Direct Radioiodination Research Articles

Green synthesis, radioiodination and in vivo biodistribution of 5-(2-hydroxyphenyl)-2,4-dihydro-3H-pyrazol-3-one derivatives as potential candidates for lung imaging

Lung targeting was developed by synthesising pyrazolone derivatives 6a-f under solvent-free and thermal conditions by reacting azo coumarins 4a-c with hydrazines 5a and b using maltose as a biodegradable catalyst. Different spectral data characterized the synthesized agents as proton-NMR, FT-IR, and mass spectra. Direct radioiodination with iodine-131 was performed and optimized to reach the highest radiochemical purities (92 ± 0.47 to 98 ± 0.21%) using chloramine-T, a moderate oxidizing agent. The 131I-pyrazolone derivatives were confirmed based on HRMS. Furthermore, radioiodinated nitro-derivatives accumulated well in the lung of normal mice during in vivo evaluation, and the better uptake was for nitrophenyl-derivative 7f, about 30.06 ± 0.04% at 30 min after injection. Consequently, synthesized radioiodinated derivatives may be employed as prospective tracers for lung perfusion scans.

Effect of Water on Direct Radioiodination of Small Molecules/Peptides Using Copper-Mediated Iododeboronation in Water-Alcohol Solvent.

Direct radioiodination of peptides using copper-mediated iododeboronation is a promising radiosynthetic method for solving issues of classical direct radiolabeling, such as toxicity of the organotin precursor (iododestannylation) or formation of radio byproducts (by electrophilic iodination of a tyrosine residue). However, the parameters for optimizing the reaction conditions for various peptides are not completely understood. In particular, considering peptide solubility, the effects of water-containing solvents on labeling efficiency should be thoroughly investigated. Herein, we describe the effect of water on copper-mediated radioiododeboronation and the key factors for ensuring the successful radiolabeling of small molecules and peptides in water-organic solvents. 125I-labeled substrates containing peptides ([125I]m/p-IBTA) were obtained with high radiochemical conversions (RCCs: >95%) using an alcohol solvent, and a decrease in these RCCs was observed with increasing water content in the methanol solvent. Additionally, when using water-methanol solvents, a difference in RCC due to the substituent effect was also observed. However, the RCCs can be improved without the use of other additives by adjusting the copper catalyst and time of the labeling reaction or by utilizing substituent effects. This study contributes to the improvement of the design of boronic peptide precursors and radiolabeling protocols using copper-mediated iododeboronation.

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One of the most clinically relevant molecular aberrations in breast cancer is overexpression of human epidermal growth factor receptor type 2 (HER2). We aimed to develop a radiolabeled tyrosine kinase inhibitor for HER2-targeted breast cancer imaging. In this study, a radioiodinated analog (^125/131I-IBA-CP) of the HER2-selective inhibitor CP724,714 was prepared and evaluated in HER2-positive or -negative subcutaneous human breast cancer xenografts. <b>Methods:</b> The CP724,714 analog IBA-CP was synthesized and assayed for its inhibitory activities against HER2 and 6 other tyrosine kinases. ^125/131I-IBA-CP was prepared using a copper-mediated radioiodination method with enhanced labeling yield and molar activity. In vitro biologic activity, including specific and nonspecific binding of ¹³¹I-IBA-CP to its HER2 kinase target, was assessed in different cell lines. In vivo small-animal ¹²⁵I-IBA-CP SPECT imaging and biodistribution studies were conducted on mice bearing HER2-positive, HER2-negative, or epidermal growth factor receptor (EGFR)-positive tumors. Nonradioactive IBA-CP and the EGFR inhibitor erlotinib were used as blocking agents to investigate the binding specificity and selectivity of ^125/131I-IBA-CP toward HER2 in vitro and in vivo. Additionally, ^125/131I-ICP was prepared by direct radioiodination of CP724,714 for comparison with ^125/131I-IBA-CP. <b>Results:</b> IBA-CP displayed superior in vitro inhibitory activity (half-maximal inhibitory concentration, 16 nM) and selectivity for HER2 over 6 other cancer-related tyrosine kinases. ^125/131I-IBA-CP was prepared in a typical radiochemical yield of about 65% (decay-corrected), radiochemical purity of more than 98%, and molar activity of 42 GBq/μmol at the end of synthesis. SPECT imaging revealed significantly higher uptake of ¹²⁵I-IBA-CP than of ¹²⁵I-ICP in the HER2-positive MDA-MB-453 tumors. Uptake in the HER2-negative MCF-7 tumors was much lower. Binding of ¹²⁵I-IBA-CP in the MDA-MB-453 tumors was blocked by coinjection with an excess amount of IBA-CP, but not by erlotinib. <b>Conclusion:</b> The radiolabeled HER2-selective inhibitor ^125/131I-IBA-CP is a promising probe for in vivo detection of HER2-positive tumors.

