| 1. |
- Garousi, Javad, et al.
(författare)
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Comparative evaluation of tumor targeting using the anti-HER2 ADAPT scaffold protein labeled at the C-terminus with indium-111 or technetium-99m
- 2017
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Ingår i: Scientific Reports. - : NATURE PUBLISHING GROUP. - 2045-2322. ; 7
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Tidskriftsartikel (refereegranskat)abstract
- ABD-Derived Affinity Proteins (ADAPTs) is a novel class of engineered scaffold proteins derived from an albumin-binding domain of protein G. The use of ADAPT6 derivatives as targeting moiety have provided excellent preclinical radionuclide imaging of human epidermal growth factor 2 (HER2) tumor xenografts. Previous studies have demonstrated that selection of nuclide and chelator for its conjugation has an appreciable effect on imaging properties of scaffold proteins. In this study we performed a comparative evaluation of the anti-HER2 ADAPT having an aspartate-glutamate-alanine-valine-aspartate-alanine-asparagine-serine (DEAVDANS) N-terminal sequence and labeled at C-terminus with (99)mTc using a cysteine-containing peptide based chelator, glycine-serine-serine-cysteine (GSSC), and a similar variant labeled with In-111 using a maleimido derivative of 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) chelator. Both (99)mTc-DEAVDANS-ADAPT6-GSSC and In-111-DEAVDANS-ADAPT6-GSSC-DOTA accumulated specifically in HER2-expressing SKOV3 xenografts. The tumor uptake of both variants did not differ significantly and average values were in the range of 19-21% ID/g. However, there was an appreciable variation in uptake of conjugates in normal tissues that resulted in a notable difference in the tumor-to-organ ratios. The In-111-DOTA label provided 2-6 fold higher tumor-to-organ ratios than (99)mTc-GSSC and is therefore the preferable label for ADAPTs.
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| 2. |
- Garousi, Javad, et al.
(författare)
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The use of radiocobalt as a label improves imaging of EGFR using DOTA-conjugated Affibody molecule
- 2017
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Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 7
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Tidskriftsartikel (refereegranskat)abstract
- Several anti-cancer therapies target the epidermal growth factor receptor (EGFR). Radionuclide imaging of EGFR expression in tumours may aid in selection of optimal cancer therapy. The In-111-labelled DOTA-conjugated Z(EGFR:2377) Affibody molecule was successfully used for imaging of EGFR-expressing xenografts in mice. An optimal combination of radionuclide, chelator and targeting protein may further improve the contrast of radionuclide imaging. The aim of this study was to evaluate the targeting properties of radiocobalt-labelled DOTA-Z(EGFR:2377). DOTA-Z(EGFR:2377) was labelled with Co-57 (T-1/2 = 271.8 d), Co-55 (T-1/2 = 17.5 h), and, for comparison, with the positron-emitting radionuclide Ga-68 (T-1/2 = 67.6 min) with preserved specificity of binding to EGFR-expressing A431 cells. The long-lived cobalt radioisotope Co-57 was used in animal studies. Both Co-57-DOTA-Z(EGFR:2377) and Ga-68-DOTA-Z(EGFR:2377) demonstrated EGFR-specific accumulation in A431 xenografts and EGFR-expressing tissues in mice. Tumour-to-organ ratios for the radiocobalt-labelled DOTA-Z(EGFR:2377) were significantly higher than for the gallium-labelled counterpart already at 3 h after injection. Importantly, Co-57-DOTA-Z(EGFR:2377) demonstrated a tumour-to-liver ratio of 3, which is 7-fold higher than the tumour-to-liver ratio for (68)GaDOTA-Z(EGFR:2377). The results of this study suggest that the positron-emitting cobalt isotope 55Co would be an optimal label for DOTA-Z(EGFR:2377) and further development should concentrate on this radionuclide as a label.
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| 3. |
- Güler, Rezan, et al.
