Reduction of renal activity retention of radiolabeled albumin binding domain‑derived affinity proteins using a non‑residualizing label strategy compared with a cleavable glycine‑leucine‑glycine‑lysine‑linker (original) (raw)
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Molecules, 2020
Albumin binding domain-Derived Affinity ProTeins (ADAPTs) are small (5 kDa) engineered scaffold proteins that are promising targeting agents for radionuclide-based imaging. A recent clinical study has demonstrated that radiolabeled ADAPTs can efficiently visualize human epidermal growth factor receptor 2 (HER2) expression in breast cancer using SPECT imaging. However, the use of ADAPTs directly labeled with radiometals for targeted radionuclide therapy is limited by their high reabsorption and prolonged retention of activity in kidneys. In this study, we investigated whether a co-injection of lysine or gelofusin, commonly used for reduction of renal uptake of radiolabeled peptides in clinics, would reduce the renal uptake of [99mTc]Tc(CO)3-ADAPT6 in NMRI mice. In order to better understand the mechanism behind the reabsorption of [99mTc]Tc(CO)3-ADAPT6, we included several compounds that act on various parts of the reabsorption system in kidneys. Administration of gelofusine, lysine,...
Influence of Several Compounds and Drugs on the Renal Uptake of Radiolabeled Affibody Molecules
Molecules, 2020
Affibody molecules are the most studied class of engineered scaffold proteins (ESPs) in radionuclide molecular imaging. Attempts to use affibody molecules directly labelled with radiometals for targeted radionuclide therapy were hampered by the high uptake and retention of radioactivity in kidneys. Several promising strategies have been implemented to circumvent this problem. Here, we investigated whether a pharmacological approach targeting different components of the reabsorption system could be used to lower the uptake of [99mTc]Tc-ZHER:2395 affibody molecule in kidneys. Pre-injection of probenecid, furosemide, mannitol or colchicine had no influence on activity uptake in kidneys compared to the control group. Mice pre-injected with maleate and fructose had 33% and 51% reduction in the kidney-associated activity, respectively, compared to the control group. Autoradiography images showed that the accumulation of activity after [99mTc]Tc-ZHER2:2395 injection was in the renal cortex...
European Journal of Nuclear Medicine and Molecular Imaging, 2008
Purpose Affibody molecules are low molecular weight proteins (7 kDa), which can be selected to bind to tumour-associated target proteins with subnanomolar affinity. Because of rapid tumour localisation and clearance from nonspecific compartments, Affibody molecules are promising tracers for molecular imaging. Earlier, 99mTc-labelled Affibody molecules demonstrated specific targeting of tumour xenografts. However, the biodistribution was suboptimal either because of hepatobiliary excretion or high renal uptake of the radioactivity. The goal of this study was to optimise the biodistribution of Affibody molecules by chelator engineering. Materials and methods Anti-HER2 ZHER2:342 Affibody molecules, carrying the mercaptoacetyl-glutamyl-seryl-glutamyl (maESE), mercaptoacetyl-glutamyl-glutamyl-seryl (maEES) and mercaptoacetyl-seryl-glutamyl-glutamyl (maSEE) chelators, were prepared by peptide synthesis and labelled with 99mTc. The tumour-targeting capacity of these conjugates was compared with each other and with the best previously available conjugate, 99mTc-maEEE-ZHER2:342, in nude mice bearing SKOV-3 xenografts. The tumour-targeting capacity of the most promising conjugate, 99mTc-maESE-ZHER2:342, was compared with radioiodinated ZHER2:342. Results All novel conjugates demonstrated successful tumour targeting and a low degree of hepatobiliary excretion. The renal uptakes of serine-containing conjugates, 33 ± 5, 68 ± 21 and 71 ± 10%IA/g, for99mTc-maESE-ZHER2:342, 99mTc-maEES-ZHER2:342 and 99mTc-maSEE-ZHER2:342, respectively, were significantly reduced in comparison with 99mTc-maEEE-ZHER2:342 (102 ± 13%IA/g). For 99mTc-maESE-ZHER2:342, a tumour uptake of 9.6 ± 1.8%IA/g and a tumour-to-blood ratio of 58 ± 6 were reached at 4 h p.i. Conclusions A combination of serine and glutamic acid residues in the chelator sequence confers increased renal excretion and relatively low renal uptake of 99mTc-labelled Affibody molecules. In combination with preserved targeting capacity, this improved imaging of targets in abdominal area.
