ABSTRACT
BACKGROUND: Relatively poor penetration and retention in tumor tissue has been documented for large molecule drugs including therapeutic antibodies and recombinant immunoglobulin constant region (Fc)-fusion proteins due to their large size, positive charge, and strong target binding affinity. Therefore, when designing a large molecular drug candidate, smaller size, neutral charge, and optimal affinity should be considered. METHODS: We engineered a recombinant protein by molecular engineering the second domain of VEGFR1 and a few flanking residues fused with the Fc fragment of human IgG1, which we named HB-002.1. This recombinant protein was extensively characterized both in vitro and in vivo for its target-binding and target-blocking activities, pharmacokinetic profile, angiogenesis inhibition activity, and anti-tumor therapeutic efficacy. RESULTS: HB-002.1 has a molecular weight of ~80 kDa, isoelectric point of ~6.7, and an optimal target binding affinity of <1 nM. The pharmacokinetic profile was excellent with a half-life of 5 days, maximal concentration of 20.27 µg/ml, and area under the curve of 81.46 µg·days/ml. When tested in a transgenic zebrafish embryonic angiogenesis model, dramatic inhibition in angiogenesis was exhibited by a markedly reduced number of subintestinal vessels. When tested for anti-tumor efficacy, HB-002.1 was confirmed in two xenograft tumor models (A549 and Colo-205) to have a robust tumor killing activity, showing a percentage of inhibition over 90% at the dose of 20 mg/kg. Most promisingly, HB-002.1 showed a superior therapeutic efficacy compared to bevacizumab in the A549 xenograft model (tumor inhibition: 84.7% for HB-002.1 versus 67.6% for bevacizumab, P<0.0001). CONCLUSIONS: HB-002.1 is a strong angiogenesis inhibitor that has the potential to be a novel promising drug for angiogenesis-related diseases such as tumor neoplasms and age-related macular degeneration.
Subject(s)
Adenocarcinoma/drug therapy , Angiogenesis Inhibitors/pharmacology , Colorectal Neoplasms/drug therapy , Immunoglobulin Fc Fragments/pharmacology , Lung Neoplasms/drug therapy , Neovascularization, Pathologic/drug therapy , Neovascularization, Physiologic/drug effects , Recombinant Fusion Proteins/pharmacology , Vascular Endothelial Growth Factor A/antagonists & inhibitors , Vascular Endothelial Growth Factor Receptor-1/pharmacology , Adenocarcinoma/pathology , Animals , Cell Line, Tumor , Cell Proliferation/drug effects , Colorectal Neoplasms/pathology , Drug Design , Female , Human Umbilical Vein Endothelial Cells/physiology , Immunoglobulin Fc Fragments/chemistry , Immunoglobulin Fc Fragments/therapeutic use , Immunoglobulin G/chemistry , Lung Neoplasms/pathology , Mice , Mice, Inbred BALB C , Recombinant Fusion Proteins/pharmacokinetics , Recombinant Fusion Proteins/therapeutic use , Vascular Endothelial Growth Factor A/chemistry , Vascular Endothelial Growth Factor A/pharmacology , Vascular Endothelial Growth Factor Receptor-1/chemistry , Vascular Endothelial Growth Factor Receptor-1/pharmacokinetics , Vascular Endothelial Growth Factor Receptor-1/therapeutic use , Zebrafish/embryologyABSTRACT
PURPOSE: The mTOR kinase inhibitor AZD8055 inhibits both mTORC1 and mTORC2 leading to disruption of glucose metabolism and proliferation pathways. This study assessed the impact of single and multiple doses of AZD8055 on the uptake of the glucose metabolism marker 2-deoxy-2-[(18) F]fluoro-D-glucose ([(18) F]FDG) and the proliferation marker 3'-deoxy-3'-[(18) F]fluorothymidine ([(18) F]FLT) in U87-MG glioma xenografts. PROCEDURES: Mice bearing U87-MG tumours received either vehicle or AZD8055 (20 mg/kg) once daily p.o. Mice were imaged with either [(18) F]FDG or [(18) F]FLT PET to assess treatment response. Comparisons were made between in vivo imaging and ex vivo histopathology data. RESULTS: Tumour uptake of [(18) F]FDG was reduced by 33 % 1 h after a single dose of AZD8055 and by 49 % following 4 days of dosing. These changes coincided with suppression of the mTOR pathway biomarkers pS6 and pAKT. In contrast, the effect of AZD8055 on [(18) F]FLT uptake was inconsistent. CONCLUSIONS: The very rapid change in [(18) F]FDG uptake following acute AZD8055 treatment suggests that this could be used as an early mechanistic biomarker of metabolic changes resulting from mTOR inhibition. The utility of [(18) F]FLT for measuring the anti-proliferative effect of AZD8055 remains unclear.