[18F]RS-127445 radiosynthesis and evaluation as a 5-HT2B receptor PET radiotracer in rat brain.

Serotonin (5-hydroxytryptamine, 5-HT) is a neurotransmitter involved in many physiological and pathological mechanisms through its numerous receptors. Among these, the 5-HT2B receptor is known to play a key role in multiple brain disorders but remains poorly understood. Positron emission tomography (PET) can contribute to a better understanding of pathophysiological mechanisms regulated by the 5-HT2B receptor. To develop the first PET radiotracer for the 5-HT2B receptor, RS-127445, a well-known 5-HT2B receptor antagonist, was labeled with fluorine-18. [18F]RS-127445 was synthesized in a high radiochemical purity and with a good molar activity and radiochemical yield. Preliminary PET scans in rats showed good brain penetration of [18F]RS-127445. However, competition experiments and in vitro autoradiography showed high non-specific binding, especially to brain white matter.

Discovery of MT-1207: A Novel, Potent Multitarget Inhibitor as a Promising Clinical Candidate for the Treatment of Hypertension.

Herein, a series of novel arylpiperazine (piperidine) derivatives were designed, synthesized, and evaluated for mechanisms of action through in vitro and in vivo studies. The most promising compound, II-13 (later named as MT-1207), is a potent α1 and 5-HT2A receptor antagonist with remarkable IC50 in the picomolar level. Importantly, in the in vivo assay, II-13 achieved an effective blood pressure (BP) reduction in the 2K2C rat model without damaging renal function. Compound II-13, with its significant advantages in terms of pharmacological effects, pharmacokinetic parameters, and a large safety window, was extensively investigated. Moreover, data also showed that compound II-13 had fewer side effects in a postural BP assay and could prevent the onset of postural hypotension. Together, these results suggested that compound II-13 is a highly potent antihypertensive drug candidate with multitarget mechanisms of action in preclinical models. Currently, MT-1207 is in phase II hypertensive clinical trials in China.

Semisynthetic Transformations on (+)-Boldine Reveal a 5-HT2A/2CR Antagonist.

Aporphine alkaloids have shown affinity for serotonin receptors (5-HTRs), and there has been a recent upsurge of interest in aporphines as 5-HT2CR ligands. 1,2,9,10-Tetraoxygenated aporphine alkaloids in particular have demonstrated good affinity for 5-HTRs. In continued efforts to understand the impacts of structural modification of the 1,2,9,10-tetraoxygenated aporphine template on affinity, selectivity, and activity at 5-HT2R subtypes, we used (+)-boldine (8) as a semisynthetic feedstock in the preparation of C-2-alkoxylated (+)-predicentrine analogues. Compound 10n, which contains a benzyloxy group at C-2, has been identified as a novel 5-HT2CR ligand with strong affinity (4 nM) and moderate selectivity versus 5-HT2BR and 5-HT2AR (12-fold and 6-fold, respectively). Compound 10n functions as an antagonist at 5-HT2A and 5-HT2C receptors. Computational experiments indicate that several hydrophobic interactions as well as strong H-bond and salt bridge interactions between the protonated amine moiety in 10n and Asp134 are responsible for the potent 5-HT2CR affinity of this compound. Furthermore, compound 10n displays favorable predicted drug-like characteristics, which is encouraging toward future optimization.

Novel serotonin 5-HT2A receptor antagonists derived from 4-phenylcyclohexane-5-spiro-and 5-methyl-5-phenyl-hydantoin, for use as potential antiplatelet agents.

