| 1. |
- Abouzayed, Ayman, et al.
(författare)
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The GRPR Antagonist [Tc-99m]Tc-maSSS-PEG(2)-RM26 towards Phase I Clinical Trial : Kit Preparation, Characterization and Toxicity
- 2023
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Ingår i: Diagnostics. - : MDPI AG. - 2075-4418. ; 13:9, s. 1611-
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Tidskriftsartikel (refereegranskat)abstract
- Gastrin-releasing peptide receptors (GRPRs) are overexpressed in the majority of primary prostate tumors and in prostatic lymph node and bone metastases. Several GRPR antagonists were developed for SPECT and PET imaging of prostate cancer. We previously reported a preclinical evaluation of the GRPR antagonist [Tc-99m]Tc-maSSS-PEG2-RM26 (based on [D-Phe(6), Sta(13), Leu(14)-NH2]BBN(6-14)) which bound to GRPR with high affinity and had a favorable biodistribution profile in tumor-bearing animal models. In this study, we aimed to prepare and test kits for prospective use in an early-phase clinical study. The kits were prepared to allow for a one-pot single-step radiolabeling with technetium-99m pertechnetate. The kit vials were tested for sterility and labeling efficacy. The radiolabeled by using the kit GRPR antagonist was evaluated in vitro for binding specificity to GRPR on PC-3 cells (GRPR-positive). In vivo, the toxicity of the kit constituents was evaluated in rats. The labeling efficacy of the kits stored at 4 degrees C was monitored for 18 months. The biological properties of [Tc-99m]Tc-maSSS-PEG2-RM26, which were obtained after this period, were examined both in vitro and in vivo. The one-pot (gluconic acid, ethylenediaminetetraacetic acid, stannous chloride, and maSSS-PEG(2)-RM26) single-step radiolabeling with technetium-99m was successful with high radiochemical yields (>97%) and high molar activities (16-24 MBq/nmol). The radiolabeled peptide maintained its binding properties to GRPR. The kit constituents were sterile and non-toxic when tested in living subjects. In conclusion, the prepared kit is considered safe in animal models and can be further evaluated for use in clinics.
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| 2. |
- Honarvar, Hadis, et al.
(författare)
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Position for site-specific attachment of a DOTA chelator to synthetic affibody molecules has a different influence on the targeting properties of 68Ga-Compared to 111in-labeled conjugates
- 2014
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Ingår i: Molecular Imaging. - : SAGE Publications. - 1535-3508 .- 1536-0121. ; 13:10
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Tidskriftsartikel (refereegranskat)abstract
- Affibody molecules, small (7 kDa) scaffold proteins, are a promising class of probes for radionuclide molecular imaging. Radiolabeling of Affibody molecules with the positron-emitting nuclide 68Ga would permit the use of positron emission tomography (PET), providing better resolution, sensitivity, and quantification accuracy than single-photon emission computed tomography (SPECT). The synthetic anti-HER2 ZHER2:S1 Affibody molecule was conjugated with DOTA at the N-terminus, in the middle of helix 3, or at the Cterminus. The biodistribution of 68Ga-and 111In-labeled Affibody molecules was directly compared in NMRI nu/nu mice bearing SKOV3 xenografts. The position of the chelator strongly influenced the biodistribution of the tracers, and the influence was more pronounced for 68Ga-labeled Affibody molecules than for the 111In-labeled counterparts. The best 68Ga-labeled variant was 68Ga-[DOTA-A1]-ZHER2:S1, which provided a tumor uptake of 13 ± 1 %ID/g and a tumor to blood ratio of 39 ± 12 at 2 hours after injection. 111In-[DOTA-A1]-ZHER2:S1 and 111In-[DOTA-K58]-ZHER2:S1 were equally good at this time point, providing a tumor uptake of 15 to 16 %ID/g and a tumor to blood ratio in the range of 60 to 80. In conclusion, the selection of the best position for a chelator in Affibody molecules can be used for optimization of their imaging properties. This may be important for the development of Affibody-based and other protein-based imaging probes.
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| 3. |
- Friedman, Mikaela, et al.
