Synthesis of 18F-labelled radiopharmaceuticals for CNS imaging : From radiosynthesis development to GMP production

Positron emission tomography (PET) is an imaging technique that can be used to follow various biological processes, such as receptor function, in a living body. PET utilisesradiopharmaceuticals, tracers containing short-lived positron-emitting radioisotopes. The aim of this thesis was to develop labelling methods using fluorine-18 and 18F-labelled tracers in order to translate 18F-labelling methods to clinical radiopharmaceutical production. Traditionally, 18F-fluorination has been divided into electrophilic and nucleophilic fluorination with “[18F]F+” and [18F]fluoride ([18F]F-), respectively. This thesis used the recently developed approach of transition metal-mediated 18F-fluorination with copper and ruthenium. In transition metal-mediated fluorination chemistry, the reactivity of electrophilic fluorination and high molar activity of nucleophilic fluorination can be combined. The conditions for copper-mediated 18F-fluorination, including [18F]fluoride activation, were optimised and the method utilised for production of the norepinephrine transporter tracer [18F]NS12137 and the dopamine transporter tracer [18F]CFT. To evaluate the usefulness of the new methodology compared to traditional electrophilic 18F-fluorination, [18F]CFT was also produced via the electrophilic pathway. Ruthenium-mediated 18F-fluorination was applied successfully to the production of a new cannabinoid subtype 1 receptor tracer (CB1R), [18F]FPATPP. The tracer was evaluated in mice and [18F]FPATPP found to be specific for CB1R with high uptake in the CB1R-rich areas in the brain. These studies showed the suitability of transition metal-mediated 18F-fluorination in the development of PET tracers. Another CB1R tracer, [18F]FMPEP-d2, was translated to clinical production according to good manufacturing practices (GMP) utilising an in-house built synthesis device. The production was followed for 5 years.Positroniemissiotomografia (PET) on kuvantamistekniikka, jolla voidaan seurata biologisia prosesseja, kuten reseptorien toimintaa, elävässä kohteessa. PETkuvantamisessa käytetään lyhytikäisillä positronisäteilevillä radionuklideilla leimattuja radiolääkkeitä, merkkiaineita. Väitöskirjatutkimuksen tavoitteena oli kehittää 18F-leimausmenetelmiä ja 18F-leimattuja merkkiaineita sekä siirtää menetelmät kliiniseen radiolääketuotantoon. 18F-fluorileimausreaktiot jaetaan elektrofiiliseen fluoraukseen ”[18F]F+”:lla ja nukleofiiliseen fluoraukseen [18F]fluoridilla ([18F]F-). Tässä väitöskirjatutkimuksessa sovellettiin uutta siirtymämetallivälitteistä 18F-leimausta käyttäen hyväksi kuparia ja ruteniumia. Siirtymämetallivälitteisessä 18F-leimauskemiassa yhdistetään elektrofiilisen fluorileimauksen reaktiivisuus ja nukleofiilisella fluorileimauksella saavutettava korkea molaarinen aktiivisuus. Työssä optimoitiin kuparivälitteisen 18F-leimauksen reaktio-olosuhteet ja [18F]F-:n aktivointiolosuhteet. Menetelmää käytettiin noradrenaliinin kuljettajaproteiinimerkkiaineen [18F]NS12137:n ja dopamiinin kuljettajaproteiinimerkkiaineen [18F]CFT:n leimaussynteeseissä. Uuden kuparivälitteisen leimausmenetelmän käytettävyyttä arvioitiin vertaamalla menetelmää elektrofiiliseen 18F-leimaukseen valmistamalla [18F]CFT:tä myös elektrofiilisesti. Ruteniumvälitteistä leimausmenetelmää käytettiin onnistuneesti uuden kannabinoidireseptori 1 (CB1R) -merkkiaineen, [18F]FPATPP:n, valmistuksessa. [18F]FPATPP:n toimivuus CB1R-merkkiaineena testattiin terveillä hiirillä. Merkkiaine osoittautui spesifiseksi CB1R:lle, kertyen aivoalueille, joissa on runsaasti CB1R:ta. Osatyöt osoittavat siirtymämetallivälitteisen 18F-leimauskemian sopivan uusien PET-merkkiaineiden synteesikehitykseen. Toinen CB1R-merkkiaine, [18F]FMPEP-d2, siirrettiin kliiniseen tuotantoon Hyvät tuotantotavat (Good Manufacturing Practices, GMP) -ohjeistuksien mukaisesti. Merkkiaineen tuotannossa käytettiin itserakennettua synteesilaitetta, ja tuotantoa seurattiin 5 vuoden ajan