BACKGROUND: Antiplatelet drugs have been used in the treatment of acute coronary syndromes and for the prevention of recurrent events. Unfortunately, many patients remain resistant to the available antiplatelet treatment. Therefore, there is a clinical need to synthesize novel antiplatelet agents, which would be associated with different pathways of platelet aggregation, to develop an alternative or additional treatment for resistant patients. Recent studies have revealed that 5-HT2A receptor antagonists could constitute alternative antiplatelet therapy. METHODS: Based on the structures of the conventional 5-HT2A receptor ligands, two series of compounds with 4-phenylcyclohexane-5-spiro- or 5-methyl-5-phenyl-hydantoin core linked to various arylpiperazine moieties were synthesized and their affinity for 5-HT2A receptor was assessed. Further, we evaluated their antagonistic potency at 5-HT2A receptors using isolated rat aorta and cells expressing human 5-HT2A receptors. Finally, we studied their anti-aggregation effect and compared it with ketanserin and sarpogrelate, the reference 5-HT2A receptor antagonists. Moreover, the structure-activity relationships were studied following molecular docking to the 5-HT2A receptor model. RESULTS: Functional bioassays revealed some of the synthesized compounds to be moderate antagonists of 5-HT2A receptors. Among them, 13, 8-phenyl-3-(3-(4-phenylpiperazin-1-yl)propyl)-1,3-diazaspiro[4.5]decane-2,4-dione, inhibited collagen stimulated aggregation (IC50 = 27.3 µM) being more active than sarpogrelate (IC50 = 66.8 µM) and comparable with ketanserin (IC50 = 32.1 µM). Moreover, compounds 2-5, 9-11, 13, 14 inhibited 5-HT amplified, ADP- or collagen-induced aggregation. CONCLUSIONS: Our study confirmed that the 5-HT2A antagonists effectively suppress platelet aggregation and remain an interesting option for the development of novel antiplatelet agents with an alternative mechanism of action.

Design, Synthesis, and Pharmacological Characterization of Heterobivalent Ligands for the Putative 5-HT2A/mGlu2 Receptor Complex.

We report the synthesis of the first series of heterobivalent ligands targeting the putative heteromeric 5-HT2A/mGlu2 receptor complex, based on the 5-HT2A antagonist MDL-100,907 and the mGlu2 ago-PAM JNJ-42491293. The functional properties of monovalent and heterobivalent ligands were characterized in 5-HT2A-, mGlu2/Gqo5-, 5-HT2A/mGlu2-, and 5-HT2A/mGlu2/Gqo5-expressing HEK293 cells using a Ca2+ imaging assay and a [3H]ketanserin binding assay. Pronounced functional crosstalk was observed between the two receptors in 5-HT2A/mGlu2 and 5-HT2A/mGlu2/Gqo5 cells. While the synthesized monovalent ligands retained the 5-HT2A antagonist and mGlu2 ago-PAM functionalities, the seven bivalent ligands inhibited 5-HT-induced responses in 5-HT2A/mGlu2 cells and both 5-HT- and Glu-induced responses in 5-HT2A/mGlu2/Gqo5 cells. However, no definitive correlation between the functional potency and spacer length of the ligands was observed, an observation substantiated by the binding affinities exhibited by the compounds in 5-HT2A, 5-HT2A/mGlu2, and 5-HT2A/mGlu2/Gqo5 cells. In conclusion, while functional crosstalk between 5-HT2A and mGlu2 was demonstrated, it remains unclear how these heterobivalent ligands interact with the putative receptor complex.

Synthesis and biological evaluation of thioadatanserin and its dialkylated products as partial 5-HTR1A agonists and 5-HTR2A antagonists for potential use in depression and anxiety disorders.

Thionation of adatanserin hydrochloride (2) with Lawesson's reagent in toluene/triethylamine afforded novel compound, (3r,5r,7r)-N-(2-(4-(pyrimidin-2-yl)piperazin-1-yl)ethyl)adamantane-1-carbothioamide (thioadatanserin, 3) in 84-90% isolated yield. Thioadatanserin underwent a tandem double alkylation with methyl iodide and benzyl bromide in NaH/THF to produce novel dialkylated products 6 and 7 respectively. The single X-ray crystal structure of 7 was determined to be 1-(2-((E- ((3r,5r,7r)-adamantan-1-yl)benzylthio)methylene)amino)ethyl)-1-benzyl-4- (pyrimidin-2-yl)piperazin-1-ium bromide showing that the piperazine ring adopts a chair-like configuration that is not co-planar with the pyrimidine ring. Thioadatanserin emerged as a dual potent partial agonist with activity against 5-HTR1A (EC50 6.7 nM) and antagonist activity against 5-HTR2A (IC50 62.3 nM) and was selective over 5-HTR2C receptor (IC50 > 3333 nM) in the PathHunter® ß-arrestin assays.

