Chelating Agent DOTA Research Articles

A Novel Small Cyclic Peptide-Based 68Ga-Radiotracer for Positron Emission Tomography Imaging of PD-L1 Expression in Tumors.

In the tumor microenvironment, programmed death protein 1 and programmed death protein ligand 1 (PD-L1) signaling pathways help tumors escape the immune system. We designed a gallium-68 (68Ga)-labeled small-molecule peptide-targeting PD-L1 and used positron emission tomography/computed tomography (PET/CT) to detect and dynamically monitor the expression level of PD-L1 in tumors. S-Cyclo(ETSK)-SF-NH2 (SETSKSF) is a cyclic peptide inhibitor comprising seven amino-acid residues. We connected it with the chelating agent DOTA, labeled DOTA-SETSKSF, with the short half-life nuclide Ga-68, and measured the stability of 68Ga-2,2',2″-(10-(2-((S)-1-((3S,6S,9S,18S)-18-((S)-1-((S)-1-amino-1-oxo-3-henylpropan-2-ylamino)-3-hydroxy-1-oxopropan-2-ylcarbamoyl)-6-((R)-1-hydroxyethyl)-3-(hydroxymethyl)-2,5,8,12-tetraoxo-1,4,7,13-tetraazacyclooctadecan-9-ylamino)-3-ydroxy-1-oxopropan-2-ylamino)-2-oxoethyl)-1,4,7,10-tetraazacyclododecane-1,4,7-triyl)triacetic acid (68Ga-DOTA-SETSKSF) in normal saline (NS), phosphate-buffered saline (PBS), and fetal bovine serum (FBS) in vitro. We conducted the 68Ga-DOTA-SETSKSF affinity test, cell-specific uptake experiments, time-combined experiments, western blotting, and laser confocal experiments to confirm the expression and localization of PD-L1 at the cell level and determine the uptake. Biodistribution and imaging experiments were performed using the H1975, B16F10, and A549 tumor models. 68Ga-DOTA-SETSKSF was successfully synthesized, and the radiochemical purity was >99% after purification. The in vitro stability of 68Ga-DOTA-SETSKSF was >95% in NS, PBS, and FBS at 37 °C after 4 h of incubation. Cell-binding experiments confirmed that 68Ga-DOTA-SETSKSF exhibited high uptake in H1975 tumors with high PD-L1 expression and low uptake in A549 tumors with low PD-L1 expression. The clear half-life (T1/2) of 68Ga-DOTA-SETSKSF from the blood was 14.48 ± 3.26 min. The percentages of the injected dose per gram of tissue (%ID/g) for H1975 and A549 tumors were 5.29 ± 0.21 and 0.89 ± 0.10 at 1 h after injection, respectively. The H1975 tumor-to-muscle and tumor-to-blood ratios were 41.79 ± 5.81 and 4.75 ± 0.19 at 4 h, respectively. Apart from the H1975 tumor, the kidney and the bladder showed high accumulation because 68Ga-DOTA-SETSKSF was excreted through the urinary system. PET/CT images showed high accumulation of 68Ga-DOTA-SETSKSF in H1975 tumors and low uptake in A549 tumors, which was consistent with the results of biodistribution experiments. 68Ga-DOTA-SETSKSF is convenient to prepare, has high stability, can be used to monitor the expression of PD-L1, and has an extremely high clinical application value.

Preparation and in vitro evaluation of radiolabeled HA-PLGA nanoparticles as novel MTX delivery system for local treatment of rheumatoid arthritis