Radiosynthesis of the norepinephrine transporter tracer [F-18]NS12137 via copper-mediated F-18-labelling

[F-18]NS12137 (exo-3-[(6-[F-18]fluoro-2-pyridyl)oxy]8-azabicyclo[3.2.1]octane) is a highly selective norepinephrine transporter (NET) tracer. NETs are responsible for the reuptake of norepinephrine and dopamine and are linked to several neurodegenerative and neuropsychiatric disorders. The aim of this study was to develop a copper-mediated F-18-fluorination method for the production of [F-18]NS12137 with straightforward synthesis conditions and high radiochemical yield and molar activity. [F-18]NS12137 was produced in two steps. Radiofluorination of [F-18]NS12137 was performed via a copper-mediated pathway starting with a stannane precursor and using [F-18]F- as the source of the fluorine-18 isotope. Deprotection was performed via acid hydrolysis. The radiofluorination reaction was nearly quantitative as was the deprotection based on HPLC analysis. The radiochemical yield of the synthesis was 15.1 +/- 0.5%. Molar activity of [F-18]NS12137 was up to 300 GBq/mu mol. The synthesis procedure is straightforward and can easily be automated and adapted for clinical production

Fast and efficient copper-mediated 18F-fluorination of arylstannanes, aryl boronic acids, and aryl boronic esters without azeotropic drying

BackgroundCopper-mediated radiofluorination is a straightforward method to produce a variety of [18F]fluoroarenes and [18F]fluoroheteroarenes. To minimize the number of steps in the production of 18F-labelled radiopharmaceuticals, we have developed a short and efficient azeotropic drying-free 18F-labelling method using copper-mediated fluorination. Our goal was to improve the copper-mediated method to achieve wide substrate scope with good radiochemical yields with short synthesis time.ResultsSolid phase extraction with Cu (OTf)2 in dimethylacetamide is a suitable activation method for [18F]fluoride. Elution efficiency with Cu (OTf)2 is up to 79% and radiochemical yield (RCY) of a variety of model molecules in the crude reaction mixture has reached over 90%. Clinically relevant molecules, norepinephrine transporter tracer [18F]NS12137 and monoamine transporter tracer [18F]CFT were produced with 16.5% RCY in 98 min and 5.3% RCY in 64 min, respectively.ConclusionsCu (OTf)2 is a suitable elution agent for releasing [18F]fluoride from an anion exchange cartridge. The method is fast and efficient and the Cu-complex is customizable after the release of [18F]fluoride. Alterations in the [18F]fluoride elution techniques did not have a negative effect on the subsequent labelling reactions. We anticipate this improved [18F]fluoride elution technique to supplant the traditional azeotropic drying of [18F]fluoride in the long run and to concurrently enable the variations of the copper-complex.</div

Cannabinoid Receptor Type 1 in Parkinson's Disease : A Positron Emission Tomography Study with [F-18]FMPEP-d(2)