Design, Synthesis, and Biological Evaluation of New Peripheral 5HT2A Antagonists for Nonalcoholic Fatty Liver Disease.

Nonalcoholic fatty liver disease (NAFLD) is increasingly prevalent worldwide, causing serious liver complications, including nonalcoholic steatohepatitis. Recent findings suggest that peripheral serotonin (5-hydroxytryptamine, 5HT) regulates energy homeostasis, including hepatic lipid metabolism. More specifically, liver-specific 5HT2A knockout mice exhibit alleviated hepatic lipid accumulation and hepatic steatosis. Here, structural modifications of pimavanserin (CNS drug), a 5HT2A antagonist approved for Parkinson's disease, led us to synthesize new peripherally acting 5HT2A antagonists. Among the synthesized compounds, compound 14a showed good in vitro activity, good liver microsomal stability, 5HT subtype selectivity, and no significant inhibition of CYP and hERG. The in vitro and in vivo blood-brain barrier permeability study proved that 14a acts peripherally. Compound 14a decreased the liver weight and hepatic lipid accumulation in high-fat-diet-induced obesity mice. Our study suggests new therapeutic possibilities for peripheral 5HT2A antagonists in NAFLD.

Targeting the 5-HT2C Receptor in Biological Context and the Current State of 5-HT2C Receptor Ligand Development.

Serotonin (5-HT) 5-HT2C receptor (5-HT2CR) is recognized as a critical mediator of diseaserelated pathways and behaviors based upon actions in the central nervous system (CNS). Since 5-HT2CR is a class A G protein-coupled receptor (GPCR), drug discovery efforts have traditionally pursued the activation of the receptor through synthetic ligands with agonists proposed for the treatment of obesity, substance use disorders and impulse control disorders while antagonists may add value for the treatment of anxiety, depression and schizophrenia. The most significant agonist discovery to date is the FDAapproved anti-obesity medication lorcaserin. In recent years, efforts towards developing other mechanisms to enhance receptor function have resulted in the discovery of Positive Allosteric Modulators (PAMs) for the 5-HT2CR, with several molecule series now reported. The biological significance and context for signaling and function of the 5-HT2CR, and the current status of 5-HT2CR agonists and PAMs are discussed in this review.

Synthesis and activity of functionalizable derivatives of the serotonin (5-HT) 5-HT2A receptor (5-HT2AR) antagonist M100907.

The approach of tethering together two known receptor ligands, to be used as molecular probes for the study of G protein-coupled receptor (GPCR) systems, has proven to be a valuable approach. Selective ligands that possess functionality that can be used to link to other ligands, are useful in the development of novel antagonists and agonists. Such molecules can also be attached to reporter molecules, such as fluorophores, for the study of GPCR dimerization and its role in signaling. The highly selective serotonin (5-HT) 5-HT2A receptor (5-HT2AR) antagonist M100907 (volinanserin) is of clinical interest in the treatment of neurological and mental health disorders. Here, we synthesized the most active (+)-M100907 enantiomer as well as a series of derivatives that possessed either an alkyne or an azide. The triazole resulting from the dipolar cycloaddition of these groups did not interfere with the ability of the bivalent ligand to act as an antagonist. Thus, we have synthesized a number of compounds which will prove useful in elucidating the role of the 5-HT2AR in the central nervous system.

Synthesis and biological evaluation of a series of novel pyridinecarboxamides as potential multi-receptor antipsychotic drugs.