Copper-mediated radioiodination and radiobromination via aryl boronic precursor and its application to 125I/77Br–labeled prostate-specific membrane antigen imaging probes

Prostate-specific membrane antigen (PSMA), expressed in prostate cancer cells, is being investigated extensively worldwide as a target for imaging and therapy of prostate cancer. Various radioiodinated PSMA imaging probes have been developed, and their structure has a peptidomimetic urea-based skeleton as a pharmacophore. For direct radioiodination of molecules containing these peptidomimetic structures, prior studies performed radioiododestannylation or electrophilic radioiodination of tyrosine residues. However, although these radiolabeling methods are frequently used, there are some issues with precursor toxicity and by-product production. Therefore, it is required to investigate a radiolabeling method that can be used for the radiosynthesis of radioiodinated PSMA imaging probes with urea-based peptidomimetic structures. We recently reported that copper-mediated radioiodination via a boronic precursor is an effective method for directly labeling a peptide. This radiohalogenation method was expected to be an effective method for radiosynthesis of PSMA imaging probes with a peptidomimetic structure. In this study, to confirm that this labeling method applies to the synthesis of the PSMA imaging probe, we synthesized PSMA imaging probes labeled with 125I and 77Br ([125I]mIB-PS and [77Br]mBrB-PS) using a copper-mediated radiohalogenation via common boronic precursors and investigated optimal boronic precursor and labeling conditions. As a result, the radiochemical yields of [125I]mIB-PS and [77Br]mBrB-PS were improved to > 93% at room temperature by optimizing the structure of the boronic precursor. We demonstrate that copper-mediated nucleophilic radiochemistry using a boronic precursor is a promising radiosynthetic method of PSMA imaging probes. Although we focused on the synthesis of PSMA imaging probes, the results in this study will also be useful for the synthesis of various radioiodine or radiobromine-labeled bioactive molecules.

Advancing 89Zr-immuno-PET in neuroscience with a bispecific anti-amyloid-beta monoclonal antibody - The choice of chelator is essential.

