SYNTHESIS AND QUALITY ASSESSMENT OF THE POSITRON EMITTER RADIOTRACER 2-[18F]-FLUOROETHYLCHOLINE, A [18F]-LABELLED CHOLINE ANALOGUE FOR TUMOURS IMAGING

Introduction: [18F]-labelled choline analogues, such as 2-[18F]-Fluoroethylcholine ( [18F]-FECH ), have suggested to be the new generation of choline derivatives for the imaging of prostate and brain tumours. In fact, tumour cells with enhanced proliferation rate exhibit an improved choline uptake due to the increased membrane phospholipids biosynthesis. The aim of this dissertation is the development and validation of a reliable automatic synthesis of [18F]-FECH. Moreover, in order to assure a safe employment of the radiotracer in a research clinical trial, particular attention was paid to establish the proper quality controls required. Materials and Methods: [18F]-FECH was synthesized with varying synthetic approaches (one step and two steps reaction, preparative HPLC or solid phase extraction purification) and by using two different automatic synthesizers (FXF-N Tracer Lab Synthesizer and Mx Tracer Lab). In the first step, cyclotron produced [18F]-Fluoride was reacted for 5 minutes at 80°C, with a 20 mg/ml solution of 1,2- bis(tosyloxy)ethane (diOTsEt) in acetonitrile in order to obtain the [18F]-EtOTs intermediate. The reaction was performed in anhydrous condition and in presence of tetrabutylammonium (TBA) as transfer phase catalyst. In the second step, without any further purification, [18F]-EtOTs was reacted for 10 minutes at 100° C in the same reactor, with a 88.6 mg/ml solution of N,N-dimethylethanolamine (DMAE) in acetonitrile. The reaction mixture was then purified by SPE or HPLC and then eluted with 16 ml of physiological solution. The effect of different precursors (Br2Et, diNosEt and BrOtsEt) on the reactions was also assessed but with lower success. Quality controls on the final product were performed by means of TLC, GC and HPLC equipped with conductimetric, UV and radiometric detectors. Sterility and absence of bacterial endotoxin were assessed by following the European Pharmacopeia guidelines. Results: [18F]-FECH was prepared with a yield between 17±2 and 41 ±2 % uncorrected for decay in a time ranging between 33 and 55 minutes, depending on the synthetic approach and on the synthesizer used. Radiochemical purity was always greater than 99 % with both free [18F]-fluoride and [18F]-EtOTs less than 0.5 % of the total radioactivity. The DMAE concentration always resulted less than 15 mg/L and was detected with either GC or HPLC with conductimetric detection. The main chemical impurity was due to dimethylmorpholinium (dMM), a cationic by-product derived from the direct reaction and cyclization of diOTsEt and DMAE. The dMM concentration was detected by HPLC with conductimetric detection and always resulted less than 180 mg/L. The absence of tosylated by-products or free OTs- anions was assessed by HPLC with UV detection. Residual solvents and TBA concentrations respected the limits fixed for an injectable radiopharmaceutical preparation and were detected by means of GC in the same analysis conditions used for DMAE. All the sample analyzed were sterile. Conclusions: [18F]-FECH radiopharmaceutical was synthesized with an high yield and radiochemical purity by using a automatic synthesizers. The complete elimination of the dMM by-product from the final solution was not possible by means of a solid phase extraction as a purification method. A research clinical trial was activated and, up to now, more than 150 patients were safely examined

Methods for preparation and administration of lutetium-177 oxodotreotide 3.7 GBq: proceedings from an Italian advisory board

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Curcumin-Based β-Diketo Ligands for Ga3+: Thermodynamic Investigation of Potential Metal-Based Drugs

