Positron emission tomography (PET) imaging of neuroblastoma and melanoma with 64 Cu-SarAr immunoconjugates (original) (raw)
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Bioconjugate Chemistry, 2008
Radiolabeled anti-carcinoembryonic antigen (CEA) antibodies have the potential to give excellent images of a wide variety of human tumors, including tumors of the colon, breast, lung, and medullar thyroid. In order to realize the goals of routine and repetitive clinical imaging with anti-CEA antibodies, it is necessary that the antibodies have high affinity for CEA, low cross reactivity and uptake in normal tissues, and low immunogenicity. The humanized anti-CEA antibody hT84.66-M5A (M5A) fulfills these criteria with an affinity constant >10 10 M −1 , no reactivity with CEA crossreacting antigens found in normal tissues, and >90% human protein sequence. A further requirement for routine clinical use of radiolabeled antibodies is a versatile method of radiolabeling that allows the use of multiple radionuclides that differ in their radioemissions and half-lives. We describe a versatile bifunctional chelator, DO3A-VS (1, 4, 7-tris(carboxymethyl)-10-(vinylsulfone)-1, 4, 7, 10tetraazacyclododecane) that binds a range of radiometals including 111 In for gamma-ray imaging and 64 Cu for Positron Emission Tomography (PET), and which can be conjugated with negligible loss of immunoreactivity either to sulfhydryls (SH) in the hinge region of lightly reduced immunoglobulins or surface lysines (NH) of immunoglobulins.
Comparison of bifunctional chelates for 64Cu antibody imaging
European Journal of Nuclear Medicine and Molecular Imaging, 2010
Purpose Improved bifunctional chelates (BFCs) are needed to facilitate efficient 64 Cu radiolabeling of monoclonal antibodies (mAbs) under mild conditions and to yield stable, target-specific agents. The utility of two novel BFCs, 1-Oxa-4,7,10-triazacyclododecane-5-S-(4-isothiocyanatobenzyl)-4,7,10-triacetic acid (p-SCN-Bn-Oxo-DO3A) and 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1 (15),11,13-triene-4-S-(4-isothiocyanatobenzyl)-3,6,9-triacetic acid (p-SCN-Bn-PCTA), for mAb imaging with 64 Cu were compared to the commonly used S-2-(4-isothiocyanatobenzyl)-1,4,7,10-tetraazacyclododecane-tetraacetic acid (p-SCN-Bn-DOTA). Methods The BFCs were conjugated to trastuzumab, which targets the HER2/neu receptor. 64 Cu radiolabeling of the conjugates was optimized. Receptor binding was analyzed using flow cytometry and radioassays. Finally, PET imaging and biodistribution studies were done in mice bearing either HER2/neu-positive or HER2/neu-negative tumors.
64Cu-p-NH2-Bn-DOTA-hu14.18K322A, a PET Radiotracer Targeting Neuroblastoma and Melanoma
Journal of Nuclear Medicine, 2012
The hu14.18K322A variant of the GD2-targeting antibody hu14.18 has been shown to elicit a level of antibody-dependent cellmediated cytotoxicity toward human neuroblastoma cells similar to that of the parent antibody. However, hu14.18K322A exhibited a decreased complement activation and associated pain, the dose-limiting toxicity in neuroblastoma immunotherapy. PET with a radiolabeled analog of the same antibody used in treatment will provide insight into the ability of hu14.18K322A to reach its target, as well as nontarget uptake that may cause side effects. Such antibody radiotracers might also provide a method for measuring GD2 expression in tumors, thus enabling the prediction of response to anti-GD2 therapy for individual patients. Methods: The conjugation of hu14.18K322A with p-NH 2 -Bn-DOTA was accomplished using N-(3-dimethylaminopropyl)-N9-ethylcarbodiimide with subsequent 64 Cu radiolabeling at 37°C for 30 min. Immunoreactivity of the conjugate was assessed by a dose-escalation blocking experiment measuring binding to purified GD2 versus GD1b as a negative control. Cell uptake and biodistribution studies in M21 (GD2-positive) and PC-3 (GD2-negative) tumor models were performed, as was small-animal PET/CT of M21 and PC-3 tumor-bearing mice. Results: The labeling of 64 Cu-p-NH 2 -Bn-DOTA-hu14.18K322A was achieved at more than 95% radiochemical purity and a specific activity of 127-370 MBq/mg (3.4-10 mCi/mg) after chromatographic purification. Preliminary in vitro data demonstrated a greater than 6-fold selectivity of binding to GD2 versus GD1b and dose-dependent inhibition of binding by unmodified hu14.8K322A. In vivo data, including small-animal PET/CT, showed significant GD2-positive tumor-targeting ability, with a persistent 2-fold-higher uptake of radiotracer than in GD2-negative tumors. Conclusion: 64 Cu-p-NH 2 -Bn-DOTA-hu14.18K322A represents a novel PET radiotracer to facilitate clinical investigations of anti-GD2 immunotherapies and to complement other imaging modalities in the staging and treatment of neuroblastoma.
