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Carolyn J. Anderson - One of the best experts on this subject based on the ideXlab platform.
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new bifunctional chelator p scn phpr ne3ta for Copper 64 synthesis peptidomimetic conjugation radiolabeling and evaluation for pet imaging
Inorganic Chemistry, 2016Co-Authors: Yongkang Gai, Carolyn J. Anderson, Lingyi Sun, Wenqi Hui, Qin Ouyang, Guangya Xiang, Dexing ZengAbstract: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 64Cu-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 64Cu under mild conditions. The conjugate 64Cu-NE3TA-PEG4-LLP2A showed significantly higher specific activity, compared with 64Cu-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 chelator for 64Cu-based radiophar...
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nanogels from metal chelating crosslinkers as versatile platforms applied to Copper 64 pet imaging of tumors and metastases
Theranostics, 2015Co-Authors: Jacques Lux, Carolyn J. Anderson, Alexander G White, Minnie Chan, Adah AlmutairiAbstract:Metals are essential in medicine for both therapy and diagnosis. We recently created the first metal-chelating nanogel imaging agent, which employed versatile, reproducible chemistry that maximizes chelation stability. Here we demonstrate that our metal chelating crosslinked nanogel technology is a powerful platform by incorporating (64)Cu to obtain PET radiotracers. Polyacrylamide-based nanogels were crosslinked with three different polydentate ligands (DTPA, DOTA, NOTA). NOTA-based nanogels stably retained (64)Cu in mouse serum and accumulated in tumors in vivo as detected by PET/CT imaging. Measurement of radioactivity in major organs ex vivo confirmed this pattern, revealing a high accumulation (12.3% ID/g and 16.6% ID/g) in tumors at 24 and 48 h following administration, with lower accumulation in the liver (8.5% ID/g at 24 h) and spleen (5.5% ID/g). Nanogels accumulated even more efficiently in metastases (29.9% and 30.4% ID/g at 24 and 48 h). These metal-chelating nanogels hold great promise for future application as bimodal PET/MRI agents; chelation of β-emitting radionuclides could enable radiation therapy.
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abstract 4923 gallium 68 and Copper 64 labeled llp2a conjugates for pet ct imaging of integrin α4β1 in melanoma
Cancer Research, 2014Co-Authors: Wissam Beaino, Carolyn J. AndersonAbstract:Introduction: Melanoma is a malignant tumor derived from epidermal melanocytes, and it is known for its therapeutic resistance, aggressive clinical behavior, and predisposition for late metastasis. Integrin α4β1 is a transmembrane non-covalent heterodimer overexpressed in melanoma tumors that plays an important role in tumor growth, angiogenesis and metastasis by promoting adhesion and migration of cancer cells. There has been increasing interest in targeting this receptor for cancer imaging and therapy. In this study we evaluated three conjugates of a peptidomimetic ligand, LLP2A known to have high binding affinity for α4β1 for PET-CT imaging and potential radiotherapy. Methods: LLP2A was conjugated to three different chelators, CB-TE1A1P, NODAGA and DOTA, for 68Ga and 64Cu labeling. The conjugates were synthesized by solid phase peptide synthesis, purified by RP-HPLC and verified by LC-MS mass spectrometry. 68Ga and 64Cu labeling was done in acetate buffer pH 4 for 68Ga and pH 6.5 for 64Cu at 70°C for 20 min. High Specific activity (1 mCi/µg) and radiopurity (>98%) were achieved. Saturation binding and competitive binding assays with B16F10 melanoma cells determined the binding affinity of the compounds. The biodistributions of the LLP2A conjugates were evaluated in B16F10 subcutaneous tumor bearing C57BL/6 mice. PET-CT imaging was performed at 2, 4 and 24 h post-injection for the 64Cu tracers and 1h post-injection for 68Ga tracer. Results: The competitive binding assay indicated that Ga-NODAGA-PEG4-LLP2A had higher affinity (IC50 = 0.68, Ki = 0.11 nM) compared to Ga-DOTA-PEG4-LLP2A (IC50 = 9.37, Ki = 1.56 nM); however, biodistribution showed similar tumor uptake for 68Ga-NODAGA-PEG4-LLP2A and 68Ga-DOTA-PEG4-LLP2A (8.7 ± 1.3 %ID/g and 9.1 ± 0.9 %ID/g respectively) but less renal and liver retention for the DOTA conjugate. Both 64Cu-labeled CB-TE1A1P-PEG4-LLP2A and NODAGA-PEG4-LLP2A showed high affinity to α4β1 integrin with a comparable Kd (0.28 nM vs 0.23 nM) and Bmax (296 fmol/mg vs 243 fmol/mg). The tumor uptake at 2 h post-injection was comparable for the two probes but 64Cu-CB-TE1A1P-PEG4-LLP2A had higher uptake compared to 64Cu-NODAGA-PEG4-LLP2A (16.9 ± 2.2 %ID/g vs13.4 ± 1.7 %ID/g). Tumor to muscle and tumor to blood ratios from biodistribution and PET-CT images were significantly higher for 64Cu-CB-TE1A1P-PEG4-LLP2A compared to 64Cu-NODAGA-PEG4-LLP2A. In addition, liver and kidney uptake was 2-fold lower for 64Cu-CB-TE1A1P-PEG4-LLP2A. Conclusion: These data demonstrate that 68Ga-labeled DOTA, NODAGA and 64Cu-labeled CB-TE1A1P, NODAGA LLP2A conjugates are excellent imaging agents for melanoma or other α4β1-positive tumors, with 64Cu-CB-TE1A1P-PEG4-LLP2A being the best. These data also suggest the potential for radiotherapy and radiotheranostics using radioisotopes like Lu-177, Y-90 and Cu-67. Citation Format: Wissam Beaino, Carolyn J. Anderson. Gallium-68- and Copper-64-labeled LLP2A conjugates for PET-CT imaging of integrin α4β1 in melanoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4923. doi:10.1158/1538-7445.AM2014-4923
