The localisation and micro-mapping of copper and other trace elements in breast tumours using a synchrotron micro-XRF system

Trace elements have critical roles in cancer biology. The quantity and distribution of the elements Cl, Ca, K, P, S, Ti, Fe, Cu and Zn in samples of primary breast cancer have been assessed. The samples were formalin fixed tissue specimens formatted as microarrays of cores 1.0 mm diameter and 10 μm thick each. The data were obtained using a synchrotron X-ray fluorescence microprobe system. The spatial resolution of elemental maps was approximately 20 μm. Maps were compared with light transmission images of the samples and then the images were stained for cancer. The synchrotron system proved successful in producing data that could be mapped into high-resolution images where clear structure could be identified. Correlation of these distributions with the concentrations of cancer cells was achieved in some samples.     

Compton suppression method and epithermal NAA in the determination of nutrients and heavy metals in Nigerian food and beverages

We used in this study Compton suppression method and epithermal neutron activation analysis to determine the concentration of nutrients and heavy metals in Nigerian food and beverages. The work was performed at the University of Texas TRIGA Reactor by short, medium, and long irradiation protocols, using thermal flux of 1.4×10¹² n cm^?2 s^?1 and epithermal flux of 1.4×10¹¹ n cm^?2 s^?1. Application of Compton suppression method has reduced interferences from Compton scattered photons thereby allowing easy evaluation of Na, Cl, Ca, Cu, Mn, Mg, Co, Cr, Rb, Fe, and Se. The epithermal NAA method has enabled determination of Cd, As, Ba, Sr, Br, I, and V with little turn-around time. Quality Control and Quality Assurance of the method was tested by analyzing four Standard Reference Materials (non-fat powdered milk, apple leaves, citrus leaves, and peach leaves) obtained from National Institute for Standards and Technology. Our results show that sorghum, millet, and maize have high values of Zn, Mn, Fe, low values of Cd, As, and Se. Powdered milks, rice, beans, and soybeans were found to have moderate amounts of all the elements. Tobacco recorded high content of Cd, Mn, and As, whereas tea, tsobo leaves, Baobab leaves, and okro seed have more As values than others. However, biscuits, macaroni, spaghetti, and noodles show lower concentrations of all the elements. The distribution of these nutrients and heavy metals in these food and beverages shows the need to fortify biscuits and pastas with micro and macro-nutrients and reduce the use of tobacco, tea, tsobo leaves, Baobab leaves, and Okro seed to avoid intake of heavy elements.     

Investigation of the natZn(p,x)62Zn nuclear process up to 70 MeV: a new 62Zn/62Cu generator

The excitation function of the ^natZn(p,x)⁶²Zn nuclear process was measured by the stacked-foil technique up to a proton energy of 70 MeV to obtain accurate data for production of the ‘mother nuclide’ (⁶²Zn) of the PET related β⁺ emitting radioisotope ⁶²Cu. Investigations were also made on the ⁶⁶Zn(p,x)⁶²Zn and ^natZn(p,xn)⁶⁶Ga processes and on the ⁶⁶Zn(p,n)⁶⁶Ga reaction using ^natZn and highly enriched ⁶⁶Zn. The excitation functions were compared with the published data. Thick target yields for the ^natZn(p,x)⁶²Zn and ^natCu(p,xn)⁶²Zn processes were also calculated up to 70 MeV. On the basis of these calculations the ^natZn+p process results in higher yield for ⁶²Zn above 50 MeV than the ^natCu+p process. The latter process is presently used for practical production of ⁶²Zn. In an energy window from 70 to 30 MeV the available EOB yield of the ^natZn+p reactions is around 19 mCi/μA h (0.7 GBq/μAh) that makes the ^natZn(p,x)⁶²Zn process a good candidate for routine generator production.     

Simultaneous detection of As and Se in polyester resin skin phantoms

It is known that As and Se act as metabolic antagonists. Hence, an improvement in assessing As-related health risks can be achieved by simultaneous quantitative measurements of both As and Se levels in the human body. In this paper, the simultaneous detection of trace concentrations of As and Se in polyester resin skin phantoms was demonstrated. The experiments were performed with a commercial miniature X-ray tube and silicon PiN detector X-ray fluorescence (XRF) system. No significant overlap between the K_α peaks of the two elements was observed. Minimum detection limits of (1.05±0.02) μg As g^?1 and (0.88±0.02) μg Se g^?1 were found.     

