Radiopharmaceutical for differential diagnosis of tuberculoma: Synthesis of 2−[11C]cyano-isonicotinic acid hydrazide

The radiochemical synthesis of 2−[¹¹C]cyano-isonicotinic acid hydrazide was accomplished. Carbon-11 labelled cyano-group was introduced at the 2-position of the pyridine ring of 1-methoxy-4-methoxycarbonyl pyridinium methyl sulfate via a Reissert-Kaufmann type reaction. The reaction was performed on a solid support (silica gel) to yield no-carrier-added methyl 2−[¹¹C]cyano-isonicotinate in (32.4 ± 12%) (EOB) yield. This method is unique for the incorporation of [¹¹C]HCN to base sensitive substrates. The carbon-11 labelled methyl ester was treated with hydrazine hydrate to obtain 2−[¹¹C]cyano-isonicotinic acid hydrazide. The final radiochemical yield was 10% (EOB) and the synthesis time was approximately 35 min.     

Stabilization of [99mTc]HMPAO—1. Ethanolic preparation

The use of [^99mTc]HMPAO for cerebral blood flow imaging has been hampered by the short useful life time of the labelled radiopharmaceutical. Preparation of [^99mTc]HMPAO in 85% ethanol was found to increase the in vitro stability (92% radiochemical purity at 90 min) in comparison to the aqueous preparation (79% at 90 min). Biological distribution studies in mice indicated similar brain uptake (4.42 ± 1.02 and 4.12 ± 0.82% per gram at 20 min) and brain:blood ratio (0.97 ± 0.27 and 1.03 ± 0.20 at 20 min) of the ethanolic (2 h after preparation) and aqueous (15 min after preparation) formulations of [^99mTc]HMPAO respectively. The improved in vitro stability of [^99mTc]HMPAO prepared in 85% ethanol may prove useful in instances where previously the 30 min time limit precluded its use.     

Application of [11C]SA4503 to selection of novel σ1 selective agonists

IntroductionThe σ₁ ligands are considered to be a new class of potential therapeutic agents for several types of central nervous system disorder. Carbon-11-labeled 1-[2-(3,4-dimethoxyphenyl)ethyl]-4-(3-phenylpropyl)piperazine ([¹¹C]SA4503) was shown to be a promising PET ligand for mapping σ₁ receptors, and was applied to measure receptor occupancy with several therapeutic drugs in the living human brain. In this study, we applied this technique for preclinical in vivo screening of novel σ₁ selective agonists.MethodsSix newly synthesized piperazine derivatives containing arylalkylamine groups and cyclohexylamine derivatives containing phenyl groups were selected and tested for their in vivo σ₁ receptor binding with [¹¹C]SA4503. The test compounds were administered by intravenous co-injection or oral administration. The in vivo receptor binding of [¹¹C]SA4503 was evaluated by a tissue dissection method at a single time point.ResultsOur in vivo screen identified the most promising candidate of novel σ₁ agonist in the piperazine derivatives. Some correlations between in vitro affinity and in vivo receptor blocking rate were observed when considering oral bioavailability. In vivo receptor blocking of piperazine derivatives after oral administration may be predictable by simple co-injection study.ConclusionLigand selection with [¹¹C]SA4503 by the in vivo receptor binding assay was performed successfully. This technique is a practical and high-throughput method that can directly evaluate blood–brain barrier permeability, receptor binding, and bioavailability of drug candidates at the same time.     

Solid state support for the synthesis of [1−11C]-putrescine

A novel solid state support was demonstrated using [¹¹C]-HCN for the radiosynthesis of [1−¹¹C]-putrescine by the Michael addition reaction to acrylonitrile. The silica gel support allowed near quantitative trapping of the [¹¹C]-HCN and its efficient use under anhydrous conditions. The absence of water during the addition reaction eliminated by-product formation and reduced the overall synthesis and purification time required for the preparation of the radiopharmaceutical. A radiochemical yield of 53 ± 4 was achieved for purified product within 40 min of EOB. The process can be automated for the routine synthesis of [1-¹¹C]-putrescine radiopharmaceutical.     

Inhalation of [123I]alpha1-protease inhibitor: toward a new therapeutic concept of alpha1-protease inhibitor deficiency?