Radiosynovectomy is a technique used to decrease inflammation of the synovial tissue by intraarticular injection of a β-emitting radionuclide, such as 177Lu, which is suitable for radiotherapy due to its decay characteristics. Drug-encapsulating nanoparticles based on poly lactic‑co‑glycolic acid (PLGA) polymer are a suitable option to treat several arthritic diseases, used as anti-inflammatory drugs transporters of such as methotrexate (MTX), which has been widely used in the arthritis treatment (RA), and hyaluronic acid (HA), which specifically binds the CD44 and hyaluronan receptors overexpressed on the inflamed synovial tissue cells. The 1,4,7,10‑Tetraazacyclododecane‑1,4,7,10‑tetraacetic acid (DOTA) was used as complexing agent of Lutetium-177 for radiotherapy porpoises. The aim of this research was to synthesize 177Lu-DOTA-HA-PLGA(MTX) as a novel, smart drug delivery system with target-specific recognition, potentially useful in radiosynovectomy for local treatment of rheumatoid arthritis. The polymeric nanoparticle system was prepared and chemically characterized. The MTX encapsulation and radiolabelling were performed with suitable characteristics for its in vitro evaluation. The HA-PLGA(MTX) nanoparticle mean diameter was 167.6 nm ± 57.4 with a monomodal and narrow distribution. Spectroscopic techniques demonstrated the effective conjugation of HA and chelating agent DOTA to the polymeric nanosystem. The MTX encapsulation was 95.2% and the loading efficiency was 6%. The radiochemical purity was 96 ± 2%, determined by ITLC. Conclusion: 177Lu-DOTA-HA-PLGA(MTX) was prepared as a biocompatible polymeric PLGA nanoparticle conjugated to HA for specific targeting. The therapeutic nanosystem is based on bi-modal mechanisms using MTX as a disease-modifying antirheumatic drug (DMARD) and 177Lu as a radiotherapeutic component. The 177Lu-DOTA-HA-PLGA(MTX) nanoparticles showed properties suitable for radiosynovectomy and further specific targeted anti-rheumatic therapy.

A new microdispersed albumin derivative potentially useful for radio-guided surgery of occult breast cancer lesions

This paper describes a new nuclear imaging agent, 2-(4-isothiocyanatobenzyl)−1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid of human albumin (HAC), potentially suitable for application in the Radio-guided Occult Lesion Localization (ROLL) of non-palpable mammalian cancerous lesions, as a tool to overtake the short radio-signal half-life of the technetium-99m based radiopharmaceutical currently used. This conjugate is a microsized powder aggregate, water-insoluble between pH 3 and 8.5, obtained by conjugating the protein with the macrocyclic chelating agent DOTA through a one-pot reaction in aqueous medium. The product has been fully characterized and is stable to the thermal conditions adopted for labeling; after radiolabeling with longer half-life radionuclides such as 177Lu or 111In, it has shown radiochemical purity (RCP) >90% and resulted stable when stored in saline or plasma for 6 days at 37 °C. A μPET/CT study, performed in vivo on adult female rats, showed that the radioactivity of HAC labeled with 64Cu remained located in the mammary glands for at least 40 h, without diffusion or drainage in healthy tissues or in the lymphatic circulation. This new imaging agent might make the ROLL procedure more accessible, safe and flexible, promoting a significant time and cost reduction of this intervention. Moreover, HAC might also be used in other radio-guided surgical procedures in oncology.

Ultrasmall graphene oxide based T1 MRI contrast agent for in vitro and in vivo labeling of human mesenchymal stem cells

Herein, we report on development of a two-dimensional nanomaterial graphene oxide (GO)-based T1 magnetic resonance imaging (MRI) contrast agent (CA) for in vitro and in vivo labeling of human mesenchymal stem cells (hMSCs). The CA was synthesized by PEGylation of ultrasmall GO, followed by conjugation with a chelating agent DOTA and then gadolinium(III) to form GO-DOTA-Gd complexes. Thus-prepared GO-DOTA-Gd complexes exhibited significantly improved T1 relaxivity, and the r1 value was 14.2 mM−1s−1 at 11.7 T, approximately three times higher than Magnevist, a commercially available CA. hMSCs can be effectively labeled by GO-DOTA-Gd, leading to remarkably enhanced cellular MRI effect without obvious adverse effects on proliferation and differentiation of hMSCs. More importantly, in vivo experiment revealed that intracranial detection of 5×105 hMSCs labeled with GO-DOTA-Gd is achieved. The current work demonstrates the feasibility of the GO-based T1 MRI CA for stem cell labeling, which may find potential applications in regenerative medicine.

Indium 111-labeled J591 anti-PSMA antibody for vascular targeted imaging in progressive solid tumors.