Drug Discovery Today, 2012
Radionuclide imaging of molecular targets for cancer therapy is likely to be a powerful tool for patient stratification and response monitoring, allowing more personalized cancer treatment. Radiolabeled proteins and peptides are a promising class of imaging probes for visualization of molecular targets in vivo. However, hepatic uptake and hepatobiliary excretion of radioactivity can decrease imaging contrast, reducing the detection sensitivity of hepatic and extrahepatic abdominal metastases, respectively. In this article, we review factors that influence the hepatic uptake of radioactivity (e.g. the chemical nature of radiocatabolites and physicochemical properties of targeting peptides and linkers) to provide input for the rational design of peptide-based imaging probes.
In Vitro and In Vivo Evaluation of a 18F-Labeled High Affinity
2013
Expression of the gastrin-releasing peptide receptor (GRPR) in prostate cancer suggests that this receptor can be used as a potential molecular target to visualize and treat these tumors. We have previously investigated an antagonist analog of bombesin (D-Phe-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH 2 , RM26) conjugated to 1,4,7triazacyclononane-N,N',N''-triacetic acid (NOTA) via a diethylene glycol (PEG 2) spacer (NOTA-P2-RM26) labeled with 68 Ga and 111 In. We found that this conjugate has favorable properties for in vivo imaging of GRPR-expression. The focus of this study was to develop a 18 F-labelled PET agent to visualize GRPR. NOTA-P2-RM26 was labeled with 18 F using aluminum-fluoride chelation. Stability, in vitro binding specificity and cellular processing tests were performed. The inhibition efficiency (IC 50) of the [ nat F]AlF-NOTA-P2-RM26 was compared to that of the nat Ga-loaded peptide using 125 I-Tyr 4-BBN as the displacement radioligand. The pharmacokinetics and in vivo binding specificity of the compound were studied. NOTA-P2-RM26 was labeled with 18 F within 1 h (60-65% decay corrected radiochemical yield, 55 GBq/µmol). The radiopeptide was stable in murine serum and showed high specific binding to PC-3 cells. [ nat F]AlF-NOTA-P2-RM26 showed a low nanomolar inhibition efficiency (IC 50 =4.4±0.8 nM). The internalization rate of the tracer was low. Less than 14% of the cell-bound radioactivity was internalized after 4 h. The biodistribution of [ 18 F]AlF-NOTA-P2-RM26 demonstrated rapid blood clearance, low liver uptake and low kidney retention. The tumor uptake at 3 h p.i. was 5.5±0.7 %ID/g, and the tumor-to-blood,-muscle and-bone ratios were 87±42, 159±47, 38±16, respectively. The uptake in tumors, pancreas and other GRPR-expressing organs was significantly reduced when excess amount of non-labeled peptide was co-injected. The low uptake in bone suggests a high in vivo stability of the Al-F bond. High contrast PET image was obtained 3 h p.i. The initial biological results suggest that [ 18 F]AlF-NOTA-P2-RM26 is a promising candidate for PET imaging of GRPR in vivo.
European journal of medicinal chemistry, 2014
Affibody molecules constitute a class of small (7 kDa) scaffold proteins that can be engineered to have excellent tumor targeting properties. High reabsorption in kidneys complicates development of affibody molecules for radionuclide therapy. In this study, we evaluated the influence of the composition of cysteine-containing C-terminal peptide-based chelators on the biodistribution and renal retention of (188)Re-labeled anti-HER2 affibody molecules. Biodistribution of affibody molecules containing GGXC or GXGC peptide chelators (where X is G, S, E or K) was compared with biodistribution of a parental affibody molecule ZHER2:2395 having a KVDC peptide chelator. All constructs retained low picomolar affinity to HER2-expressing cells after labeling. The biodistribution of all (188)Re-labeled affibody molecules was in general comparable, with the main observed difference found in the uptake and retention of radioactivity in excretory organs. The (188)Re-ZHER2:V2 affibody molecule with a...
Chemical design of radiolabeled antibody fragments for low renal radioactivity levels
Cancer research, 1999
The renal uptake of radiolabeled antibody fragments presents a problem in targeted imaging and therapy. We hypothesized that the renal radioactivity levels of radiolabeled antibody fragments could be reduced if radiolabeled compounds of urinary excretion were released from glomerularly filtered antibody fragments before they were incorporated into renal cells by the action of brush border enzymes, present on the lumen of renal tubules. 3'-[131I]Iodohippuryl N(epsilon)-maleoyl-L-lysine ([131I]HML) was conjugated with a thiolated Fab fragment because the glycyl-lysine sequence in HML is a substrate for a brush border enzyme and metaiodohippuric acid is released by cleavage of the linkage. Fab fragments were also radiolabeled by direct radioiodination (125I-Fab) or by conjugation with meta-[125I]-iodohippuric acid via an amide bond [N-(5-maleimidopentyl) 3'-iodohippuric acid amide ([125I]MPH-Fab)] or an ester bond [maleimidoethy 3'-iodohippurate ([125I]MIH-Fab)] by procedur...