The alpha1-protease inhibitor (alpha1-Pi) is separated from human serum and is therefore extremely expensive. Because only 2%-3% concentrates in the lung after intravenous administration, inhalational therapy for alpha1-Pi deficiency would seem likely to be better. The aims of this study were therefore to determine the pattern of deposition of inhaled alpha1-Pi labeled with 123I and measure the amount deposited in the lungs. METHODS: Eighteen patients with congenital severe alpha1-Pi deficiency were enrolled in the study. The low-specific-activity 123I-labeled alpha1-Pi aerosol (median particle size +/- SD, 3.9 +/- 2.5 microm) was generated by an air pressure-driven nebulizer. The patients inhaled for an average of 23.6 +/- 8.9 min. Static scintigrams in two projections were acquired immediately after (T1) and 1 (T2), 4 (T3), and 24 h (T4) after inhalation. The patients were divided into the following three groups according to their forced expiratory volume in 1 s (FEV1): group I, < or =40% of predicted normal (n = 8); group II, 40% < FEV1 < or = 60% of predicted normal (n = 4); group III, >60% of predicted normal (n = 6). RESULTS: The absolute percentage uptake values of alpha1-Pi in group I were 12.4 for T1, 7.3 for T2, 4.6 for T3, and 1.2 for T4; in group II the values were 13.0, 9.6, 6.2, and 2.0, respectively; and in group III, 14.6, 11.4, 6.5, and 3.6, respectively. Differences between the groups were generally statistically significant. Between T1 and T2, the probability value was <0.05 for group I versus group II, <0.006 for group I versus group III, and <0.39 for group II versus group III. Between T1 and T3, the probability value was <0.29 for group I versus group II, <0.22 for group I versus group III, and <0.94 for group II versus group III. Retention (between T1 and T4) was also dependent on the grade of the disease: P < 0.2 for group I versus group II, P < 0.001 for group I versus group III, and P < 0.02 for group II versus group III. Grading of the uptake pattern by three independent experienced investigators (87% agreement) revealed a peripheral deposition that was group dependent. We found that greater peripheral deposition corresponded with lower lung functional impairment: P < 0.5 for group I versus group II, P < 0.01 for group I versus group III, and P < 0.08 for group II versus group III. Degradation also corresponded with functional impairment: P < 0.05 for group I versus group II, P < 0.006 for group I versus group III, and P < 0.3 for group II versus group III. CONCLUSION: The results of this study show that sufficient amounts of alpha1-Pi can be deposited in the periphery of the lung by inhalation at least in patients with low-grade disease. Inhalation of alpha1-Pi may thus represent a new and more convenient route of drug administration.     

PET Imaging of CRF1 with [11C]R121920 and [11C]DMP696: is the target of sufficient density?

AIM: Overstimulation of the CRF type 1 receptor (CRF1) is implicated in anxiety and depressive disorders. The aim of this study was to investigate the in vivo binding characteristics of [11C]R121920 and [11C]DMP696 in the nonhuman primate for application in positron emission tomography (PET) studies of CRF1. METHODS: PET imaging with the two novel CRF1 radioligands was performed in baboon. In vitro binding studies for CRF1 were performed in postmortem brain tissue of baboon and human to assess sufficiency of receptor density for PET. RESULTS: Both [11C]R121920 and [11C]DMP696 distributed rapidly and uniformly throughout the brain. Washout was comparable across brain regions, without differences in volume of distribution between regions reported to have high and low in vitro CRF1 binding. Membrane-enriched tissue homogenate assay using [(125)I]Tyr(0)-sauvagine and specific CRF1 antagonists CP154,526 and SN003 in human occipital cortex yielded maximal binding (Bmax) of 63.3 and 147.3 fmol/mg protein, respectively, and in human cerebellar cortex yielded Bmax of 103.6 and 64.6 fmol/mg protein, respectively. Dissociation constants (K(D)) were subnanomolar. In baboon, specific binding was not detectable in the same regions; therefore, Bmax and K(D) were not measurable. Autoradiographic results were consistent except there was also detectable CRF1-specific binding in baboon cerebellum. CONCLUSION: Neither [11C]R121920 nor [11C]DMP696 demonstrated quantifiable regional binding in vivo in baboon. In vitro results suggest CRF1 density in baboon may be insufficient for PET. Studies in man may generate more promising results due to the higher CRF1 density compared with baboon in cerebral cortex and cerebellum.     

