Suzanne V. Smith

Capture of Radioactive Cesium and Iodide Ions from Water by Using Titanate Nanofibers and Nanotubes

Radioactive Cs and I ions are the products of uranium fission, and can be easily dissolved in water during an accident at a nuclear reactor, such as those that occurred at Chernobyl in 1986, at Three Mile Island in Pennsylvania in 1979, and in 2011 at Fukushima, Japan. In 2009, leaks of radioactive materials such as Cs and I isotopes also occurred during minor accidents at nuclear power stations in Britain, Germany, and the U.S. These leaks have raised concerns about exposure levels in the nearby communities because it is feared that these fission products could make their way into the food chain when present in waste water. Radioactive iodine is also used in the treatment of thyroid cancer, and, as a result, radioactive wastewater is discharged by a large number of medical research institutions. The wide use of radioisotopes requires effective methods to manage radioactive waste, and methods currently used are complex and extremely costly. Herein we demonstrate a potentially cost-effective method to remediate Cs and I ions from contaminated water by using the unique chemistry of titanate nanotubes and nanofibers, which can not only chemisorb these ions but efficiently trap them for safe disposal. Inorganic cation exchangers, such as crystalline silicotitanates, zeolites, clay minerals, layered Zr phosphates, and layered sulfide frameworks, have been studied for separation of Cs ions from nuclear wastewater and safe disposal of the exchanged cations because of the ability of these exchangers to withstand intense radiation and elevated temperatures, in addition to their high ion-exchange capacity. Because ion exchange in materials is usually a reversible process, except in micas, the radioactive ions in the exchanger may be released to water. Titanates are refractory mineral substances that are very stable with respect to radiation and chemical, thermal, and mechanical changes. Titanate nanofibers and nanotubes (with chemical formula Na2Ti3O7) can be easily synthesized at low cost under hydrothermal conditions. These materials possess a layered structure in which TiO6 octahedra are the basic structural units (Figure S1 in the Supporting Information). These layers carry negative charges and are approximately two oxygen atoms thick. Na ions are situated between the layers and can be exchanged with other cations. In the present study, we show how trititanate nanofibers (T3NF) and nanotubes (T3NT) can be used to efficiently remove radioactive Cs ions from aqueous solution by cation exchange. Figure 1a shows that the nanotubular T3NT can remove 80% of Cs ions from solutions with Cs concentrations up to 250 ppm. The ions can be completely removed when the Cs ion concentration is below 80 ppm. In contrast, the fibril T3NF has a comparatively lower absorption capacity than

Positron emission tomography (PET) imaging of neuroblastoma and melanoma with 64Cu-SarAr immunoconjugates

The advancement of positron emission tomography (PET) depends on the development of new radiotracers that will complement 18F-FDG. Copper-64 (64Cu) is a promising PET radionuclide, particularly for antibody-targeted imaging, but the high in vivo lability of conventional chelates has limited its clinical application. The objective of this work was to evaluate the novel chelating agent SarAr (1-N-(4-aminobenzyl)-3, 6,10,13,16,19-hexaazabicyclo[6.6.6] eicosane-1,8-diamine) for use in developing a new class of tumor-specific 64Cu radiopharmaceuticals for imaging neuroblastoma and melanoma. The anti-GD2 monoclonal antibody (mAb) 14.G2a, and its chimeric derivative, ch14.18, target disialogangliosides that are overexpressed on neuroblastoma and melanoma. Both mAbs were conjugated to SarAr using carbodiimide coupling. Radiolabeling with 64Cu resulted in >95% of the 64Cu being chelated by the immunoconjugate. Specific activities of at least 10 μCi/μg (1 Ci = 37 GBq) were routinely achieved, and no additional purification was required after 64Cu labeling. Solid-phase radioimmunoassays and intact cell-binding assays confirmed retention of bioactivity. Biodistribution studies in athymic nude mice bearing s.c. neuroblastoma (IMR-6, NMB-7) and melanoma (M21) xenografts showed that 15–20% of the injected dose per gram accumulated in the tumor at 24 hours after injection, and only 5–10% of the injected dose accumulated in the liver, a lower value than typically seen with other chelators. Uptake by a GD2-negative tumor xenograft was significantly lower (<5% injected dose per gram). MicroPET imaging confirmed significant uptake of the tracer in GD-2-positive tumors, with minimal uptake in GD-2-negative tumors and nontarget tissues such as liver. The 64Cu-SarAr-mAb system described here is potentially applicable to 64Cu-PET imaging with a broad range of antibody or peptide-based imaging agents.