Performance of a 62Zn/62Cu microgenerator in kit-based synthesis and delivery of [62Cu]Cu–ETS for PET perfusion imaging

The performance of a commercially produced ⁶²Zn/⁶²Cu microgenerator system, and an associated kit-based radiopharmaceutical synthesis method, was evaluated for clinical site production of [⁶²Cu]Cu–ETS (ethylglyoxal bis(thiosemicarbazonato)copper(II)), an investigational agent for PET perfusion imaging. Using 37 generators, containing 1.84±0.23 GBq ⁶²Zn at 9:00 AM on the day of clinical use, a total of 45 patient doses of [⁶²Cu]Cu–ETS (672±172 MBq) were delivered without difficulty. ⁶²Cu elution yields were high (approximately 90%), accompanied by extremely low ⁶²Zn breakthrough (<0.001%). Radiopharmaceutical preparation, from the start-of-elution to time-of-injection, consumed less than five minutes. The ⁶²Zn/⁶²Cu microgenerator was a dependable source of short-lived positron-emitting ⁶²Cu, and the kit-based synthesis proved to be rapid, robust, and highly reliable for “on-demand” delivery of [⁶²Cu]Cu–ETS for PET perfusion imaging.     

Evaluation of the chromium contamination at Ribeirão dos Bagres,Franca (SP), Brazil,by the 210Pb method

This paper presents a chronological study focusing the use of ²¹⁰Pb in four sediment cores collected at Ribeirão dos Bagres in the region of Franca, São Paulo State, Brazil. When evaluating the sedimentation rate by using the ²¹⁰Pb method, it was possible to monitor historical changes occurring in the concentrations of chromium present in the sediments. In order to do so, several specific parameters were determined in sediments, as chromium concentration, major oxides, organic matter, grain size, and chemical and physical–chemical parameters of waters. The sediments possessed a Cr concentration range of 0.83–26.25 mg/kg that is below the world's average of shales. The sedimentation rates obtained from the ²¹⁰Pb analysis were between 0.07 and 0.77 g cm⁻² year⁻¹, while the linear sedimentation rate was from 0.6 to 3.2 cm/year. The adoption of the CIC (Constant Initial Concentration) ²¹⁰Pb model allowed characterize older sediments, aged 177 years, and younger sediments, aged 4 years. The organic matter in the sediments was more abundant in the first few centimeters of practically all profiles, and the analysis of oxides showed that silica is the major constituent of the sediments, in which there is predominance of sand and silt. In addition, the analysis of the physical and chemical parameters of waters indicated that they are suitable for navigation and natural harmony landscape.     

Methodology for labeling proteins and peptides with lead-212 (212Pb)

IntroductionAlpha particles possess an exquisite degree of cytotoxicity when employed for targeted α-particle therapy (TAT) or radioimmunotherapy (RIT). ²¹²Pb, which acts as an in vivo generator of the α-emitting nuclide ²¹²Bi has shown great promise in pre-clinical studies when used to label the HER2 binding antibody, trastuzumab. Currently, the first RIT clinical trial employing ²¹²Pb radiolabeled trastuzumab is in progress. This report provides detailed current protocol operations and steps that were generated for use in the clinical trial as well as the relevant pre-clinical experimentation, and describes in detail the labeling of proteins or peptides with ²¹²Pb as provided via a ²²⁴Ra based generator system.Methods²¹²Pb was eluted from the ²²⁴Ra/²¹²Pb generator using hydrochloric acid (2 M). The generator eluate was evaporated and digested with nitric acid (8 M) followed by extraction of the ²¹²Pb with dilute nitric acid (0.1 M). The dilute nitric acid solution of ²¹²Pb was used to label the immunoconjugate Trastuzumab-TCMC (2-(4-isothiocyanatobenzyl-1,4,7,10-tetraaza-1,4,7,10,tetra-(2-carbamonylmethyl)-cyclododecane) at pH 5.5.ResultsElution of ²¹²Pb from the generator was efficient yielding > 90% of available ²¹²Pb. Trastuzumab-TCMC was efficiently labeled with a radiochemical yield of 94% ± 4% (n = 7) by ITLC and an isolated yield of 73% ± 3% (n = 7).ConclusionsThe results show the feasibility of generating radioimmunoconjugates and peptide conjugates for use as in vivo α generator systems in the clinic. The technology holds promise in applications involving the treatment of minimal disease such as micrometastases and residual tumor after surgical debulking, hematological cancers, infections, and compartmental cancers, such as ovarian cancer.     