Preliminary evaluation of 1′-[18F]fluoroethyl-β-D-lactose ([18F]FEL) for detection of pancreatic cancer in nude mouse orthotopic xenografts

IntroductionEarly detection of pancreatic cancer could save many thousands of lives. Non-invasive diagnostic imaging, including PET with [¹⁸F]FDG, has inadequate resolution for detection of small (2–3 mm) pancreatic tumours. We demonstrated the efficacy of PET imaging with an ¹⁸F-labelled lactose derivative, [¹⁸F]FEDL, that targets HIP/PAP, a biomarker that is overexpressed in the peritumoural pancreas. We developed another analogue, 1-[¹⁸F]fluoroethyl lactose ([¹⁸F]FEL), which is simpler to synthesise, for the same application. We conducted a preliminary evaluation of the new probe and its efficacy in detecting orthotopic pancreatic carcinoma xenografts in mice.MethodsXenografts were developed in nude mice by injecting L3.6pl/GL⁺ pancreatic carcinoma cells into the pancreas of each mouse. Tumour growth was monitored by bioluminescence imaging (BLI); accuracy of BLI tumour size estimates was verified by MRI in two representative mice. When the tumour size reached approximately 2–3 mm, the animals were injected with [¹⁸F]FEL (3.7 MBq) and underwent static PET/CT scans. Blood samples were collected at 2, 5, 10, 20 and 60 min after [¹⁸F]FEL injection to track blood clearance. Following imaging, animals were sacrificed and their organs and tumours/pancreatic tissue were collected and counted on a gamma counter. Pancreas, including tumour, was frozen, sliced and used for autoradiography and immunohistochemical analysis of HIP/PAP expression.ResultsTumour growth was rapid, as observed by BLI and MRI. Blood clearance of [¹⁸F]FEL was bi-exponential, with half-lives of approximately 3.5 min and 40 min. Mean accumulation of [¹⁸F]FEL in the peritumoural pancreatic tissue was 1.29 ± 0.295 %ID/g, and that in the normal pancreas of control animals was 0.090 ± 0.101 %ID/g. [¹⁸F]FEL was cleared predominantly by the kidneys. Comparative analysis of autoradiographic images and immunostaining results demonstrated a correlation between [¹⁸F]FEL binding and HIP/PAP expression.Conclusion[¹⁸F]FEL may be useful for non-invasive imaging of early-stage pancreatic tumours by PET. The results warrant further studies.     

The remotely-controlled preparation of a 11C-labelled radiopharmaceutical—[1−11C]acetate

Remotely-controlled apparatus is described for the preparation of the radiopharmaceutical, [1−¹¹C]acetate, from cyclotron-produced [¹¹C]carbon dioxide according to established radiochemistry. This apparatus features a multi-ported reaction vessel (fitted with an electrical stirrer), twelve solenoid valves (to direct fluid flows and hydraulically-powered syringes), one hydraulic oil pump and one heated water-bath (all operated at 24 V d.c.). These components are controlled either with a rotary-switch or with an “Apple II” microcomputer acting through a digital output card. An important advantage of the use of this apparatus over the use of manually-controlled apparatus is that it results in a much reduced radiation dose to the operator. Moreover it has been shown that [1−¹¹C]acetate can be prepared much more efficiently with the remotely-controlled apparatus than with corresponding manually-controlled apparatus. Thus the overall efficiencies (radiochemical yields uncorrected for decay) for the conversion of [¹¹C]carbon dioxide into [1−¹¹C]acetate for injection are 24 ± 8, 39 ± 7 and 47 ±8% for manual, remote rotary-switch and remote microcomputer control, respectively. The high efficiency and consistent performance of the remotely-controlled apparatus have been found to permit useful flexibility in the design of clinical experiments with [1−¹¹C]acetate and positron emission tomography.     

