In vivo phage display screening for tumor vascular targets in glioblastoma identifies a llama nanobody against dynactin-1-p150Glued

Autor: Jari P. Lappalainen, Erwin van Wijk, Ilse Roodink, Stef J.F. Letteboer, William P.J. Leenders, Seppo Ylä-Herttuala, Joost J.C. Verhoeff, Ronald Roepman, Kiek Verrijp, Petri I. Mäkinen, Jos M.H. Raats, Sanne A. M. van Lith
Přispěvatelé: Radiotherapy
Rok vydání: 2016
Předmět:
Zdroj: Oncotarget
Oncotarget, 7(44), 71594-71607. Impact Journals
Oncotarget, 7(44), 71594. Impact Journals
ISSN: 1949-2553
Popis: // Sanne A.M. van Lith 1, * , Ilse Roodink 1, 5, * , Joost J.C. Verhoeff 2 , Petri I. Makinen 3 , Jari P. Lappalainen 3 , Seppo Yla-Herttuala 3, 4 , Jos Raats 5 , Erwin van Wijk 6 , Ronald Roepman 7 , Stef J. Letteboer 7 , Kiek Verrijp 1 , William P.J. Leenders 1 1 Department of Pathology, RadboudUMC, 6500 HB, Nijmegen, The Netherlands 2 Department of Radiotherapy, Amsterdam Medical Center, 1100 DD, Amsterdam, The Netherlands 3 Department of Biotechnology and Molecular Medicine, University of Eastern Finland, FI-70211, Kuopio, Finland 4 Science Service Center and Gene Therapy Unit, Kuopio University Hospital, 70210 Kuopio, Finland 5 Modiquest BV, LSP, Molenstraat 110, 5342 CC, Oss, The Netherlands 6 Department of Otorhinolaryngology, RadboudUMC, 6500 HB, Nijmegen, The Netherlands 7 Department of Genetics, RadboudUMC, 6500 HB, Nijmegen,The Netherlands * These authors contributed equally to this work Correspondence to: William Leenders, email: william.leenders@radboudumc.nl Keywords: glioma, stroma, targeting, nanobody, macrophages Received: June 30, 2016 Accepted: September 19, 2016 Published: September 26, 2016 ABSTRACT Diffuse gliomas are primary brain cancers that are characterised by infiltrative growth. Whereas high-grade glioma characteristically presents with perinecrotic neovascularisation, large tumor areas thrive on pre-existent vasculature as well. Clinical studies have revealed that pharmacological inhibition of the angiogenic process does not improve survival of glioblastoma patients. Direct targeting of tumor vessels may however still be an interesting therapeutic approach as it allows pinching off the blood supply to tumor cells. Such tumor vessel targeting requires the identification of tumor-specific vascular targeting agents (TVTAs). Here we describe a novel TVTA, C-C7, which we identified via in vivo biopanning of a llama nanobody phage display library in an orthotopic mouse model of diffuse glioma. We show that C-C7 recognizes a subpopulation of tumor blood vessels in glioma xenografts and clinical glioma samples. Additionally, C-C7 recognizes macrophages and activated endothelial cells in atherosclerotic lesions. By using C-C7 as bait in yeast-2-hybrid (Y2H) screens we identified dynactin-1-p150Glued as its binding partner. The interaction was confirmed by co-immunostainings with C-C7 and a commercial anti-dynactin-1-p150Glued antibody, and via co-immunoprecipitation/western blot studies. Normal brain vessels do not express dynactin-1-p150 Glued and its expression is reduced under anti-VEGF therapy, suggesting that dynactin-1-p150 Glued is a marker for activated endothelial cells. In conclusion, we show that in vivo phage display combined with Y2H screenings provides a powerful approach to identify tumor-targeting nanobodies and their binding partners. Using this combination of methods we identify dynactin-1-p150 Glued as a novel targetable protein on activated endothelial cells and macrophages.
Databáze: OpenAIRE
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