Fully Automated Methods for [18F]FDG and [18F]-NaF Productions Using Explora FDG4: Validation and Reliability of Multi-subsequent Preparations for Clinical Applications.

BACKGROUND: The worldwide usage of [18F]-sodium fluoride in clinical applications has increased the interest in the facility of its production. The development of a new automated method for multi-preparations of [18F]-NaF and [18F]FDG on an Explora FDG4 module is described. Explora FDG4 is one of the most widely used synthesizers for FDG production in daily routine use and is specifically designed to run up to four different productions with a single module. Therefore, slight modifications are carried out in order to increase the potential of the synthesizer to perform more radiopharmaceuticals. METHODS: A fully automated method for multi-preparations of [18F]-NaF and [18F]FDG using Explora FDG4 was developed. Slight modifications to the Explora's hardware and software configuration were applied. A new elution vial for NaF preparation was installed and connected to the free position to MVP1. Quality control was carried out using the standard analytical methods applied for GMP production. RESULTS: This modification successfully provides preparation of [18F]-NaF without affecting the daily FDG production using one set preparation. [18F]-NaF was obtained in a high radiochemical yield (>90%, n=100) in 10 min total preparation time. The quality control results for both obtained products, FDG (RCP >95%) and NaF (RCP >98%), showed that the radiopharmaceuticals were in compliance with USP and Ph.Eur. specifications and compatible with clinical applications. CONCLUSION: A rapid and simple method for multi preparations of [18F]-NaF and [18F]FDG using a single Explora module was designed. Yet, the chemistry module has the potential to generate more radiopharmaceuticals to decrease the cost of preparation of [18F]-NaF compared to the cassette-based synthesizers, reducing radiation exposure resulting from manual preparations and increasing the reproducibility of [18F]-NaF preparation.

Fluorine walk: The impact of fluorine in quinolone amides on their activity against African sleeping sickness.

Human African Trypanosomiasis, also known as African sleeping sickness, is caused by the parasitic protozoa of the genus Trypanosoma. If there is no pharmacological intervention, the parasites can cross the blood-brain barrier (BBB), inevitably leading to death of the patients. Previous investigation identified the quinolone amide GHQ168 as a promising lead compound having a nanomolar activity against T. b. brucei. Here, the role of a fluorine substitution at different positions was investigated in regard to toxicity, pharmacokinetics, and antitrypanosomal activity. This 'fluorine walk' led to new compounds with improved metabolic stability and consistent activity against T. b. brucei. The ability of the new quinolone amides to cross the BBB was confirmed using an 18F-labelled quinolone amide derivative by means of ex vivo autoradiography of a murine brain.

Initial Clinical Investigation of [18F]Tetrafluoroborate PET/CT in Comparison to [124I]Iodine PET/CT for Imaging Thyroid Cancer.

AIM: Recently, [F]tetrafluoroborate ([F]TFB) has been introduced as a versatile PET probe for imaging the human sodium/iodide symporter activity. This pilot study aimed to compare [F]TFB-PET/CT with [I]NaI-PET/CT imaging in thyroid cancer patients. METHODS: Nine patients with newly diagnosed differentiated thyroid cancer underwent both [F]TFB- and [I]NaI-PET/CT after total thyroidectomy. PET/CT scans were visually analyzed for the presence of remnant thyroid tissue and for metastatic lesions on a patient and lesion basis. For semiquantitative analysis, thyroid remnant/tumor to blood pool ratios were calculated. RESULTS: All patients presented with positive [F]TFB and [I]NaI-PET/CT scans. Retention of I in remnant thyroid tissue was significantly higher as compared with [F]TFB (P < 0.01). In a lesion-based analysis, both tracers identified an almost equal number of foci with [F]TFB depicting a total of 41 foci and I a total of 40 foci, respectively. In 6 of 9 patients, both radiopharmaceuticals returned an identical number of foci. Two I-positive benign thyroid remnants were missed by [F]TFB-PET/CT in a single patient. In another case, both tracers identified different thyroid remnant tissues in the cervical region. Notably, [F]TFB demonstrated additional (I-negative) cervical lymph node metastases in 2 patients, leading to an overall agreement between the radiotracers of 91% (74/81 foci). DISCUSSION: In this pilot study, [F]TFB-PET was not inferior to [I]NaI-PET for detecting thyroid cancer and its metastases and was able to detect [I]NaI-PET-negative viable differentiated thyroid cancer metastases. Further clinical evaluation as a PET tracer for imaging thyroid pathophysiology and human sodium/iodide symporter expressing neoplasms is highly warranted.

