A new anti-glioma therapy, AG119: pre-clinical assessment in a mouse GL261 glioma model

Abstract

Background: High grade gliomas (HGGs; grades III and IV) are the most common primary brain tumors in adults, and their malignant nature ranks them fourth in incidence of cancer death. Standard treatment for glioblastomas (GBM), involving surgical resection followed by radiation and chemotherapy with temozolomide (TMZ) and the anti-angiogenic therapy bevacizumab, have not substantially improved overall survival. New therapeutic agents are desperately needed for this devastating disease. Here we study the potential therapeutic agent AG119 in a pre-clinical model for gliomas. AG119 possesses both anti-angiogenic (RTK inhibition) and antimicrotubule cytotoxic activity in a single molecule.

Methods: GL261 glioma-bearing mice were either treated with AG119, anti-VEGF (vascular endothelial growth factor) antibody, anti c-Met antibody or TMZ, and compared to untreated tumor-bearing mice. Animal survival was assessed, and tumor volumes and vascular alterations were monitored with morphological magnetic resonance imaging (MRI) and perfusion-weighted imaging, respectively.

Results: Percent survival of GL261 HGG-bearing mice treated with AG119 was significantly higher (p < 0.001) compared to untreated tumors. Tumor volumes (21-31 days following intracerebral implantation of GL261 cells) were found to be significantly lower for AG119 (p < 0.001), anti-VEGF (p < 0.05) and anti-c-Met (p < 0.001) antibody treatments, and TMZ-treated (p < 0.05) mice, compared to untreated controls. Perfusion data indicated that both AG119 and TMZ were able to reduce the effect of decreasing perfusion rates significantly (p < 0.05 for both), when compared to untreated tumors. It was also found that IC50 values for AG119 were much lower than those for TMZ in T98G and U251 cells.

Conclusions: These data support further exploration of the anticancer activity AG119 in HGG, as this compound was able to increase animal survival and decrease tumor volumes in a mouse GL261 glioma model, and that AG119 is also not subject to methyl guanine transferase (MGMT) mediated resistance, as is the case with TMZ, indicating that AG119 may be potentially useful in treating resistant gliomas.

Fig. 1

Survival data for treated and untreated GL261 glioma-bearing mice. AG119 (n = 7), TMZ (n = 5), anti-c-Met antibody (n = 5), and anti-VEGF antibody (n = 5) treated GL261 glioma-bearing mice, compared to untreated controls (UT) (n = 5). When comparing AG119 (***p < 0.001; p = 0.0003) or TMZ (**p < 0.01; p = 0.0016) to UT, there was a significant increase in survival

Fig. 2

a Tumor volumes (mm³) of GL261 glioma-bearing animals either untreated (UT) or treated with anti-VEGF (n = 5) or anti-c-Met (n = 5) antibody therapies, TMZ (n = 5), or AG119 (n = 7), as measured by MRI. There was a significant decrease in tumors treated with either AG119 (***p < 0.001; p = 0.00067), TMZ (*p < 0.05; p = 0.03392), anti-VEGF (*p < 0.05; p = 0.01540) or ant-c-Met (**p < 0.01; p = 0.00539) when compared to untreated mice. b-f T₂-weighted MR images of GL261 glioma-bearing mice. Tumors are outlined in red based on tumor boundary contrast with ‘normal’ brain tissue. b Representative untreated mouse (vehicle control) (21 days following GL261 cell implantation). c Anti-VEGF antibody treated mouse at 27 days after cell implantation. d Anti-c-Met antibody treated mouse at 28 days following cell implantation. ei-v TMZ-treated mice at 28–30 days following cell implantation. fi-v AG119-treated mice at 26–29 days following cell implantation. Mice were treated when tumor volumes were >10 mm³. Tumor volumes were measured by adding tumor areas in multiple 1 mm image slices. Each image is obtained from different mice in each treatment group

Fig. 3

a Measured decreases in perfusion rates (ml/100 g x min) in untreated, TMZ- or AG119-treated GL261 mouse gliomas. Perfusion rates were measured using an arterial spin label perfusion MRI method, and relative decreases in tumors were obtained compared to the contralateral side. There were significant increases in tumor perfusion rates in both AG119 and TMZ-treated tumors (*p < 0.05), compared to untreated tumors, which had substantially decreased perfusion rates. b-d Perfusion MR images of untreated (b), TMZ (c), and AG119-treated (d) GL261 glioma-bearing mice. T2-weighted morphological images are in the top panels (i), and perfusion maps are in the bottom panels (ii). Dark region in panel Bii depicts decreased perfusion rates in an untreated tumor, which is not as severe in treated tumors (panels cii and dii). Tumors are outlined in red

Fig. 4

Cytotoxic effect of AG119 on temozolomide (TMZ)-resistant cells. Cells, U251 (TMZ-sensitive; MGMT⁻) and T98G (TMZ-resistant; MGMT⁺), were treated with AG119 or TMZ for 48 h and viability determined with Presto Blue. Data are mean IC₅₀ values (μM) ± SEM, n = 4–9 independent experiments