Development of methodology for synthesis of [α-C]-labeled phenethylamine

A one solvent two-step procedure for the synthesis of [α-C]-labeled phenethylamine is described. The method employs crown ethers to aid in the solubilization of sodium cyanide in tetrahydrofuran (THF) for the formation of phenylacetonitrile from benzyl chloride. Reduction of phenylacetonitrile to phenethylamine was accomplished with borane using the same solvent. The overall yield of phenethylamine was 44.5% based on cyanide. The procedure is amenable to hot cell conditions for eventual [α-¹¹C]- labeling of phenethylamine. Dimethylsulfoxide (DMSO) was found to be a less desirable solvent for this reaction due to its participation in the Kornblum oxidation.     

Improved Synthesis of 2'-deoxy-2'-[18F]fluoro-5-Methyl-1-β-DArabinofuranosyluracil ([18F]FMAU)

[18F]FMAU is an established PET probe used to monitor cellular proliferation. For clinical applications, a fully automated cGMP-compliant radiosynthesis would be preferred. However, the current synthesis of [18F]FMAU requires HBr activation of the sugar prior to the coupling with silylated uracil. This multiple step procedure makes the development of an automated protocol difficult and complicated. In this study, we report the use of Friedel-Crafts catalysts for an improved synthesis of [18F]FMAU, which also includes a significantly simplified one-pot reaction condition.     

Optimization of precursor synthesis,formulation and stability of 1′-[18 F]fluoroethyl-β-D-lactose ([18 F]FEL) for preclinical studies in detection of pancreatic cancer

Introduction1′-[¹⁸ F]Fluoroethyl-β-d-lactose ([¹⁸ F]FEL) is a new PET imaging agent for early detection of pancreatic cancer and hepatocellular carcinoma. We previously reported the syntheses of [¹⁸ F]FEL using a bromo- and a tosyl- precursor, followed by an improved method using a nosyl-precursor. However, some steps in the synthesis of the precursor appeared to be problematic producing low yields. Here, we report on an optimized method for synthesis of the precursor and production of [¹⁸ F]FEL; we also describe [¹⁸ F]FEL’s formulation and stability.MethodsAcetylation of d-lactose 1 was performed following a literature procedure to obtain 1′,2′,3′,6′,2,3,4,6-d-lactose octa-acetate 2a/2b. Bromination of 2a/2b was performed using HBr/acetic acid to produce 1'-bromo-2′,3′,6′,2,3,4,6-hepta-O-acetyl-α-d-lactose 3. Coupling of 3 with ethylene glycol was performed in the presence of Ag-tosylate and an excess of ethylene glycol to produce 4a. Compound 4a was reacted with p-nitrophenylsulfonyl chloride to produce the nosyl derivative 5. Radiofluorination of 5 was performed using K[¹⁸ F]fluoride/kryptofix to obtain 6, which was purified by HPLC and hydrolyzed with Na-methoxide to produce 7.ResultsCompound 2 (2a/2b) was obtained in 83% yield as a mixture of two anomeric products. Compound 3 was obtained from the 2a/2b mixture in 80% yield as one product. Coupling of 3 with ethylene glycol produced 4a in 90% yield. Compound 5 was obtained in 64% yield, and radiofluorination of 5 produced 6 in 62.5% ± 7.5% yields (n = 8). Hydrolysis of 6 with Na-methoxide produced 7 in 42.0% ± 7.0% yield (n = 8) from the end of bombardment.ConclusionsA simple 4-step synthesis of the precursor, compound 5, has been achieved with improved yields. A new formulation of [¹⁸ F]FEL has been developed that allows the product to remain stable at ambient temperature for use in animal studies. This improved synthesis of the precursor and stable formulation of [¹⁸ F]FEL should be useful for routine production of the radiotracer and its preclinical and, possibly, clinical applications.     