(författare)
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Directed evolution to low nanomolar affinity of a tumor-targeting epidermal growth factor receptor-binding affibody molecule
- 2008
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Ingår i: Journal of Molecular Biology. - : Elsevier BV. - 0022-2836 .- 1089-8638. ; 376:5, s. 1388-1402
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Tidskriftsartikel (refereegranskat)abstract
- The epidermal growth factor receptor 1 (EGFR) is overexpressed in various malignancies and is associated with a poor patient prognosis. A small, receptor-specific, high-affinity imaging agent would be a useful tool in diagnosing malignant tumors and in deciding upon treatment and assessing the response to treatment. We describe here the affinity maturation procedure for the generation of Affibody molecules binding with high affinity and specificity to EGFR. A library for affinity maturation was constructed by rerandomization of selected positions after the alignment of first-generation binding variants. New binders were selected with phage display technology, using a single oligonucleotide in a single-library effort, and the best second-generation binders had an approximately 30-fold improvement in affinity (K(d)=5-10 nM) for the soluble extracellular domain of EGFR in biospecific interaction analysis using Biacore. The dissociation equilibrium constant, K(d), was also determined for the Affibody with highest affinity using EGFR-expressing A431 cells in flow cytometric analysis (K(d)=2.8 nM). A retained high specificity for EGFR was verified by a dot blot assay showing staining only of EGFR proteins among a panel of serum proteins and other EGFR family member proteins (HER2, HER3, and HER4). The EGFR-binding Affibody molecules were radiolabeled with indium-111, showing specific binding to EGFR-expressing A431 cells and successful targeting of the A431 tumor xenografts with 4-6% injected activity per gram accumulated in the tumor 4 h postinjection.
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| 4. |
- Löfblom, John, et al.
(författare)
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Affibody molecules : Engineered proteins for therapeutic, diagnostic and biotechnological applications
- 2010
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Ingår i: FEBS Letters. - : Wiley. - 0014-5793 .- 1873-3468. ; 584:12, s. 2670-2680
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Forskningsöversikt (refereegranskat)abstract
- Affibody molecules are a class of engineered affinity proteins with proven potential for therapeutic, diagnostic and biotechnological applications. Affibody molecules are small (6.5 kDa) single domain proteins that can be isolated for high affinity and specificity to any given protein target. Fifteen years after its discovery, the Affibody technology is gaining use in many groups as a tool for creating molecular specificity wherever a small, engineering compatible tool is warranted. Here we summarize recent results using this technology, propose an Affibody nomenclature and give an overview of different HER2-specific Affibody molecules. Cumulative evidence suggests that the three helical scaffold domain used as basis for these molecules is highly suited to create a molecular affinity handle for vastly different applications.
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| 5. |
- Nordberg, Erika, et al.
(författare)
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In vivo and in vitro uptake of In-111, delivered with the affibody molecule(Z(EGFR:955))(2), in EGFR expressing tumour cells
- 2008
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Ingår i: Oncology Reports. - : Spandidos Publications. - 1021-335X .- 1791-2431. ; 19:4, s. 853-857
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Tidskriftsartikel (refereegranskat)abstract
- The epidermal growth factor receptor, EGFR, is overexpressed in many carcinomas. Targeting this receptor with radionuclides is important for imaging and therapy applications in nuclear medicine. We investigated the in vitro and in vivo properties of a new high affinity EGFR binding affibody molecule, (Z(EGFR:955))(2), when conjugated with CHXA"-DTPA and labelled with In-111. The binding time patterns and retention studies were performed using cultured squamous carcinoma A431 cells that overexpress EGFR. In the in vivo studies, female BALB/c nu/nu mice carrying tumours from xenografted A431 cells were used. The in vitro studies showed EGFR specific binding, high uptake and good retention of In-111 when delivered as [In-111](Z(EGFR:955))(2). The retention after 72 h of incubation was 38.0 +/- 1.15% of the initial level. The biodistribution study showed a tumour specific In-111 uptake of 3.8 +/- 1.4% of injected dose per gram turnout tissue 4 h post-injection. The tumour to blood ratio was 9.1 and the tumours could easily be visualized with a gamma camera at this time-point. In-111 delivered with [In-111](Z(EGFR:955))(2) gave an EGFR specific uptake and the results indicated that the (Z(EGFR:955))(2) affibody molecule is a candidate for radionuclide-based tumour imaging. Potential therapy applications are discussed.
