Inhibition of Autophagy Enhances Anti-Angiogenic Efficacy in Intracranial Glioma

点击翻译按钮获取中文摘要

DESCRIPTION (provided by applicant): Malignant gliomas are highly aggressive, invasive, and vascular tumors for which clinical prognosis remains extremely poor. Gliomas are highly vascular, but current anti-angiogenic therapy (AAT) induces only a transitory clinical benefit. Evidence suggests that gliomas adapt to AAT, by activation of alternative angiogenic pathways and increased invasion. We hypothesize that AAT can be combined with molecular targeting (AAT+MT), to disrupt the adaptive mechanisms that enable tumors to survive AAT alone. A key target is autophagy, a degradative pathway that may enable tumor cell survival during hypoxic/nutrient stress. Autophagy prevents toxic protein and organelle accumulation and provides energy and biosynthetic substrates. Autophagic pathways are linked to pathways controlling cell death, and autophagic inhibition may induce cell death. We have developed a comprehensive dual bolus perfusion MRI method that enables us to visualize tumor perfusion and growth in vivo, in mice with intracranial gliomas. This method first utilizes dynamic contrast enhanced (DCE) MRI to provide maps of Ktrans, an index of vascular permeability. Then dynamic susceptibility contrast MRI (DSC-MRI) is done to measure cerebral blood flow (CBF). This dual bolus method suggests that AAT+MT therapy with the anti-angiogenic receptor tyrosine kinase inhibitor Cediranib (AstraZeneca) and the autophagy inhibitor quinacrine can have synergistic cytotoxic effects. Quinacrine is an anti-malarial agent which interrupts autophagosome function and has excellent blood brain barrier permeability. Preliminary results show anti-angiogenic/anti-tumor efficacy in vivo with resulting extended survival. Specific Aim 1 will characterize the dose-dependent in vitro responses of mouse (4C8) and human (U87) glioma cells to Cediranib and quinacrine, singly and in combination, under normal and oxygen glucose deprived conditions. These experiments will test hypotheses that a) hypoxic/nutrient stress plays a role in the synergism between Cediranib and quinacrine; b) induction of autophagy occurs with Cediranib under conditions of hypoxic/nutrient stress; and c) combined late-stage autophagic inhibition via quinacrine effectively induces autophagosome accumulation and tumor cell death. Specific Aim 2 will investigate the effects of single and combined Cediranib and quinacrine treatment in vivo, employing the intracranial mouse 4C8 and U87 malignant glioma models. These experiments test the hypothesis that combined administration of the two agents results in a synergistic anti-vascular/anti-tumor therapeutic efficacy. Dual bolus DCE/DSC MRI will be used to assess tumor growth and angiogenesis. Immunohistochemistry will be used ex vivo, to quantify tumor cell apoptosis, necrosis, autophagy, hypoxia and microvessel density. Both Specific Aims will employ immunoblot with cell and tissue lysates to quantify cellular markers and investigate underlying mechanisms. These preliminary studies are required to justify more detailed studies of the drug efficacy and the mechanisms of AAT+MT in treating malignant glioma, defined in a R01-type application.