Temporal variations of 7Be, 210Pb and 137Cs in moss samples over 14 month period

Concentrations of ²¹⁰Pb, ⁷Be and ¹³⁷Cs in moss samples were continuously measured, using low-background HPGe spectrometer at Novi Sad, Serbia (45°14′45″N, 19°51′35″E). Weekly data collected over 14 month period from January 2007 to March 2008 are presented and discussed. Measured values of ⁷Be activity concentrations in dry moss samples are ranged from 201 to 920 Bq/kg showing prominent increase in summer and autumn season. Data for ²¹⁰Pb, ranged from 347 to 885 Bq/kg do not show such trend. The average concentration of ¹³⁷Cs is 8.9 Bq/kg. It is shown that yield of ⁷Be can be estimated using average values of activity concentrations. Precipitation amount and duration of precipitation are measured and their possible influence on activity concentrations measured in moss samples was considered.     

Characteristics of proton beam scanning dependent on Li target thickness from the viewpoint of heat removal and material strength for accelerator-based BNCT

This study demonstrates the characterization of proton spot scanning on a Li target assembly for accelerator-based BNCT from the viewpoint of heat removal and material strength. These characteristics are investigated as to their dependence on the Li target thickness, considering that the Cu backing plate has more suitable heat removal properties than Li. Two situations are considered in this paper, i.e. the cyclic operation of the spot scanning, and a stalled spot scanning cycle where the proton beam stays focused on a single position on the Li target.It was found that the maximum of the Li temperature and the strain of the Cu backing increase as the cycle period increases. A cycle period less than 120 ms (over 8.3 Hz of frequency) enables the Li temperature to be kept below 150 °C and a cycle of less than 115 ms (8.7 Hz) keeps the Cu strain below the critical value for a 230 μm thick Li target, though the values are evaluated conservatively. Against expectation, the Li temperature and Cu strain are larger for a 100 μm thick target than for a 230 μm target. The required cycle period in this case is 23 ms (43 Hz) for maintaining a reasonable Li temperature and 9 ms (110 Hz) to prevent Cu fatigue fracture.For a stall in the spot scanning cycle, the Cu temperature increases as the beam shutdown time increases. The time for Cu to reach its melting point is estimated to be 4.2 ms at the surface, 20 ms at 1 mm depth, for both of 100 and 230 μm thick targets. At least 34 ms is estimated to be enough to make a hole on Cu backing plate. A beam shutdown mechanism with a response time of about 20 ms is therefore required.     

Enrichment and vertical profiles of 210Po and 210Pb in monazite areas of coastal Karnataka

A study on radiation level and radionuclide distribution in the environment of coastal Karnataka has revealed the presence of low-level monazite deposit in the Ullal beach area. The paper presents systematic studies on the distribution, enrichment and vertical profiles of ²¹⁰Po and ²¹⁰Pb, important daughter products of ²³⁸U, in Ombattu Kere, Summer Sand and the Bhagavathi Temple region of the Ullal beach area of coastal Karnataka. Sand samples collected at different depths from these locations were analyzed for ²¹⁰Po and ²¹⁰Pb activities to understand the distribution, enrichment and vertical profiles of these radionuclides in monazite area. The activity of ²¹⁰Po in the Ullal region is found to vary from 1.7 to 43.2 Bq kg^?1 with a mean value of 11.2 Bq kg^?1 and that of ²¹⁰Pb varies from 1.0 to 66.7 Bq kg^?1 with a mean value of 19.1 Bq kg^?1. The mean ²¹⁰Po/²¹⁰Pb ratio was observed to be 0.6. The absorbed gamma dose rate in the region varies in the range 39–460 nGy h^?1 with a mean value of 193 nGy h^?1.     