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| 6. |
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| 7. |
- Steffen, A.C, et al.
(författare)
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Affibody mediated tumor targeting of HER-2 expressing xenografts in mice
- 2006
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Ingår i: European Journal of Nuclear Medicine and Molecular Imaging. - : Springer Science and Business Media LLC. - 1619-7070 .- 1619-7089. ; 33:6, s. 631-638
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Tidskriftsartikel (refereegranskat)abstract
- PURPOSE: Targeted delivery of radionuclides for diagnostic and therapeutic applications has until recently largely been limited to receptor ligands, antibodies and antibody-derived molecules. Here, we present a new type of molecule, a 15-kDa bivalent affibody called (Z(HER2:4))(2), with potential for such applications. The (Z(HER2:4))(2) affibody showed high apparent affinity (K (D)=3 nM) towards the oncogene product HER-2 (also called p185/neu or c-erbB-2), which is often overexpressed in breast and ovarian cancers. The purpose of this study was to investigate the in vivo properties of the new targeting agent. METHODS: The biodistribution and tumour uptake of the radioiodinated (Z(HER2:4))(2) affibody was studied in nude mice carrying tumours from xenografted HER-2 overexpressing SKOV-3 cells. RESULTS: The radioiodinated (Z(HER2:4))(2) affibody was primarily excreted through the kidneys, and significant amounts of radioactivity were specifically targeted to the tumours. The blood-borne radioactivity was, at all times, mainly in the macromolecular fraction. A tumour-to-blood ratio of about 10:1 was obtained 8 h post injection, and the tumours could be easily visualised with a gamma camera at this time point. CONCLUSION: The results indicate that the (Z(HER2:4))(2) affibody is an interesting candidate for applications in nuclear medicine, such as radionuclide-based tumour imaging and therapy.
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| 8. |
- Tolmachev, Vladimir A., et al.
(författare)
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Optical Spectra of Composite One-Dimensional Photonic Crystals With Extended Stop Bands Based on a Si-Air Structure
- 2015
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Ingår i: Journal of Lightwave Technology. - 0733-8724 .- 1558-2213. ; 33:17, s. 3577-3583
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Tidskriftsartikel (refereegranskat)abstract
- The optical spectra of composite one-dimensional photonic crystals (1-D PCs), fabricated by microstructuring of Si, are investigated. The composite PC is based on a Si-air structure and consists of two periodic 1-D PCs with various lattice constants a with a(1) = 2.7 mu m and a(2) = 4 mu m and various filling fractions f. The PCs are fabricated using photolithography followed by anisotropic chemical etching of (1 1 0) Si. Reflection and transmission spectra of this structure are measured using a Fourier transform infrared spectrometer combined with an IR microscope. The transmission spectra obtained demonstrate extended photonic stop bands (SBs), with characteristic transmission bands between the SBs. Theoretical and experimental transmission results indicate that the position of the extended SBs does not depend on the sequence of the individual PCs within the composite structure. In general, the calculated electric field distribution in the composite structure is similar to that of the individual PC components.
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| 9. |
- Tolmachev, Vladimir, et al.