BackgroundJ591 is a monoclonal antibody that targets the external domain of the prostate-specific membrane antigen (PSMA). Besides prostate cancer cells, it also targets the neovasculature of non-prostate solid tumors. We provide an analysis of the antibody mass-dose dependency of lesion uptake and normal tissue retention, together with an assessment of lesion detectability using 111In-J591 imaging, compared with conventional imaging in patients with a variety of solid tumors.MethodsTwenty patients in six cohorts received fixed amounts (5, 10, 20, 40, 60, and 100 mg) of J591 in a phase I trial. A maximum of four administrations per patient was given, with each administration separated by 3 weeks. All antibody administrations included 370 MBq (10 mCi) of 111In labeled to 2 mg of J591 via the chelating agent DOTA. Three whole body (WB) gamma camera scans with at least one SPECT scan, along with multiple WB count-rate measurements and blood samples, were obtained for all patients. The effect of escalating antibody mass on lesion uptake and normal tissue retention was evaluated using lesion, liver, serum, and WB residence times and ratios thereof for each treatment cycle. Lesion detectability using 111In-J591 imaging was compared to the standard imaging on a lesion-by-lesion basis.ResultsA total of 170 lesions in 20 patients were detected by standard or 111In-J591 imaging. 111In-J591 targeted both skeletal and soft tissue diseases in all tumor types. 111In-J591 imaging identified 74% (20/27) of skeletal lesions, 53% (18/34) of nodes, and 64% (70/109) of other soft tissue/organ lesions. There was increasing 111In-J591 uptake in lesions with increasing antibody mass-dose, coupled with decreasing retention in the liver for increments up to 20 mg, and no significant change at higher antibody mass.ConclusionsRadiolabeled J591 antibody has potential as a targeting agent for solid tumor vasculature and lesion detection. Bone and soft tissue lesions arising from tumors of diverse origin were targeted by the anti-PSMA antibody J591. For the detection of lesions in these tumors by J591 antibody scans, an antibody mass of 20 mg is adequate. The optimal time of imaging is 5 to 7 days post-injection.

A novel PET imaging using ⁶⁴Cu-labeled monoclonal antibody against mesothelin commonly expressed on cancer cells.

Mesothelin (MSLN) is a 40-kDa cell differentiation-associated glycoprotein appearing with carcinogenesis and is highly expressed in many human cancers, including the majority of pancreatic adenocarcinomas, ovarian cancers, and mesotheliomas, while its expression in normal tissue is limited to mesothelial cells lining the pleura, pericardium, and peritoneum. Clone 11-25 is a murine hybridoma secreting monoclonal antibody (mAb) against human MSLN. In this study, we applied the 11-25 mAb to in vivo imaging to detect MSLN-expressing tumors. In in vitro and ex vivo immunochemical studies, we demonstrated specificity of 11-25 mAb to membranous MSLN expressed on several pancreatic cancer cells. We showed the accumulation of Alexa Fluor 750-labeled 11-25 mAb in MSLN-expressing tumor xenografts in athymic nude mice. Then, 11-25 mAb was labeled with 64Cu via a chelating agent DOTA and was used in both in vitro cell binding assay and in vivo positron emission tomography (PET) imaging in the tumor-bearing mice. We confirmed that 64Cu-labeled 11-25 mAb highly accumulated in MSLN-expressing tumors as compared to MSLN-negative ones. The 64Cu-labeled 11-25 mAb is potentially useful as a PET probe capable of being used for wide range of tumors, rather than 18F-FDG that occasionally provides nonspecific accumulation into the inflammatory lesions.

Optimized solid phase-assisted synthesis of dendrons applicable as scaffolds for radiolabeled bioactive multivalent compounds intended for molecular imaging.

Dendritic structures, being highly homogeneous and symmetric, represent ideal scaffolds for the multimerization of bioactive molecules and thus enable the synthesis of compounds of high valency which are e.g., applicable in radiolabeled form as multivalent radiotracers for in vivo imaging. As the commonly applied solution phase synthesis of dendritic scaffolds is cumbersome and time-consuming, a synthesis strategy was developed that allows for the efficient assembly of acid amide bond-based highly modular dendrons on solid support via standard Fmoc solid phase peptide synthesis protocols. The obtained dendritic structures comprised up to 16 maleimide functionalities and were derivatized on solid support with the chelating agent DOTA. The functionalized dendrons furthermore could be efficiently reacted with structurally variable model thiol-bearing bioactive molecules via click chemistry and finally radiolabeled with 68Ga. Thus, this solid phase-assisted dendron synthesis approach enables the fast and straightforward assembly of bioactive multivalent constructs for example applicable as radiotracers for in vivo imaging with Positron Emission Tomography (PET).