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Copper 64 labeled cb te1k1p conjugates of mcp 1 for pet imaging of macrophages in tuberculosis granulomas
The Journal of Nuclear Medicine, 2014Co-Authors: Dexing Zeng, Wissam Beaino, Joshua T Mattila, Majiong Jiang, Brian J Lopresti, Marie Coleman, Joanne L Flynn, Carolyn J. AndersonAbstract:1210 Objectives Higher levels of activated macrophages in granulomas indicate that the lesion is more “active” leading to disease progression. MCP-1 (also called chemokine ligand-2) is the natural ligand of Chemokine Receptor 2 that is predominantly expressed on monocyte-derived macrophages and found in high concentrations in activated macrophages. The newly developed CB-TE1KP chelator was attached to MCP-1 via strain-promoted azide-alkyne cycloaddition (SPAAC), and the resulting CB-TE1K1P-MCP-1 conjugates were labeled with Cu-64 under mild conditions for PET imaging of macrophages in TB granulomas Methods MCP-1 was functionalized with PEG-azido by mixing with azido-PEG4-NHS in PBS buffer. After purification, the azido-MPC-1 conjugates “clicked” with the CB-TE1K1P-PEG4-DBCO via SPAAC. The purified MCP-1 conjugates with varied numbers of chelators were labeled with Cu-64 under mild conditions, and then were injected i.v. into the TB infected monkeys for PET imaging. Biotin-MCP-1 was prepared by mixing azido-MCP-1 with DBCO-PEG4-biotin and used for the corresponding flow cytometry Results The conjugation yields were nearly quantitative due to reaction with 25-fold excess DBCO-chelator/biotin. Flow cytometry results indicated that MCP-1 bound to macrophages, and over modification of MCP-1 (over 3 moieties/MCP-1) dramatically compromised its affinity. Cu-64 labeling of CB-TE1K1P-MCP-1 conjugates was achieved at 37 °C in 30 min with a SA of 10-15 µCi/µg. The corresponding PET imaging studies in TB infected monkeys demonstrated significant uptake in the TB granulomas with a standard uptake value (SUV) of 1.0, and a ratio of granuloma:normal lung of 6.7 Conclusions The first cross-bridged chelator attached MCP-1 were successfully prepared and radiolabeled with Cu-64 under mild conditions, and the resulting 64Cu-MCP-1 conjugates showed promise for PET imaging of macrophages in the TB infected monkey Research Support Bill & Melinda Gates Foundation #1034408
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Roles of Atox1 and p53 in the trafficking of Copper-64 to tumor cell nuclei: Implications for cancer therapy
JBIC Journal of Biological Inorganic Chemistry, 2014Co-Authors: Wissam Beaino, Yunjun Guo, Albert J. Chang, Carolyn J. AndersonAbstract:Owing to its cytotoxicity, free Copper is chelated by protein side chains and does not exist in vivo. Several chaperones transport Copper to various cell compartments, but none have been identified that traffic Copper to the nucleus. Copper-64 decays by β + and β − emission, allowing positron emission tomography and targeted radionuclide therapy for cancer. Because the delivery of 64Cu to the cell nucleus may enhance the therapeutic effect of Copper radiopharmaceuticals, elucidation of the pathway(s) involved in transporting Copper to the tumor cell nucleus is important for optimizing treatment. We identified Atox1 as one of the proteins that binds Copper in the nucleus. Mouse embryonic fibroblast cells, positive and negative for Atox1, were used to determine the role of Atox1 in 64Cu transport to the nucleus. Mouse embryonic fibroblast Atox1+/+ cells accumulated more 64Cu in the nucleus than did Atox1−/− cells. HCT 116 colorectal cancer cells expressing p53 (+/+) and not expressing p53 (−/−) were used to evaluate the role of this tumor suppressor protein in 64Cu transport. In cells treated with cisplatin, the uptake of 64Cu in the nucleus of HCT 116 p53+/+ cells was greater than that in HCT 116 p53−/− cells. Atox1 expression increased in HCT 116 p53+/+ and p53−/− cells treated with cisplatin; however, Atox1 localized to the nuclei of p53+/+ cells more than in the p53−/− cells. The data presented here indicate that Atox1 is involved in Copper transport to the nucleus, and cisplatin affects nuclear transport of 64Cu in HCT 116 cells by upregulating the expression and the nuclear localization of Atox1.
Paul S Donnelly - One of the best experts on this subject based on the ideXlab platform.
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macrocyclic bifunctional chelators and conjugation strategies for Copper 64 radiopharmaceuticals
Advances in Inorganic Chemistry, 2016Co-Authors: Brett M Paterson, Paul S DonnellyAbstract:The positron-emitting radionuclide Copper-64 is of interest in the development of new molecular imaging agents for use with positron emission tomography. This account highlights the development and application of macrocyclic chelators that have been designed to form complexes of Copper-64 that are exceptionally stable in vivo and can be readily conjugated to biologically active molecules and nanoparticles. “Click chemistry” is discussed as a conjugation technique that is bioorthogonal with both pre- and postlabeling capabilities. Enzyme-mediated site-specific strategies are introduced as new conjugation techniques with the potential to improve homogeneity, reproducibility, and the likelihood for retention of full biological function of targeted tracers for noninvasive diagnostic imaging.
