Mechanistic interrogation of combination Bevacizumab/dual PI3K/mTOR inhibitor response in Glioblastoma implementing novel MR and PET imaging biomarkers.


Resistance to bevacizumab (BEV) in glioblastoma (GBM) is believed to occur via activation of molecular networks including the mTOR/PI3K pathway. Implementing an MRI/PET molecular imaging biomarker approach, we sought to interrogate response to combining BEV with the mTOR/PI3K inhibitor BEZ235.


Tumors were established by orthotopically implanting U87MG-luc2 in mice. Animals were treated with BEZ235 and/or BEV, and imaged using diffusion weighted-MRI, T2 weighted (T2w), and T2* weighted (T2*w) before and following delivery of superparamagnetic iron oxide (SPIO) contrast. Maps for changes in relaxation rates: ΔR2, ΔR2* and apparent diffusion coefficient (ADC) were calculated. Vessel Size Index (VSI) and micro vessel density index (MDI) were derived. 3´-deoxy-3´-[18F]fluorothymidine ([18F]FLT)- and O-(2-[18F]fluoroethyl)-L-tyrosine ([18F]FET) PET was further performed and tumor endothelium/proliferation markers assessed by immunohistochemistry.


Treatment with BEV resulted in a pronounced decrease in tumor volume (T2w MRI). No additive effect on tumour volume was observed in BEV/BEZ235 combination compared with BEV monotherapy. Ki67 proliferation index staining and [18F]FLT uptake studies were used to support observations. Using ΔR2* and ΔR2 values respectively, BEZ235 + BEV combination significantly reduced tumor microvessel volume in comparison to BEV alone. Decreased MDI was further observed in the combination group; supported by von Willebrand Factor (vWF) immunohistochemistry. We observed decreased [18F]FET uptake following BEV, but failed to observe further reduced [18F]FET uptake in the combination cohort. vWF IHC analysis showed mean tumor vessel size increased in all cohorts. Conclusions: Assessing MR imaging biomarker parameters together with [18F]FET and [18F]FLT PET, informed drug combination mechanism of action and provided clues as to potential clinical response. Translation of a BEZ35/BEV combination regimen could support reduction of peritumoral edemaobviating the requirement for steroids. Implementing hypothesis driven molecular imaging studies facilitates the interrogation of drug response in the pre-clinic. These data may more accurately predict the clinical potential of novel therapeutic approaches in oncology.