Background The endocannabinoid system is a widespread neuromodulatory system affecting several biological functions and processes. High densities of type 1 cannabinoid (CB1) receptors and endocannabinoids are found in basal ganglia, which makes them an interesting target group for drug development in basal ganglia disorders such as Parkinson's disease (PD). Objective The aim of this study was to investigate CB1 receptors in PD with [F-18]FMPEP-d(2) positron emission tomography (PET) and the effect of dopaminergic medication on the [F-18]FMPEP-d(2) binding. Methods The data consisted of 16 subjects with PD and 10 healthy control subjects (HCs). All participants underwent a [F-18]FMPEP-d(2) high-resolution research tomograph PET examination for the quantitative assessment of cerebral binding to CB1 receptors. To investigate the effect of dopaminergic medication on the [F-18]FMPEP-d(2) binding, 15 subjects with PD underwent [F-18]FMPEP-d(2) PET twice, both on and off antiparkinsonian medication. Results [F-18]FMPEP-d(2) distribution volume was significantly lower in the off scan compared with the on scan in basal ganglia, thalamus, hippocampus, and amygdala (P < 0.05). Distribution volume was lower in subjects with PD off than in HCs globally (P < 0.05), but not higher than in HCs in any brain region. Conclusions Subjects with PD have lower CB1 receptor availability compared with HCs. PD medication increases CB1 receptor toward normal levels. (c) 2022 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder SocietyPeer reviewe

Safety, biodistribution and radiation dosimetry of 18 F-rhPSMA-7.3 in healthy adult volunteers

This first-in-human study investigated the safety, biodistribution and radiation dosimetry of the novel 18F-labeled radiohybrid prostate-specific membrane antigen (rhPSMA) positron emission tomography (PET) imaging agent, 18F-rhPSMA-7.3. Methods: Six healthy volunteer subjects (3 males, 3 females) underwent multiple whole-body PET acquisitions at scheduled time points up to 248 minutes after the administration of 18F-rhPSMA-7.3 (mean activity 220; range, 210-228 MBq). PET scans were conducted in three separate sessions and subjects were encouraged to void between sessions. Blood and urine samples were collected for up to 4 hours post-injection to assess metabolite-corrected radioactivity in whole blood, plasma and urine. Quantitative measurements of 18F radioactivity in volumes of interest (VOIs) over target organs were determined directly from the PET images at 8 time points and normalized time-activity concentration curves were generated. These normalized cumulated activities were then inputted into the OLINDA/EXM package to calculate the internal radiation dosimetry and the subjects' effective dose. Results: 18F-rhPSMA-7.3 was well tolerated. One adverse event (mild headache, not requiring medication) was considered possibly related to 18F-rhPSMA-7.3: because of the temporal association with 18F-rhPSMA-7.3 injection, a causal relationship could not be excluded. The calculated effective dose was 0.0141 mSv/MBq when using a 3.5-hour voiding interval. The organs with the highest absorbed dose per unit of administered radioactivity were the adrenals (mean absorbed dose, 0.1835 mSv/MBq), the kidneys (mean absorbed dose, 0.1722 mSv/MBq), the submandibular glands (mean absorbed dose, 0.1479 mSv) and the parotid glands (mean absorbed dose, 0.1137 mSv/MBq). At the end of the first scanning session (mean time, 111 min post-injection), an average of 7.2% (range, 4.4-9.0%) of the injected radioactivity of 18F-rhPSMA-7.3 was excreted into urine. Conclusion: The safety, biodistribution and internal radiation dosimetry 18F-rhPSMA-7.3 are considered favorable for PET imaging

Automated GMP production and long-term experience in radiosynthesis of CB(1)tracer [F-18]FMPEP-d(2)

Here, we describe the development of an in-house-built device for the fully automated multistep synthesis of the cannabinoid CB(1)receptor imaging tracer (3R,5R)-5-(3-([F-18]fluoromethoxy-d(2))phenyl)-3-(((R)-1-phenylethyl)amino)-1-(4-(trifluoromethyl)phenyl)pyrrolidin-2-one ([F-18]FMPEP-d(2)), following good manufacturing practices. The device is interfaced to a HPLC and a sterile filtration unit in a clean room hot cell. The synthesis involves the nucleophilic(18)F-fluorination of an alkylating agent and its GC purification, the subsequent(18)F-fluoroalkylation of a precursor molecule, the semipreparative HPLC purification of the(18)F-fluoroalkylated product, and its formulation for injection. We have optimized the duration and temperature of the(18)F-fluoroalkylation reaction and addressed the radiochemical stability of the formulated product. During the past 5 years (2013-2018), we have performed a total of 149 syntheses for clinical use with a 90% success rate. The activity yield of the formulated product has been 1.0 +/- 0.4 GBq starting from 11 +/- 2 GBq and the molar activity 600 +/- 300 GBq/mu mol at the end of synthesis