(författare)
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Increasing thermal stability and improving biodistribution of VEGFR2-binding affibody molecules by a combination of in silico and directed evolution approaches
- 2020
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Ingår i: Scientific Reports. - : Nature Research. - 2045-2322. ; 10:1
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Tidskriftsartikel (refereegranskat)abstract
- The family of vascular endothelial growth factor (VEGF) ligands and their interactions with VEGF receptors (VEGFRs) play important roles in both pathological and physiological angiogenesis. Hence, agonistic and antagonistic ligands targeting this signaling pathway have potential for both studies on fundamental biology and for development of therapies and diagnostics. Here, we engineer VEGFR2-binding affibody molecules for increased thermostability, refolding and improved biodistribution. We designed libraries based on the original monomeric binders with the intention of reducing hydrophobicity, while retaining high affinity for VEGFR2. Libraries were displayed on bacteria and binders were isolated by fluorescence-activated cell sorting (FACS). In parallel, we used an automated sequence- and structure-based in silico algorithm to identify potentially stabilizing mutations. Monomeric variants isolated from the screening and the in silico approach, respectively, were characterized by circular dichroism spectroscopy and biosensor assays. The most promising mutations were combined into new monomeric constructs which were finally fused into a dimeric construct, resulting in a 15 degrees C increase in melting temperature, complete refolding capability after heat-induced denaturation, retained low picomolar affinity and improved biodistribution profile in an in vivo mouse model. These VEGFR2-binding affibody molecules show promise as candidates for further in vivo studies to assess their suitability as molecular imaging and therapeutic agents.
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| 4. |
- Honarvar, Hadis, 1984-, et al.
(författare)
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Evaluation of HER2-specific peptide ligand for its employment as radiolabeled imaging probe
- 2018
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Ingår i: Scientific Reports. - : NATURE PUBLISHING GROUP. - 2045-2322. ; 8
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Tidskriftsartikel (refereegranskat)abstract
- HER2 transmembrane receptor is an important target in immunotherapy treatment of breast and gastroesophageal cancer. Molecular imaging of HER2 expression may provide essential prognostic and predictive information concerning disseminated cancer and aid in selection of an optimal therapy. Radiolabeled low molecular weight peptide ligands are particularly attractive as probes for molecular imaging, since they reach and bind to the target and clear from non-target organs and blood stream faster than bulky antibodies. In this study, we evaluated a potential HER2-imaging probe, an A9 nonapeptide, derived from the trastuzumab-Fab portion. Its cellular uptake was investigated by mass spectrometry analysis of the cytoplasmic cellular extracts. Moreover, based on in-silico modeling, DTPA chelator was conjugated to N-terminus of A9. In-111-labeled A9 demonstrated nanomolar affinity to HER2-expressing BT474 cells and favorable biodistribution profile in NMRI mice. This study suggests that the peptide A9 represents a good lead candidate for development of molecular probe, to be used for imaging purposes and for the delivery of cytotoxic agents.
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| 5. |
- Mitran, Bogdan, et al.
(författare)
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Selection of an optimal macrocyclic chelator improves the imaging of prostate cancer using cobalt-labeled GRPR antagonist RM26
- 2019
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Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 9
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Tidskriftsartikel (refereegranskat)abstract
- Gastrin-releasing peptide receptors (GRPRs) are promising targets in oligometastatic prostate cancer. We have recently used 55Co (T1/2 = 17.5 h) as a label for next day PET imaging of GRPR expression obtaining high imaging contrast. The radionuclide-chelator combination can significantly influence the biodistribution of radiopeptides. Therefore, in this study, we hypothesized that the properties of 55Co-labeled PEG2-RM26 can be improved by identifying the optimal macrocyclic chelator. All analogues (X-PEG2-RM26, X = NOTA,NODAGA,DOTA,DOTAGA) were successfully labeled with radiocobalt with high yields and demonstrated high stability. The radiopeptides bound specifically and with picomolar affinity to GRPR and their cellular processing was characterized by low internalization. The best binding capacity was found for DOTA-PEG2-RM26. Ex vivo biodistribution in PC-3 xenografted mice was characterized by rapid blood clearance via renal excretion. Tumor uptake was similar for all conjugates at 3 h pi, exceeding the uptake in all other organs. Higher kidney uptake and longer retention were associated with N-terminal negative charge (DOTAGA-containing conjugate). Tumor-to-organ ratios increased over time for all constructs, although significant chelator-dependent differences were observed. Concordant with affinity measurements, DOTA-analog had the best retention of activity in tumors, resulting in the highest tumor-to-blood ratio 24 h pi, which translated into high contrast PET/CT imaging (using 55Co).
