Novel radiolabeled affibodies for targeted imaging and therapy

Novel radiolabeled affibodies for targeted imaging and therapy 2019-2022
Acronym: Bilateral agreement no.3698/13.09.2018 Romanian Academy- Hungarian Academy of Sciences
Project director: Stefan Szedlacsek
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The project is agreed as a bilateral collaboration between IBAR and ATOMKI, and the University of Debrecen and IFIN-HH participate in this project voluntarily.

The project is agreed as a bilateral collaboration between IBAR and ATOMKI, and the University of Debrecen and IFIN-HH participate in this project voluntarily. There are two main directions intended by the proposed project: receptor mapping and therapy, using an affibody against the HER2 receptor, combined with a suitable radioisotope. In this sense, the specific objectives are: 

  1. expression and purification of affibody; 
  2. establishing labeling procedures;
  3. ex vivo and/or in vivo testing of optimal compounds;
Stefan Szedlacsek, Dr.
Stefan Szedlacsek, Dr.

Head of Department

Prof. Dr. Stefan Szedlacsek is the Head of the Enzymology Department at the Institute of Biochemistry of the Romanian Academy. He holds a PhD degree in Biotechnology from Polytechnical University of Bucharest as well as a MSc in Organic Synthesis (Polytechnical University, Bucharest) and MSc in Mathematics (University of Bucharest). As a visiting scientist, he performed research in the field of cholesterol metabolism at the University of Illinois at Urbana-Champaign (USA), where he succeeded to evidence a new pathway in the metabolism of oxysterols.  He is an “Alexander von Humboldt” fellow and worked in Germany, in the Institute of Biochem More...

Adina-Gabriela Puiu, PhD Student
Adina-Gabriela Puiu, PhD Student

Research Assistant

Adina has a Bachelor's degree in Biology and a Master's degree in Applied Genetics and Biotechnology from the Faculty of Biology, University of Bucharest. She is a PhD student and Research Assistant at the Institute of Bio More...

Andrei-Mihai Vasilescu, PhD Student
Andrei-Mihai Vasilescu, PhD Student

Research Assistant

Andrei-Mihai Vasilescu is a researcher in the Institute of Biochemistry of the Romanian Academy. Andrei-Mihai is currently working in Enzymology in the Enzymology.

Horea Szedlacsek
Horea Szedlacsek

Research Assistant

Horea Szedlacsek is a researcher in the Institute of Biochemistry of the Romanian Academy. Horea is currently working in Enzymology in the Enzymology.

Rodica-Aura Badea, Dr.
Rodica-Aura Badea, Dr.

Scientific Researcher

Rodica-Aura Badea is a researcher in the Institute of Biochemistry of the Romanian Academy. Rodica-Aura is currently working in Cell Signaling Research in the Enzymology.

Simona Călăraș
Simona Călăraș

Research Assistant

The Romanian team The Hungarian team

Dr.Stefan Szedlacsek, Romanian Head of Project

Dr.Horea Szedlacsek

Dr.Rodica-Aura Badea

Asc.Simona Calaras

PhD Student Adina-Gabriela Puiu

PhD Student Andrei Mihai Vasilescu

Dr. Szűcs Zoltán , Institute for Nuclear Research (Atomki) Debrecen

Dr. Gyula Tircsó, University of Debrecen

PhD Student, Károly Brezovcsik, University of Debrecen

The project comprises three different stages: 

I.Preparatory experiments with the main objective being to obtain a stable chemical compound, in turn being subdivided in:

I.1. Smale-scale expression, purification and optimization of the refolding of the anti-HER2 affibody (IBAR). 

I.2. Optimized methodology for radioisotope production (ATOMKI & IFIN-HH). 

I.3. Comparison among various radiolabeling scenarious using the proposed radioisotopes with chelator agents like: new derivates of DOTA, NODAGA, etc to fit the N-terminal cysteine link to the anti-HER2 affibody (IBAR &ATOMKI, UD) 

II.In vivo testing with the objective of testing the behavior of the most promising radiolabeled affibody on cell lines (murine and human kind) This stage subdivides in:

II.1 Large-scale expression, purification and labeling of of anti-HER2 affibody (IBAR)

II.2. Optimizitation of the radiopharmaceutical compounds (IBAR,ATOMKI & IFIN-HH) II.3In vitro radiopharmaceutical testing (IBAR, ATOMKI&IFIN-HH)

III.In vitro testing with the main objective: biodistribution analysis and image quality enhancement using the most promising radiolabeled affibody against HER2 compound (IBAR, ATOMKI &IFN-HH).

The 3,9-PC2ABnpCO2H platform was selected for bioconjugation and for further in vivo studies.

Anti-HER2 affibody was used as a highly specific binding protein, which was expressed in a prokaryotic system, purified by affinity chromatography followed by RP-HPLC. 

The specificity of the affibody for HER2 receptors was proved following its conjugation with AlexaFluor555 dye. Finally the affibody was conjugated with the selected 3,9-PC2ABnpCO2H BFC and [52Mn]Mn(II) labelling was performed.

The labelled conjugate was tested in a pilot study by PET imaging using MDA-MB (HER2+) and 4T1 (HER2−) tumour-bearing mice. 

The results demonstrate that the [[52Mn]Mn(3,9-PC2ABnpMA)(H2O)]-Cys-HER2-affibody conjugate is an efficient and specific molecular vehicle for targeting and visualizing HER2+ tumours by 52Mn-based PET imaging.

Published papers: Synthesis and characterization of a novel [52Mn]Mn-labelled affibody based radiotracer for HER2+ targeting” Váradi, B., Brezovcsik, K., Garda, Z., Madarasi, E., Szedlacsek, H., Badea, R.-A., Vasilescu, A.-M., Puiu, A.-G., Ionescu, A., Sima, L.-E., Munteanu, C., V.-A., Călăraș, S., Vagner, A., Szikra, D., Toàn, N., M., Nagy, T., Szűcs, Z., Szedlacsek, S., E., Nagy, G., Tircsó, Inorganic Chemistry Frontiers, Accepted: 29 May 2023; First published: 06 Jun 2023; DOI https://doi.org/10.1039/D3QI00356F;

Meeting Abstract: Labelling anti-HER2-affibodies with Mn-52 via pyclen based bifunctional ligands: from ligand design to in vivo PET/MR experiments”; Varadi, B, ; Garda, Z; Madarasi, E; Brezovcsik, K; Vagner, A; Nagy, T; Garai, I ; Ngo, TM; Puiu, AG ; Vasilescu, AM; Szucs, Z ; Szedlacsek, SE; Nagy, G; Tircso, G at Conference Meeting 35th Annual Meeting of the European-Association-of-Nuclear-Medicine (EANM); Barcelona, Spain; Oct. 15-19, 2022; https://www.webofscience.com/wos/woscc/full-record/WOS:000857046600592

Invited Lecturer in the Fulbright Visiting Scholar Program for the Chemistry Seminar Series at Clarkson University, NY (USA); oral presentations: "Molecular Vehicles for targeted diagnostics and Therapy of Cancer”; 2023 January 20-21;