In previous study, a series of benzamides was identified as potent antipsychotic agents. As a continuation of the program to discover novel antipsychotics, herein we reported the evaluation of a series of pyridinecarboxamide derivatives. The most promising compound 7h not only held good activities on dopamine D2, serotonin 5-HT1A and 5-HT2A receptors, but also exhibited low potency for α1A, H1 and 5-HT2C receptors, indicating a low propensity of side effects like orthostatic hypotension and weight gain. Furthermore, 7h exhibited more potent antipsychotic-like effect than aripiprazole in behavioral studies. The preliminary results were promising enough for further research around this scaffold.

5-HT1A and 5-HT2A receptors affinity, docking studies and pharmacological evaluation of a series of 8-acetyl-7-hydroxy-4-methylcoumarin derivatives.

In this work we describe the synthesis, docking studies and biological evaluation of a focused library of novel arylpiperazinyl derivatives of 8-acetyl-7-hydroxy-4-methylcoumarin. The new compounds were screened for their 5-HT1A and 5-HT2A receptor affinity. Among the evaluated compounds, six displayed high affinities to 5-HT1A receptors (4a-0.9 nM, 6a-0.5 nM, 10a-0.6 nM, 3b-0.9 nM, 6b-1.5 nM, 10b-1 nM). Compound 6a and 10a bearing a bromo- or methoxy- substituent in ortho position of the piperazine phenyl ring, were identified as potent antagonists of the 5-HT1A receptors. In the tail suspension test, mice injected with 6a showed a dose-dependent increase in depressive-like behavior that was related to a decrease in locomotor activity. Compound 10a did not decrease or prolong immobility time nor did it affect home cage activity. Molecular docking studies using 5-HT1A and 5-HT2A homology models revealed structural basis of the high affinity of ortho-substituted derivatives and subtle changes in amino acid interactions patterns depending on the length of the alkyl linker.

Homology Modeling Inspired Synthesis of 5-HT2A Receptor Inhibitors: A Diazepine Analogue of the Atypical Antipsychotic JL13.

BACKGROUND: The benzoxazepine JL13 is an analogue of the clozapine family of antipsychotic agents which target the 5-HT2A receptor, and has showed promise as an atypical antipsychotic agent. Based on the dearth of clinically effective anti-psychotic agents available, we sought to design and chemically synthesize additional analogues. METHODS: Structure function analysis was conducted using state of art computational methods, which were designed to highlight new candidates for chemical synthesis. Efficient syntheses were then conducted and the products screened for affinity to the receptor. RESULTS: Among many new analogues prepared, an aza analogue demonstrated seventeen times greater affinity for the receptor than JL13. CONCLUSION: An efficient synthetic route to an aza-analogue of JL13 was developed and will allow rapid modifications of the core and synthesis of related libraries.

4-Phenyl quinoline derivatives as potential serotonin receptor ligands with antiproliferative activity.

Antagonists of signaling receptors are often effective non-toxic therapeutic agents. Over the years, there have been evidences describing the role of serotonin or 5-hydroxytryptamine (5-HT) in development of cancer. Although there are reports on the antiproliferative effects of some serotonin receptor antagonists, there are very few investigations related to understanding their structure-activity relationships. In this study, we report the screening of a library of 4-phenyl quinoline derivatives for their antiproliferative activities. Preliminary docking studies indicated that these ligands had the ability to bind to two of the serotonin receptors, 5-HT1B and 5-HT2B. The results of the in silico experiments were validated by performing in vitro studies on MCF-7 breast cancer cell line. The ethylpiperazine derivatives showed maximum toxicity against this cancer cell line. The compounds inhibited Calcium ion efflux (induced by serotonin) and ERK activation. One of the most active 4-phenyl quinoline derivatives (H3a) also induced apoptosis, thereby, suggesting the use of this scaffold as a potential anticancer drug.

Structure-Based Scaffold Repurposing for G Protein-Coupled Receptors: Transformation of Adenosine Derivatives into 5HT2B/5HT2C Serotonin Receptor Antagonists.