ATP-dependent copper transport by the Menkes protein in membrane vesicles isolated from cultured Chinese hamster ovary cells

The Menkes (MNK) protein is a vital component of copper homeostasis in mammalian cells. In this paper we provide the first biochemical evidence that the MNK protein functions as a copper‐translocating P‐type ATPase in mammalian cells. The enzyme activity in membrane vesicles prepared from Chinese hamster ovary cells overexpressing MNK was ATP‐dependent, correlated with the amount of MNK and followed Michaelis‐Menten kinetics with respect to copper. The copper transport was observed only under reducing conditions suggesting MNK transports Cu(I). This study opens the way to detailed structure‐function studies and assessment of functional MNK derived from patients with Menkes disease.

Synthesis of a new cage ligand, SarAr, and its complexation with selected transition metal ions for potential use in radioimaging

A new hexaazamacrobicyclic cage ligand, 1-N-(4-aminobenzyl)-3,6,10,13,16,19-hexaazabicyclo[6.6.6]eicosane-1,8-diamine (SarAr) has been designed for conjugation to proteins. SarAr was synthesised and characterised by microanalyses, 1H NMR and electrospray mass spectrometry. The complexation of selected transition metal ions (Cu(II), Ni(II) and Co(II) at 10−6 M) by SarAr was complete within 30 min over pH 6 to 8. The [64Cu(SarAr)]2+ complex was investigated with a view to applications in radioimaging. The [64Cu(sar)]2+ complex was found to be stable in human plasma for at least 174 h and biodistribution studies in mice, showed that the [64Cu(SarAr)]2+complex was rapidly excreted through the renal system unlike the free 64Cu2+. Overall, the simple synthesis, ready complexation behaviour of SarAr, the kinetic inertness of the [Cu(SarAr)]2+ complex to dissociation of 64Cu and its facile elimination from mice make it an attractive prospect for use in nuclear medicine.

Insights into the mechanisms of copper tolerance of a population of black-banded rainbowfish (Melanotaenia nigrans) (Richardson) exposed to mine leachate, using 64/67Cu

This study investigates the mechanisms of copper tolerance of a population of black-banded rainbowfish (Melanotaenia nigrans) (Richardson). The population has been exposed to elevated copper concentrations for over 40 years, due to leachate from the Rum Jungle uranium/copper mine. At the time of collection the 96 h EC(50) of exposed [E] fish was 8.3 times higher than that of reference [R] fish. The bioconcentration of 64/67Cu in fish was used to investigate the mechanism of copper tolerance in E fish. Both E and R fish were exposed to low (L(Cu), 30 microg Cu l(-1)) and elevated (E(Cu), 300 microg Cu l(-1)) copper concentrations for 24 and 48 h, respectively. Radioactivity was measured at seven or eight time points in four tissue sections: head (including gills, heart and brain), internal organs (including gastrointestinal tract, liver, kidneys and gonads), muscle and whole body. One-compartment bioconcentration models were fit to data and compared using an F-test. Copper concentrations in all tissue sections were significantly (P<0.05) less (up to 50%) in E fish compared with the respective tissue sections of R fish when exposed to both L(Cu) and E(Cu). The exception was copper accumulation in the internal organs, which was not significantly different between E and R fish exposed to E(Cu). The mechanism of copper tolerance was concluded to be reduced copper uptake in the gills, rather than increased binding or elimination. Allozyme electrophoresis was performed to determine if genetic selection had occurred in the E fish population. Allozyme frequencies at the AAT-1 and GPI-1 loci were significantly (P<0.05) different between E and R fish. Heterozygosity was reduced in E fish compared with that of R fish. Collectively these results suggest that genetic selection may have occurred in the E fish population. Consequently, the selection of allozymes less sensitive to copper may be another mechanism of copper tolerance of E fish. This is the first study on the mechanisms of copper tolerance in a wild fish population that has been exposed to elevated copper concentrations. These findings aid the understanding of metal tolerance in fish and emphasise the importance of sample selection and its implication for toxicity testing.