The accelerated approval of the monoclonal antibody (mAb) aducanumab as a treatment option for Alzheimer's Disease and the continued discussions about its efficacy have shown that a better understanding of immunotherapy for the treatment of neurodegenerative diseases is needed. 89Zr-immuno-PET could be a suitable tool to open new avenues for the diagnosis of CNS disorders, monitoring disease progression, and assessment of novel therapeutics. Herein, three different 89Zr-labeling strategies and direct radioiodination with 125I of a bispecific anti-amyloid-beta aducanumab derivate, consisting of aducanumab with a C-terminal fused anti-transferrin receptor binding single chain Fab fragment derived from 8D3 (Adu-8D3), were compared ex vivo and in vivo with regard to brain uptake and target engagement in an APP/PS1 Alzheimer's disease mouse model and wild type animals.Methods: Adu-8D3 and a negative control antibody, based on the HIV specific B12 antibody also carrying C-terminal fused 8D3 scFab (B12-8D3), were each conjugated with NCS-DFO, NCS-DFO*, or TFP-N-suc-DFO-Fe-ester, followed by radiolabeling with 89Zr. 125I was used as a substitute for 124I for labeling of both antibodies. 30 µg of radiolabeled mAb, corresponding to approximately 6 MBq 89Zr or 2.5 MBq 125I, were injected per mouse. PET imaging was performed 1, 3 and 7 days post injection (p.i.). All mice were sacrificed on day 7 p.i. and subjected to ex vivo biodistribution and brain autoradiography. Immunostaining on brain tissue was performed after autoradiography for further validation.Results: Ex vivo biodistribution revealed that the brain uptake of [89Zr]Zr-DFO*-NCS-Adu-8D3 (2.19 ±0.12 %ID/g) was as high as for its 125I-analog (2.21 ±0.15 %ID/g). [89Zr]Zr-DFO-NCS-Adu-8D3 and [89Zr]Zr-DFO-N-suc-Adu-8D3 showed significantly lower uptake (< 0.65 %ID/g), being in the same range as for the 89Zr-labeled controls (B12-8D3). Autoradiography of [89Zr]Zr-DFO*-NCS-Adu-8D3 and [125I]I-Adu-8D3 showed an amyloid-beta related granular uptake pattern of radioactivity. In contrast, the [89Zr]Zr-DFO-conjugates and the control antibody groups did not show any amyloid-beta related uptake pattern, indicating that DFO is inferior for 89Zr-immuno-PET imaging of the brain in comparison to DFO* for Adu-8D3. This was confirmed by day 7 PET images showing only amyloid-beta related brain uptake for [89Zr]Zr-DFO*-NCS-Adu-8D3. In wild type animals, such an uptake was not observed. Immunostaining showed a co-localization of all administered Adu-8D3 conjugates with amyloid-beta plaques.Conclusion: We successfully demonstrated that 89Zr-immuno-PET is suitable for imaging and quantifying amyloid-beta specific brain uptake using a bispecific aducanumab brain shuttling antibody, Adu-8D3, but only when using the novel chelator DFO*, and not DFO, for labeling with 89Zr.

Copper-mediated radioiodination reaction through aryl boronic acid or ester precursor and its application to direct radiolabeling of a cyclic peptide.

A copper-mediated radioiodination using aryl boronic precursors is attracting attention as a solution to oxidative iododestannylation and nickel-mediated radioiodination drawbacks. The copper-mediated radiolabeling method allows radioiodination at room temperature with stable aryl boronic precursors without preparing complex starting materials or reagents and can be performed in a reaction vessel exposed to air. This method has good potential in radiochemistry; however, studies on the scope of copper-mediated radioiodination through boronic precursors are insufficient. In particular, few reports have demonstrated the effect of protecting groups on radiolabeling efficiency. Therefore, the effect of the protecting group of aryl boronic acids on the copper-mediated radioiodination was investigated. In addition, this method, which does not require heating, is expected to be useful for direct radiolabeling of peptides. Thus, we attempted direct radioiodination of c(RGDyk) as an example. The resulting radioiodination method was well tolerated in various substrates and was unaffected by the pinacol ester-type protecting group. Also, c(RGDyk) was labeled with 125 I via copper-mediated radioiodination using an aryl boronic acid precursor. The reaction time and yield were improved, compared with the indirect method. Furthermore, the large difference in polarity between the boronic acid precursor and the radiolabeled compound facilitated purification.