Inorganic chemistry, 2016
Bifunctional chelators play an important role in developing metallic radionuclide-based radiopharmaceuticals. In this study, a new bifunctional ligand, p-SCN-PhPr-NE3TA, was synthesized and conjugated to a very late antigen-4 targeting peptidomimetic, LLP2A, for evaluating its application in (64)Cu-based positron emission tomography (PET) imaging. The new ligand exhibited strong selective coordination of Cu(II), leading to a robust Cu complex, even in the presence of 10-fold Fe(III). The LLP2A conjugate of p-SCN-PhPr-NE3TA was prepared and successfully labeled with (64)Cu under mild conditions. The conjugate (64)Cu-NE3TA-PEG4-LLP2A showed significantly higher specific activity, compared with (64)Cu-NOTA-PEG4-LLP2A, while maintaining comparable serum stability. Subsequent biodistribution studies and PET imaging in mice bearing B16F10 xenografts confirmed its favorable in vivo performance and high tumor uptake with low background, rendering p-SCN-PhPr-NE3TA a promising bifunctional ch...
RSC Advances, 2017
is a monopicolinate cyclam previously presented as a better 64 Cu chelator than dota, nota and other chelators with an improved biodistribution and in vivo resistance to transchelation. This study aimed to determine whether te1pa could improve the in vivo stability of 64 Cu chelation concerning radioimmunoconjugates in order to obtain better contrast in PET imaging. te1pa was activated on its remaining acid function to obtain a N-hydroxysulfosuccinimide ester and was then conjugated to the F6 mouse IgG1a (F6 mAb), directed against CEA (carcinoembryonic antigen), leading to the F6-te1pa immunoconjugate. F6-te1pa was compared to F6-C-dota, i.e. F6 mAb conjugated with a Cfunctionalized dota which is the only chelator used nowadays in preclinical trials for 64 Cu PET imaging. Immunoconjugates were radiolabeled with 64 Cu showing an equivalent conjugation rate of 1 ligand per mAb. The study of the complexation kinetics highlighted a relatively fast process and 64 Cu-F6-te1pa, exhibiting a specific activity of 69.3 AE 28.9 MBq mg À1 , was proved to be inert since only 4.3% of radioactivity was transchelated from the ligand to EDTA (50 000 equiv., overnight) used as a competitor. All these results are comparable with C-functionalized dota. However, in vivo studies carried out in LS174T tumor-bearing nude mice showed a limited transchelation of superoxide dismutase (SOD) into the liver; 1.6% for 64 Cu-F6-te1pa after 24 h post-injection, compared to 4.3% for 64 Cu-F6-C-dota. The uptake of 64 Cu-F6-te1pa in tumors and radioactivity distribution in organs after 24 and 48 h was satisfactory and equivalent to various standards presented in the literature. Finally, PET-phenotypic images obtained with 64 Cu-F6-te1pa at 24 h post-injection showed an excellent contrast between tumors and the healthy tissues around, which agrees well with the results of the biodistribution. The usefulness of te1pa for PET phenotypic imaging using 64 Cu has been validated. The synthesis of a bifunctional derivative of te1pa will be the next step of this work to keep the ligand properties intact.
Copper-64-labeled antibodies for PET imaging
Journal of nuclear medicine : official publication, Society of Nuclear Medicine, 1992
In the imaging of tumors using radiolabeled monoclonal antibodies, the use of PET gives increased sensitivity over conventional gamma camera imaging techniques. Copper-64, a positron-emitting radionuclide, has been labeled to 1A3, an anticolorectal carcinoma monoclonal antibody, and its fragments 1A3-F(ab')2 utilizing the bifunctional chelate Br-benzyl-TETA. The 64Cu-labeled intact 1A3 and 1A3-F(ab')2 have been evaluated as potential imaging agents for PET. Biodistribution studies of 64Cu-benzyl-TETA-1A3 and 64Cu-benzyl-TETA-1A3-F(ab')2 in tumor-bearing hamsters were compared with those of 111In-Br phi HBED-1A3, 111In-Br phi HBED-1A3-F(ab')2 and 125I-labeled intact 1A3 and 1A3-F(ab')2. Tumor uptake of 64Cu-labeled intact 1A3 and fragments in the hamster model was superior to both 111In- and 125I-labeled intact 1A3 and fragments. Human dosimetry data for 64Cu- and 123I-labeled 1A3 and 1A3-F(ab')2 were calculated from biodistribution data in rats. High kidney u...