(författare)
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Affibody molecules for epidermal growth factor receptor targeting in vivo : aspects of dimerization and labeling chemistry
- 2009
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Ingår i: Journal of Nuclear Medicine. - : Society of Nuclear Medicine. - 0161-5505 .- 1535-5667 .- 2159-662X. ; 50:2, s. 274-283
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Tidskriftsartikel (refereegranskat)abstract
- Noninvasive detection of epidermal growth factor receptor (EGFR) expression in malignant tumors by radionuclide molecular imaging may provide diagnostic information influencing patient management. The aim of this study was to evaluate a novel EGFR-targeting protein, the ZEGFR:1907 Affibody molecule, for radionuclide imaging of EGFR expression, to determine a suitable tracer format (dimer or monomer) and optimal label. METHODS: An EGFR-specific Affibody molecule, ZEGFR:1907, and its dimeric form, (ZEGFR:1907)2, were labeled with 111In using benzyl-diethylenetriaminepentaacetic acid and with 125I using p-iodobenzoate. Affinity and cellular retention of conjugates were evaluated in vitro. Biodistribution of radiolabeled Affibody molecules was compared in mice bearing EGFR-expressing A431 xenografts. Specificity of EGFR targeting was confirmed by comparison with biodistribution of non-EGFR-specific counterparts. RESULTS: Head-to-tail dimerization of the Affibody molecule improved the dissociation rate. In vitro, dimeric forms demonstrated superior cellular retention of radioactivity. For both molecular set-ups, retention was better for the 111In-labeled tracer than for the radioiodinated counterpart. In vivo, all conjugates accumulated specifically in xenografts and in EGFR-expressing tissues. The retention of radioactivity in tumors was better in vivo for dimeric forms; however, the absolute uptake values were higher for monomeric tracers. The best tracer, 111In-labeled ZEGFR:1907, provided a tumor-to-blood ratio of 100 (24 h after injection). CONCLUSION: The radiometal-labeled monomeric Affibody molecule ZEGFR:1907 has a potential for radionuclide molecular imaging of EGFR expression in malignant tumors.
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| 10. |
- Tolmachev, Vladimir, et al.
(författare)
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HEHEHE-Tagged Affibody Molecule May Be Purified by IMAC, Is Conveniently Labeled with [Tc-99m(CO)(3)](+), and Shows Improved Biodistribution with Reduced Hepatic Radioactivity Accumulation
- 2010
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Ingår i: Bioconjugate chemistry. - : American Chemical Society (ACS). - 1043-1802 .- 1520-4812. ; 21:11, s. 2013-2022
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Tidskriftsartikel (refereegranskat)abstract
- Affibody molecules are a class of small (ca. 7 kDa) robust scaffold proteins suitable for radionuclide molecular imaging of therapeutic targets in vivo. A hexahistidine tag at the N-terminus streamlines development of new imaging probes by enabling facile purification using immobilized metal ion affinity chromatography (IMAC), as well as convenient [Tc-99m(CO)(3)](+)-labeling. However, previous studies in mice have demonstrated that Affibody molecules labeled by this method yield higher liver accumulation of radioactivity, compared to the same tracer lacking the hexahistidine tag and labeled by an alternative method. Two variants of the HER2-binding Affibody molecule Z(HER2:342) were made in an attempt to create a tagged tracer that could be purified by immobilized metal affinity chromatography, yet would not result in anomalous hepatic radioactivity accumulation following labeling with [Tc-99m(CO)(3)](+). In one construct, the hexahistidine tag was moved to the C-terminus. In the other construct, every second histidine residue in the hexahistidine tag was replaced by the more hydrophilic glutamate, resulting in a HEHEHE-tag. Both variants, denoted Z(HER2:342)-H-6 and (HE)(3)-Z(HER2:342), respectively, could be efficiently purified using IMAC and stably labeled with [Tc-99m(CO)(3)](+) and were subsequently compared with the parental H-6-Z(HER2:342) having an N-terminal hexahistidine tag. All three variants were demonstrated to specifically bind to HER2-expressing cells in vitro. The hepatic accumulation of radioactivity in a murine model was 2-fold lower with [Tc-99m(CO)(3)](+)-Z(HER2:342)-H-6 compared to [Tc-99m(CO)(3)](+)-H-6-Z(HER2:342), and more than 10-fold lower with [Tc-99m(CO)(3)](+)-(HE)(3)-Z(HER2:342). These differences translated into appreciably superior tumor-to-liver ratio for [Tc-99m(CO)(3)](+)-(HE)(3)-Z(HER2:342) compared to the alternative conjugates. This information might be useful for development of other scaffold-based molecular imaging probes.
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