High‐resolution animal PET imaging of prostate cancer xenografts with three different 64Cu‐labeled antibodies against native cell‐adherent PSMA

The prostate specific membrane antigen (PSMA) is expressed by virtually all prostate cancers and represents an ideal target for diagnostic and therapeutic strategies. This article compares the in vivo behavior and tumor uptake of three different radiolabeled anti-PSMA monoclonal antibodies (mAbs) and corresponding F(ab)(2) and Fab fragments thereof. The mAbs 3/A12, 3/F11, and 3/E7 and fragments of 3/A12 were conjugated with the chelating agent DOTA and radiolabeled with 64Cu. For the microPET imaging studies, SCID mice bearing PSMA-positive C4-2 and PSMA-negative DU 145 prostate cancer xenografts were used. Each animal received 20-30 microg radiolabeled mAb or fragment corresponding to an activity of 8-14 MBq. Imaging was performed 3, 24, and 48 hr post-injection. After the last scan, mice were sacrificed and tracer in vivo biodistribution was measured by gamma-counting. Static microPET images of mice with PSMA-positive tumors revealed a high uptake of the mAbs in the C4-2 tumors at 24 and 48 hr after tracer injection and only a minimal distribution in the DU 145 tumors and other organs. In contrast, the F(ab)(2) and Fab fragments of 3/A12 were detected at a high extend in the kidney but not in the C4-2 tumors. These results were confirmed by gamma counting of dissected organs after the final imaging. Due to the high and specific uptake of the 64Cu-labeled mAbs in PSMA-positive tumors, these antibodies represent excellent tools for prostate cancer imaging.

Antibody Mass Escalation Study in Patients with Castration-Resistant Prostate Cancer Using 111In-J591: Lesion Detectability and Dosimetric Projections for 90Y Radioimmunotherapy

J591, a monoclonal antibody that targets the external domain of the prostate-specific membrane antigen, has potential as an agent for radioimmunotherapy. A pilot trial was performed in patients with prostate cancer using repetitive administrations of escalating masses of J591. An analysis was performed to assess lesion detectability by (111)In-J591 gamma-camera imaging compared with standard imaging methods and the effect of increasing antibody mass on lesion detectability, biodistribution, and dosimetry. Fourteen patients with metastatic prostate cancer received escalating amounts (10, 25, 50, and 100 mg) of J591 in a series of administrations each separated by 3 wk. All antibody administrations included a fixed amount of the radiolabeled antibody (111)In-1,4,7,10-tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid-J591 ((111)In-DOTA-J591) (2 mg of J591 labeled with 185 MBq [5 mCi] of (111)In via the chelating agent DOTA). Three whole-body gamma-camera scans with at least 1 SPECT scan together with multiple whole-body counting-rate measurements and serum activity-concentration measurements were obtained in all patients. Images were analyzed for distribution and lesion targeting. Estimates of clearance rates and liver and lesion uptake were made for each treatment cycle. These estimates were used to generate dosimetric projections for radioimmunotherapy with (90)Y-labeled J591. A total of 80 lesions in 14 patients were detected. Both skeletal and soft-tissue diseases were targeted by the antibody as seen on (111)In-J591 scans. The antibody localized to 93.7% of skeletal lesions detected by conventional imaging. Clearance of radioactivity from the whole body, serum, and liver was dependent on antibody mass. Normalized average values of the ratio of residence times between lesion and liver for 10, 25, 50, and 100 mg of antibody were 1.0, 1.9, 3.2, and 4.0. Dosimetric projections for radioimmunotherapy with (90)Y-labeled J591 suggested similar absorbed doses to lesions for treatment at the maximally tolerated activity (MTA), irrespective of antibody mass. However, absorbed doses to liver at the MTA would be antibody mass-dependent with estimates of 20, 10, 7, and 5 Gy for 10, 25, 50, and 100 mg of J591. The proportion of the amount of antibody increased in lesions and decreased in the liver with increasing mass of administered antibody up to a dose of 50 mg. Proportional hepatic uptake continued to decrease with increasing antibody mass up to 100 mg. The optimal antibody mass for radioimmunotherapy would therefore appear to be greater than or equal to 50 mg.

90Y-DOTA-Lanreotide: A potential agent for targeted therapy

Several human tumors such as neuroendocrine tumors, medullary thyroid carcinoma, etc., express somatostatin receptors which specifically bind somatostatin and its analogues such as lanreotide, octreotide, etc. In order to prepare a therapeutic agent for targeting such tumors, attempts were made to prepare 90Y-DOTA-Lanreotide. Lanreotide could be successfully conjugated with the macrocyclic chelating agent DOTA (1,4,7,10-tetraaza cyclododecane tetracetic acid) which forms stable complexes with 90Y. 90Y-DOTA-Lanreotide could be prepared in >98% radiochemical purity and remained stable for 72 hours at room temperature. The tracer showed specific binding to A431 cells. Biodistribution studies in C57BL6 mice bearing melanoma showed ∼1.3% uptake pergram of tumor at 24-hour p.i.

