This article has been accepted for publication and undergone full peer review but has not been through the copyediting, typesetting, pagination and proofreading process which may lead to differences between this version and the Version of Record. Please cite this article as doi: 10.1002/ijc.32845 Research article Novel engineered TRAIL-based chimeric protein strongly inhibits tumor growth and bypasses TRAIL resistance Piotr Rozga 1* , Damian Kloska 2 , Sebastian Pawlak 1 , Malgorzata Teska-Kaminska 1 , Marlena Galazka 1 , Katarzyna Bukato 1 , Anna Pieczykolan 1 , Albert Jaworski 1 , Anna Molga- Kaczmarska 1 , Aleksandra Kopacz 2 , Bogna Badyra 2 , Neli Kachamakova-Trojanowska 2 , Olga Zolnierkiewicz 2 , Marta Targosz-Korecka 3 , Katarzyna Poleszak 1 , Michal Szymanik 1 , Bartlomiej Zerek 1 , Jerzy Pieczykolan 1# , Alicja Jozkowicz 2# , Anna Grochot-Przeczek 2# 1 Department of Drug Discovery, Adamed Pharma S.A., Pienkow, 05-152 Czosnow, Poland 2 Department of Medical Biotechnology, Faculty of Biochemistry Biophysics and Biotechnology, Jagiellonian University, 30-387 Krakow, Poland 3 Department of Physics of Nanostructures and Nanotechnology, Institute of Physics, Jagiellonian University, 30-387 Krakow, Poland * Corresponding author Piotr Rozga, Department of Drug Discovery, Adamed Pharma S.A., Pienkow, Mariana Adamkiewicza 6a, 05-152 Czosnow, Poland, Phone: +48227327876, Fax: +48227327860, E-mail: piotr.rozga@adamed.com # contributed equally as senior authors Running title: A novel TRAIL-based fusion protein efficiently kills tumors Key words: TRAIL, apoptosis, chimeric protein, anticancer therapy, antiangiogenic therapy Novelty and impact: DR agonists, applied as single agents, are therapeutically ineffective. We developed a new anticancer fusion protein, AD-O51.4, comprising VEGFA-derived positively charged peptides fused to a reengineered TRAIL. AD-O51.4 exerts both direct cytotoxic and antiangiogenic effects, bypasses resistance to TRAIL-induced cell death, exhibits very high anticancer activity, and causes no symptoms of drug toxicity in mice and monkeys. Our data demonstrate that AD-O51.4 is a promising candidate for clinical trials. This article is protected by copyright. All rights reserved.