Curcumin is known for its therapeutic properties; among these, antioxidant, anti-inflammatory and anti-cancer ones stand out. Besides, curcumin metal complexes have shown widespread application in medicine and can be exploited as lead structures for developing metal-based drugs. Unfortunately, curcumin is poorly bioavailable, mainly due to its instability in physiological conditions; this weakness is tightly connected to the presence of the β-diketo moiety undergoing tautomeric equilibrium. Stability and metal-chelating ability can be tuned by modulating the electronic effects and steric hindrance close to the β-diketo moiety; in addition, formation of a metal complex shifts the tautomeric equilibrium towards the β-keto–enol form and increases stability in biological media. Among the metals used in clinical therapy, gallium nitrate has shown to have significant antitumor activity against non-Hodgkin lymphoma and bladder cancer, thus indicating that gallium-based drugs have potential for further development as antineoplastic agents with improved therapeutic activity. Curcuminoids have demonstrated high affinity for gallium(III), allowing the formation of stable positively charged M:L 1:2 β-diketonate complexes that benefit from the therapeutic activity of both the metal and the ligand. Seven new curcumin derivatives were synthesized and completely characterized. The new derivatives retain the solvent-dependent keto–enol tautomerism, with the prevalence of the diketo form in aqueous solution. Enhanced stability in simulated physiological conditions was observed in comparison to the lead compound curcumin. The presence of Ga3+ anticipates the dissociation of the enolic proton, allowing chelate complex formation, and simultaneously it shifts the tautomeric equilibrium towards the keto–enol form. A complete 1H/13C NMR and UV–Vis study was performed to define the metal-to-ligand stoichiometry ratio and the overall stability constants. In addition, we demonstrated that some of the derivatives have increased antiproliferative activity on colon cancer cells compared to curcumin and antioxidant properties. On the whole, the synthesized curcumin-based molecules may act as new gallium(III) chelators with improved stability with respect to curcumin and could open interesting perspectives for the development of novel therapeutic agents for cancer

Development of a Potential Gallium-68-Labelled Radiotracer Based on DOTA-Curcumin for Colon-Rectal Carcinoma: From Synthesis to In Vivo Studies

Colorectal cancer is the third most commonly occurring cancer in men and the second most commonly occurring cancer in women worldwide. We have recently reported that curcuminoid complexes labelled with gallium-68 have demonstrated preferential uptake in HT29 colorectal cancer and K562 lymphoma cell lines compared to normal human lymphocytes. In the present study, we report a new gallium-68-labelled curcumin derivative (68Ga-DOTA-C21) and its initial validation as marker for early detection of colorectal cancer. The precursor and non-radioactive complexes were synthesized and deeply characterized by analytical methods then the curcuminoid was radiolabelled with gallium-68. The in vitro stability, cell uptake, internalization and efflux properties of the probe were studied in HT29 cells, and the in vivo targeting ability and biodistribution were investigated in mice bearing HT29 subcutaneous tumour model. 68Ga-DOTA-C21 exhibits decent stability (57 \ub1 3% after 120 min of incubation) in physiological media and a curcumin-mediated cellular accumulation in colorectal cancer cell line (121 \ub1 4 KBq of radiotracer per mg of protein within 60 min of incubation). In HT29 tumour-bearing mice, the tumour uptake of 68Ga-DOTA-C21 is 3.57 \ub1 0.3% of the injected dose per gram of tissue after 90 min post injection with a tumour to muscle ratio of 2.2 \ub1 0.2. High amount of activity (12.73 \ub1 1.9% ID/g) is recorded in blood and significant uptake of the radiotracer occurs in the intestine (13.56 \ub1 3.3% ID/g), lungs (8.42 \ub1 0.8% ID/g), liver (5.81 \ub1 0.5% ID/g) and heart (4.70 \ub1 0.4% ID/g). Further studies are needed to understand the mechanism of accumulation and clearance; however, 68Ga-DOTA-C21 provides a productive base-structure to develop further radiotracers for imaging of colorectal cancer

Labelling of

In spite of the hazard due to the radiation exposure, preparation of 90Y- and 177Lu-labelled radiopharmaceuticals is still mainly performed using manual procedures. In the present study the performance of a commercial automatic synthesizer based on disposable cassettes for the labelling of 177Lu- and 90Y-DOTA-conjugated biomolecules (namely, DOTATOC and PSMA-617) was evaluated and compared to a manual and a semiautomated approach. The dose exposure of the operators was evaluated as well. More than 300 clinical preparations of both 90Y- and 177Lu-labelled radiopharmaceuticals have been performed using the three different methods. The mean radiochemical yields for 90Y-DOTATOC were 96.2±4.9%, 90.3±5.6%, and 82.0±8.4%, while for 177Lu-DOTATOC they were 98.3%  ± 0.6, 90.8%  ± 8.3, and 83.1±5.7% when manual, semiautomated, and automated approaches were used, respectively. The mean doses on the whole hands for yttrium-90 preparations were 0.15±0.4 mSv/GBq, 0.04±0.1 mSv/GBq, and 0.11±0.3 mSv/GBq for manual, semiautomated, and automated synthesis, respectively, and for lutetium-177 preparations, they were 0.02±0.008 mSv/GBq, 0.01±0.03 mSv/GBq, and 0.01±0.02 mSv/GBq, respectively. In conclusion, the automated approach guaranteed reliable and reproducible preparations of pharmaceutical grade therapeutic radiopharmaceuticals in a decent RCY. The radiation exposure of the operators remained comparable to the manual approach mainly due to the fact that a dedicated shielding was still not available for the system

