Tuning Cellular Rhythms: A New Hope for Glioblastoma Treatment

In vertebrates, including humans, the circadian system controls a number of functions, including food cycles, sleep/wake cycles, and rhythms of activity and rest. An elevated risk of cancer, inflammatory conditions, and metabolic diseases can all be brought on by disruption of this system. The very aggressive grade 4 brain tumor known as glioblastoma (GBM) has a terrible prognosis, with a median life of only 12 to 15 months, and it is resistant to standard therapy. In culture, GBM cells continue to have a working circadian rhythm. Researchers assessed the impact of pharmacological circadian clock modification and molecular clock disruption on GBM cells in order to investigate more potent therapies with fewer adverse effects.

Tests were conducted on GBM-derived cells using specific inhibitors that target the cytosolic kinases glycogen synthase kinase-3 (GSK-3) and casein kinase 1 ε/δ (CK1ε/δ), along with a cryptochrome protein stabilizer and Per2 knockdown. GSK-3 inhibitor CHIR99021 dramatically impacted cell survival, migration, distribution of the cell cycle, and clock protein expression. In comparison to control cells, these cells also displayed elevated reactive oxygen species and changed lipid droplet content. Cell viability was enhanced by the combination of CHIR99021 and temozolomide more so than by temozolomide alone. Per2 disruption also had an influence on GBM migration and cell cycle progression, indicating that GBM cell biology is significantly impacted by pharmacological manipulation or disruption of the molecular clock.

The circadian system uses transcriptional-translational feedback loops (TTFLs) and secondary loops with nuclear receptors to control circadian rhythms in gene expression, which occur roughly every 24 hours. The serine/threonine protein kinase GSK-3 is involved in cell motility, death, and proliferation. In a number of studies, pharmacological inhibitors of GSK-3 and CK1δ/ε have demonstrated potential for controlling cellular clocks. Greater GSK-3β activity has been seen in GBM, and treating this kinase may have therapeutic benefits.

Researchers postulated that genetic disruption of clock genes and pharmacological regulation of the circadian mechanism could have a major impact on the biology of GBM. The effects of GSK-3 inhibitor CHIR99021, cryptochrome protein stabilizer KL001, CK1ε/δ inhibitor PF670462, and Per2 disruption were examined in GBM T98G cells. The findings showed that CHIR99021, with IC50 values of 8.6 μM for T98G cells, significantly affected cell viability. In migratory experiments, this inhibitor also decreased vimentin expression, a protein associated with tumor cell movement, and wound closure. Owing to these noteworthy consequences, CHIR99021 was chosen for additional studies to examine its influence on the biology of GBM.

Effect of the pharmacological modulation and Per2 KD on GBM cell proliferation and migration.

Glioblastoma (GBM) is an extremely vascularized and invasive brain tumor that has a dismal prognosis; less than 5% of patients survive for five years after diagnosis, and the median survival time is 12 to 15 months. A potential treatment approach for high-grade gliomas is to target the circadian clock. Using the CK1ε/δ inhibitor (PF670462), GSK-3 inhibitor (CHIR99021), and CRY protein stabilizer (KL001), we examined the effects of pharmaceutical circadian clock modification on GBM cell viability, proliferation, and migration.

It was also examined how GSK-3 inhibition affected metabolic metrics, clock protein expression, and cell cycle distribution. The cell cycle distribution was significantly altered by CHIR99021 treatment, showing a larger proportion of dead cells and an increase in S-phase cells. Increased quantities of nuclear REV-ERB and BMAL1 proteins were detected by Western blot and immunocytochemistry, indicating disturbed clock protein expression. Furthermore, CHIR99021-treated cells showed altered lipid droplet content and elevated reactive oxygen species (ROS) levels, suggesting metabolic reprogramming.

The results of cell viability were better with combination treatment of CHIR99021 and temozolomide (TMZ) than with TMZ alone. This suggests that regular chemotherapy could be combined with circadian clock modification. The impact of Per2 knockdown on circadian disruption on GBM cell migration and cell cycle progression provides more evidence for the circadian clock's biological significance in GBM.

Overall, our results indicate that GBM cell survival, migration, and metabolism are strongly impacted by targeting the circadian clock through pharmacological modulation or genetic disruption. These findings highlight the possibility of boosting treatment efficacy while reducing negative effects by combining circadian clock inhibitors with already available treatments. To clarify the underlying mechanisms and improve therapy approaches, more research is required.

REFERENCE :

Wagner, P.M., Fornasier, S.J. & Guido, M.E. Pharmacological Modulation of the Cytosolic Oscillator Affects Glioblastoma Cell Biology. Cell Mol Neurobiol 44, 51 (2024). https://doi.org/10.1007/s10571-024-01485-2 

IMAGE REFERENCE :

https://link.springer.com/article/10.1007/s10571-024-01485-2#citeas

https://www.moffitt.org/endeavor/archive/what-is-glioblastoma-and-why-is-it-so-deadly/