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Copper 64 labeled sar conjugated grp receptor antagonists as new pet tracers for imaging of prostate cancer
The Journal of Nuclear Medicine, 2014Co-Authors: Eleni Gourni, Brett M Paterson, Paul S Donnelly, Del Pozzo Luigi, Emilie Kheirallah, Jean Claude Reubi, Christiane Smerling, Helmut R MaeckeAbstract:1082 Objectives The gastrin-releasing peptide receptor (GRPr) appears to be one of the most important molecular targets for the visualization of prostate cancer and is currently being targeted with radiolabeled bombesin derivatives. The present study aims at developing statine-based bombesin receptor antagonists suitable for labeling with 64Cu for PET imaging. Methods The potent GRPr antagonist D-Phe-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH2, was conjugated to the sarcophagine (3,6,10,13,16,19-hexaazabicyclo[6.6.6] icosane=Sar) derivative MeCOSar via PEG4 (LE1) and PEG2 (LE2) spacers and radiolabeled with 64Cu. IC(50) values were determined using 125I-Tyr4-BN as radioligand. Antagonist potency was determined by Ca2+-flux studies. In vivo-studies were performed in PC-3 tumor-bearing nude mice. Results Both conjugates exhibited high affinity to GRPr ( natCu-LE1: 1.4±0.1 nM and natCu-LE2: 3.8±0.6 nM). Ca2+-flux measurements confirmed their antagonist properties. The peptides were stably radiolabeled with 64Cu, with labelling yields >95% and specific activities of about 100 MBq/nmol. Biodistribution revealed specific targeting of the tumor and GRPr-positive organs, Cu-64-LE1 exhibiting the highest tumor uptake of 19.6 ± 4.7 % IA/g at 1 h p.i.. Biodistribution at later time points showed increasing tumor-to-background ratios with time. This was illustrated by the acquisition of PET images at 4 and 24 h p.i. showing high tumor-to-normal tissue contrast. Conclusions This study demonstrates the high affinity of the MeCOSar-PEGx-bombesin conjugates to GRPr. The stable encapsulation of 64Cu by MeCOSar, the long half-life of 64Cu and the suitable biodistribution profile of the 64Cu-labeled peptides lead to PET images of high contrast and to a potential translation into the clinic. Research Support This work was supported by the German Consortium for Translational Cancer Research (DKTK).
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peptide targeted Copper 64 radiopharmaceuticals
Current Topics in Medicinal Chemistry, 2011Co-Authors: Michelle T, Paul S DonnellyAbstract:Peptide targeted ⁶⁴Cu-labelled diagnostic agents for positron emission tomography are viable candidates for molecular imaging of cancer. In a clinical setting, optimal image quality relies on selective tumor uptake of the ⁶⁴Cu-labelled radiotracer. The three components of the radiotracer construct--the chelate group, linker and targeting peptide--all influence the biodistribution of the ⁶⁴Cu-labelled radiotracer in vivo. Low or moderate Cu complex stability in vivo results in transmetallation of ⁶⁴Cu to endogenous proteins, giving rise to high background activity. The length and the nature of the linker group affect the affinity of the ⁶⁴Cu-labelled radiotracer for the target receptor. Variations in the peptide sequence can impact on the metabolic stability and therefore the bioavailability and tumor retention of the ⁶⁴Cu-labelled radiotracer in vivo. Lastly, the hydrophilicity of the construct can influence radiotracer metabolism and clearance pathways. Recent advances in the field of peptide targeted ⁶⁴Cu-labelled radiopharmaceuticals involve GRPR-targeted and αvβ3 integrin receptor-targeted constructs. These constructs are based on the bombesin peptide sequence and the RGD recognition motif respectively. These examples are reviewed as case studies in the optimisation of ⁶⁴Cu radiotracer design.
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versatile new bis thiosemicarbazone bifunctional chelators synthesis conjugation to bombesin 7 14 nh2 and Copper 64 radiolabeling
Inorganic Chemistry, 2010Co-Authors: Brett M Paterson, John A Karas, Denis B Scanlon, Jonathan M White, Paul S DonnellyAbstract:New bifunctional derivatives of diacetyl-bis(4-methylthiosemicarbazone) (H2atsm) have been prepared by a selective transamination reaction of a new dissymmetric bis(thiosemicarbazone) precursor H2L1. The new derivatives contain an aliphatic carboxylic acid (H2L2 and H2L3), t-butyl carbamate (H2L4), or ammonium ion (H2L5) functional group. The new ligands and Copper(II) complexes have been characterized by NMR spectroscopy, mass spectrometry, and microanalysis. The complex CuII(L4) was structurally characterized by X-ray crystallography and shows the metal center to be in an N2S2 distorted square planar coordination geometry. Electrochemical measurements show that the Copper(II) complexes undergo a reversible reduction attributable to a Cu(II)/Cu(I) process. The ligands and the Copper(II) complexes featuring a carboxylic acid functional group have been conjugated to the tumor targeting peptide bombesin(7−14)-NH2. The bifunctional peptide conjugates were radiolabeled with Copper-64 in the interest of develo...
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versatile new bis thiosemicarbazone bifunctional chelators synthesis conjugation to bombesin 7 14 nh2 and Copper 64 radiolabeling
Inorganic Chemistry, 2010Co-Authors: Brett M Paterson, John A Karas, Denis B Scanlon, Jonathan M White, Paul S DonnellyAbstract:New bifunctional derivatives of diacetyl-bis(4-methylthiosemicarbazone) (H(2)atsm) have been prepared by a selective transamination reaction of a new dissymmetric bis(thiosemicarbazone) precursor H(2)L(1). The new derivatives contain an aliphatic carboxylic acid (H(2)L(2) and H(2)L(3)), t-butyl carbamate (H(2)L(4)), or ammonium ion (H(2)L(5)) functional group. The new ligands and Copper(II) complexes have been characterized by NMR spectroscopy, mass spectrometry, and microanalysis. The complex Cu(II)(L(4)) was structurally characterized by X-ray crystallography and shows the metal center to be in an N(2)S(2) distorted square planar coordination geometry. Electrochemical measurements show that the Copper(II) complexes undergo a reversible reduction attributable to a Cu(II)/Cu(I) process. The ligands and the Copper(II) complexes featuring a carboxylic acid functional group have been conjugated to the tumor targeting peptide bombesin(7-14)-NH(2). The bifunctional peptide conjugates were radiolabeled with Copper-64 in the interest of developing new positron emission tomography (PET) imaging agents. The conjugates were radiolabeled with Copper-64 rapidly in high radiochemical purity (>95%) at room temperature under mild conditions and were stable in a cysteine and histidine challenge study.
