NGR (Asn-Gly-Arg)-targeted delivery of coagulase to tumor vasculature arrests cancer cell growth

Khaled Seidi,Davoud Farajzadeh,Amir Daei Farshchi Adli,Rana Jahanban-Esfahlan,Babak Minofar,Nosratollah Zarghami,Heidi A Neubauer,Richard Moriggl,Peyman Zare,Mehran Mesgari Abbasi,Hassan Monhemi,Tahereh Javaheri,Ramezan Behzadi

doi:10.1038/s41388-018-0213-4

Abstract

Induction of selective thrombosis and infarction in tumor-feeding vessels represents an attractive strategy to combat cancer. Here we took advantage of the unique coagulation properties of staphylocoagulase and genetically engineered it to generate a new fusion protein with novel anti-cancer properties. This novel bi-functional protein consists of truncated coagulase (tCoa) and an NGR (GNGRAHA) motif that recognizes CD13 and αvβ3 integrin receptors, targeting it to tumor endothelial cells. Herein, we report that tCoa coupled by its C-terminus to an NGR sequence retained its normal binding activity with prothrombin and avβ3 integrins, as confirmed in silico and in vitro. Moreover, in vivo biodistribution studies demonstrated selective accumulation of FITC-labeled tCoa-NGR fusion proteins at the site of subcutaneously implanted PC3 tumor xenografts in nude mice. Notably, systemic administration of tCoa-NGR to mice bearing 4T1 mouse mammary xenografts or PC3 human prostate tumors resulted in a significant reduction in tumor growth. These anti-tumor effects were accompanied by massive thrombotic occlusion of small and large tumor vessels, tumor infarction and tumor cell death. From these findings, we propose tCoa-NGR mediated tumor infarction as a novel and promising anti-cancer strategy targeting both CD13 and integrin αvβ3 positive tumor neovasculature.

Highlights

Tumor infarction mediated by targeted delivery of coagulating proteins represents an appealing and cost effective strategy to combat a broad range of cancers [1]
The truncated coagulase (tCoa)-NGRprothrombin complex derived from our molecular dynamics (MD) simulation closely resembles a previously reported complex of prothrombin and another staphylocoagulase variant [21], suggesting that our engineered fusion protein adopts a similar biologically-relevant conformation
Our results indicated macroscopic evidence of thrombosis and hemorrhaging at the site of the subcutaneously implanted PC3 tumor xenografts treated with tCoa-NGR fusion protein, while no thrombotic changes were detected in the tumor xenografts of mice that were treated with tCoa (Fig. 5a), suggesting that tCoa-NGR effectively mediated selective blood coagulation in the tumor vasculature

Summary

Introduction

Tumor infarction mediated by targeted delivery of coagulating proteins represents an appealing and cost effective strategy to combat a broad range of cancers [1]. Austria Institute of Animal Breeding and Genetics, University of Veterinary Medicine Vienna, 1210 Vienna, Austria Medical University of Vienna, 1090 Vienna, Austria Department of Clinical Biochemistry and Laboratory Medicine, Faculty of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran Iranian National Science Foundation, Tehran, Iran studies have described the use of a truncated form of the human coagulation-inducing protein tissue factor (tTF) conjugated to diverse tumor endothelial moieties such as NGR (Asn-Gly-Arg) peptides. We have further developed this tumorspecific neovascular targeting approach by using bi-specific NGR peptides that recognize both CD13 and αvβ integrin, in order to lower the dose escalation and achieve more physiologic targeting of tumor neovasculature to avoid potential negative side effects

Methods

Results

Conclusion