Radiolabeled Chalcone Derivatives as Potential Radiotracers for β-Amyloid Plaques Imaging

Natural products often provide a pool of pharmacologically relevant precursors for the development of various drug-related molecules. In this review, the research performed on some radiolabeled chalcone derivatives characterized by the presence of the α-β unsaturated carbonyl functional group as potential radiotracers for the imaging of β-amyloids plaques will be summarized. Chalcones’ structural modifications and chemical approaches which allow their radiolabeling with the most common SPECT (Single Photon Emission Computed Tomography) and PET (Positron Emission Tomography) radionuclides will be described, as well as the state of the art regarding their in vitro binding affinity and in vivo biodistribution and pharmacokinetics in preclinical studies. Moreover, an explanation of the rationale behind their potential utilization as probes for Alzheimer’s disease in nuclear medicine applications will be provided

Metal binding ability of curcumin derivatives: a theoretical vs. experimental approach

Theoretical calculations employing DFT at the B3LYP/6-311G++** level are used to investigate the tautomeric equilibrium in curcumin derivatives. The solvent effect is evaluated using the CPCM continuum solvation method. The results are compared with experimental data obtained from the X-ray crystal structure of K2A23 and UV-vis data. The KE tautomer is more stable in a vacuum and in the solid state, while in water the DK tautomer reaches a population of 90%. In agreement with spectroscopic data, theoretical calculations predict a slight prevalence of the DK form in non-aqueous solvent systems. The ability to chelate metal ions [Fe3+, Ga3+ and Cu2+] is then explored by means of 1H, 13C NMR and UV-Vis spectroscopy. From the calculation of the overall stability constants of metal complexes and 1H NMR titrations with Ga3+, it is clear that the more stable species has a 1 : 2 M/L molar ratio. The curcuminoid coordinates the metal ion through the keto–enol function in the dissociated form; in addition 2D 1H 13C NMR experiments suggest the involvement of carboxylic oxygen in metal coordination it was found in the solid state for the complex [Ga(K2A33)2]PF6. The rate of the complexation reaction is strongly influenced by the type of substituent on the aromatic ring of the curcuminoid (K2A33 ≈ K2A23 ≫ K2A21). In addition DPPH assay evidences how antioxidant ability of curcumin derivatives is mainly due to the presence of a phenolic group and metal coordination by a keto–enolic moiety does not affect it, especially for K2A21

Efficient automated one-step synthesis of 2-[18F]fluoroethylcholine for clinical imaging: optimized reaction conditions and improved quality controls of different synthetic approaches

[(18)F]-labelled choline analogues, such as 2-[(18)F]fluoroethylcholine ((18)FECH), have suggested to be a new class of choline derivatives highly useful for the imaging of prostate and brain tumours. In fact, tumour cells with enhanced proliferation rate usually exhibit an improved choline uptake due to the increased membrane phospholipids biosynthesis. The aim of this study was the development of a high yielding synthesis of (18)FECH. The possibility of shortening the synthesis time by reacting all the reagents in a convenient and rapid one-step reaction was specially considered

Semiautomated labelling and fractionation of yttrium-90 and lutetium-177 somatostatin analogues using disposable syringes and vials

The treatment of tumours expressing somatostatin receptors with yttrium-90 (90Y)-labelled and lutetium-177 (177Lu)-labelled somatostatin analogues is one of the most interesting therapeutic approaches adopted in nuclear medicine in recent years. However, the process of synthesis and fractionation of these radiopharmaceuticals is still mainly carried out manually despite the high radiation exposure to the operators and the need to comply with good manufacturing practices. In this study a semiautomatic synthesizer [automatic dose dispenser (ADD-2)] using only disposable syringes and vials has been presented