Adenosine derivatives developed to activate adenosine receptors (ARs) revealed micromolar activity at serotonin 5HT2B and 5HT2C receptors (5HTRs). We explored the structure-activity relationship at 5HT2Rs and modeled receptor interactions in order to optimize affinity and simultaneously reduce AR affinity. Depending on N6 substitution, small 5'-alkylamide modification maintained 5HT2BR affinity, which was enhanced upon ribose substitution with rigid bicyclo[3.1.0]hexane (North (N)-methanocarba), e.g., N6-dicyclopropylmethyl 4'-CH2OH derivative 14 (Ki 11 nM). 5'-Methylamide 23 was 170-fold selective as antagonist for 5HT2BR vs 5HT2CR. 5'-Methyl 25 and ethyl 26 esters potently antagonized 5HT2Rs with moderate selectivity in comparison to ARs; related 6-N,N-dimethylamino analogue 30 was 5HT2R-selective. 5' position flexibility of substitution was indicated in 5HT2BR docking. Both 5'-ester and 5'-amide derivatives displayed in vivo t1/2 of 3-4 h. Thus, we used G protein-coupled receptor modeling to repurpose nucleoside scaffolds in favor of binding at nonpurine receptors as novel 5HT2R antagonists, with potential for cardioprotection, liver protection, or central nervous system activity.

Asymmetric total synthesis of (+)-O-methylasparvenone, a rare nitrogen-free serotonin 2C receptor antagonist.

The first enantioselective synthesis of the fungal metabolite (+)-O-methylasparvenone was achieved in eight steps and 22% overall yield from inexpensive 3,4,5-trimethoxybenzaldehyde dimethyl acetal. Key steps include (i) early-stage asymmetric alkynylation of an aromatic aldehyde with a propiolate, (ii) intramolecular Friedel-Crafts acylation, and (iii) site-selective cleavage of an aryl methyl ether.

Reformulating a Pharmacophore for 5-HT2A Serotonin Receptor Antagonists.

Several pharmacophore models have been proposed for 5-HT2A serotonin receptor antagonists. These typically consist of two aromatic/hydrophobic moieties separated by a given distance from each other, and from a basic amine. Although specified distances might vary, the models are relatively similar in their general construction. Because our preliminary data indicated that two aromatic (hydrophobic) moieties might not be required for such action, we deconstructed the serotonin-dopamine antipsychotic agent risperidone (1) into four smaller structural fragments that were thoroughly examined in 5-HT2A receptor binding and functional (i.e., two-electrode voltage clamp (TEVC) and intracellular calcium release) assays. It was apparent that truncated risperidone analogues behaved as antagonists. In particular, 6-fluoro-3-(1-methylpiperidin-4-yl)benzisoxazole (4) displayed high affinity for 5-HT2A receptors (Ki of ca. 12 nM) relative to risperidone (Ki of ca. 5 nM) and behaved as a potent 5-HT2A serotonin receptor antagonist. These results suggest that multiple aromatic (hydrophobic) moieties are not essential for high-affinity 5-HT2A receptor binding and antagonist activity and that current pharmacophore models for such agents are very much in need of revision.

Synthesis and biological investigation of new equatorial (ß) stereoisomers of 3-aminotropane arylamides with atypical antipsychotic profile.