Cannabinoid Receptor Type 1 in Parkinson's Disease: A Positron Emission Tomography Study with [F-18]FMPEP-d(2)

Background: The endocannabinoid system is a widespread neuromodulatory system affecting several biological functions and processes. High densities of type 1 cannabinoid (CB1) receptors and endocannabinoids are found in basal ganglia, which makes them an interesting target group for drug development in basal ganglia disorders such as Parkinson's disease (PD). Objective: The aim of this study was to investigate CB1 receptors in PD with [18F]FMPEP-d2 positron emission tomography (PET) and the effect of dopaminergic medication on the [18F]FMPEP-d2 binding. Methods: The data consisted of 16 subjects with PD and 10 healthy control subjects (HCs). All participants underwent a [18F]FMPEP-d2 high-resolution research tomograph PET examination for the quantitative assessment of cerebral binding to CB1 receptors. To investigate the effect of dopaminergic medication on the [18F]FMPEP-d2) binding, 15 subjects with PD underwent [18F]FMPEP-d2 PET twice, both on and off antiparkinsonian medication. Results: [18F]FMPEP-d2 distribution volume was significantly lower in the off scan compared with the on scan in basal ganglia, thalamus, hippocampus, and amygdala (P off than in HCs globally (P Conclusions: Subjects with PD have lower CB1 receptor availability compared with HCs. PD medication increases CB1 receptor toward normal levels. (c) 2022 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society</p

Cannabinoid Type 1 Receptors Are Upregulated During Acute Activation of Brown Adipose Tissue

Activating brown adipose tissue (BAT) could provide a potential approach for the treatment of obesity and metabolic disease in humans. Obesity is associated with upregulation of the endocannabinoid system, and blocking the cannabinoid type 1 receptor (CB1R) has been shown to cause weight loss and to decrease cardiometabolic risk factors. These effects may be mediated partly via increased BAT metabolism, since there is evidence that CB1R antagonism activates BAT in rodents. To investigate the significance of CB1R in BAT function, we quantified the density of CB1R in human and rodent BAT using the positron emission tomography radioligand [F-18]FMPEP-d(2) and measured BAT activation in parallel with the glucose analog [F-18]fluorodeoxyglucose. Activation by cold exposure markedly increased CB1R density and glucose uptake in the BAT of lean men. Similarly, 3-receptor agonism increased CB1R density in the BAT of rats. In contrast, overweight men with reduced BAT activity exhibited decreased CB1R in BAT, reflecting impaired endocannabinoid regulation. Image-guided biopsies confirmed CB1R mRNA expression in human BAT. Furthermore, CB1R blockade increased glucose uptake and lipolysis of brown adipocytes. Our results highlight that CB1Rs are significant for human BAT activity, and the CB1Rs provide a novel therapeutic target for BAT activation in humans

2,6-Bis(1,4,7,10-tetraazacyclododecan-1-ylmethyl)pyridine and Its Benzene Analog as Nonmetallic Cleaving Agents of RNA Phosphodiester Linkages

2,6-Bis(1,4,7,10-tetraazacyclododecan-1-ylmethyl)pyridine (11a) and 1,3-bis(1,4,7,10-tetraazacyclododecan-1-ylmethyl)benzene (11b) have been shown to accelerate at 50 mmol·L−1 concentration both the cleavage and mutual isomerization of uridylyl-3′,5′-uridine and uridylyl-2′,5′-uridine by up to two orders of magnitude. The catalytically active ionic forms are the tri- (in the case of 11b) tetra- and pentacations. The pyridine nitrogen is not critical for efficient catalysis, since the activity of 11b is even slightly higher than that of 11a. On the other hand, protonation of the pyridine nitrogen still makes 11a approximately four times more efficient as a catalyst, but only for the cleavage reaction. Interestingly, the respective reactions of adenylyl-3′,5′-adenosine were not accelerated, suggesting that the catalysis is base moiety selective