Search

Cannabinoid type 2 receptors (CB2R) have emerged as promising targets for the diagnosis and therapy of brain pathologies. However, no suitable radiotracers for accurate CB2R mapping have been found to date, limiting the investigation of the CB2 receptor expression using positron emission tomography (PET) imaging. In this work, we report the evaluation of the invivo expression of CB2R with [11C]A-836339 PET after cerebral ischemia and in two rat models of neuroinflammation, first by intrastriatal LPS and then by AMPA injection. PET images and invitro autoradiography showed a lack of specific [11C]A-836339 uptake in these animal models demonstrating the limitation of this radiotracer to image CB2 receptor under neuroinflammatory conditions. Further, using immunohistochemistry, the CB2 receptor displayed a modest expression increase after cerebral ischemia, LPS and AMPA models. Finally, [18F]DPA-714-PET and immunohistochemistry demonstrated decreased neuroinflammation by a selective CB2R agonist, JWH133. Taken together, these findings suggest that [11C]A-836339 is not a suitable radiotracer to monitor invivo CB2R expression by using PET imaging. Future studies will have to investigate alternative radiotracers that could provide an accurate binding to CB2 receptors following brain inflammation.

The translocator protein 18kDa (TSPO) is today a validated target for a number of therapeutic applications, but also a well-recognized diagnostic/imaging biomarker for the evaluation of inflammatory related-disease state and progression, prompting the development of specific and dedicated TSPO ligands worldwide. For this purpose, pyrazolo[1,5-a]pyrimidine acetamides constitute a unique class of high affinity and selectivity TSPO ligands; it includes DPA-714, a fluorine-containing derivative that has also been labelled with the positron-emitter fluorine-18, and is nowadays widely used as a Positron Emission Tomography imaging probe. Recently, to prevent defluorination issues encountered invivo with this tracer, a first series of analogues was reported where the oxygen atom bridging the phenyl ring of the core structure and the fluorinated moiety was replaced with a more robust linkage. Among this new series, CfO-DPA-714 was discovered as a highly promising TSPO ligand. Herein, a novel series of fluorinated analogues of the latter molecule were synthesized and invitro characterized, where the pharmacomodulation at the amide position of the molecule was explored. Thirteen compounds were thus prepared from a common key-ester intermediate (synthesized in 7 steps from 4-iodobenzoate - 11% overall yield) and a set of commercially available amines and obtained with moderate to good yields (23-81%) and high purities (>95%). With one exception, all derivatives displayed nanomolar to subnanomolar affinity for the TSPO and also high selectivity versus the CBR (Ki (CBR)/Ki (TSPO)>103). Within this series, three compounds showed better Ki values (0.25, 0.26 and 0.30nM) than that of DPA-714 (0.91nM) and CfO-DPA-714 (0.37nM), and favorable lipophilicity for brain penetration (3.67.4&nbsp

Parkinson’s disease (PD) is a slowly progressing neurodegenerative disorder caused by loss of dopaminergic neurons in the substantia nigra (SN), leading to severe impairment in motor and non-motor functions. Endogenous subventricular zone (SVZ) neural stem cells constantly give birth to new cells which might serve as a possible source for regeneration in the adult brain. However, neurodegeneration is accompanied by neuroinflammation and dopamine depletion, potentially compromising regeneration. We therefore employed in vivo imaging methods to study striatal deafferentation ([123I]Ioflupane SPECT, DaTscanTM) and neuroinflammation in SN and striatum ([18F]DPA-714 PET) in the intranigral 6-hydroxydopamine (6-OHDA) PD mouse model. Additionally, we transduced cells in the SVZ with a lentivirus encoding firefly luciferase and followed migration of progenitor cells in the SVZ – olfactory bulb (OB) axis via bioluminescence imaging (BLI) under disease and control conditions. We found that activation of microglia in the SN is an acute process coming along with the degeneration of dopaminergic cell bodies in the SN. Dopaminergic deafferentation of the striatum does not influence the generation of Dcx+ neuroblasts in the SVZ, but generates chronic astrogliosis in the nigrostriatal system.

SSR180575 (1) is a high-affinity (0.83nM) TSPO 18kDa ligand belonging to the pyridazino[4,5-b]indole-1-acetamides family. Herein we describe the synthesis and invitro characterization of two series of new fluorinated analogues. Eleven compounds (out of fifteen) displayed nanomolar to subnamolar affinities (0.30-8.1nM) and high selectivities (Ki(CBR)/Ki(TSPO)>10(3)). Two derivatives stand out as promising candidates for drug development or use as PET probes for invivo neuroinflammation imaging, once fluorine-18-labelled.

DPA-714 (N,N-diethyl-2-(2-(4-(2-fluoroethoxy)phenyl)-5,7-dimethylpyrazolo[1,5-a]pyrimidin-3-yl)acetamide) is a recentlydiscovered fluorinated ligand of the translocator protein 18 kDa (TSPO). Labelled with the short-lived positron emitterfluorine-18, this structure is today the radioligand of reference for in vivo imaging of microglia activation andneuroinflammatory processes with positron emission tomography. In the present work, an isotopically tritium-labelledversion was developed ([3H]DPA-714), in order to access high resolution in vitro and ex vivo microscopic autoradiographystudies, repeated and long-lasting receptor binding studies and in vivo pharmacokinetic determination at late time points.Briefly, DPA-714 as reference, and its 3,5-dibrominated derivative as precursor for labelling, were both prepared from DPA-713 in nonoptimized 32% (two steps) and 10% (three steps) yields, respectively. Reductive debromination using deuteriumgas and Pd/C as catalyst in methanol, performed at the micromolar scale, confirmed the regioselective introduction of twodeuterium atoms at the meta positions of the phenyl ring. Tritiodebromination was analogously performed using no-carriertritium gas. HPLC purification provided >96% radiochemically pure [3H]DPA-714 (7 GBq) with a 2.1 TBq/mmol specificradioactivity. Interestingly, additional hydrogen-for-tritium exchanges were also observed at the 5-methyl and 7-methylpositions of the pyrazolo[1,5-a]pyrimidine, opening novel perspectives in the labelling of compounds featuring thisheterocyclic core.