Michael J Welch - One of the best experts on this subject based on the ideXlab platform.
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a semi automated system for the routine production of Copper 64
Applied Radiation and Isotopes, 2012Co-Authors: Maiko Kume, Michael J Welch, Paul Carey, Gregory G Gaehle, Evelyn Madrid, Thomas Voller, William H Margenau, Suzanne E. LapiAbstract:Abstract An automated system for the production of high specific activity 64 Cu via the irradiation of electroplated enriched 64 Ni targets has been developed. We have been operating this system continually on a biweekly or weekly basis for more than two years. Since the inception of this automated production system, (October 1, 2008), we have had 145 productions, produced 53562 mCi and shipped out 25629 mCi of this isotope to external users. We routinely produce over 400 mCi of this isotope per batch with a specific activity of 14,000±7600 mCi/μmol for distribution to some 12–15 centers each production.
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in vivo evaluation of Copper 64 labeled monooxo tetraazamacrocyclic ligands
Nuclear Medicine and Biology, 2004Co-Authors: Xiankai Sun, Arthur E Martell, Michael J Welch, Joonyoung Kim, Carolyn J. AndersonAbstract:Copper-64 (T(1/2)=12.7 h; beta(+): 0.653 MeV, 17.4%; beta(-): 0.578 MeV, 39%) has applications in positron emission tomography (PET) imaging and radiotherapy, and is conveniently produced on a biomedical cyclotron. Tetraazamacrocyclic ligands are the most widely used bifunctional chelators (BFCs) for attaching Copper radionuclides to antibodies and peptides due to their relatively high kinetic stability. In this paper, we evaluated three monooxo-tetraazamacrocyclic ligands with different ring sizes and oxo group positions. H1 [1,4,7,10-tetraazacyclotridecan-11-one], H2 [1,4,8,11-tetraazacyclotetradecan-5-one] and H3 [1,4,7,10-tetraazacyclotridecan-2-one] were radiolabeled with (64)Cu in high radiochemical yields under mild conditions. The three (64)Cu-labeled complexes are all +1 charged, as determined by their electrophoretic mobility. While they demonstrated >95% stability in rat serum out to 24 h, both biodistribution and microPET imaging studies revealed high uptake and long retention of the compounds in major clearance organs (e.g., blood, liver and kidney), which suggests that (64)Cu dissociated from the complexes in vivo. Of the three complexes, (64)Cu-2(+), which has a cyclam backbone (1,4,8,11-tetraazacyclotetradecane), exhibited the lowest nontarget organ accumulation. The data from these studies may invalidate the candidacy of the monooxo-tetraazamacrocyclics as BFCs for Copper radiopharmaceuticals. However, the data presented here suggest that neutral or negatively charged Cu(II) complexes of tetraazamacrocyclic ligands with a cyclam backbone (tetradecane) are optimal for Copper radiopharmaceutical applications.
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in vivo behavior of Copper 64 labeled methanephosphonate tetraaza macrocyclic ligands
Journal of Biological Inorganic Chemistry, 2003Co-Authors: Xiankai Sun, Melinda Wuest, Zoltan Kovacs, Dean A Sherry, Ramunas J Motekaitis, Zheng Wang, Arthur E Martell, Michael J Welch, Carolyn J. AndersonAbstract:Copper-64 ( T(1/2)=12.7 h; beta(+): 0.653 MeV, 17.4%; beta(-): 0.578 MeV, 39%) is produced in a biomedical cyclotron and has applications in both imaging and therapy. Macrocyclic chelators are widely used as bifunctional chelators to bind Copper radionuclides to antibodies and peptides owing to their relatively high kinetic stability. In this paper, we evaluated three tetraaza macrocyclic ligands with two, three, and four pendant methanephosphonate functional groups. DO2P [1,4,7,10-tetraazacyclododecane-1,7-di(methanephosphonic acid)], DO3P [1,4,7,10-tetraazacyclododecane-1,4,7-tri(methanephosphonic acid)], and DOTP [1,4,7,10-tetraazacyclododecane-1,4,7,10-tetra(methanephosphonic acid)] were all radiolabeled with (64)Cu in high radiochemical yields. Copper-64-labeled DO2P and DOTP were highly stable in rat serum out to 24 h, while (64)Cu-DO3P remained 73% intact, with the remainder possibly forming a (64)Cu(.)2DO3P dimer by 24 h. The biodistribution experiments were performed in normal Sprague-Dawley rats. Of the three complexes, (64)Cu-DO2P demonstrated the most optimal clearance through the blood and liver. Copper-64-DO3P and (64)Cu-DOTP exhibited higher liver uptake and longer retention of liver activity, possibly because of the large negative charge of the complexes under physiological conditions. All three (64)Cu-labeled complexes showed high accumulation in bone, likely due to the binding of the methanephosphonate groups to hydroxyapatite. These results suggest that this series of methanephosphonate macrocyclic ligands may be useful as potential bone-imaging agents. The thermodynamic stability constants of the Cu(II) complexes with these three ligands were determined, and were found to be significantly higher than those of their acetate analogues. The Cu(II)-DO2P complex exhibited the highest stability constant among divalent transition metal ion DO2P complexes. Metabolism studies of (64)Cu-DO2P in rat liver suggest that the DO2P ligand may be used as a bifunctional chelator for Copper radionuclides in radiodiagnostic or radiotherapeutic studies.
