Technical Aspect of the Automated Synthesis and Real-Time Kinetic Evaluation of [¹¹C]SNAP-7941

Abstract

Positron emission tomography (PET) is an essential molecular imaging technique providing insights into pathways and using specific targeted radioligands for in vivo investigations. Within this protocol, a robust and reliable remote-controlled radiosynthesis of [ 11 C]SNAP-7941, an antagonist to the melanin-concentrating hormone receptor 1, is described. The radiosynthesis starts with cyclotron produced [ 11 C]CO 2 that is subsequently further reacted via a gas-phase transition to [ 11 C]CH 3 OTf. Then, this reactive intermediate is introduced to the precursor solution and forms the respective radiotracer. Chemical as well as the radiochemical purity are determined by means of RP-HPLC, routinely implemented in the radiopharmaceutical quality control process. Additionally, the molar activity is calculated as it is a necessity for the following real-time kinetic investigations. Furthermore, [ 11 C]SNAP-7941 is applied to MDCKII-WT and MDCKII-hMDR1 cells for evaluating the impact of P-glycoprotein (P-gp) expression on cell accumulation. For this reason, the P-gp expressing cell line (MDCKII-hMDR1) is either used without or with blocking prior to experiments by means of the P-gp substrate (±)-verapamil and the results are compared to the ones observed for the wildtype cells. The overall experimental approach demonstrates the importance of a precise time-management that is essential for every preclinical and clinical study using PET tracers radiolabelled with short-lived nuclides, such as carbon-11 (half-life: 20 min). Video Link The video component of this article can be found at https://www.jove.com/video/59557/ 11 C]SNAP-7941. µPET imaging in rats demonstrated low brain accumulation, which increased dramatically after administration of the P-gp inhibitor tariquidar 11. These data suggested that [ 11 C]SNAP-7941 is a substrate of this efflux transporter system impeding ligand binding to central MCHR1. Unfortunately, there is still a lack of adequate in vitro models enabling the prediction of BBB penetration in an early stage of tracer development.

Pitfalls and solutions of the fully-automated radiosynthesis of [11C]metoclopramide

“…The radiotracer formulation part of the module was washed with water and ethanol including the bulb and product collection vial. The preparation procedure was performed similarly for the GE TRACERlab™ FX C Pro and TRACERlab™ FX2 C. A detailed protocol for the module preparation was published recently (Vraka et al 2019).

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“…The automatic radiosynthesis was then started by delivering 116 ± 17 GBq of [ 11 C]CO 2 to the carbon dioxide trap and confirmation of the end of delivery. The gas phase transformation of [ 11 C]CO 2 via the formation of [ 11 C]CH 4 produced approximately 50 GBq of [ 11 C]CH 3 I and has been described in detail elsewhere (Vraka et al 2019; Pichler et al 2018a; Philippe et al 2018; Larsen et al 1997). [ 11 C]CH 3 OTf was produced by passing [ 11 C]CH 3 I gas through the silver triflate column at 200 °C.…”

Discovery of melanin‐concentrating hormone receptor 1 in brown adipose tissue

“…A link between BAT and the MCH system is further supported by neuronal MCH projections to brainstem regions harboring sympathetic neurons involved in BAT activation. 32 As a result, PET-tracers visualizing MCHR1 were developed [33][34][35][36][37][38][39][40][41][42] to enable its noninvasive in vivo quantification and monitoring of related pathologies, as well as to support drug development.…”

“…Taking advantage of the specific MCHR1 antagonist SNAP-7941, 43 we developed the radiolabeled analogue [ 11 C]SNAP-7941 and its fluoroethylated derivative [ 18 F]FE@SNAP as specific PET-tracers targeting the MCHR1 in the central nervous system. [33][34][35][36][37][38][39][40][41] However, during a preclinical evaluation, both tracers were found to be P-glycoprotein substrates. 35,40,44 Unexpectedly, nonnegligible MCHR1-tracer uptake in BAT of rats was repeatedly detected via small animal PET (μPET) (Fig.…”