Improved synthesis of 2′-deoxy-2′-[18F]-fluoro-1-β-d-arabinofuranosyl-5-iodouracil ([18F]-FIAU)

An improved synthesis of 2′-[¹⁸F]-fluoro-2′-deoxy-1-β-d-arabinofuranosyl-5-iodouracil ([¹⁸F]-FIAU) has been developed. The method utilizes trimethylsilyl trifluoromethanesulfonate (TMSOTf) catalyzed coupling of 2-deoxy-2-[¹⁸F]-fluoro-1,3,5-tri-O-benzoyl-d-arabinofuranose with 2,4-bis(trimethylsilyloxy)-5-iodouracil to yield the protected dibenzoyl-[¹⁸F]-FIAU. Dibenzoyl-[¹⁸F]-FIAU was deprotected with sodium methoxide to yield a mixture of α- and β-anomers in a ratio of 1:1, which were purified by HPLC. The procedure described in this article eliminates the need for HBr activation of the sugar prior to coupling with silylated iodouracil and is suitable for automation. The total reaction time was about 110 min, starting from [¹⁸F]-fluoride. The average isolated yield of the required β-anomer was 10±6% (decay corrected) with average specific activity of 125 mCi/μmol.     

Synthesis of carbon-11-labeled piperidine ring of N-[ω-(6-methoxynaphthalen-1-yl)alkyl] derivatives as new selective PET σ1 receptor probes

Carbon-11-labeled piperidine ring of N-[ω-(6-methoxynaphthalen-1-yl)alkyl] derivatives were first designed and synthesized as new selective PET σ₁ receptor probes. The target tracers were prepared by O-[¹¹C]methylation of their corresponding phenolic hydroxyl precursors using [¹¹C]CH₃OTf under basic conditions and isolated by a simplified SPE method in 40–50% radiochemical yields based on [¹¹C]CO₂ and decay corrected to EOB. The overall synthesis time from EOB was 15–20 min, the radiochemical purity was >99%, and the specific activity at EOS was 111–185 GBq/μmol.     

Re-evaluation of in vivo selectivity of [(11)C]SA4503 to σ(1) receptors in the brain: Contributions of emopamil binding protein

IntroductionCarbon-11-labeled 1-[2-(3,4-dimethoxyphenyl)ethyl]-4-(3-phenylpropyl)piperazine ([¹¹C]SA4503) was shown to be a promising PET ligand for mapping σ₁ receptors, and was applied to human subjects. However, an in vitro study indicated that SA4503 also binds to the emopamil binding protein (EBP), vertebral Δ8-Δ7 sterol isomerase. To our knowledge, no information is available about the possibility of [¹¹C]SA4503 binding to EBP in the brain in vivo.MethodsTo confirm the selectivity of [¹¹C]SA4503, we carried out an in vivo blocking experiment using high-affinity EBP and σ₁ selective blocker.ResultsThe brain uptake of [¹¹C]SA4503 was dose-dependently decreased by SA4503 and the high-affinity σ₁ blockers haloperidol, ifenprodil, and trifluperidol. On the other hand, the effects of the high-affinity EBP blockers tamoxifen and trifluoperazine were negligible.ConclusionsOur results confirmed the σ₁-selective binding of [¹¹C]SA4503 in the brain.     

Microwave-assisted one-pot radiosynthesis of 2'-deoxy-2'-[(18)F]fluoro-5-methyl-1-β-d-arabinofuranosyluracil ([(18)F]-FMAU)

Objectives[¹⁸F]-FMAU is a PET tracer being evaluated for imaging cell proliferation. Current multi-step procedures of [¹⁸F]-FMAU synthesis are time-consuming, resulting in low radiochemical yield and inconvenient applications for the clinic. We have previously reported the use of Friedel-Crafts catalysts for an improved synthesis of [¹⁸F]-FMAU. In this study, we investigated the efficiency of microwave-assisted radiosynthesis of [¹⁸F]-FMAU in comparison with conventional thermal conditions.MethodsA simplified one-pot synthesis of [¹⁸F]-FMAU was developed under microwave conditions. Various reaction times, temperatures, and microwave powers were systematically explored to optimize the coupling reaction of 2-deoxy-2-[¹⁸F]fluoro-1,3,5-tri-O-benzoyl-d-arabinofuranose ([¹⁸F]-sugar) and bis-2,4-(trimethylsilyloxy)-5-methyluracil (silylated uracil) in the presence of a Friedel-Crafts catalyst, trimethylsilyl trifluoromethanesulfonate (TMSOTf).ResultsMicrowave significantly enhanced the coupling efficiency of [¹⁸F]-sugar and silylated uracil by reducing the reaction time to 10 min (6-fold reduction as compared to conventional heating) at 95 °C. Base hydrolysis followed by high-performance liquid chromatography purification produced the desired [¹⁸F]-FMAU. The overall radiochemical yield was 20 ± 4% (decay corrected, n = 3). Radiochemical purity was > 99% and specific activity was > 400 mCi/μmol. The α/β anomer ratio was 1:2. The radiosynthesis time was about 90 min from the end of bombardment.ConclusionsA reliable microwave-assisted approach has been developed for routine synthesis of [¹⁸F]-FMAU. The new approach affords a simplified process with shorter synthesis time and higher radiochemical yield as compared to conventional heating. A fully automated microwave-assisted synthesis of [¹⁸F]-FMAU can be readily achieved under new reaction conditions.     