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radiolabeling and in vivo behavior of Copper 64 labeled cross bridged cyclam ligands
Journal of Medicinal Chemistry, 2002Co-Authors: Xiankai Sun, Melinda Wuest, Ramunas J Motekaitis, Arthur E Martell, Michael J Welch, Andrew C Boswell, Gary R Weisman, Edward H Wong, David P Reed, Carolyn J. AndersonAbstract:Macrocyclic chelators and their metal complexes have widespread applications in the biomedical sciences, including radiopharmaceutical chemistry. The use of Copper radionuclides in radiopharmaceuticals is increasing. Macrocyclic chelators have been found to have enhanced in vivo stability over acyclic chelators such as ethylenediaminetetraacetic acid (EDTA) and diethylenetriaminepentaacetic acid (DTPA). The currently used chelators of choice for labeling Copper radionuclides to biological molecules are analogues of TETA (1,4,8,11-tetraazacyclotetradecane-1,4,8,11-tetraacetic acid); however, recent reports have demonstrated evidence of in vivo instability of the radio-Cu(II)−TETA complexes. A new class of structurally reinforced macrocycles, the “cross-bridged” cyclam derivatives, form highly stable complexes with Cu(II) that are resistant to dissociation in strong acid. Here, we evaluate a series of 64Cu(II) cross-bridged macrocyclic complexes for biological stability and in vivo behavior. The ligands eva...
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Copper 64 diacetyl bis n4 methylthiosemicarbazone an agent for radiotherapy
Proceedings of the National Academy of Sciences of the United States of America, 2001Co-Authors: Jason S Lewis, Judith M. Connett, Richard Laforest, Thomas L Buettner, Shengkwei Song, Yasuhisa Fujibayashi, Michael J WelchAbstract:Systemic administration of hypoxia-selective 64Cu-diacetyl-bis(N4-methylthiosemicarbazone) (64Cu-ATSM) has increased significantly the survival time of hamsters bearing human GW39 colon cancer tumors. Radiotherapy experiments were performed in animals bearing either 7-day-old (0.5–1.0 g) or 15-day-old (1.5–2.0 g) tumors. Studies compared animals treated with a single dose of 0, 4, 6, 7, 8, or 10 mCi of 64Cu-ATSM (1 Ci = 37 GBq) with or without the vasodilator hydralazine. A multiple dose regimen of 3 × 4 mCi at 72-h intervals was studied also. Single doses of >6 mCi of 64Cu-ATSM and the dose-fractionation protocol significantly increased the survival time of the hamsters compared with controls. The highest dose, 10 mCi of 64Cu-ATSM, increased survival to 135 days in 50% of animals bearing 7-day-old tumors, 6-fold longer than control animals' survival (20 days), with only transient leucopenia and thrombocytopenia but no overt toxicity. Human absorbed doses were calculated from hamster biodistribution; the dose-critical organs were the lower large intestine (1.43 ± 0.19 rad/mCi) and upper large intestine (1.20 ± 0.38 rad/mCi). High-resolution MRI and positron-emission tomography using a therapeutic administration of 10 mCi were used to monitor tumor volume and morphology and to assess tumor dosimetry accurately, giving a tumor dose of 81 ± 7.5 rad/mCi. 64Cu-ATSM has increased the survival time of tumor-bearing animals significantly with no acute toxicity and thus is a promising agent for radiotherapy.
Xiankai Sun - One of the best experts on this subject based on the ideXlab platform.
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Copper 64 labelled fingolimod for pet imaging of the sphingolipid 1 phosphate axis in prostate cancer
The Journal of Nuclear Medicine, 2015Co-Authors: Hancheng Cai, Saleh Ramezani, Preston Christensen, Guiyang Hao, Xiankai SunAbstract:1123 Objectives The sphingolipid-1-phosphate axis including sphingosine kinase (SPhK) and sphingosine-1-phosphate receptors (S1PRs) is implicated in human cancer development and progression.1 Fingolimod (FTY720), a synthetic sphingosine analogue, is an FDA-approved immunmodulating drug, which has been documented as anti-tumor agent by targeting SPhK/S1PRs.2 In this work, we report the development of Copper-64 labelled FTY720 for PET imaging of the sphingolipid signaling axis in prostate cancer. Methods To be labeled with 64Cu, FTY720 was conjugated with a chelator,p-SCN-Bn-NOTA. After the in vitro stability and lipophilicity evaluation of the radiotracer, preliminary small animal PET/CT imaging with 64Cu-NOTA-FTY720 was performed in two prostate cancer xenograft models, the SPhK overexpressing PC-3 and the low SPhK expressing LNCaP. The imaging specificity was tested by a blockade experiment of co-injecting the radiotracer with FTY720. Results The conjugate NOTA-FTY720, was obtained in 50.1% yield and characterized by LC-MS, and 1H-NMR. NOTA-FTY720 was labeled with 64Cu in 94.7% radiochemical yield. After HPLC purification and reconstitution with 8% ethanol in saline, 64Cu-NOTA-FTY720 was obtained with > 99.5% radiochemical purity and a specific activity of 403-806 mCi/μmol. The radiotracer remained > 99.0% intact within 24 h incubation in rat serum and its log P value was 1.07. As expected, the radiotracer showed higher uptake in PC-3 xenografts than that in LnCaP xenografts on in vivo PET/CT images at 24 h. Both tumors’ uptakes in the presence of FTY720 were significantly reduced, indicating that the imaging is specifically targeting sphingolipid-1-phosphate axis. Conclusions A radiotracer for noninvasive assessment of the sphingolipid-1-phosphate axis was successfully developed by derivatizing an FDA-approved immunomodulating drug. The preliminary evaluation of the radiotracer 64Cu-NOTA-FTY720 demonstrated its potential utility in prostate cancer imaging and therapy. Further imaging studies and biological validation are ongoing.