A novel dipeptide coupled with pyrazine-oxadiazole derivative as a potential antitubercular agent: Synthesis, radioiodination and bioevaluation

Tuberculosis (TB) is a disease caused by Mycobacterium and usually attack the lung. Synthesis of new dipeptide derivatives attached to antitubercular active heterocyclic rings like pyrazine and 1,3,4-oxadiazole called ethyl 2-(2-(5-((pyrazin-2-ylamino) methyl)-1,3,4-oxadiazol-2-ylthio) acetamido) acetamido)-3-(4-hydroxyphenyl) propanoate (EPOGTP) and iodinated EPOGTP are reported. The compounds have been characterized by mass, FT-IR and 1H NMR spectroscopy. Their in vitro investigation against Mycobacterium tuberculosis cell line indicated good IC50value of 210 μg/ml for EPOGTP and 86 μg/ml for iodo-EPOGTP. For study the biodisriution, the direct radioiodination of EPOGTP with iodine-131 using mild oxidizing agent, N-Bromosuccinimide (NBS), was performed and optimized for obtaining the maximum radiochemical purity (97.3 ± 0.47%). Then, the in vivo biodistribution in healthy mice showed good accumulation of radioiodinated EPOGTP in lung of about 41.83 ± 0.23% (the percentage of injected dose per gram of organ) at 15 min post-injection. As a conclusion, the synthetized dipeptide and its iodinated derivative could be further evaluated as a potential antitubercular agents.

Radiolabeled IgG antibodies: Impact of various labels on neonatal Fc receptor binding.

The number of therapeutic antibodies in research and development as well as their complexity increases from year to year. Novel therapeutic protein formats, such as Fc-fusions, bispecific, or multivalent antibodies, are currently in preclinical and clinical development. Therefore, the need for biodistribution and imaging studies, eg, with radiolabeled proteins are very high. However, the labeling process or the label itself can have an impact on binding to cellular receptors, eg, to neonatal Fc receptor (FcRn), which can lead to altered PK properties compared with the unlabeled antibody. FcRn affinity chromatography allows the assessment of immunoglobulin G (IgG) samples with respect to their pH-dependent FcRn interaction. We analyzed IgGs with different types of labels, namely, direct iodination with 125 I; chelating agents, such as DOTA and DOTAM; and [3 H]propionate. Direct radio-iodination leads to shifts in FcRn column retention time, which might indicate a potentially faster clearance. Furthermore, high conjugation ratios of chelator lower the affinity to FcRn successively and thus may influence the lysosomal degradation of the antibody in endothelial cells. In contrast, IgGs labeled with [3 H]propionate did not show any timeshifts in FcRn affinity chromatography. This article is based on the oral presentation at the IIS 2018 Prague and highlights the importance of an affinity chromatography for characterization of potential changes in affinity to FcRn itself or charge and hydrophobicity.

Radiosynthesis of microtubule-targeted theranostic methyl N-[5-(3'-radiohalobenzoyl)-1H-benzimidazol-2-yl]carbamates.

Microtubules are a target for a broad spectrum of drugs used as chemotherapeutics to treat hematological malignancies and solid tumors. Most of these drugs have significant dose-limiting toxicities including peripheral neuropathies that can be debilitating and permanent. In an ongoing effort to develop safer and more effective drugs, benzimidazole-based compounds are being developed as replacement for vincristine and similar agents. In this report, we describe radiosyntheses of novel microtubule-targeting methyl N-[5-(3'-radiohalobenzoyl)-1H-benzimidazol-2-yl]carbamates 4 that are intended as potential imaging agents and molecular radiotherapeutics. 125 I- and 131 I-radiolabeled derivatives were prepared either by direct radioiodination of methyl N-(6-benzoyl-1H-benzimidazol-2-yl)carbamate 1 or radioiododestannylation of the corresponding stannane precursor 3. The direct radioiodination was conducted in a solution of 1 in triflic acid and produced after ~1hour at elevated temperatures and HPLC purification on average 62% of the no-carrier added products 125 I-4 and 131 I-4. Radioiododestannylation of 3'-trimethylstannane 3 proceeded with ease at room temperature in the presence of H2 O2 as the oxidant and produced no-carrier-added 125 I-4 and 131 I-4 in high isolated yields, on average 85%. The radiohalodestannylation protocol is universal and can be applied to other radiohalides including 124 I to produce 124 I-4, a positron emission tomography agent, and 211 At to produce 211 At-4, an α-particle emitting radiotherapeutic.