Nuclear Medicine and Biology, 2011
Introduction-Use of copper radioisotopes in antibody radiolabeling is challenged by reported loss of the radionuclide from the bifunctional chelator used to label the protein. The objective of this study was to investigate the relationship between the thermodynamic stability of the 64 Cucomplexes of five commonly used bifunctional chelators (BFCs) and the biodistribution of an antibody labeled with 64 Cu using these chelators in tumor-bearing mice.
Evaluation of novel bifunctional chelates for the development of Cu-64-based radiopharmaceuticals
Nuclear Medicine and Biology, 2008
Background: Currently available bifunctional chelates (BFCs) for attaching Cu-64 to a targeting molecule are limited by either their radiolabeling conditions or in vivo stability. With the goal of identifying highly effective BFCs, we compared the properties of two novel BFCs, 1-oxa-4,7,10-triazacyclododecane-S-5-(4-nitrobenzyl)-4,7,10-triacetic acid (p-NO 2 -Bn-Oxo) and 3,6,9,15-tetraazabicyclo[9.3.1] pentadeca-1(15),11,13-triene-S-4-(4-nitrobenzyl)-3,6,9-triacetic acid (p-NO 2 -Bn-PCTA), with the commonly used S-2-(4-nitrobenzyl)-1,4,7,10-tetraazacyclododecanetetraacetic acid (p-NO 2 -Bn-DOTA). Methods: p-NO 2 -Bn-DOTA, p-NO 2 -Bn-Oxo and p-NO 2 -Bn-PCTA were each radiolabeled with Cu-64 under various conditions to assess the reaction kinetics and robustness of the radiolabeling. Stability of each Cu-64 BFC complex was evaluated at low pH and in serum. Small animal positron emission tomography imaging and biodistribution studies in mice were undertaken. Results: p-NO 2 -Bn-Oxo and p-NO 2 -Bn-PCTA possessed superior reaction kinetics compared to p-NO 2 -Bn-DOTA under all radiolabeling conditions; N98% radiochemical yields were achieved in b5 min at room temperature even when using near stoichiometric amounts of BFC. Under nonideal conditions, such as low or high pH, high radiochemical yields were still achievable with the novel BFCs. The radiolabeled compounds were stable in serum and at pH 2 for 48 h. The imaging and biodistribution of the Cu-64-radiolabeled BFCs illustrated differences between the BFCs, including preferential clearance via the kidneys for the p-NO 2 -Bn-PCTA Cu-64 complex. Conclusions: The novel BFCs facilitated efficient Cu-64 radiolabeling under mild conditions to produce stable complexes at potentially high specific activities. These BFCs may find wide utility in the development of Cu-64-based radiopharmaceuticals.
Pharmaceutical Biology, 1995
biological behavior (Mausner, 1993). Physical properties that are important to consider include the radionuclide half-life, the type, energy and branching ratio of particulate radiation and the gamma-ray energies and abundances. The physical half-life should be matched with the in vivo pharmacokinetics of the molecule under consideration. Time-dose fractionation is also an important criterion . For equal radioactivity concentrations in the target, radionuclides with long half lives will produce a lower absorbed dose rate than those with short half lives. The type of particulate emission also must be considered. Although Auger and low-energy conversion electrons can be potentially lethal , this effect can best be realized with intranuclear localization of the radionuclide, which does not generally occur with radiolabeled MAbs. Beta particles are less densely ionizing and have a range longer than a's or the Auger and conversion electrons so that the distribution requirements are less restrictive for RIT of bulky disease or for macrometastases. The gamma-ray energies and abundances are also important physical properties, because the presence of gamma rays allows the possibility of external imaging.
Monoclonal antibodies for copper-64 PET dosimetry and radioimmunotherapy
Cancer Biology & Therapy, 2011
Copper-64 emits b þ and bparticles suitable for positron emission tomography and radioimmunotherapy (RIT) of cancer. Copper-64-labelled antibodies have caused complete responses in laboratory animal RIT studies at far lower radiation doses than traditionally prescribed. The intracellular localization of copper radioisotopes may lead to cytotoxic effects by mechanisms beyond ionizing radiation damage. The purpose of this research was to develop a model using both internalizing and non-internalizing antibodies for direct comparison in future RIT studies using the same animal model of cancer. The monoclonal antibodies, cBR96 and cT84.66, were conjugated with N-hydroxysulfosuccinimidyl DOTA. All conjugates retained high immunoreactivity and labelled efficiently with 64 Cu with high specific activity and radiochemical purity. Twenty-four hour biodistributions determined in LS174T tumour-bearing nude mice demonstrated low organ and high tumour uptakes for both monoclonal antibodies. This model constitutes a promising system for elucidating whether internalization of 64 Cu is responsible for an enhanced tumour cytotoxicity in vivo.