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click chemistry strategy for labeling antibodies with Copper 64 via a cross bridged tetraazamacrocyclic chelator scaffold
Bioconjugate Chemistry, 2015Co-Authors: Amit Kumar, Saleh Ramezani, Guiyang Hao, Li Liu, Jer Tsong Hsieh, Xiankai SunAbstract:We report a click-chemistry based modular strategy for antibody labeling with (64)Cu (t1/2 = 12.7 h; β(+) 0.656 MeV, 17.4%; β(-) 0.573 MeV, 39%; EC 43%) under ambient condition utilizing a cross-bridged tetraazamacrocyclic (CB-TE2A) analogue, which otherwise requires harsh conditions that make the CB-TE2A analogues under-utilized for protein labeling despite the fact that they form kinetically inert Copper complexes with high in vivo stability. Our strategy involves prelabeling a CB-TE2A based scaffold (CB-TE2A-1C) with (64)Cu and its subsequent reaction with an antibody via the tetrazine-norbornene mediated click chemistry. The effectiveness of this strategy was demonstrated by labeling two monoclonal antibodies, an anti-PSMA antibody (YPSMA-1) and a chimeric anti-phosphatidylserine antibody (Bavituximab). The immunoreactivity of the antibodies remained unchanged after the tetrazine modification and click-chemistry (64)Cu labeling. To further demonstrate the practicality of the modular (64)Cu labeling strategy, we tested positron emission tomography (PET) imaging of tumor with the (64)Cu-labeled bavituximab in a mouse xenograft model. The tumor visualization and uptake of the labeled antibody exhibited the versatility of the click-chemistry strategy.
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preparation and biological evaluation of Copper 64 labeled tyr3 octreotate using a cross bridged macrocyclic chelator
Clinical Cancer Research, 2004Co-Authors: Jennifer E Sprague, Xiankai Sun, Gary R Weisman, Edward H Wong, Yijie Peng, Samuel Achilefu, Carolyn J. AndersonAbstract:Purpose: Somatostatin receptors (SSTr) are expressed on many neuroendocrine tumors, and several radiotracers have been developed for imaging these types of tumors. For this reason, peptide analogues of somatostatin have been well characterized. Copper-64 (t 1/2 = 12.7 hours), a positron emitter suitable for positron emission tomography (PET) imaging, was shown recently to have improved in vivo clearance properties when chelated by the cross-bridged tetraazamacrocycle 4,11-bis(carboxymethyl)-1,4,8,11-tetraazabicyclo(6.6.2)hexadecane (CB-TE2A) compared with 1,4,8,11-tetraazacyclotetradecane-1,4,8,11-tetraacetic acid (TETA). Experimental Design: CB-TE2A and TETA were conjugated to the somatostatin analogue tyrosine-3-octreotate (Y3-TATE) for evaluation of CB-TE2A as a bifunctional chelator of 64 Cu. The in vitro affinity of each compound for SSTr was determined using a homologous competitive binding assay. In vivo characteristics of both radiolabeled compounds were examined in biodistribution and microPET studies of AR42J tumor-bearing rats. Results: Cu-CB-TE2A-Y3-TATE ( K d = 1.7 nmol/L) and Cu-TETA-Y3-TATE ( K d = 0.7 nmol/L) showed similar affinities for AR42J derived SSTr. In biodistribution studies, nonspecific uptake in blood and liver was lower for 64 Cu-CB-TE2A-Y3-TATE. Differences increased with time such that, at 4 hours, blood uptake was 4.3-fold higher and liver uptake was 2.4-fold higher for 64 Cu-TETA-Y3-TATE than for 64 Cu-CB-TE2A-Y3-TATE. In addition, 4.4-times greater tumor uptake was detected with 64 Cu-CB-TE2A-Y3-TATE than with 64 Cu-TETA-Y3-TATE at 4 hours postinjection. MicroPET imaging yielded similar results. Conclusions: CB-TE2A appears to be a superior in vivo bifunctional chelator of 64 Cu for use in molecular imaging by PET or targeted radiotherapy due to both improved nontarget organ clearance and higher target organ uptake of 64 Cu-CB-TE2A-Y3-TATE compared with 64 Cu-TETA-Y3-TATE.
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in vivo evaluation of Copper 64 labeled monooxo tetraazamacrocyclic ligands
Nuclear Medicine and Biology, 2004Co-Authors: Xiankai Sun, Arthur E Martell, Michael J Welch, Joonyoung Kim, Carolyn J. AndersonAbstract:Copper-64 (T(1/2)=12.7 h; beta(+): 0.653 MeV, 17.4%; beta(-): 0.578 MeV, 39%) has applications in positron emission tomography (PET) imaging and radiotherapy, and is conveniently produced on a biomedical cyclotron. Tetraazamacrocyclic ligands are the most widely used bifunctional chelators (BFCs) for attaching Copper radionuclides to antibodies and peptides due to their relatively high kinetic stability. In this paper, we evaluated three monooxo-tetraazamacrocyclic ligands with different ring sizes and oxo group positions. H1 [1,4,7,10-tetraazacyclotridecan-11-one], H2 [1,4,8,11-tetraazacyclotetradecan-5-one] and H3 [1,4,7,10-tetraazacyclotridecan-2-one] were radiolabeled with (64)Cu in high radiochemical yields under mild conditions. The three (64)Cu-labeled complexes are all +1 charged, as determined by their electrophoretic mobility. While they demonstrated >95% stability in rat serum out to 24 h, both biodistribution and microPET imaging studies revealed high uptake and long retention of the compounds in major clearance organs (e.g., blood, liver and kidney), which suggests that (64)Cu dissociated from the complexes in vivo. Of the three complexes, (64)Cu-2(+), which has a cyclam backbone (1,4,8,11-tetraazacyclotetradecane), exhibited the lowest nontarget organ accumulation. The data from these studies may invalidate the candidacy of the monooxo-tetraazamacrocyclics as BFCs for Copper radiopharmaceuticals. However, the data presented here suggest that neutral or negatively charged Cu(II) complexes of tetraazamacrocyclic ligands with a cyclam backbone (tetradecane) are optimal for Copper radiopharmaceutical applications.