Radioiodinated Small-Molecule Tyrosine Kinase Inhibitor for HER2-Selective SPECT Imaging.

One of the most clinically relevant molecular aberrations in breast cancer is overexpression of human epidermal growth factor receptor type 2 (HER2). We aimed to develop a radiolabeled tyrosine kinase inhibitor for HER2-targeted breast cancer imaging. In this study, a radioiodinated analog (125/131I-IBA-CP) of the HER2-selective inhibitor CP724,714 was prepared and evaluated in HER2-positive or -negative subcutaneous human breast cancer xenografts. Methods: The CP724,714 analog IBA-CP was synthesized and assayed for its inhibitory activities against HER2 and 6 other tyrosine kinases. 125/131I-IBA-CP was prepared using a copper-mediated radioiodination method with enhanced labeling yield and molar activity. In vitro biologic activity, including specific and nonspecific binding of 131I-IBA-CP to its HER2 kinase target, was assessed in different cell lines. In vivo small-animal 125I-IBA-CP SPECT imaging and biodistribution studies were conducted on mice bearing HER2-positive, HER2-negative, or epidermal growth factor receptor (EGFR)-positive tumors. Nonradioactive IBA-CP and the EGFR inhibitor erlotinib were used as blocking agents to investigate the binding specificity and selectivity of 125/131I-IBA-CP toward HER2 in vitro and in vivo. Additionally, 125/131I-ICP was prepared by direct radioiodination of CP724,714 for comparison with 125/131I-IBA-CP. Results: IBA-CP displayed superior in vitro inhibitory activity (half-maximal inhibitory concentration, 16 nM) and selectivity for HER2 over 6 other cancer-related tyrosine kinases. 125/131I-IBA-CP was prepared in a typical radiochemical yield of about 65% (decay-corrected), radiochemical purity of more than 98%, and molar activity of 42 GBq/μmol at the end of synthesis. SPECT imaging revealed significantly higher uptake of 125I-IBA-CP than of 125I-ICP in the HER2-positive MDA-MB-453 tumors. Uptake in the HER2-negative MCF-7 tumors was much lower. Binding of 125I-IBA-CP in the MDA-MB-453 tumors was blocked by coinjection with an excess amount of IBA-CP, but not by erlotinib. Conclusion: The radiolabeled HER2-selective inhibitor 125/131I-IBA-CP is a promising probe for in vivo detection of HER2-positive tumors.

Dual-Modality Immuno-PET and Near-Infrared Fluorescence Imaging of Pancreatic Cancer Using an Anti-Prostate Stem Cell Antigen Cys-Diabody.