Cholinergic activity and levodopa-induced dyskinesia: a multitracer molecular imaging study.

OBJECTIVE: To investigate the association between levodopa-induced dyskinesias and striatal cholinergic activity in patients with Parkinson's disease. METHODS: This study included 13 Parkinson's disease patients with peak-of-dose levodopa-induced dyskinesias, 12 nondyskinetic patients, and 12 healthy controls. Participants underwent 5-[123I]iodo-3-[2(S)-2-azetidinylmethoxy]pyridine single-photon emission computed tomography, a marker of nicotinic acetylcholine receptors, [123I]N-ω-fluoropropyl-2ß-carbomethoxy-3ß-(4-iodophenyl)nortropane single-photon emission computed tomography, to measure dopamine reuptake transporter density and 2-[18F]fluoro-2-deoxyglucose positron emission tomography to assess regional cerebral metabolic activity. Striatal binding potentials, uptake values at basal ganglia structures, and correlations with clinical variables were analyzed. RESULTS: Density of nicotinic acetylcholine receptors in the caudate nucleus of dyskinetic subjects was similar to that of healthy controls and significantly higher to that of nondyskinetic patients, in particular, contralaterally to the clinically most affected side. INTERPRETATION: Our findings support the hypothesis that the expression of dyskinesia may be related to cholinergic neuronal excitability in a dopaminergic-depleted striatum. Cholinergic signaling would play a role in maintaining striatal dopaminergic responsiveness, possibly defining disease phenotype and progression.

Validation of a [Al18F]PSMA-11 preparation for clinical applications.

Imaging prostate-specific membrane antigen (PSMA) using positron emission tomography (PET) has been presented so far as the most sensitive and specific with regard to prostate cancer detection, in particular in high-risk prostate cancer patients. Currently, it mainly features Gallium-68 (68Ga) labeled PSMA ligands, notably [68Ga]Glu-urea-Lys(Ahx)-HBED-CC ([68Ga]-PSMA-11) and [68Ga]DOTAGA-FFK (Sub-KuE termed ([68Ga]PSMA-I&T). However, 68Ga has several shortcomings as radionuclide including a short half-life and non-ideal energies. This has motivated consideration of 18F-labeled analogues for PET imaging of prostate cancer. Here, we describe a simple synthesis and validation of a fluorine-18 labeled Glu-urea-Lys(Ahx)-HBED-CC ([Al18F]PSMA-11) for nuclear medicine applications. An efficient method for preparation of [Al18F]PSMA-11 was developed and validated (according to Pharm Eur) for routinely clinical applications. [Al18F]PSMA-11 was reproducibly obtained in radiochemical yields of 84 ± 6% (n = 15) and > 98% radiochemical purity using an improved one-step radiofluorination in aqueous solution. The total (production/preparation) time, including purification, pharmacological formulation of the isolated product and the quality control of the injectable solution was less than 60min. The [Al18F]PSMA-11 was stable over 4h in 1% EtOH/saline selected as injection solution. The solution was sterile, non-pyrogenic and ready for clinical applications after sterile filtration through a 0.22µm membrane filter under sterile conditions. In addition, [Al18F]PSMA-11 exhibited higher uptake and retention in PMSA-expressing LNCap prostate cells as compared to its clinically established 68Ga-labeled analogues [68Ga]PSMA-11 and [68Ga]PSMA-I&T as well as to [68Ga]NOTA-Bn-PSMA. The simple and fast preparation of [Al18F]PSMA-11 combined with its favorable pharmacological properties warrant its translation to a clinical setting. CONCLUSION: The facile and high-yielding radiosynthesis of [Al18F]PSMA-11as well as its promising in vitro and in-vivo characteristics makes it worthy of clinical development for PET imaging of prostate cancer.

Combined [(18)F]DPA-714 micro-positron emission tomography and autoradiography imaging of microglia activation after closed head injury in mice.

