title: Feasibility of Affibody Molecule-Based PNA-Mediated Radionuclide Pretargeting of Malignant Tumors.
abstract: Affibody molecules are small (7 kDa), non-immunoglobulin scaffold proteins with a potential as targeting agents for radionuclide imaging of cancer. However, high renal re-absorption of Affibody molecules prevents their use for radionuclide therapy with residualizing radiometals. We hypothesized that the use of Affibody-based peptide nucleic acid (PNA)-mediated pretargeting would enable higher accumulation of radiometals in tumors than in kidneys. To test this hypothesis, we designed an Affibody-PNA chimera ZHER2:342-SR-HP1 containing a 15-mer HP1 PNA recognition tag and a complementary HP2 hybridization probe permitting labeling with both (125)I and (111)In. (111)In-ZHER2:342-SR-HP1 bound specifically to HER2-expressing BT474 and SKOV-3 cancer cells in vitro, with a KD of 6±2 pM for binding to SKOV-3 cells. Specific high affinity binding of the radiolabeled complementary PNA probe (111)In-/(125)I-HP2 to ZHER2:342-SR-HP1 pre-treated cells was demonstrated. (111)In-ZHER2:342-SR-HP1 demonstrated specific accumulation in SKOV-3 xenografts in BALB/C nu/nu mice and rapid clearance from blood. Pre-saturation of SKOV-3 with non-labeled anti-HER2 Affibody or the use of HER2-negative Ramos xenografts resulted in significantly lower tumor uptake of (111)In-ZHER2:342-SR-HP1. The complementary PNA probe (111)In/(125)I-HP2 accumulated in SKOV-3 xenografts when ZHER2:342-SR-HP1 was injected 4 h earlier. The tumor accumulation of (111)In/(125)I-HP2 was negligible without ZHER2:342-SR-HP1 pre-injection. The uptake of (111)In-HP2 in SKOV-3 xenografts was 19±2 %ID/g at 1 h after injection. The uptake in blood and kidneys was approximately 50- and 2-fold lower, respectively. In conclusion, we have shown that the use of Affibody-based PNA-mediated pretargeting enables specific delivery of radiometals to tumors and provides higher radiometal concentration in tumors than in kidneys.
authors.affiliation: 1. Department of Immunology, Genetics and Pathology, Uppsala University, Uppsala, Sweden.
authors.forename: Hadis
authors.initials: H
authors.lastname: Honarvar
authors.affiliation: 2. School of Biotechnology, Division of Protein Technology, KTH Royal Institute of Technology, Stockholm, Sweden.
authors.forename: Kristina
authors.initials: K
authors.lastname: Westerlund
authors.affiliation: 1. Department of Immunology, Genetics and Pathology, Uppsala University, Uppsala, Sweden.
authors.forename: Mohamed
authors.initials: M
authors.lastname: Altai
authors.affiliation: 3. Section of Nuclear Medicine and PET, Department of Surgical Sciences, Uppsala University, Uppsala, Sweden.
authors.forename: Mattias
authors.initials: M
authors.lastname: Sandström
authors.affiliation: 4. Preclinical PET Platform, Department of Medicinal Chemistry, Uppsala University, Uppsala, Sweden.
authors.forename: Anna
authors.initials: A
authors.lastname: Orlova
authors.affiliation: 1. Department of Immunology, Genetics and Pathology, Uppsala University, Uppsala, Sweden.
authors.forename: Vladimir
authors.initials: V
authors.lastname: Tolmachev
authors.affiliation: 2. School of Biotechnology, Division of Protein Technology, KTH Royal Institute of Technology, Stockholm, Sweden.
authors.forename: Amelie Eriksson
authors.initials: AE
authors.lastname: Karlström
mesh_terms: D000818:Animals; D004195:Disease Models, Animal; D064593:Heterografts; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D009369:Neoplasms / Q000175:diagnosis* / Q000628:therapy*; D020135:Peptide Nucleic Acids / Q000493:pharmacokinetics*; D011859:Radiography / Q000379:methods*; D011868:Radioisotopes / Q000493:pharmacokinetics*; D011878:Radiotherapy / Q000379:methods*; D013194:Staining and Labeling / Q000379:methods*
chemical_list: D020135:Peptide Nucleic Acids; D011868:Radioisotopes
keywords: Affibody; HER2; peptide nucleic acid; radionuclide pretargeting; scaffold protein