Pancreatic cancer has a high mortality rate due to late diagnosis and the tendency to invade surrounding tissues and metastasize at an early stage. A molecular imaging agent that enables both presurgery antigen-specific PET (immuno-PET) and intraoperative near-infrared fluorescence (NIRF) guidance might benefit diagnosis of pancreatic cancer, staging, and surgical resection, which remains the only curative treatment. Methods: We developed a dual-labeled probe based on A2 cys-diabody (A2cDb) targeting the cell-surface prostate stem cell antigen (PSCA), which is expressed in most pancreatic cancers. Maleimide-IRDye800CW was site-specifically conjugated to the C-terminal cys-tag (A2cDb-800) without impairing integrity or affinity (half-maximal binding, 4.3 nM). Direct radioiodination with 124I (124I-A2cDb-800) yielded a specific activity of 159 ± 48 MBq/mg with a radiochemical purity exceeding 99% and 65% ± 4.5% immunoreactivity (n = 3). In vivo specificity for PSCA-expressing tumor cells and biodistribution of the dual-modality tracer were evaluated in a prostate cancer xenograft model and compared with single-labeled 124I-A2cDb. Patient-derived pancreatic ductal adenocarcinoma xenografts (PDX-PDACs) were grown subcutaneously in NSG mice and screened for PSCA expression by immuno-PET. Small-animal PET/CT scans of PDX-PDAC-bearing mice were obtained using the dual-modality 124I-A2cDb-800 followed by postmortem NIRF imaging with the skin removed. Tumors and organs were analyzed ex vivo to compare the relative fluorescent signals without obstruction by other organs. Results: Specific uptake in PSCA-positive tumors and low nonspecific background activity resulted in high-contrast immuno-PET images. Concurrent with the PET studies, fluorescent signal was observed in the PSCA-positive tumors of mice injected with the dual-tracer 124I-A2cDb-800, with low background uptake or autofluorescence in the surrounding tissue. Ex vivo biodistribution confirmed comparable tumor uptake of both 124I-A2cDb-800 and 124I-A2cDb. Conclusion: Dual-modality imaging using the anti-PSCA cys-diabody resulted in high-contrast immuno-PET/NIRF images of PDX-PDACs, suggesting that this imaging agent might offer both noninvasive whole-body imaging to localize PSCA-positive pancreatic cancer and fluorescence image-guided identification of tumor margins during surgery.

In vitro evaluation of concentration, labeling effectiveness and stability for 131 I-labeled radioimmunoassay ligand using real-time detection technology.

Radioimmunoassay belongs to the analytical method enabling highly specific and sensitive quantification of molecules. The verification of the real-time radioimmunoassay technology usefulness for ligand-quality characteristics evaluation such as concentration, influence of radiolabeling on binding affinity and stability was estimated. The anti-epidermal growth factor receptor antibody 131 I-cetuximab was employed as the ligand antibody. The concentration of 131 I-cetuximab was derived from the shape of binding curves coming from the ligand-receptor interaction. The binding curves also allowed the estimation of 131 I-cetuximab binding affinity for different radiolabeling procedures (incubation times 1, 5, and 10minutes) in stability testing up to 96hours at 4°C. The stability testing also included comparative analysis by size exclusion high-performance liquid chromatography. The assessment of cetuximab concentrations using real-time method showed acceptable accordance between real and calculated values. The real-time method revealed that 1-minute radiolabeling proved to be the optimal incubation time for direct radioiodination of cetuximab. Stability testing showed the significant change in radioligand affinity by one order at the longest incubation times (72 and 96hours). Characterization of stability and binding behavior of radiolabeled monoclonal antibodies by the verified real-time method before use in other assays may be employed to eliminate variability and suboptimal antibody performance.

Radioiodination and Biological Evaluation of Epinephrine as a Possible Adrenergic Receptors Imaging Agent

An adopted method for the preparation of high radiochemical purity [ 125 I] iodoepinephrine was developed in order to characterize the binding properties of adrenergic receptors. Direct radioiodination of epinephrine was carried out using chloramine-T as oxidizing agent. The reaction proceeds well within 15 min at ambient room temperature up to 25+1oC and afforded a radiochemical yield up to 94%. Different chromatographic techniques (electrophoresis and pap er chromatography) were used to evaluate the radiochemical yield and purity of the labeled product. Biodistribution studies were carried out in normal Albino Swiss mice and the results showed rapid and high cardiac uptake of 125 I-epinephrine. The result in dicates the possibility of using radioiodinated epinephrine as myocardial imaging agent.

Preparation and biodistribution of radioiodinated analgin as a possible inflammatory imaging agent

Direct radioiodination of analgin with 125I was carried out using chloramine-T as an oxidizing agent. The labeling conditions were optimized. When performed with 50 μg of analgin and 10 μg of chloramine-T at pH 9 for 15 min at room temperature, the reaction gives 97.5 ± 1.2% radiochemical yield of 125I-analgin. The radiochemical yield of the final product was determined by electrophoresis and HPLC. Biodistribution studies were carried out in normal Albino Swiss mice and in those with septic and aseptic inflammations at different time intervals after administration of radioiodinated analgin. It was found that radioiodinated analgin differentiates between septic and aseptic inflammations and can be used to detect inflammation.