BACKGROUND: Traumatic brain injury (TBI) is a major cause of death and disability. Neuroinflammation contributes to acute damage after TBI and modulates long-term evolution of degenerative and regenerative responses to injury. The aim of the present study was to evaluate the relationship of microglia activation to trauma severity, brain energy metabolism, and cellular reactions to injury in a mouse closed head injury model using combined in vivo PET imaging, ex vivo autoradiography, and immunohistochemistry. METHODS: A weight-drop closed head injury model was used to produce a mixed diffuse and focal TBI or a purely diffuse mild TBI (mTBI) in C57BL6 mice. Lesion severity was determined by evaluating histological damage and functional outcome using a standardized neuroscore (NSS), gliosis, and axonal injury by immunohistochemistry. Repeated intra-individual in vivo µPET imaging with the specific 18-kDa translocator protein (TSPO) radioligand [(18)F]DPA-714 was performed on day 1, 7, and 16 and [(18)F]FDG-µPET imaging for energy metabolism on days 2-5 after trauma using freshly synthesized radiotracers. Immediately after [(18)F]DPA-714-µPET imaging on days 7 and 16, cellular identity of the [(18)F]DPA-714 uptake was confirmed by exposing freshly cut cryosections to film autoradiography and successive immunostaining with antibodies against the microglia/macrophage marker IBA-1. RESULTS: Functional outcome correlated with focal brain lesions, gliosis, and axonal injury. [(18)F]DPA-714-µPET showed increased radiotracer uptake in focal brain lesions on days 7 and 16 after TBI and correlated with reduced cerebral [(18)F]FDG uptake on days 2-5, with functional outcome and number of IBA-1 positive cells on day 7. In autoradiography, [(18)F]DPA-714 uptake co-localized with areas of IBA1-positive staining and correlated strongly with both NSS and the number of IBA1-positive cells, gliosis, and axonal injury. After mTBI, numbers of IBA-1 positive cells with microglial morphology increased in both brain hemispheres; however, uptake of [(18)F]DPA-714 was not increased in autoradiography or in µPET imaging. CONCLUSIONS: [(18)F]DPA-714 uptake in µPET/autoradiography correlates with trauma severity, brain metabolic deficits, and microglia activation after closed head TBI.

A Novel Way To Radiolabel Human Butyrylcholinesterase for Positron Emission Tomography through Irreversible Transfer of the Radiolabeled Moiety.

The enzyme butyrylcholinesterase (BChE) is known to be involved in the detoxification of xenobiotics in blood plasma and is associated with the progress of neurodegenerative disorders, diabetes type 2, obesity, and diseases of the cardiovascular system. In the present study, we developed carbamate-based inhibitors serving as positron emission tomography (PET) radiotracers with (18) F and (11) C as radioisotopes to visualize BChE distribution. These inhibitors are radiolabeled at the carbamate site and transfer this moiety onto BChE, which thus results in covalent and permanent radiolabeling of the enzyme. There are no comparable radiotracers for cholinesterases described to date. By ex vivo autoradiography experiments on mice brain slices and kinetic investigations, selective and covalent transfer of the radiolabeled carbamate moiety onto BChE was proven. These tracers might provide high resolution of BChE distribution in vivo to enable investigations into the pathophysiological mechanisms of diseases associated with alterations in BChE occurrence.

Improved synthesis of [¹8F]FS-PTAD as a new tyrosine-specific prosthetic group for radiofluorination of biomolecules.

A novel prosthetic group, 4-(p-([(18)F]fluorosulfonyl)phenyl)-1,2,4-triazoline-3,5-dione ([(18)F]FS-PTAD) for site-specific radiofluorination of tyrosine residue in small molecules is described. Coupling of [(18)F]FS-PTAD with L-tyrosine, N-acetyl-L-tyrosine methyl amide and phenol as model compounds were achieved in buffered aqueous solution at room temperature, resulting in the corresponding fluorinated tyrosine and phenol derivatives. The total synthesis time including radiosynthesis, HPLC purification and formulation was less than 60 min (n=15) with ≥98% radio chemical purity. An initial in vitro evaluation of [(18)F]FS-PTAD-tyrosine in glioma cell lines revealed moderate uptake.

Synthesis and biological activity of some 3-(4-(substituted)-piperazin-1-yl)cinnolines.

