Recommendations for In Vitro and In Vivo Testing of Magnetic Nanoparticle Hyperthermia Combined with Radiation Therapy

Magnetic nanoparticle (MNP)-mediated hyperthermia (MH) coupled with radiation therapy (RT) is a novel approach that has the potential to overcome various practical difficulties encountered in cancer treatment. In this work, we present recommendations for the in vitro and in vivo testing and application of the two treatment techniques. These recommendations were developed by the members of Working Group 3 of COST Action TD 1402: Multifunctional Nanoparticles for Magnetic Hyperthermia and Indirect Radiation Therapy (“Radiomag”). The purpose of the recommendations is not to provide definitive answers and directions but, rather, to outline those tests and considerations that a researcher must address in order to perform in vitro and in vivo studies. The recommendations are divided into 5 parts: (a) in vitro evaluation of MNPs; (b) in vitro evaluation of MNP-cell interactions; (c) in vivo evaluation of the MNPs; (d) MH combined with RT; and (e) pharmacokinetic studies of MNPs. Synthesis and characterization of the MNPs, as well as RT protocols, are beyond the scope of this wor

Investigation of p-SCN-Bn-DOTA-trastuzumab labeled with radioactive and non-radioactive lutetium and yttrium: a crucial step for future applications

The significance lies in preparing stable, trastuzumab-immunoconjugate through the utilization of non-radioactive LuCl3 and YCl3, via p-SCN-Bn-DOTA. This approach is crucial to determine potential physicochemical alterations in immunoconjugate structure following metal binding. Post-conjugation, employing a 1:20 molar ratio, freeze-drying was performed to obtain stable immunoconjugates for subsequent analysis. Several chemical methods were employed to characterize antibody stability and retained immunoreactivity within the formulated immunoconjugates. Proof of protein integrity came from SDS-PAGE electrophoresis, with uniform fragment intensities (25 kDa for light chain, 50 kDa for heavy chain) indicating antibody non-degradation (1). IR and Raman spectroscopy verified secondary structural changes, with the presence of characteristic amide bands in both spectra indicating the retention of native secondary structure (2). Employing MALDI-TOF-MS, 4.9 p-SCN-Bn-DOTA molecules were determined per antibody molecule. The promising outcomes from non-radioactive labeling provide an opportunity for potential labeling with radioactive lutetium-177 and yttrium-90, each with a specific activity of 200 µCi/mL. Radioisotopes were incubated with p-SCN-Bn-DOTA-trastuzumab for an hour at 40 ºC. Evaluation of radiochemical purity and stability was conducted using the ITLC-SG system. Optimal mobile phases, specifically 0.4 M methanol:sodium acetate (1:1) for yttrium-90 and 0.9% NaCl for lutetium-177, facilitated thorough examination. Remarkable radiolabeling efficiency was achieved, >96% for yttrium-90 and >99% for lutetium-177. Stability assessments after 72 hours demonstrated greater stability in 177Lu-p-SCN-Bn-DOTA-trastuzumab (<1.5% lutetium-177 release) compared to the 90Y-labeled counterpart (<17% yttrium-90 release). This study demonstrates the successful development of radioimmunoconjugates, positioning this agent for potential application in vivo investigations

Status Report and Beam Time Request for Experiment AD-4

Summary of current status and plans for October 200

Biological Effects of Antiprotons Are Antiprotons a Candidate for Cancer Therapy?

2009 Status Report of AD-4 Experimen

COST Action CA19114, Network for Optimized Astatine labelled Radiopharmaceuticals

Cancer is a major health concerns for European citizens. Thus, the main research aim of this Network for Optimized Astatine labeled Radiopharmaceuticals (NOAR) COST Action is to successfully demonstrate that one of the most promising radionuclides for Targeted Alpha Therapy (TAT), namely astatine-211, can become the European standard for treatment of certain cancerous pathologies. To this end, an efficient networking is essential among all European stakeholders interested in promoting astatine-211 for medical applications. NOAR COST Action brings together European and international excellence labs, astatine-211 production centers, hospitals, industry and patient associations from more than 20 countries, thus covering the whole value chain of innovation: production, chemistry, radiochemistry, biology, preclinical and clinical research and delivery of radiopharmaceuticals to patients. A European web portal will be created containing information for patients, practitioners, researchers, Industry and as a contact point for National and European patient associations. The idea is to gather forces at the European level in order to implement actions to leverage hurdles to the development of this powerful radionuclide and to identify pathologies in which it will be particularly relevant. A special emphasis will be given to train a new generation of young researchers and PhD students, promoting interdisciplinary competencies through international and inter-sectoral mobility. The long-term goal of this project is to make Astatine-211 technology available to all European citizen