Selective mono-radioiodination and characterization of a Cell-Penetrating Peptide: L-Tyr-Maurocalcine

Abstract Mono-and poly- iodinated peptides form frequently during radioiodination procedures. However, the formation of a single species in its mono-iodinated form is essential for quantitative studies such as determination of tissue concentration or image quantification. Therefore, the aim of the present study was to define the optimal experimental conditions in order to exclusively obtain the mono-iodinated form of L-maurocalcine (L-MCa). L-MCa is an animal venom toxin which was shown to act as a cell-penetrating peptide. In order to apply the current direct radioiodination technique using oxidative agents including chloramine T, Iodo-Gen® or lactoperoxidase, an analogue of this peptide containing a tyrosine residue (Tyr-L-MCa) was synthesized and was shown to fold/oxidize properly. The enzymatic approach using lactoperoxidase/H2O2 was found to be the best method for radioiodination of Tyr-L-MCa. MALDI-TOF mass spectrometry analyses were then used for identification of the chromatographic eluting components of the reaction mixtures. We observed that the production of different radioiodinated species depended upon the reaction conditions. Our results successfully described the experimental conditions of peptide radioiodination allowing the exclusive production of the mono-iodinated form with high radiochemical purity and without the need for a purification step. Mono-radioiodination of L-Tyr-MCa will be crucial for future quantitative studies, investigating the mechanism of cell penetration and in vivo biodistribution.

Improved Tumor Targeting of Anti-HER2 Nanobody Through N-Succinimidyl 4-Guanidinomethyl-3-Iodobenzoate Radiolabeling

Nanobodies are approximately 15-kDa proteins based on the smallest functional fragments of naturally occurring heavy chain-only antibodies and represent an attractive platform for the development of molecularly targeted agents for cancer diagnosis and therapy. Because the human epidermal growth factor receptor type 2 (HER2) is overexpressed in breast and ovarian carcinoma, as well as in other malignancies, HER2-specific Nanobodies may be valuable radiodiagnostics and therapeutics for these diseases. The aim of the present study was to evaluate the tumor-targeting potential of anti-HER2 5F7GGC Nanobody after radioiodination with the residualizing agent N-succinimidyl 4-guanidinomethyl 3-(125/131)I-iodobenzoate (*I-SGMIB). The 5F7GGC Nanobody was radiolabeled using *I-SGMIB and, for comparison, with N(ε)-(3-*I-iodobenzoyl)-Lys(5)-N(α)-maleimido-Gly(1)-GEEEK (*I-IB-Mal-d-GEEEK), another residualizing agent, and by direct radioiodination using IODO-GEN ((125)I-Nanobody). The 3 labeled Nanobodies were evaluated in affinity measurements, and paired-label internalization assays were performed on HER2-expressing BT474M1 breast carcinoma cells and in paired-label tissue distribution measurements in mice bearing subcutaneous BT474M1 xenografts. *I-SGMIB-Nanobody was produced in 50.4% ± 3.6% radiochemical yield and exhibited a dissociation constant of 1.5 ± 0.5 nM. Internalization assays demonstrated that intracellular retention of radioactivity was up to 1.5-fold higher for *I-SGMIB-Nanobody than for coincubated (125)I-Nanobody or *I-IB-Mal-d-GEEEK-Nanobody. Peak tumor uptake for *I-SGMIB-Nanobody was 24.50% ± 9.89% injected dose/g at 2 h, 2- to 4-fold higher than observed with other labeling methods, and was reduced by 90% with trastuzumab blocking, confirming the HER2 specificity of localization. Moreover, normal-organ clearance was fastest for *I-SGMIB-Nanobody, such that tumor-to-normal-organ ratios greater than 50:1 were reached by 24 h in all tissues except lungs and kidneys, for which the values were 10.4 ± 4.5 and 5.2 ± 1.5, respectively. Labeling anti-HER2 Nanobody 5F7GGC with *I-SGMIB yields a promising new conjugate for targeting HER2-expressing malignancies. Further research is needed to determine the potential utility of *I-SGMIB-5F7GGC labeled with (124)I, (123)I, and (131)I for PET and SPECT imaging and for targeted radiotherapy, respectively.