Purpose: The purpose of the study was to validate [18F]DPA-714, a translocator protein (TSPO)18 kDa radioligand, as a probe to non-invasively quantify the inflammatory state in inflammatorybowel disease (IBD) animal models. Procedures: Quantitative positron emission tomography (PET) imaging of intestinal inflammationwas conducted with 2-deoxy-2-[18F]fluoro-D-glucose ([18F]FDG) a glucose metabolismsurrogate marker and [18F]DPA-714 a ligand of the 18 kDa TSPO, on two IBD models. The firstmodel was induced using dextran sodium sulfate (DSS), creating global inflammation in thecolon. The second model was induced by rectally administering trinitrobenzenesulfonic acid(TNBS), creating local and acute inflammation. Results: The level of inflammation was analyzed using PET imaging on days 7 and 8. The analysisobtained with [18F]DPA-714, yielded a significant difference between the DSS treated (0.50±0.17%ID/cc) and non-treated rats (0.35±0.15%ID/cc). [18F]FDG on the other hand did not yield asignificant difference.We did observe a mean glucose consumption in the colon increase from 0.40±0.11%ID/cc to 0.54±0.17%ID/cc. In the TNBSmodel, the uptake level of [18F]DPA-714 increasedsignificantly from 0.46±0.23%ID/cc for the non-treated group, to 1.30±0.62%ID/cc for those treated.PET signal was correlated with increased TSPO expression at cellular level. Conclusions: Results indicate that [18F]DPA-714 is suitable for studying inflammation in IBDmodels. [18F]DPA-714 could be a good molecular probe to non-invasively evaluate the level andlocalization of inflammation. Moreover, in vivo imaging using this TSPO ligand is potentially apowerful tool to stage and certainly to follow the evolution and therapeutic efficiency atmolecular level within this disease family.

DPA-C5yne, the lead compound of a novel series of DPA-714 derivatives in which the fluoroethoxy chain linked to thephenylpyrazolopyrimidine scaffold has been replaced by a fluoroalkyn-1-yl moiety, is a high affinity (Ki: 0.35 nM) and selectiveligand targeting the translocator protein 18 kDa. In the present work, DPA-C5yne was labelled with no-carrier-added [18F]fluoride based on a one-step tosyloxy-for-fluorine nucleophilic substitution reaction, purified by cartridge and HPLC, andformulated as an i.v. injectable solution using a TRACERLab FX N Pro synthesizer. Typically, 4.3–5.2GBq of [18F]DPA-C5yne,ready-to-use, chemically and radiochemically pure (> 95%), was obtained with specific radioactivities ranging from 55 to110GBq/μmol within 50–60 min, starting from a 30GBq [18F]fluoride batch (14–17%). LogP and LogD of [18F]DPA-C5yne weremeasured using the shake-flask method and values of 2.39 and 2.51 were found, respectively. Autoradiography studiesperformed on slices of ((R,S)-α-amino-3-hydroxy-5-methyl-4-isoxazolopropionique (AMPA)-lesioned rat brains showed a hightarget-to-background ratio (1.9 ± 0.3). Selectivity and specificity of the binding for the translocator protein was demonstratedusing DPA-C5yne (unlabelled), PK11195 and Flumazenil (central benzodiazepine receptor ligand) as competitors. Furthermore,DPA-C5yne proved to be stable in plasma at 37°C for at least 90min.

The TSPO (translocator protein), also known as the peripheral benzodiazepine receptor, is upregulated in the brain of subjects suffering from neurodegenerative disorders such as Alzheimer's, Parkinson's and Huntington's disease. Moreover, this overexpression has been proved to be linked to microglia activation making thus the TSPO a marker of choice of neuroinflammatory processes and therefore a potential target for the development of radioligands for positron emission tomography imaging. The discovery of selective TSPO ligands and their labelling with the short-lived positron-emitter isotopes carbon-11 and fluorine-18 emerged in the mid-1980s with the preparation of the 3-isoquinolinecarboxamide [(11) C]PK11195. To date, an impressive number of promising compounds-[(11) C]PK11195-challengers-have been developed; some radioligands-for example, [(11) C]PBR28, [(11) C]DPA-713, [(18) F]FEDAA1106 and [(18) F]DPA-714-are currently used in clinical trials. As illustrated in this review, the methodologies applied for the preparation of these compounds remain mainly [(11) C]methylations using [(11) C]MeI or [(11) C]MeOTf and SN 2-type nucleophilic aliphatic [(18) F]fluorinations-two processes illustrating the state-of-the-art arsenal of reactions that involves these two short-lived radioisotopes-but alternative processes, such as [(11) C]carbonylations using [(11) C]CO and [(11) C]COCl2 as well as SN Ar-type nucleophilic [(18) F]fluorinations, have also been reported and as such, reviewed herein.