Tetrazine-trans-cyclooctene ligations are remarkably fast and selective reactions even at low micro-molar concentrations. In bioorthogonal radiochemistry, tools that enable conjugation of radioactive probes to pre-targeted vectors are of great interest. Herein, we describe the successful development of the first (11)C-labelled tetrazine and its reaction with trans-cyclooctenol.

The serotonin (5-hydroxytryptamine [5-ΗΤ]) 7 receptor (5-HT 7 R) is the most recently discovered 5-HT receptor, and its physiologic and possible pathophysiologic roles are not fully elucidated. So far, no suitable 5-HT 7 R PET radioligand is available, thus limiting the investigation of this receptor in the living brain. Here, we present the radiosynthesis and in vivo evaluation of piperazine-1-yl]butyl}p-1,3-dihydro-2H-indol-2-one) and piperazine-1-yl]butyl}-1,3-dihydro-2H-indol-2-one) as selective 5-HT 7 R PET radioligands in the pig brain. The 5-HT 7 R distribution in the postmortem pig brain is also assessed. Methods: In vitro autoradiography with the 5-HT 7 R selective radioligand 3 H-labeled (R)-3-(2-(2-(4-methylpiperidin-1-yl) ethyl)pyrrolidine-1-sulfonyl)phenol (SB-269970) was performed on pig brain sections to establish the 5-HT 7 R binding distribution. Radiolabeling of 5-HT 7 R selective compounds was performed in an automated synthesis module in which we conducted either palladiummediated cross coupling ( 11 C-Cimbi-712) or conventional O-methylation ( 11 C-Cimbi-717) using 11 C-MeI and 11 C-MeOTf, respectively. After intravenous injection of the radioligands in Danish Landrace pigs, the in vivo brain distribution of the ligands was studied. Specific binding of 11 C-Cimbi-712 and 11 C-Cimbi717 to 5-HT 7 R was investigated by intravenous administration of SB-269970 before a second PET scan. Results: High 5-HT 7 R density was found in the thalamus and cortical regions of the pig brain by autoradiography. The radiosynthesis of both radioligands succeeded after optimization efforts (radiochemical yield, ∼20%-30% at the end of synthesis). Timeactivity curves of 11 C-Cimbi-712 and 11 C-Cimbi-717 showed high brain uptake and distribution according to 5-HT 7 R distribution, but the tracer kinetics of 11 C-Cimbi-717 were faster than 11 C-Cimbi-712. Both radioligands were specific for 5-HT 7 R, as binding could be blocked by pretreatment with SB-269970 for 11 C-Cimbi-717 in a dose-dependent fashion. For 11 C-Cimbi-717, nondisplaceable binding potentials of 6.4 ± 1.2 (n 5 6) were calculated in the thalamus. Conclusion: Both 11 C-Cimbi-712 and 11 C-Cimbi-717 generated a specific binding in accordance with 5-HT 7 R distribution and are potential PET radioligands for 5-HT 7 R. 11 C-Cimbi-717 is the better candidate because of the more reversible tracer kinetics, and this radioligand showed a dose-dependent decline in cerebral binding after receptor blockade. Thus, 11 C-Cimbi-717 is currently the most promising radioligand for investigation of 5-HT 7 R binding in the living human brain. The serotonergic system plays a key modulatory role in the brain and is a target for many drug treatments for brain disorders either through reuptake blockade or interactions with one or more of the 14 subtypes of 5-hydroxytryptamine (5-HT) receptors. Our knowledge about the behavior of the 5-HT system in vivo is still scattered, and most of the understanding is derived from animal models. However, the use of imaging techniques such...