Preparation of bicyclic nitrogen intermediates as useful scaffolds for the synthesis of new biologically active compounds.
Creators: name: SALVI, MATTEO
Publisher: name: Universit� degli Studi di Milano
Year: 2014

description: The discovery of the orexin (also known as hypocretin) neuropeptide system determined a major advancement in the understanding of sleep and arousal states and its study emerged as one of the new key fields of investigation in neuroscience. The orexin signaling system is highly conserved across species and it is based on two hypotalamic peptides, orexin-A (OXA) and orexin-B (OXB). The orexin peptides were found to bind to the previously identified orphan 7TM G-protein coupled receptors, orexin receptor-1 (OX1R) and orexin receptor-2 (OX2R). Many compelling evidences, suggest that the orexin system is strongly implicated in sleep disorders, being, for instance, a strong genetic linkage between orexin hypofunction and regulation of wakefulness. Besides sleep and arousal, the orexinergic system has an important role in controlling other features of CNS functions, i.e. anxiety, depression, feeding, drug addiction and so on. Combined genetic and pharmacological approaches indicate that orexin signaling may represent a confluence of sleep, feeding and reward pathways. Therefore, many efforts to obtain selective orexin receptor antagonists (SORAs) towards OX1R or OX2R were attempted. Nevertheless, dual orexin receptor antagonists (DORAs), maybe due to a somehow synergistic effect produced by their action on both receptors, seem to be the most promising agents for treating insomnia.
Some of these orexin receptor antagonists known in literature, result of research done by several pharmaceutical companies, contain as a fundamental motif the ethylendiaminic sequence NCCN.
On these basis, we planned to insert the NCCN motif in a bicyclic framework such as the norbornane (bicyclo[2.2.1]heptane) in order to obtain a markedly rigid structure, in the order to confer specific features to the final derivatives.
Three distinct NCCN-containing scaffolds (different regiochemistry) were proposed at the beginning of the Ph.D. project: TYPE I scaffold, TYPE II scaffold and TYPE III scaffold.
After a detailed research in the literature, we found that the TYPE I scaffold was already successfully been employed in molecules with orexin antagonism activity, so we decided to focus the research only on TYPE II and TYPE III scaffolds.
The TYPE II and TYPE III scaffolds were synthetized in both exo/endo-stereochemistry and subsequently were functionalized. The functionalization of both nitrogen atoms of the NCCN motif with proper substituents allowed for the preparation of a small library of new compounds.
From TYPE II scaffold exploration were obtained 11 endo-derivatives (9a-k) and 9 exo-derivatives (12a-i) as racemic mixtures. While for the TYPE III scaffold were conduced a most targeted exploration with the synthesis of 2 exo-derivatives (23a,b) and 5 endo-derivatives (24a-e), as racemic mixtures.
In support to the synthetic exploration, a computational study was conduced in order to identify a Pharmacophoric Model able to indicate compounds with potential biological activity toward the orexin receptors. The first remark emerged for the TYPE II scaffold derivatives was the best fitting in the Pharmacophoric Model of the compounds with the exo-stereochemistry. In the case of the TYPE III scaffold derivatives emerged that the conformation adopted from the endo-analogues accommodate in best manner with the Pharmacophore features.
Subsequently, the biological functional activity of the TYPE II and TYPE III scaffolds derivatives toward the orexin receptors were assayed using a FLIPR protocol.
From the biological activity evaluation it emerged that the TYPE II derivatives with the exo-stereochemistry were active at nanomolar level towards both OX1R and OX2R, while the endo- assessment shown no antagonism. On the contrary, the endo-TYPE III scaffold derivatives showed nanomolar activity for both the orexin receptors (even if generally most active towards OX1R).
These results confirm the observation extrapolated from the computational study conduced.
The two copounds 12e and 24b emerged from the respective series as powerful antagonist towards both the orexin receptors and selected for further evaluations. First, this two compound were subjected to chiral HPLC separation in order to study the stereospecificity of their interaction and to determine the respective eutomer. For both the compounds only one of the obtained enantiomers retain low nanomolal antagonist activity, that is compounds (+)-12e and (+)-24b.
The two compounds (+)-12e and (+)-24b were selected for further investigation in order to obtain a complete biological characterization. The drug to drug interaction (DDI) potentials were verified in a CYP inhibition test on the major isoforms and the two compounds were further assessed for their pharmacokinetic profiles in rats.
From biological data and pharmacokinetic measurements the lead cmpds (+)-12e and (+)-24b are promising DORAs and they will be evaluated in further pharmacological tests to assess their potential as new treatment for insomnia.