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| 6. |
- Myrhammar, Anders, et al.
(författare)
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Evaluation of an antibody-PNA conjugate as a clearing agent for antibody-based PNA-mediated radionuclide pretargeting
- 2020
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Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 10:1
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Tidskriftsartikel (refereegranskat)abstract
- Radionuclide molecular imaging of cancer-specific targets is a promising method to identify patients for targeted antibody therapy. Radiolabeled full-length antibodies however suffer from slow clearance, resulting in high background radiation. To overcome this problem, a pretargeting system based on complementary peptide nucleic acid (PNA) probes has been investigated. The pretargeting relies on sequential injections of primary, PNA-tagged antibody and secondary, radiolabeled PNA probe, which are separated in time, to allow for clearance of non-bound primary agent. We now suggest to include a clearing agent (CA), designed for removal of primary tumor-targeting agent from the blood. The CA is based on the antibody cetuximab, which was conjugated to PNA and lactosaminated by reductive amination to improve hepatic clearance. The CA was evaluated in combination with PNA-labelled trastuzumab, T-ZHP1, for radionuclide HER2 pretargeting. Biodistribution studies in normal mice demonstrated that the CA cleared ca. 7 times more rapidly from blood than unmodified cetuximab. Injection of the CA 6 h post injection of the radiolabeled primary agent [131I]I-T-ZHP1 gave a moderate reduction of the radioactivity concentration in the blood after 1 h from 8.5 ± 1.8 to 6.0 ± 0.4%ID/g. These proof-of-principle results could guide future development of a more efficient CA.
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| 7. |
- Rinne, Sara S., et al.
(författare)
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Ga-66-PET-imaging of GRPR-expression in prostate cancer : production and characterization of [Ga-66]Ga-NOTA-PEG(2)-RM26
- 2021
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Ingår i: Scientific Reports. - : Springer Nature. - 2045-2322. ; 11
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Tidskriftsartikel (refereegranskat)abstract
- Molecular imaging of the gastrin-releasing peptide receptor (GRPR) could improve patient management in prostate cancer. This study aimed to produce gallium-66 (T-1/2=9.5 h) suitable for radiolabeling, and investigate the imaging properties of gallium-66 labeled GRPR-antagonist NOTA-PEG(2)-RM26 for later-time point PET-imaging of GRPR expression. Gallium-66 was cyclotron-produced using a liquid target, and enriched [Zn-66]Zn(NO3)(2). In vitro, [Ga-66]Ga-NOTA-PEG(2)-RM26 was characterized in GRPR-expressing PC-3 prostate cancer cells. In vivo, specificity test and biodistribution studies were performed 3 h and 22 h pi in PC-3 xenografted mice. microPET/MR was performed 3 h and 22 h pi. Biodistribution of [Ga-66]Ga-NOTA-PEG(2)-RM26 was compared with [Ga-68]Ga-NOTA-PEG(2)-RM26 3 h pi. [Ga-66]Ga-NOTA-PEG(2)-RM26 was successfully prepared with preserved binding specificity and high affinity towards GRPR. [Ga-66]Ga-NOTA-PEG(2)-RM26 cleared rapidly from blood via kidneys. Tumor uptake was GRPR-specific and exceeded normal organ uptake. Normal tissue clearance was limited, resulting in no improvement of tumor-to-organ ratios with time. Tumors could be clearly visualized using microPET/MR. Gallium-66 was successfully produced and [Ga-66]Ga-NOTA-PEG(2)-RM26 was able to clearly visualize GRPR-expression both shortly after injection and on the next day using PET. However, delayed imaging did not improve contrast for Ga-labeled NOTA-PEG(2)-RM26.
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| 8. |
- Rinne, Sara S., et al.