Effective separation method of 64Cu from 67Ga waste product with a solvent extraction and chromatography

A simple chemical process with a solvent extraction was investigated as an effective separation method for ⁶⁴Cu radionuclide from waste production, which is collected as solution after extracting ⁶⁷Ga and recovering ⁶⁸Zn target materials. For the production of radionuclide ⁶⁷Ga, the enriched ⁶⁸Zn material electroplated on Cu backing plate is usually exposed to energetic protons. The protons produce ⁶⁷Ga including other radionuclides, such as ⁵⁷Ni, ^57,55Co, ^64,67Cu by several nuclear reactions. After extracting ⁶⁷Ga and recovering ⁶⁸Zn through several steps of chemical processes, the residual solution is usually discarded even though it contains other species of radioisotopes. In this study, a simple chemical process having a high separation efficiency of ⁶⁴Cu from the waste solution was investigated. With this method, a promising radiotracer as a diagnostic in PET and a therapeutic in radio-immunotherapy, ⁶⁴Cu was estimated to be produced as high as 1,200 mCi at EOB within 3 h chemical processing after extraction of ⁶⁷Ga and ⁶⁸Zn.     

Simultaneous production of high specific activity 64Cu and 61Co with 11.4 MeV protons on enriched 64Ni nuclei

The ⁶⁴Cu and ⁶¹Co radionuclides were produced simultaneously by irradiation of enriched ⁶⁴Ni on a low energy proton-only cyclotron. Nickel targets were prepared by electrodeposition of enriched ⁶⁴Ni (>95%) on Au backing at thicknesses of 25–225 mg/cm² with efficiencies >99%. Irradiations up to 30 μA for 8 h were performed with 11.4 MeV protons using a water-cooled target mounting. Radiochemical separation of ⁶⁴Cu and ⁶¹Co from ⁶⁴Ni was performed by chromatography of the chlorocomplexes in a single step using an anion exchange resin column with a yield >95%. Using this method, the Ni target material was recovered and re-plated for subsequent production runs with an overall efficiency >96%. The excitation function for the ⁶⁴Ni(p,n)⁶⁴Cu reaction was measured and compared with published values. Experimental thick target saturation yields of 159 mCi/μA for ⁶⁴Cu and 715 μCi/μA for ⁶¹Co were achieved. Typical specific activities of ⁶⁴Cu were found to be 18.8±3.3 Ci/μmol.     

Near-threshold 7Li(p,n)7Be neutrons on the practical conditions using thick Li-target and Gaussian proton energies for BNCT

The near threshold ⁷Li(p,n)⁷Be neutrons generated by incident proton energy having Gaussian distribution with mean energies from 1.85 to 1.95 MeV, were studied as a practical neutron source for BNCT wherein an RFQ accelerator and a thick Li-target are used. Gaussian energy distributions with the standard deviation of 0, 10, 20 and 40 keV for mean proton energies from 1.85 to 1.95 MeV were surveyed in 0.01 MeV increments. A thick liquid Li-target whose dimensions were established in our previous experiments (i.e., 1 mm-thick with 50 mm width and 50 mm length) was considered in this study. The suitable incident proton energy and physical dimensions of Pb layer which serves as a gamma absorber and a Polyethylene layer which is used as a BDE were surveyed by means of the concepts of TPD. Dose distribution were calculated by using MCNP5. A proton beam with mean energy of 1.92 MeV and a Gaussian energy distribution with a standard deviation of 20 keV at a current of 10 mA was selected from the viewpoint of irradiation time and practically achievable proton current. The suitable thicknesses of Pb gamma absorber was estimated to be about 3 cm. The estimated thickness of the polyethylene BDE was about 24 mm for an ideal proton current of 13 mA, and was 18 mm for a practical proton current of 10 mA.     