Radiolabeling Antibodies with Holmium-166

We report the preliminary results from radiolabeling of a chelate-conjugated antibody with 166Ho produced from the beta(-)-decay of 166Dy. Ho-166 was separated from mg quantities of Dy target by reverse phase ion-exchange chromatography employing a cation exchange HPLC column and 0.085 M alpha-HIBA at pH = 4.3 as eluent. Evaporation to dryness of 166Ho fraction (up to 25 mL) and thermal decomposition of alpha-HIBA yielded 166Ho in a dry state which was then solubilized in 0.5 mL of 0.1 M HCl. Subsequent radiolabeling of CHX-B-DTPA conjugated 135-14 monoclonal antibodies with purified 166 Ho was readily achieved with approximately 80% efficiency and with a specific activity of 3-4 mCi of 166Ho per mg of protein. 166Ho-antibody conjugates are stable with regards to transferrin challenge for a period of 50 h. Further, it was shown that any Fe3+ ions present in alpha-HIBA as an impurity interfere with the labeling.

[166Dy]Dysporium/[166Ho]Holmium In Vivo generator

A novel approach for the delivery of 166Ho (t1/2 = 26.6 h) to tissue is via the in vivo decay of its 81.5 h parent, 166Dy-an in vivo generator system. A critical question for the in vivo 166Dy/166Ho generator system is whether translocation of the daughter nucleus occurs. The in vitro and in vivo integrity of the [166Dy]Dy/166Ho-DTPA complex was investigated and results indicated that no translocation of the daughter nucleus occurs subsequent to beta- decay of 166Dy. Biodistribution studies of [166Dy]Dy-DTPA showed that the ratio of 166Dy/166Ho in bone remains constant (+/- 7%) over a 20 h period, indicating no significant in vivo loss of 166Ho from the complex. Increasing the in vivo residence time of [166Dy]Dy-DTPA complex attached to HSA gave similar results.

The ionic charge of copper-64 complexes conjugated to an engineered antibody affects biodistribution

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.

Molecular imaging with copper-64 in the drug discovery and development arena

The contribution of positron emission tomography (PET) to the drug discovery and development (D3) pipeline has been inhibited by the short half-lifes of PET radioisotopes, 11C and 18F, poor availability and the high cost of infrastructure. Copper-64 (64Cu) has a 12.7 h half-life, simple yet flexible radiochemistry and imaging characteristics that make it ideal for a wider application across the D3 arena. Recent scientific breakthroughs in the production of 64Cu show that its, commercial production can be made more widely available. More importantly, for pharmaceutical research and development programmes wishing to incorporate the high sensitivity and spatial resolution of PET, but no desire to implement and maintain expensive radiochemistry infrastructure, 64Cu is an exciting option.

Optimizing radiolabeling amine-functionalized silica nanoparticles using SarAr-NCS for applications in imaging and radiotherapy

Silica nanoparticles functionalized with amine groups and in the size range of approximately 60-94 nm were produced by combining sol-gel processing and emulsion technology. Hexa-aza cage ligand SarAr-NCS was conjugated to the silica nanoparticles and subsequently radiolabeled with a solution of (57)Co(2+)-doped carrier Co(2+). The number of Co(2+) ions bound to the silica particles at pH 7 was used to determine the average number of available SarAr-NCS ligands conjugated to a silica particle. For organically modified silica particles of 94.0 and 59.5 nm diameter, the maximum number of metal binding sites was determined to be 11700 and 3270 sites per particle, respectively. For silica particles (63.5 nm peak diameter) produced using an water-in-oil emulsion, the calculated average was 4480 on the particle surface. The number of SarAr-NCS conjugated on the particles was easily controlled, potentially providing for a range of products for applications in the risk assessment of particles and theranostic imaging or radiotherapy when radiolabeled with a suitable radioisotope such as (64)Cu or (67)Cu.