Inhibition of MEK confers hypersensitivity to X-radiation in the context of BRAF mutation in a model of childhood astrocytoma

Abstract

Purpose Curative therapy for childhood glioma presents challenges when complete resection is not possible. Patients with recurrent low-grade tumors or anaplastic astrocytoma may receive radiation treatment, however, the long-term sequellae from radiation treatment can be severe. As many childhood gliomas are associated with activation of BRAF, we have explored the combination of ionizing radiation with MEK inhibition in a model of BRAF-mutant anaplastic astrocytoma. Experimental Design The regulation of TORC1 signaling by BRAF was examined in BT-40 (BRAF mutant) and BT-35 (BRAF wild type) xenografts, in a cell line derived from the BT-40 xenograft and two adult BRAF mutant glioblastoma cell lines. The effect of MEK inhibition (selumetinib), XRT (total dose10 Gy as 2 Gy daily fractions), or the combination of selumetinib and XRT was evaluated in subcutaneous BT-40 xenografts. Results Inhibition of MEK signaling by selumetinib, suppressed TORC1 signaling only in the context of the BRAF-mutant both in vitro and in vivo. Inhibition of MEK signaling in BT-40 cells or in xenografts lead to a complete suppression of FANCD2 and conferred hypersensitivity to XRT in BT-40 xenografts without increasing local skin toxicity. Conclusions Selumetinib suppressed TORC1 signaling in the context of BRAF mutation. Selumetinib caused a rapid downregulation of FANCD2 and markedly potentiated the effect of XRT. These data suggest the possibility of potentiating the effect of XRT selectively in tumor cells by MEK inhibition in the context of mutant BRAF or maintaining tumor control at lower doses of XRT that would decrease long-term sequelae.

Overcoming resistance to single-agent therapy for oncogenic BRAF gene fusions via combinatorial targeting of MAPK and PI3K/mTOR signaling pathways

“…We also observed some suppression of phosphorylated S6 which is often used as a signaling readout for the PI3K/mTOR pathway. This correlates with previous findings in BT-40 cell line where selumetinib suppressed TORC1 signaling (mTOR component), albeit in BRAF-V600E mutation background [ 32 ]. In soft agar colony formation assay, we observed significant inhibition of colony formation with trametinib across all tested BRAF-fusions (Figure 2B , p < 0.05 to 0.0001) , although we required higher concentrations for suppression compared to KIAA1549-BRAF expressing cells.…”

Identification of MEK162 as a Radiosensitizer for the Treatment of Glioblastoma

“…This interaction appears specific for the MAPK axis as it was not observed for any of the tested inhibitors in the PI3K/mTOR pathway. Previous research in glioma and other cancers have identified similar radiosensitizing interactions in monolayer cell lines addicted to the MAPK pathway by activating mutations in either KRAS or BRAF (25)(26)(27). Here we report that even in the absence of these activating mutations; MEK162 is able to enhance the irradiation response of glioma cells, but only when cultured as spheroids, underlining the importance of MEK signaling in these more physiologically relevant culture conditions.…”

Evaluation of patritumab with or without erlotinib in combination with standard cytotoxic agents against pediatric sarcoma xenograft models

“…Tumor tissues and cell lines were snap frozen and immunoblotting was performed as previously described . Antibodies used were against ERBB3 (D22C5 Rabbit mAb), phospho‐ERBB3 (Y1289; D1B5 Rabbit mAb), HRG (#2573 Rabbit Ab), and GAPDH (D16H11 rabbit Mab; loading control), which were purchased from Cell Signaling Tecnology (Danvers, MA).…”