V-79 Chinese hamster cells irradiated with antiprotons, a study of peripheral damage due to medium and long range components of the annihilation radiation

PURPOSE: Radiotherapy of cancer carries a perceived risk of inducing secondary cancer and other damage due to dose delivered to normal tissue. While expectedly small, this risk must be carefully analysed for all modalities. Especially in the use of exotic particles like pions and antiprotons, which annihilate and produce a mixed radiation field when interacting with normal matter nuclei, the biological effective dose far out of field needs to be considered in evaluating this approach. We describe first biological measurements to address the concern that medium and long range annihilation products may produce a significant background dose and reverse any benefits of higher biological dose in the target area. MATERIALS AND METHODS: Using the Antiproton Decelerator (AD) at CERN (Conseil Europeen pour la Recherche Nucleaire) we irradiated V-79 Chinese Hamster cells embedded in gelatine using an antiproton beam with fluence ranging from 4.5 x 10(8) to 4.5 x 10(9) particles, and evaluated the biological effect on cells located distal to the Bragg peak using clonogenic survival and the COMET assay. RESULTS: Both methods show a substantial biological effect on the cells in the entrance channel and the Bragg Peak area, but any damage is reduced to levels well below the effect in the entrance channel 15 mm distal to the Bragg peak for even the highest particle fluence used. CONCLUSIONS: The annihilation radiation generated by antiprotons stopping in biological targets causes an increase of the penumbra of the beam but the effect rapidly decreases with distance from the target volume. No major increase in the biological effect is found in the far field outside of the primary beam

Development of ready to use kit formulation for trastuzumab radioimmunoconjugates and identification of radiochemical purity as the first step in quality control of the final product

Aim The aim of this study is to present the part of our project dedicated to obtaining a stable, ready to use freeze dried kit formulation of antibody radioimmunoconjugates (trastuzumab immunoconjugates labelled with 90Y and 177Lu). As the first step in on-going in vitro stability of the final product and radiochemical purity determination, we used ITLC-SG method with different mobile phases. Methods Radioactive labelling of trastuzumab was performed with 90Y and 177Lu via DOTA, DTPA and 1B4M-DTPA in molar ratio 1:20. The specific activity of 1.425 mCi (90Y) and 8.150 mCi (177Lu) was achieved, using a solutions of 0.04 M HCl. Radiolabeling is performed by adding 8.5 µL of 90Y at pH 4.5-5 and 5 µL 177Lu at pH 6. Solutions with Tr-DTPA and Tr-1B4M-DTPA were incubated at room temperature for 30 min, while Tr-DOTA was incubated at 40 °C for 1 hour. Radiochemical purity of radioisotopes was tested with ITLC-SG using three mobile phases: 0.9% NaCl, 0.4 M methanol/sodium-acetate (1:1) and 0.1 M acetic buffer. The stability of radioimmunoconjugates was tested in 0.9% NaCl (177Lu) and 0.4 M methanol/sodium-acetate (1:1) (90Y), after incubation at room temperature for 1, 24, 48 and 72h. Results After choosing the most suitable mobile phase for determination of radiochemical purity by ITLC-SG of conjugates labeled with 90Y (99.87%) we used 0.4 M methanol/sodium acetate (1:1), and those with 177Lu (100%) with 0.9%. NaCl. Examination of radiochemical yield of radioimmunoconjugates showed the presence of radioactivity only at the start of the strip, due to the high Mw of Tr. The absence of radiolabeled fragments of the antibody, as well as radiolabeled chelators and free radioisotopes, proved that the stable radioimmunoconjugates were formulated. The highest yield of labeling with 90Y (>96%) is achieved in 0.4 M methanol/sodium-acetate (1:1), while with 177Lu (>99%) in 0.9% NaCl. Test stability after 24h showed the highest stability of 90Y-DOTA-Tr (>92.40%) and 177Lu-DOTA-Tr (>99.14%), with minimum released 90Y3+ (84.90%) and 177Lu-DOTA-Tr (>98.52%), with minimum released 90Y3+ (<15.10%) and 177Lu3+ (<1.48%). Conclusions After obtaining the final ready to use kit formulation, the results of the determination of radiochemical purity using ITLC-SG show a high radiolabeling efficiency (>95%), using both isotopes. However, radioactive yield with 177Lu (99%) was higher compared with 90Y (>96%). This method was used to monitor the stability of radiolabeled conjugates and after 72 hours of incubation, a small amount of free radioisotopes was released from radioimmunoconjugates (<5% of 177Lu and <25% of 90Y). The next planed step includes in vivo examinations in healthy mice and in a mouse model of HER2 positive breast tumor after the i.v. injection of radiolabeled trastuzumab radioimmunoconjugates in order to monitor and determine their pharmacokinetics and biodistribution in the whole body and critical organs/tumor