(författare)
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Increase in negative charge of Ga-68/chelator complex reduces unspecific hepatic uptake but does not improve imaging properties of HER3-targeting affibody molecules
- 2019
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Ingår i: Scientific Reports. - : Nature Publishing Group. - 2045-2322. ; 9
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Tidskriftsartikel (refereegranskat)abstract
- Upregulation of the human epidermal growth factor receptor type 3 (HER3) is a common mechanism to bypass HER-targeted cancer therapy. Affibody-based molecular imaging has the potential for detecting and monitoring HER3 expression during treatment. In this study, we compared the imaging properties of newly generated Ga-68-labeled anti-HER3 affibody molecules (HE)(3)-Z(HER3)-DOTA and (HE)(3)-Z(HER3)-DOTAGA with previously reported [Ga-68]Ga-(HE)(3)-Z(HER3)-NODAGA. We hypothesized that increasing the negative charge of the gallium-68/chelator complex would reduce hepatic uptake, which could lead to improved contrast of anti-HER3 affibody-based PET-imaging of HER3 expression. (HE)(3)-Z(HER3)-X (X = DOTA, DOTAGA) were produced and labeled with gallium-68. Binding of the new conjugates was specific in HER3 expressing BxPC-3 and DU145 human cancer cells. Biodistribution and in vivo specificity was studied in BxPC-3 xenograft bearing Balb/c nu/nu mice 3 h pi. DOTA- and DOTAGA-containing conjugates had significantly higher concentration in blood than [Ga-68]Ga-(HE)(3)-Z(HER3)-NODAGA. Presence of the negatively charged Ga-68-DOTAGA complex reduced the unspecific hepatic uptake, but did not improve overall biodistribution of the conjugate. [Ga-68]Ga-(HE)(3)-Z(HER3)-DOTAGA and [Ga-68]Ga-(HE)(3)-Z(HER3)-NODAGA had similar tumor-to-liver ratios, but [Ga-68]Ga-(HE)(3)-Z(HER3)-NODAGA had the highest tumor uptake and tumor-to-blood ratio among the tested conjugates. In conclusion, [Ga-68] Ga-(HE)(3)-Z(HER3)-NODAGA remains the favorable variant for PET imaging of HER3 expression.
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| 9. |
- Rinne, Sara S., et al.
(författare)
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Optimization of HER3 expression imaging using affibody molecules : Influence of chelator for labeling with indium-111
- 2019
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Ingår i: Scientific Reports. - : NATURE PUBLISHING GROUP. - 2045-2322. ; 9
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Tidskriftsartikel (refereegranskat)abstract
- Radionuclide molecular imaging of human epidermal growth factor receptor 3 (HER3) expression using affibody molecules could be used for patient stratification for HER3-targeted cancer therapeutics. We hypothesized that the properties of HER3-targeting affibody molecules might be improved through modification of the radiometal-chelator complex. Macrocyclic chelators NOTA (1,4,7-triazacyclononane-N,N',N ''-triacetic acid), NODAGA (1-(1,3-carboxypropyl)-4,7-carboxymethyl-1,4,7-triazacyclononane), DOTA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraaceticacid), and DOTAGA (1,4,7,10-tetraazacyclododececane, 1-(glutaric acid)-4,7,10-triacetic acid) were conjugated to the C-terminus of anti-HER3 affibody molecule Z(08698) and conjugates were labeled with indium-111. All conjugates bound specifically and with picomolar affinity to HER3 in vitro. In mice bearing HER3-expressing xenografts, no significant difference in tumor uptake between the conjugates was observed. Presence of the negatively charged In-111-DOTAGA-complex resulted in the lowest hepatic uptake and the highest tumor-to-liver ratio. In conclusion, the choice of chelator influences the biodistribution of indium-111 labeled anti-HER3 affibody molecules. Hepatic uptake of anti-HER3 affibody molecules could be reduced by the increase of negative charge of the radiometal-chelator complex on the C-terminus without significantly influencing the tumor uptake.
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| 10. |
- Tolmachev, Vladimir, et al.
(författare)
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Molecular design of radiocopper-labelled Affibody molecules
- 2018
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Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 8:1
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Tidskriftsartikel (refereegranskat)abstract
- Cu-CB-TE2A-GEEE-ZHER2:342 was 16 ± 6%ID/g and tumor-to-blood ratio was 181 ± 52. In conclusion, a combination of the cross-bridged CB-TE2A chelator and Gly-Glu-Glu-Glu spacer is preferable for radiocopper labelling of Affibody molecules and, possibly, other scaffold proteins having high renal re-absorption.
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