High tumor uptake of 64Cu: Implications for molecular imaging of tumor characteristics with copper-based PET tracers

IntroductionThe use of copper-based positron emission tomography (PET) tracers in cancer studies is increasing. However, as copper has previously been found in high concentrations in human tumor tissue in vivo, instability of PET tracers could result in tumor accumulation of non-tracer-bound radioactive copper that may influence PET measurements. Here we determine the degree of ⁶⁴Cu uptake in five commonly used human cancer xenograft models in mice. Additionally, we compare copper accumulation in tumor tissue to gene expression of human copper transporter 1 (CTR1).MethodsSmall animal PET scans were performed on five different human cancer xenograft mice models 1 h and 22 h post injection (p.i.) of ⁶⁴CuCl₂. Regions of interest (ROIs) were drawn on tumor tissue and sections of various organs on all images. Quantitative real-time PCR (qPCR) gene expression measurements of CTR1 were performed on tumor samples obtained after the 22 h scan.ResultsA relatively high tumor uptake of ⁶⁴Cu was seen in four out of five tumor types and an increase in ⁶⁴Cu accumulation was seen in three out of five tumor types between 1 h and 22 h p.i. No relationship was found between tumor uptake of ⁶⁴Cu and gene expression of CTR1.ConclusionsThe relatively high, time- and tumor type dependent ⁶⁴Cu uptake demonstrated here in five different human cancer xenograft models in mice, emphasizes the importance of validating tracer uptake and indicates that high in vivo stability of copper-based PET tracers is of particular importance because non-tracer-bound copper can accumulate in tumor tissue to a level that could potentially lead to misinterpretation of PET data.     

Radiolabeling of HTE1PA: A new monopicolinate cyclam derivative for Cu-64 phenotypic imaging. In vitro and in vivo stability studies in mice

IntroductionHTE1PA, a monopicolinate-N-alkylated cyclam-based ligand has previously demonstrated fast complexation process, high kinetic inertness and important thermodynamic and electrochemical stability with respect to natural copper. In this work we first developed a new synthetic route to obtain HTE1PA in good yields. Then, we investigated HTE1PA chelation properties towards copper-64 and assessed in vitro and in vivo stability of the resulting compound.MethodsRadiolabeling of HTE1PA with copper-64 was tested at different ligand concentrations in ammonium acetate medium. In vitro stability study was carried out by incubating [⁶⁴Cu]TE1PA complex in human serum at both 37 °C and 4 °C; chromatographic controls were performed over 24 h. Biodistribution, pharmacokinetic and hepatic metabolism of [⁶⁴Cu]TE1PA were conducted in BALC/c mice in comparison with [⁶⁴Cu]acetate and [⁶⁴Cu]DOTA, used as a reference ligand.ResultsThe promising results obtained for natural copper complexation were confirmed. HTE1PA was quantitatively radiolabeled in 15 min at room temperature. The resulting complex showed high serum stability. [⁶⁴Cu]TE1PA induced a significant uptake in the liver and kidneys at early biodistribution time point. Nevertheless, a high speed wash out was observed at 24 h leading to significantly lower uptake into the liver compared to [⁶⁴Cu]DOTA. The metabolism study was consistent with a high resistance to transchelation as the initial uptake into liver matches with the intact form of [⁶⁴Cu]TE1PA.ConclusionDespite the partial elimination of HTE1PA – as copper-64 complex – through the hepatic route, its high selectivity for copper and its resistance to transchelation make it a promising ligand for antibody radiolabeling with either copper-64 or copper-67.     

