell line-specific after SEQ-dinaciclib/SpyADI or SEQ-abemaciclib/SpyADI treatment (Fig.…”

“…The hallmark of genomic instability in GBM is related to perturbations in the S-phase and G2/M transition, with the cyclin D1-CDK4/6-Rb pathway being mostly altered [ 15 ]. Just recently, we described the successful elimination of patient-derived two-dimensional (2D)- and three-dimensional (3D)-cultured GBM cells by the global CDKi dinaciclib as well as the selective CDK4/6 inhibitor abemaciclib [ 16 , 17 ]. The aforementioned CDKi even interfered with the tryptophan catabolism and had complex effects at both the morphological and molecular levels [ 17 ].…”

The Individual Effects of Cyclin-Dependent Kinase Inhibitors on Head and Neck Cancer Cells—A Systematic Analysis

“…This has profound biological effects. In a very recent study on patient-derived glioblastoma models, we described the CDKi-induced loss of mitochondrial function in pioneering work, characterized by a multivacuolar phenotype and signs of early-methuosis [ 60 ]. Methuosis, a non-apoptotic cell death phenotype, is defined by the accumulation of large fluid-filled cytoplasmic vacuoles that originate from macropinosomes [ 61 ].…”

DDRugging glioblastoma: understanding and targeting the DNA damage response to improve future therapies

“…On the topic on non-DDR signalling kinases that impact the DDR, Riess and colleagues recently assessed targeting of the CDK family of signalling kinases given their common dysregulation in glioblastoma and the recent advancement of a new generation of clinically approved compounds [195]. Using a CDK-based monotherapy approach in various 3D glioblastoma preclinical models, they showed that CDK inhibitors could negatively affect tumour growth, but also that some CDK inhibitors were able to effectively combine with DNA damaging regimes such as radiation and temozolomide treatments.…”

“…Using a CDK‐based monotherapy approach in various 3D glioblastoma preclinical models, they showed that CDK inhibitors could negatively affect tumour growth, but also that some CDK inhibitors were able to effectively combine with DNA damaging regimes such as radiation and temozolomide treatments. However, they also showed that not all tested CDK inhibitors behaved in the same way, with some conferring antagonistic properties when combined with temozolomide, potentially through differential effects on global gene expression patterns [ 194 ]. This study further highlights the need to understand the intricate functional interplay and regulatory mechanisms within the DDR as part of preclinical studies to help maximise the therapeutic potential of such new combinatorial regimes within the clinic.…”