A new series of 6-substituted-4-methyl-3-(4-arylpiperazin-1-yl)cinnolines 8-10 were synthesized as potential antifungal agents via intramolecular cyclization of the respective 1-(2-arylhydrazono)-1-(4-arylpiperazin-1-yl)propan-2-ones 5-7, mediated by polyphosphoric acid (PPA). The amidrazones themselves were synthesized via direct interaction of the appropriate hydrazonoyl chlorides 4a-d with the corresponding N-substituted piperazine in the presence of triethylamine. The structures of the new prepared compounds were confirmed by elemental analyses, (1)H-NMR, (13)C-NMR, and ESI-HRMS spectral data. The antitumor, antibacterial, and antifungal activity of the newly synthesized compounds was evaluated.

Auger electron emitter against multiple myeloma--targeted endo-radio-therapy with 125I-labeled thymidine analogue 5-iodo-4'-thio-2'-deoxyuridine.

INTRODUCTION: Multiple myeloma (MM) is a plasma cell malignancy characterized by accumulation of malignant, terminally differentiated B cells in the bone marrow. Despite advances in therapy, MM remains an incurable disease. Novel therapeutic approaches are, therefore, urgently needed. Auger electron-emitting radiopharmaceuticals are attractive for targeted nano-irradiation therapy, given that DNA of malignant cells is selectively addressed. Here we evaluated the antimyeloma potential of the Auger electron-emitting thymidine analogue (125)I-labeled 5-iodo-4'-thio-2'-deoxyuridine ([(125)I]ITdU). METHODS: Cellular uptake and DNA incorporation of [(125)I]ITdU were determined in fluorodeoxyuridine-pretreated KMS12BM, U266, dexamethasone-sensitive MM1.S and -resistant MM1.R cell lines. The effect of stimulation with interleukin 6 (IL6) or insulin-like growth factor 1 (IGF1) on the intracellular incorporation of [(125)I]ITdU was investigated in cytokine-sensitive MM1.S and MM1.R cell lines. Apoptotic cells were identified using Annexin V. Cleavage of caspase 3 and PARP was visualized by Western blot. DNA fragmentation was investigated using laddering assay. Therapeutic efficiency of [(125)I]ITdU was proven by clonogenic assay. RESULTS: [(125)I]ITdU was shown to be efficiently incorporated into DNA of malignant cells, providing a promising mechanism for delivering highly toxic Auger radiation emitters into tumor DNA. [(125)I]ITdU had a potent antimyeloma effect in cell lines representing distinct disease stages and, importantly, in cell lines sensitive or resistant to the conventional therapeutic agent, but was not toxic for normal plasma and bone marrow stromal cells. Furthermore, [(125)I]ITdU abrogated the protective actions of IL6 and IGF1 on MM cells. [(125)I]ITdU induced massive damage in the DNA of malignant plasma cells, which resulted in efficient inhibition of clonogenic growth. CONCLUSION: These studies may provide a novel treatment strategy for overcoming resistance to conventional therapy in multiple myeloma.

New molecular markers for prostate tumor imaging: a study on 2-methylene substituted fatty acids as new AMACR inhibitors.

The development of prostate carcinoma is associated with alterations in fatty acid metabolism. α-Methylacyl-CoA racemase (AMACR) is a peroxisomal and mitochondrial enzyme that catalyses interconversion between the (S)/(R)-isomers of a range of α-methylacyl-CoA thioesters. AMACR is involved in the ß-oxidation of the dietary branched-chain fatty acids and bile acid intermediates. It is highly expressed in prostate (more than 95 %), colon (92 %), and breast cancers (44 %) but not in the respective normal or hyperplastic tissues. Thus, targeting of AMACR could be a new strategy for molecular imaging and therapy of prostate and some other cancers. Unlabeled 2-methylenacyl-CoA thioesters (12 a-c) were designed as AMACR binding ligands. The thioesters were tested for their ability to inhibit the AMACR-mediated epimerization of (25R)-THC-CoA and were found to be strong AMACR inhibitors. Radioiodinated (E)-(131) I-13-iodo-2-methylentridec-12-enoic acid ((131) I-7 c) demonstrated preferential retention in AMACR-positive prostate tumor cells (LNCaP, LNCaP C4-2wt and DU145) compared with both AMACR-knockout LNCaP C4-2 AMACR-siRNA and benign BPH1 prostate cell lines. A significant protein-bound radioactive fraction with main bands at 47 (sum of molecular weights of AMACR plus 12 c), 70, and 75 kDa was detected in LNCaP C4-2 wt cells. In contrast, only negligible amounts of protein-bound radioactivity were found in LNCaP C4-2 AMACR-siRNA cells.