A series of novel 3ß-aminotropane derivatives containing a 2-naphthalene or a 2-quinoline moiety was synthesised and evaluated for their affinity for 5-HT1A, 5-HT2A and D2 receptors. Their affinity for the receptors was in the nanomolar to micromolar range. p-Substitution (6c, 6f, 6i, 6l, 6o), as well as substitution with chlorine atoms (6g, 6h, 6i), led to a significant increase in binding affinity for D2 receptors with compounds 6f (Ki=0.6nM), 6c and 6i (Ki=0.4nM), having the highest binding affinities. m-Substituted derivatives were the most promising ligands in terms of 5-HT2A receptor binding affinity whereas 2-quinoline derivatives (10a, 10b) displayed the highest affinity for 5-HT1AR and were the most selective ligands with Ki=62.7nM and Ki=30.5nM, respectively. Finally, the selected ligands 6b, 6d, 6e, 6g, 6h, 6k, 6n and 6o, with triple binding activity for the D2, 5-HT1A and 5-HT2A receptors, were subjected to in vivo tests, such as those for induced hypothermia, climbing behaviour and the head twitch response, in order to determine their pharmacological profile. The tested ligands presented neither agonist nor antagonist properties for the 5-HT1A receptors in the induced hypothermia and lower lip retraction (LLR) tests. All tested compounds displayed antagonistic activity against 5-HT2A, with 6n and 6o being the most active. Four (6b, 6k, 6n and 6o) out of eight tested compounds could be classified as D2 antagonists. Additionally, evaluation of metabolic stability was performed for selected ligands, and introduction of halogen atoms into the benzene ring of 6h, 6k, 6n and 6o improved their metabolic stability. The project resulted in the selection of the lead compounds 6n and 6o, which had antipsychotic profiles, combining dopamine D2-receptor and 5-HT2A antagonism and metabolic stability.

Structure-Based Discovery of Novel and Selective 5-Hydroxytryptamine 2B Receptor Antagonists for the Treatment of Irritable Bowel Syndrome.

Here we employed structure-based ligand discovery techniques to explore a recently determined crystal structure of the 5-hydroxytryptamine 2B (5-HT2B) receptor. Ten compounds containing a novel chemical scaffold were identified; among them, seven molecules were active in cellular function assays with the most potent one exhibiting an IC50 value of 27.3 nM. We then systematically probed the binding characteristics of this scaffold by designing, synthesizing, and testing a series of structural modifications. The structure-activity relationship studies strongly support our predicted binding model. The binding profiling across a panel of 11 5-HT receptors indicated that these compounds are highly selective for the 5-HT2B receptor. Oral administration of compound 15 (30 mg/kg) produced significant attenuation of visceral hypersensitivity in a rat model of irritable bowel syndrome (IBS). We expect this novel scaffold will serve as the foundation for the development of 5-HT2B antagonists for the treatment of IBS.

Discovery of novel potent and selective ligands for 5-HT2A receptor with quinazoline scaffold.

A series of compounds with quinazoline scaffold were designed, synthesized and evaluated as novel potent 5-HT2A receptor ligands. N-(4-Chlorophenyl)-2-(piperazin-1-yl)quinazolin-4-amine (5o) has a Ki value of 14.04 ± 0.21 nM, with a selectivity more than 10,000 fold over 5-HT1A receptors (D1 and D2-like receptors). The functional assay showed that this compound is an antagonist to 5-HT2A receptor with an IC50 value of 1.66 µM.

New arylpiperazinylalkyl derivatives of 8-alkoxy-purine-2,6-dione and dihydro[1,3]oxazolo[2,3-f]purinedione targeting the serotonin 5-HT1A /5-HT2A /5-HT7 and dopamine D2 receptors.

To obtain potential antidepressants and/or antipsychotics, a series of new long-chain arylpiperazine derivatives of 8-alkoxy-purine-2,6-dione (10-24) and dihydro[1,3]oxazolo[2,3-f]purinedione (30-34) were synthesized and their serotonin (5-HT1A , 5-HT2A , 5-HT6 , 5-HT7 ) and dopamine (D2 ) receptor affinities were determined. The study allowed the identification of some potent 5-HT1A /5-HT7 /D2 ligands with moderate affinity for 5-HT2A sites. The binding mode of representative compounds from both chemical classes (11 and 31) in the site of 5-HT1A receptor was analyzed in computational studies. In functional in vitro studies, the selected compounds 15 and 16 showed antagonistic properties for the evaluated receptors. 8-Methoxy-7-{4-[4-(2-methoxyphenyl)-piperazin-1-yl]-butyl}-1,3-dimethyl-purine-2,6-dione (15) showed a lack of activity in terms and under the conditions of the forced swim, four plate and amphetamine-induced hyperactivity tests in mice, probably as a result of its high first pass effect in the liver.