European Journal of Nuclear Medicine and Molecular Imaging, 2009
Purpose HER2 is a transmembrane tyrosine kinase, which is overexpressed in a number of carcinomas. The Affibody molecule Z HER2:342 is a small (7 kDa) affinity protein binding to HER2 with an affinity of 22 pM. The goal of this study was to evaluate the use of ((4-hydroxyphenyl)ethyl) maleimide (HPEM) for radioiodination of Z HER2:342 and to compare the targeting properties of monomeric and dimeric forms of Z HER2:342 . Methods The biodistribution of different radioiodinated derivatives of Z HER2:342 was studied in BALB/C nu/nu mice bearing HER2-expressing SKOV-3 xenografts. Biodistributions of 125 I-PIB-Z HER2:342 and site-specifically labelled 125 I-HPEM-Z HER2:342 -C were compared. Biodistributions of monomeric 131 I-HPEM-Z HER2:342 -C and dimeric 125 I-HPEM-(Z HER2:342 ) 2 -C were evaluated using a pairedlabel method.
Reducing Renal Uptake of Radiolabeled Peptides Using Albumin Fragments
Journal of Nuclear Medicine, 2008
In most types of peptide receptor radionuclide therapy, the maximum activity dose that can be administered is limited by high and persistent renal retention of the radiolabeled peptides, which is, at least partly, mediated by the megalin receptor. Several agents that interfere with renal reabsorption of radiolabeled peptides have been identified (e.g., lysine, arginine, succinylated gelatin solution), but none of these inhibit renal reabsorption completely. Albumin, a naturally abundant megalin ligand, might be a safe and potent alternative. In this study, we analyzed the effects of albumin and fragments of albumin (FRALB) on the renal reabsorption of 111 In-diethylenetriaminepentaacetic acid (DTPA)-D-Phe 1 -octreotide ( 111 In-octreotide), [Lys 40 (aminohexoic acid-DTPA-111 In)NH 2 ]-exendin-4 ( 111 In-exendin), and 111 In-1,4,7,10-tetraazacyclododecane-N,N9,N$,N9$-tetraacetic acid (DOTA)-Glu 1 -minigastrin ( 111 In-minigastrin). Methods: The effects of albumin and FRALB on megalin-associated binding of 111 In-octreotide, 111 In-exendin, and 111 In-minigastrin were assessed in vitro using rat yolk sac epithelial (BN16) cells. In vivo, uptake and localization of 111 In-albumin and 111 In-FRALB in the kidneys of Wistar rats were determined, as well as the effect of lysine, succinylated gelatin solution, albumin, and FRALB on the kidney uptake of 111 In-octreotide, 111 In-exendin, and 111 Inminigastrin. Results: FRALB significantly reduced binding and uptake of 111 In-octreotide, 111 In-exendin, and 111 In-minigastrin by BN16 cells. In rats, renal uptake of 111 In-labeled FRALB was significantly higher than that of 111 In-labeled intact albumin (P , 0.001). FRALB administration effectively reduced renal uptake of 111 In-octreotide, 111 In-exendin, and 111 In-minigastrin. Administration of 1-2 mg of FRALB reduced renal uptake of 111 In-octreotide as efficiently as 80 mg of lysine. Conclusion: Renal uptake of 111 In-octreotide and other radiolabeled peptides in rats can be effectively reduced by administration of albumin fragments. Additional studies to identify the albumin fragments responsible for inhibition of renal peptide uptake are warranted.
Radiopharmaceuticals for renal studies: Evaluation of protein binding
Journal of Radioanalytical and Nuclear Chemistry, 1989
Determination of total in vitro protein binding was evaluated for the following radiopharmaceuticals:99mTc-diethylenetriaminepentaacetic acid (DTPA),99mTc-dimercaptosuccinic acid (DMSA),99mTc-glucoheptonate (GH) and99mTc-fosfomycin (PHO). For that they were incubated wtih human serum at 37°C. After three and sixty minutes of incubation, the bound fraction was evaluated by two different methods: gel filtration and precipitation with trichloroacetic acid (TCA). The percentage of the99mTc-kidney agents bound