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comparative in vivo stability of Copper 64 labeled cross bridged and conventional tetraazamacrocyclic complexes
Journal of Medicinal Chemistry, 2004Co-Authors: Andrew C Boswell, Xiankai Sun, Gary R Weisman, Edward H Wong, Weijun Niu, And Arnold L Rheingold, Carolyn J. AndersonAbstract:The increased use of Copper radioisotopes in radiopharmaceutical applications has created a need for bifunctional chelators (BFCs) that form stable radioCopper complexes and allow covalent attachment to biological molecules. The chelators most commonly utilized for labeling Copper radionuclides to biomolecules are analogues of 1,4,8,11-tetraazacyclotetradecane-1,4,8,11-tetraacetic acid (TETA); however, recent reports have communicated the instability of the radio-Cu(II)-TETA complexes in vivo. A class of bicyclic tetraazamacrocycles, the ethylene "cross-bridged" cyclam (CB-cyclam) derivatives, form highly kinetically stable complexes with Cu(II) and therefore may be less susceptible to transchelation than their nonbridged analogues in vivo. Herein we report results on the relative biological stabilities and identification of the resulting radiolabeled metabolites of a series of (64)Cu-labeled macrocyclic complexes. Metabolism studies in normal rat liver have revealed that the (64)Cu complex of 4,11-bis(carboxymethyl)-1,4,8,11-tetraazabicyclo[6.6.2]hexadecane ((64)Cu-CB-TE2A) resulted in significantly lower values of protein-associated (64)Cu than (64)Cu-TETA [13 +/- 6% vs 75 +/- 9% at 4 h]. A similar trend was observed for the corresponding cyclen derivatives, with the (64)Cu complex of 4,10-bis(carboxymethyl)-1,4,7,10-tetraazabicyclo[5.5.2]tetradecane ((64)Cu-CB-DO2A) undergoing less transchelation than the (64)Cu complex of 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid ((64)Cu-DOTA) [61 +/- 14% vs 90.3 +/- 0.5% protein associated (64)Cu at 4 h]. These data indicate that the structurally reinforcing cross-bridge enhances in vivo stability by reducing metal loss to protein in both the cyclam and cyclen cross-bridged (64)Cu complexes and that (64)Cu-CB-TE2A is superior to (64)Cu-CB-DO2A in that regard. These findings further suggest that a bifunctional chelator derivative of CB-TE2A is a highly desirable alternative for labeling Copper radionuclides to biological molecules for diagnostic imaging and targeted radiotherapy.
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macrocyclic bifunctional chelators and conjugation strategies for Copper 64 radiopharmaceuticals
Advances in Inorganic Chemistry, 2016Co-Authors: Brett M Paterson, Paul S DonnellyAbstract:The positron-emitting radionuclide Copper-64 is of interest in the development of new molecular imaging agents for use with positron emission tomography. This account highlights the development and application of macrocyclic chelators that have been designed to form complexes of Copper-64 that are exceptionally stable in vivo and can be readily conjugated to biologically active molecules and nanoparticles. “Click chemistry” is discussed as a conjugation technique that is bioorthogonal with both pre- and postlabeling capabilities. Enzyme-mediated site-specific strategies are introduced as new conjugation techniques with the potential to improve homogeneity, reproducibility, and the likelihood for retention of full biological function of targeted tracers for noninvasive diagnostic imaging.
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the ionic charge of Copper 64 complexes conjugated to an engineered antibody affects biodistribution
Bioconjugate Chemistry, 2015Co-Authors: Brett M Paterson, Jason L J Dearling, Vamsidhar Akurathi, Soledad Betanzoslara, Ted S Treves, Stephan D VossAbstract:The development of biomolecules as imaging probes requires radiolabeling methods that do not significantly influence their biodistribution. Sarcophagine (Sar) chelators form extremely stable complexes with Copper and are therefore a promising option for labeling proteins with (64)Cu. However, initial studies using the first-generation sarcophagine bifunctional chelator SarAr to label the engineered antibody fragment ch14.18-ΔCH2 (MW 120 kDa) with (64)Cu showed high tracer retention in the kidneys, presumably because the high local positive charge on the Cu(II)-SarAr moiety resulted in increased binding of the labeled protein to the negatively charged basal cells of the glomerulus. To test this hypothesis, ch14.18-ΔCH2 was conjugated with a series of Sar derivatives of decreasing positive charge and three commonly used macrocyclic polyaza polycarboxylate (PAC) bifunctional chelators (BFC). The immunoconjugates were labeled with (64)Cu and injected into mice, and PET/CT images were obtained at 24 and 48 h postinjection (p.i.). At 48 h p.i., ex vivo biodistribution was assessed. In addition, to demonstrate the potential of metastasis detection using (64)Cu-labeled ch14.18-ΔCH2, a preclinical imaging study of intrahepatic neuroblastoma tumors was performed. Reducing the positive charge on the Sar chelators decreased kidney uptake of Cu-labeled ch14.18-ΔCH2 by more than 6-fold, from >45 to <6% ID/g, whereas the uptake in most other tissues, including liver, was relatively unchanged. However, despite this dramatic decrease, the renal uptake of the PAC BFCs was generally lower than that of the Sar derivatives, as was the liver uptake. Uptake of (64)Cu-labeled ch14.18-ΔCH2 in neuroblastoma hepatic metastases was detected using PET.