Radioiodination and biological evaluation of salbutamol as a β2-adrenoceptor agonist

A procedure for preparing high-radiochemical-purity 125I-salbutamol was developed in order to characterize the binding properties of β2-adrenoceptors. Direct radioiodination of salbutamol was carried out using Chloramine T as an oxidizing agent. The optimized conditions for salbutamol labeling with 125I are as follows: 100 μg of salbutamol, 50 μg of Chloramine T, 150 μl of phosphate buffer (pH 7), room temperature (25 ± 1°C), 5 min. The radiochemical yield of up to 98% was attained. The radiochemical yield and purity of the labeled product were evaluated by electrophoresis and HPLC. Biodistribution studies were carried out in normal Albino Swiss mice at different time intervals after administration of 125I-salbutamol. The results indicate that the labeled compound cleared from the systematic circulation within 3 h after administration and majority of organs showed significant decrease in the 125I-salbutamol uptake. The heart and lung uptake was high. 125I-Salbutamol can be used as β-receptor imaging agent.

Recombinant fragment of an antibody tailored for direct radioiodination

Recombinant fragment of an antibody tailored for direct radioiodination

Labeling of atenolol with radioactive iodine-125 using N-bromosuccinimide and hydrogen peroxide as oxidizing agents

An adopted method for the preparation of high radiochemical purity 125I-atenolol was investigated. Direct radioiodination of atenolol was carried out using N-bromosuccinamide or hydrogen peroxide as an oxidizing agent. The reaction proceeds well within 30 min at room temperature (25 ± 1 °C) and afforded a radiochemical yield up to 97% as pure as 125I-atenolol. Different chromatographic techniques (electrophoresis, TLC and HPLC) were used to determine the radiochemical yield and purity of the labeled product. Biodistribution studies were carried out in normal Albino Swiss mice and the results indicate that 125I-atenolol can be used safely as myocardial imaging agent.

The use of closo-dodecaborate-containing linker improves targeting of HNSCC xenografts with radioiodinated chimeric monoclonal antibody U36

Radionuclide imaging of head and neck squamous cell carcinoma (HNSCC) using monoclonal antibodies (MAbs) has the potential to contribute to improved diagnosis and staging, thereby making more effective treatment possible. Chimeric monoclonal antibody U36 (cMAb U36), specific to CD44v6 antigen, is a candidate for the targeting of HNSCC. The aim of this study was to compare the influence of indirect iodination via closo-dodecaborate-based linker (DABI) with the influence of direct radioiodination on the biodistribution of the chimeric anti-CD44v6 antibody U36. The study was performed using nude mice bearing UT-SCC7 HNSCC xenografts using the paired-label method. The biodistribution of cMAb U36 labelled directly with 131I and using DABI with 125I was compared in the same animals. The influence of DABI on the tumour-to-organ ratio was evaluated. For both conjugates, radioactivity uptake in blood and organs decreased with time, except in tumours and the thyroid. DABI-labelled cMAb U36 was characterised by fast blood clearance and an elevated uptake in the liver and spleen. The use of DABI enabled a 1.5 to 2-fold improvement in the tumour-to-blood and tumour-to-organ ratios in comparison with direct radioiodination, with the exception of the liver and spleen. These results indicate that DABI is a promising linker for the coupling of radioiodine to HNSCC-targeting antibodies.