111In]‐DOTA‐Lys8‐Vasotocin: A new promising peptide for oxytocin receptor expressing tumors

AbstractOxytocin receptors (OTR) are expressed by a large variety of human tumors. We have developed a new reagent specifically designed to transfer radionuclides to OTR‐expressing tumors. We selected Lys8‐vasotocin (LVT), an analogue of oxytocin (OT), with high affinity for OTR and coupled the chelating agent DOTA. The new reagent (DOTA‐LVT) was radiolabeled with 111In and showed specific binding to OTR positive cell lines. In vivo experiments demonstrated the receptor mediated binding of 111In‐DOTA‐LVT to OTR expressing tumors. The present study has shown that the new reagent may be promising to convey radioisotopes to OTR positive tumors although further improvements are warranted.

Biodistribution and radioimmunotherapy of human breast cancer xenografts with radiometal-labeled DOTA conjugated anti-HER2/neu antibody 4D5.

HER2/neu oncogene encodes a 185 kDa trans-membrane protein which is overexpressed in 20-30% of breast and ovarian cancers and portends a poor prognosis. We have studied the targeting and therapy of this oncoprotein with 4D5, a murine monoclonal antibody which recognizes a distinct epitope on the extracelluar domain of HER2/neu. We conjugated the antibody with an active ester of the macrocyclic chelating agent DOTA, radiolabeled the conjugate with either (111)In or (90)Y, and studied the antibody distribution and therapy, respectively, in athymic mice bearing xenografts of MCF7/HER2/neu, a human breast cancer cell line transfected with the HER2/neu oncogene. For the biodistribution of (111)In-labeled DOTA-4D5, a high specificity of tumor localization (30% ID/g) was seen with a tumor-to-blood ratio of greater than 2 at 48 h postinjection. Compared to a previously published study with (125)I-labeled 4D5 in beige nude mice bearing NIH3T3/HER2/neu xenografts [De Santes et al. (1992) Cancer Res. 52, 1916-1923], (111)In-labeled 4D5 antibody gave superior antibody uptake in tumor (30% ID/g vs 17% ID/g at 48h). In the therapy study, treatment of the nude mice bearing MCF7/HER2/neu xenografts with 100 microCi (3 microg) of (90)Y-labeled DOTA-4D5 caused a 3-fold reduction of tumor growth compared to untreated controls (injected with human serum albumin) in 40 days. Treatment of animals with 100 microCi of nonspecific antibody (90)Y-labeled DOTA-Leu16 (3 microg) had no tumor growth inhibition. Treatment with unlabeled DOTA-4D5 (3 microg) had a slight effect on tumor growth compared to untreated controls. When analyzed at the level of single animals, no effect was seen in seven of nine animals; however, in two of the animals, tumor growth inhibition was observed. Although a cold antibody therapeutic effect was unexpected at this dose level (3 microg), it may be possible that in some animals that 3 microg of antibody of (90)Y-labeled DOTA-4D5 augmented tumor growth reduction. To further explore the effects of cold antibody treatment alone, animals were treated with 100 or 400 microg of unlabeled 4D5 administered in two doses. These animals showed a 1.7-1.8-fold reduction in tumor growth over 28 days, a result less than that obtained with RIT only.

Preparation and biodistribution of 111Indium and 90Yttrium labelled immunoconjugates incorporating the macrocyclic chelating agent DOTA—laboratory data : D.E. Snook, G. Rowlinson-Busza, C. Meares and A.A. Epenetos ICRF Oncology Group, Hammersmith Hospital, London, UK and Dept of Chemistry, University of California, Davis, California, USA

Preparation and biodistribution of 111Indium and 90Yttrium labelled immunoconjugates incorporating the macrocyclic chelating agent DOTA—laboratory data : D.E. Snook, G. Rowlinson-Busza, C. Meares and A.A. Epenetos ICRF Oncology Group, Hammersmith Hospital, London, UK and Dept of Chemistry, University of California, Davis, California, USA