Is ultrasonographic evaluation the the length of the kidney possible in children? Apropos of a prospective study in Madagascar children less than 1 year of age]

Renal US was prospectively performed in 124 madagascan children less than 1 year of age. Patients were examined in the prone position and maximum kidney length was measured in the longitudinal plane. These measurements and the height and weight of our patient population were compared to published tables. Kidney length and height and weight of our patient population were inferior to the previously published reference data and the growth curve of kidneys steeper than normative standards (p < 0.001). Because of the important variability in US measurement of kidney length it is not possible to definitely conclude that length and growth curve of kidneys in madagascan children are statistically different from those of the published normative standards.     

Are [O-methyl-11C]derivatives of ICI 89,406 β1-adrenoceptor selective radioligands suitable for PET?

Purpose Radioligand binding studies show that β₁-adrenoceptor (β₁-AR) density may be reduced in heart disease without down regulation of β₂-ARs. Radioligands are available for measuring total β-AR density non-invasively with clinical positron emission tomography (PET) but none are selective for β₁- or β₂-ARs. The aim was to evaluate ICI 89,406, a β₁-AR-selective antagonist amenable to labelling with positron emitters, for PET. Methods The S-enantiomer of an [O-methyl-¹¹C] derivative of ICI 89,406 ((S)-[¹¹C]ICI-OMe) was synthesised. Tissue radioactivity after i.v. injection of (S)-[¹¹C]ICI-OMe (< 2 nmol·kg⁻¹) into adult Wistar rats was assessed by small animal PET and post mortem dissection. Metabolism was assessed by HPLC of extracts prepared from plasma and tissues and by measuring [¹¹C]CO₂ in exhaled air. Results The heart was visualised by PET after injection of (S)-[¹¹C]ICI-OMe but neither unlabelled (S)-ICI-OMe nor propranolol (non-selective β-AR antagonist) injected 15 min after (S)-[¹¹C]ICI-OMe affected myocardial radioactivity. Ex vivo dissection showed that injecting unlabelled (S)-ICI-OMe, propranolol or CGP 20712A (β₁-selective AR antagonist) at high dose (> 2 μmol·kg⁻¹) before (S)-[¹¹C]ICI-OMe had a small effect on myocardial radioactivity. HPLC demonstrated that radioactivity in myocardium was due to unmetabolised (S)-[¹¹C]ICI-OMe although ¹¹C-labelled metabolites rapidly appeared in plasma and liver and [¹¹C]CO₂ was detected in exhaled air. Conclusion Myocardial uptake of (S)-[¹¹C]ICI-OMe after i.v. injection was low, possibly due to rapid metabolism in other tissues. Injection of unlabelled ligand or β-AR antagonists had little effect indicating that binding was mainly to non-specific myocardial sites, thus precluding the use of (S)-[¹¹C]ICI-OMe to assess β₁-ARs with PET.     

On polysemy of term "regeneration". Dedicated to memory of Grigori? Ivanovich Voronin]

Comparison of the meaning contents of regeneration processes in different branches of engineering and industry, and space biology and medicine brings to the conclusion that polysemy of term "regeneration" causes certain terminology difficulties in communication of members of various professional communities. To avoid ambiguity in each specific case, term "regeneration" is proposed to be accompanied by a minimum of appropriate explicative words. In the context of growing concern about the global ecological trouble, the biological regeneration processes developed for support of human life in space vehicles have much promise for designing bio-regeneration technologies to be used in preservation of environment, and industrial biosynthesis.