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single chain antibody conjugated to a cage amine chelator and labeled with positron emitting Copper 64 for diagnostic imaging of activated platelets
Molecular Pharmaceutics, 2014Co-Authors: Karen Alt, Brett M Paterson, Gojko Buncic, Katie Ardipradja, Christine Schieber, Bock Lim, Stan Poniger, Bjoern Jakoby, Xiaowei WangAbstract:Imaging of activated platelets using an activation specific anti-GPIIb/IIIa integrin single-chain antibody (scFvanti-LIBS) conjugated to a positron emitting Copper-64 complex of a cage amine sarcophagine chelator (MeCOSar) is reported. This tracer was compared in vitro to a (64)Cu(II) complex of the scFv conjugated to another commonly used macrocycle, DOTA. The scFvanti-LIBS-MeCOSar conjugate was radiolabeled with (64)Cu(II) rapidly under mild conditions and with higher specific activity than scFvanti-LIBS-DOTA. The utility of scFvanti-LIBS-MeCOSar as a diagnostic agent was assessed in vivo in a mouse model of acute thrombosis. The uptake of scFvanti-LIBS-(64)CuMeCOSar in the injured vessel was significantly higher than the noninjured vessel. Positron emission tomography (PET) was used to show accumulation of scFvanti-LIBS-(64)CuMeCOSar with high and specific uptake in the injured vessel. ScFvanti-LIBS-(64)CuMeCOSar is an excellent tool for highly sensitive in vivo detection of activated platelets in PET and has the potential to be used for early diagnosis of acute thrombotic events.
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Copper 64 labeled sar conjugated grp receptor antagonists as new pet tracers for imaging of prostate cancer
The Journal of Nuclear Medicine, 2014Co-Authors: Eleni Gourni, Brett M Paterson, Paul S Donnelly, Del Pozzo Luigi, Emilie Kheirallah, Jean Claude Reubi, Christiane Smerling, Helmut R MaeckeAbstract:1082 Objectives The gastrin-releasing peptide receptor (GRPr) appears to be one of the most important molecular targets for the visualization of prostate cancer and is currently being targeted with radiolabeled bombesin derivatives. The present study aims at developing statine-based bombesin receptor antagonists suitable for labeling with 64Cu for PET imaging. Methods The potent GRPr antagonist D-Phe-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH2, was conjugated to the sarcophagine (3,6,10,13,16,19-hexaazabicyclo[6.6.6] icosane=Sar) derivative MeCOSar via PEG4 (LE1) and PEG2 (LE2) spacers and radiolabeled with 64Cu. IC(50) values were determined using 125I-Tyr4-BN as radioligand. Antagonist potency was determined by Ca2+-flux studies. In vivo-studies were performed in PC-3 tumor-bearing nude mice. Results Both conjugates exhibited high affinity to GRPr ( natCu-LE1: 1.4±0.1 nM and natCu-LE2: 3.8±0.6 nM). Ca2+-flux measurements confirmed their antagonist properties. The peptides were stably radiolabeled with 64Cu, with labelling yields >95% and specific activities of about 100 MBq/nmol. Biodistribution revealed specific targeting of the tumor and GRPr-positive organs, Cu-64-LE1 exhibiting the highest tumor uptake of 19.6 ± 4.7 % IA/g at 1 h p.i.. Biodistribution at later time points showed increasing tumor-to-background ratios with time. This was illustrated by the acquisition of PET images at 4 and 24 h p.i. showing high tumor-to-normal tissue contrast. Conclusions This study demonstrates the high affinity of the MeCOSar-PEGx-bombesin conjugates to GRPr. The stable encapsulation of 64Cu by MeCOSar, the long half-life of 64Cu and the suitable biodistribution profile of the 64Cu-labeled peptides lead to PET images of high contrast and to a potential translation into the clinic. Research Support This work was supported by the German Consortium for Translational Cancer Research (DKTK).
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versatile new bis thiosemicarbazone bifunctional chelators synthesis conjugation to bombesin 7 14 nh2 and Copper 64 radiolabeling
Inorganic Chemistry, 2010Co-Authors: Brett M Paterson, John A Karas, Denis B Scanlon, Jonathan M White, Paul S DonnellyAbstract:New bifunctional derivatives of diacetyl-bis(4-methylthiosemicarbazone) (H2atsm) have been prepared by a selective transamination reaction of a new dissymmetric bis(thiosemicarbazone) precursor H2L1. The new derivatives contain an aliphatic carboxylic acid (H2L2 and H2L3), t-butyl carbamate (H2L4), or ammonium ion (H2L5) functional group. The new ligands and Copper(II) complexes have been characterized by NMR spectroscopy, mass spectrometry, and microanalysis. The complex CuII(L4) was structurally characterized by X-ray crystallography and shows the metal center to be in an N2S2 distorted square planar coordination geometry. Electrochemical measurements show that the Copper(II) complexes undergo a reversible reduction attributable to a Cu(II)/Cu(I) process. The ligands and the Copper(II) complexes featuring a carboxylic acid functional group have been conjugated to the tumor targeting peptide bombesin(7−14)-NH2. The bifunctional peptide conjugates were radiolabeled with Copper-64 in the interest of develo...