title: Comparative Evaluation of Affibody Molecules for Radionuclide Imaging of in Vivo Expression of Carbonic Anhydrase IX.
abstract: Overexpression of the enzyme carbonic anhydrase IX (CAIX) is documented for chronically hypoxic malignant tumors as well as for normoxic renal cell carcinoma. Radionuclide molecular imaging of CAIX would be useful for detection of hypoxic areas in malignant tumors, for patients' stratification for CAIX-targeted therapies, and for discrimination of primary malignant and benign renal tumors. Earlier, we have reported feasibility of in vivo radionuclide based imaging of CAIX expressing tumors using Affibody molecules, small affinity proteins based on a nonimmunoglobulin scaffold. In this study, we compared imaging properties of several anti-CAIX Affibody molecules having identical scaffold parts and competing for the same epitope on CAIX, but having different binding paratopes. Four variants were labeled using residualizing99mTc and nonresidualizing125I labels. All radiolabeled variants demonstrated high-affinity detection of CAIX-expressing cell line SK-RC-52 in vitro and specific accumulation in SK-RC-52 xenografts in vivo.125I-labeled conjugates demonstrated much lower radioactivity uptake in kidneys but higher radioactivity concentration in blood compared with99mTc-labeled counterparts. Although all variants cleared rapidly from blood and nonspecific compartments, there was noticeable difference in their biodistribution. The best variant for imaging of expression of CAIX in disseminated cancer was99mTc-(HE)3-ZCAIX:2 providing tumor uptake of 16.3 ± 0.9% ID/g and tumor-to-blood ratio of 44 ± 7 at 4 h after injection. For primary renal cell carcinoma, the most promising imaging candidate was125I-ZCAIX:4 providing tumor-kidney ratio of 2.1 ± 0.5. In conclusion, several clones of scaffold proteins should be evaluated to select the best variant for development of an imaging probe with optimal sensitivity for the intended application.
authors.affiliation: Department of Immunology, Genetics and Pathology, Uppsala University , SE-75285 Uppsala, Sweden.
authors.forename: Javad
authors.initials: J
authors.lastname: Garousi
authors.affiliation: Department of Immunology, Genetics and Pathology, Uppsala University , SE-75285 Uppsala, Sweden.
authors.forename: Hadis
authors.initials: H
authors.lastname: Honarvar
authors.affiliation: Division of Protein Technology, School of Biotechnology, KTH-Royal Institute of Technology , SE-106 91 Stockholm, Sweden.
authors.forename: Ken G
authors.initials: KG
authors.lastname: Andersson
authors.affiliation: Division of Molecular Imaging, Department of Medicinal Chemistry, Uppsala University , SE-751 83 Uppsala, Sweden.
authors.forename: Bogdan
authors.initials: B
authors.lastname: Mitran
authors.affiliation: Division of Molecular Imaging, Department of Medicinal Chemistry, Uppsala University , SE-751 83 Uppsala, Sweden.
authors.forename: Anna
authors.initials: A
authors.lastname: Orlova
authors.affiliation: Department of Immunology, Genetics and Pathology, Uppsala University , SE-75285 Uppsala, Sweden.|Ridgeview Instruments AB , SE-74020 Vänge, Sweden.
authors.forename: Jos
authors.initials: J
authors.lastname: Buijs
authors.affiliation: Division of Protein Technology, School of Biotechnology, KTH-Royal Institute of Technology , SE-106 91 Stockholm, Sweden.
authors.forename: John
authors.initials: J
authors.lastname: Löfblom
authors.affiliation: Department of Immunology, Genetics and Pathology, Uppsala University , SE-75285 Uppsala, Sweden.|Affibody AB , SE-171 63 Stockholm, Sweden.
authors.forename: Fredrik Y
authors.initials: FY
authors.lastname: Frejd
authors.affiliation: Department of Immunology, Genetics and Pathology, Uppsala University , SE-75285 Uppsala, Sweden.
authors.forename: Vladimir
authors.initials: V
authors.lastname: Tolmachev
mesh_terms: D000818:Animals; D000071231:Carbonic Anhydrase IX / Q000378:metabolism*; D002292:Carcinoma, Renal Cell / Q000000981:diagnostic imaging* / Q000378:metabolism*; D045744:Cell Line, Tumor; D005260:Female; D064593:Heterografts; D006801:Humans; D007680:Kidney Neoplasms / Q000000981:diagnostic imaging* / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D019275:Radiopharmaceuticals / Q000032:analysis
chemical_list: D019275:Radiopharmaceuticals; D000071231:Carbonic Anhydrase IX
keywords: CAIX; affibody molecule; imaging; radionuclide