Anthracycline-induced cytotoxicity in the GL261 glioma model system

Abstract

Glioblastoma (GBM) is a lethal astrocyte-derived tumor that is currently treated with a multi-modal approach of surgical resection, radiotherapy, and temozolomide-based chemotherapy. Alternatives to current therapies are urgently needed as its prognosis remains poor. Anthracyclines are a class of compounds that show great potential as GBM chemotherapeutic agents and are widely used to treat solid tumors outside the central nervous system. Here we investigate the cytotoxic effects of doxorubicin and other anthracyclines on GL261 glioma tumor cells in anticipation of novel anthracycline-based CNS therapies. Three methods were used to quantify dose-dependent effects of anthracyclines on adherent GL261 tumor cells, a murine cell-based model of GBM. MTT assays quantified anthracycline effects on cell viability, comet assays examined doxorubicin genotoxicity, and flow cytometry with Annexin V/PI staining characterized doxorubicin-induced apoptosis and necrosis. Dose-dependent reductions in GL261 cell viability were found in cells treated with doxorubicin (EC50 = 4.9 μM), epirubicin (EC50 = 5.9 μM), and idarubicin (EC50 = 4.4 μM). Comet assays showed DNA damage following doxorubicin treatments, peaking at concentrations of 1.0 μM and declining after 25 μM. Lastly, flow cytometric analysis of doxorubicin-treated cells showed dose-dependent induction of apoptosis (EC50 = 5.2 μM). Together, these results characterized the cytotoxic effects of anthracyclines on GL261 glioma cells. We found dose-dependent apoptotic induction; however at high concentrations we find that cell death is likely necrotic. Our results support the continued exploration of anthracyclines as compounds with significant potential for improved GBM treatments.

Successful Partnerships: Exploring the Potential of Immunogenic Signals Triggered by TMZ, CX-4945, and Combined Treatment in GL261 Glioblastoma Cells

“…This can be seen with GL261 cells in Figure 3A (white brackets and arrows). At the assayed Doxo concentrations, most (75-90%) of GL261 cells evaluated by us should be dead/dying, and concentrations were far above the described EC50 [44].…”

“…This exposure has been analyzed by confocal microscopy in GL261 cells, both untreated and treated with TMZ (2 mM), CX-4945 (25 and 100 µM), and CX-4945+TMZ (all combinations) after 24 h of treatment. Cells treated with doxorubicin (Doxo) were used as a positive control since they have been described to trigger CRT exposure [43,44]. This can be seen with GL261 cells in Figure 3A (white brackets and arrows).…”

Musashi-1 Regulates MIF1-Mediated M2 Macrophage Polarization in Promoting Glioblastoma Progression

“…To summarize, we confirm that MSI1 overexpression leads to MIF1 upregulation in vivo and that such interplay between these two factors may result in enhanced tumor growth in an in vivo mouse model. MSI1-mediated MIF1 enrichment were further substantiated by utilizing GL261-Luc, another frequently used syngeneic murine GBM model that expresses luciferase to facilitate visualization of tumor growth and responses to treatments via bioluminescence imaging [38]. At first, we used the CRISPR/Cas9 technology to knock out the MSI1 gene in the GL261-Luc cell line (Figure 3C).…”

Redox Flow Iontophoresis for Continuous Drug Delivery

“…Doxorubicin for instance is, among others, routinely used for breast, ovarian, lung, and thyroid cancer therapy but has also been shown to be effective against glioma cell lines in vitro. [30][31][32] However, due to the limited permeability of the blood-brain barrier to doxorubicin, the drug is currently not commonly used for brain cancer therapy. [33] Other promising drug candidates are for instance cisplatin for cancer therapy, the opioid peptide DAMGO to complement opioid-based analgesia, [34] 𝛾-aminobutyric acid for epileptic seizure control, [17] or macromolecules such as monoclonal antibodies.…”