Preparation of 65Zn and 134Cs sealed sources based on commercial organic and synthesized inorganic sorbents for calibration purposes of γ-detectors

The sealed source of ⁶⁵Zn was prepared by using permutit as a commercial organic resin in an anion form, while in case of the ¹³⁴Cs, the sealed source was prepared by using the synthesized 12-molybdocerate(IV) gel as an inorganic exchanger in cation form. The batch technique was carried out for loading ⁶⁵Zn from 0.2 M HCl-60% acetone medium and for loading ¹³⁴Cs from 0.005 M HNO₃ onto the appropriate exchanger. The sealed source material of ⁶⁵Zn-permutit or ¹³⁴Cs-12-molybdocerate(IV), about 100 mg, was placed inside the cavity of a holder capsule (artelone) and its activity was measured to be 56.61±2.59 kBq (1.53±0.07 μCi) for ⁶⁵Zn on 21 April 2008 and 1.48±0.089 kBq (0.04±0.0024 μCi) for ¹³⁴Cs on 5 May 2008. The self absorption was calculated by using artelone and compared with the same thickness of stainless steel capsule as the holder material. The quality control was performed on the prepared sealed sources.     

Excitation function of the 64Ni(α,p)67Cu reaction for production of 67Cu

The excitation function of the ⁶⁴Ni(α,p)⁶⁷Cu reaction was measured from threshold up to 24 MeV in order to investigate the possibility of production of the β⁻-emitting therapeutic radioisotope ⁶⁷Cu (T_1/2=61.9 h). Two stacks of thin metallic self-supporting foils of ⁶⁴Ni (enrichment 77.8%) prepared by electrolytic deposition were irradiated by α-particle beams. The radioactivity was determined via HPGe detector γ-ray spectrometry. Some ⁶⁷Ga activity (which emits the same γ-rays as ⁶⁷Cu), formed via the ^natCu(α,x)⁶⁷Ga process on trace copper impurity in the Ni foils, was also observed. Corrections were done for ⁶⁷Ga activity contribution and for the ⁶⁷Cu activity escape fraction from the thin Ni-foil. The maximum cross section of the ⁶⁴Ni(α,p)⁶⁷Cu reaction amounts to 34 mb at 22 MeV. The experimental results were compared with theoretical predictions. The integral yield of ⁶⁷Cu at 24 MeV α-particle energy, calculated from the excitation function, amounts to 544 KBq/μAh (48.5 MBq/μA at saturation). It is thus a low-yield reaction.     

5-Aminolevulinic acid-based photodynamic therapy of chordoma: In vitro experiments on a human tumor cell line

BackgroundChordomas are very rare tumors of the skull base and the sacrum. They show infiltrating and destructive growth and are known to be chemo- and radio-resistant. After surgical resection, the recurrence rate is high and overall survival limited. As current adjuvant treatments are ineffective, new treatment concepts are urgently needed. 5-aminolevulinic acid-based photodynamic therapy (5-ALA based PDT) showed promising results for malignant gliomas. However, it is unknown so far, whether chordomas accumulate protoporphyrin IX (PPIX) after application of 5-ALA and whether they are sensitive to subsequent 5-ALA based PDT.MethodsThe immortalized human chordoma cells U-CH2 were used as in vitro model. After incubation for 4 h or 6 h with different 5-ALA concentrations, PPIX accumulation was determined by flow cytometry. To assess sensitivity to PDT, chordoma cells were incubated at 30.000 cells/well (high cell density) or 15.000 cells/well (low cell density) with graded doses of 5-ALA (0–50 μg/ml) in 96-well plates and subsequently exposed to laser light of 635 nm wavelength (18.75 J/cm²). Cell survival was measured 24 h after exposure to laser light using the WST-1 assay.ResultsU-CH2 cells dose-dependently accumulated PPIX (ANOVA; p < 0.0001). PPIX fluorescence was significantly higher, when cells were incubated with 5-ALA for 6 h compared to 4 h at higher 5-ALA concentrations (ANOVA/Bonferroni; p ≤ 0.05 for ≥ 30 μg/ml 5-ALA). For both cell densities, a 5-ALA dose-dependent decline in viability was observed (ANOVA; p < 0.0001). Viability was significantly lower at higher 5-ALA concentrations, when 30.000 cells/wells were treated compared to 15.000 cells/well (ANOVA/Bonferroni; p ≤ 0.001 for ≥ 30 μg/ml 5-ALA). LD50 was 30.25 μg/ml 5-ALA.ConclusionThe human UCH-2 cell line was a very useful in vitro model to study different effects of 5-ALA based PDT. For the first time, it could be shown that human chordoma cells may be destroyed by 5-ALA/PDT.