A single drop histamine sensor based on AuNPs/MnO2 modified screen-printed electrode

In order to ensure high food quality, one of the prime importance is the detection and quantification of histamine, well known marine food poison. In this work, we constructed novel electrochemical biosensor for the detection of histamine based on gold nanoparticles decorated on manganese dioxide (Au/MnO2) and used for modification of screen-printed carbon electrode (Au/MnO2@SPCE). The constructed sensor was then used for the estimation of histamine content in a single drop. Materials used in this study were synthesized and characterized using HR-TEM, XRPD and electrochemical methods. The amperometric detection method was optimized and, under selected operating parameters (supporting electrolyte pH 6, working potential of 1 V), the proposed sensor possesses linear working range from 0.3 mu M to 5.1 mu M, with a detection limit of 0.08 mu M. The effect of selected interferences was investigated and it was found that the developed approach offers accurate, precise, selective, fast and reproducible quantification of histamine using only one drop of the sample. In the end, this work stands as a proof-of-concept of the modified electrodes and electrochemical detection as a promising and prospective approach for the applications in real-time monitoring of the food quality.Ministry of Education, Science and Technological Development of the Republic of Serbia, through Eureka project [E!13303]info:eu-repo/semantics/publishedVersio

Complementary approaches for the evaluation of biocompatibility of Y-90-labeled superparamagnetic citric acid (Fe,Er)(3)O-4 coated nanoparticles

Magnetic nanoparticles (MNPs) are of immense interest for diagnostic and therapeutic applications in medicine. Design and development of new iron oxide-based MNPs for such applications is of rather limited breadth without reliable and sensitive methods to determine their levels in body tissues. Commonly used methods, such as ICP, are quite problematic, due to the inability to decipher the origin of the detected iron, i.e. whether it originates from the MNPs or endogenous from tissues and bodily fluids. One of the approaches to overcome this problem and to increase reliability of tracing MNPs is to partially substitute iron ions in the MNPs with Er. Here, we report on the development of citric add coated (Fe,Er)(3)O-4 nanopartides and characterization of their physico-chemical and biological properties by utilization of various complementary approaches. The synthesized MNPs had a narrow (6-7 nm) size distribution, as consistently seen in atomic pair distribution function, transmission electron microscopy, and DC magnetization measurements. The particles were found to be superparamagnetic, with a pronounced maximum in measured zero-field cooled magnetization at around 90 K. Reduction in saturation magnetization due to incorporation of 1.7% Er3+ into the Fe3O4 matrix was clearly observed. From the biological standpoint, citric acid coated (Fe,Er)(3)O-4 NPs were found to induce low toxicity both in human cell fibroblasts and in zebrafish (Danio rerio) embryos. Biodistribution pattern of the MNPs after intravenous administration in healthy Wistar rats was followed by the radiotracer method, revealing that Y-90-labeled MNPs were predominantly found in liver (7533% ID), followed by lungs (16.70% ID) and spleen (2.83% ID). Quantitative agreement with these observations was obtained by ICP-MS elemental analysis using Er as the detected tracer. Based on the favorable physical, chemical and biological characteristics, citric add coated (Fe,Er)(3)O-4 MNPs could be further considered for the potential application as a diagnostic and/or therapeutic agent. This work also demonstrates that combined application of these techniques is a promising tool for studies of pharmacokinetics of the new MNPs in complex biological systems.Peer-reviewed manuscript: [https://imagine.imgge.bg.ac.rs/handle/123456789/1763]Supplementary data: [https://imagine.imgge.bg.ac.rs/handle/123456789/1764