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Inhibition of Insulin-Regulated Aminopeptidase by Imidazo[1,5-α]pyridines; Synthesis and Evaluation
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry. (Preparative Medicinal Chemistry)
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Preparative Medicinal Chemistry.ORCID iD: 0000-0002-0817-8140
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Preparative Medicinal Chemistry.ORCID iD: 0000-0001-8852-6071
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

Inhibition of Insulin-regulated Aminopeptidase (IRAP) has been shown to improve cognitive functions in several animal models. Recently, we performed a screening campaign identifying novel small-molecule based compounds acting as inhibitors of the enzymatic activity IRAP. Here we report on the chemical synthesis, structure-activity relationships (SAR) and initial characterization of physicochemical properties of a series of imidazo[1,5-α]pyridine-based inhibitors, including delineation of their mode of action as non-competitive inhibitors with a small L-leucine-based IRAP substrate. The best compound displays an pIC50 values of 6.0. We elucidate the importance of two chiral sites in these molecules and find they have little impact on the compound´s metabolic stability or physicochemical properties. The carbonyl group of a central urea moiety was initially believed to mimic substrate binding to a catalytically important Zn2+ ion in the active site, although the plausibility of this binding hypothesis is challenged by observation of excellent selectivity versus the closely related aminopeptidase N (APN). Taken together with the non-competitive inhibition pattern, we also consider an alternative model of allosteric binding.

Keywords [en]
Insulin-regulated aminopeptidase, IRAP, inhibitors
National Category
Medicinal Chemistry
Research subject
Medicinal Chemistry
Identifiers
URN: urn:nbn:se:uu:diva-406413OAI: oai:DiVA.org:uu-406413DiVA, id: diva2:1412783
Available from: 2020-03-07 Created: 2020-03-07 Last updated: 2020-03-07
In thesis
1. Inhibitors Targeting Insulin-Regulated Aminopeptidase (IRAP): Identification, Synthesis and Evaluation
Open this publication in new window or tab >>Inhibitors Targeting Insulin-Regulated Aminopeptidase (IRAP): Identification, Synthesis and Evaluation
2020 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Insulin-regulated aminopeptidase (IRAP) has emerged as a potential new therapeutic target for treatment of cognitive disorders. Inhibition of the enzymatic activity facilitates cognition in rodents. Potent and selective peptide and pseudopeptide based inhibitors have been developed, but most of them suffer from poor pharmacokinetics and blood-brain-barrier penetration. Hence, development of less-complex inhibitors with good pharmacokinetic properties are of great importance.

The aim of this thesis was to identify and optimize new small-molecule based IRAP inhibitors for use as research tools to investigate the cognitive effects of IRAP inhibition. Adaptation of an existing enzymatic assay into a screening compatible procedure allowed the evaluation of 10,500 compounds as IRAP inhibitors. The screening campaign resulted in 23 compounds displaying more than 60% inhibition. Two of these compounds, a spiro-oxindole dihydroquinazolinone and an imidazo[1,5-α]pyridine, were further investigated in terms of structure-activity relationship, physicochemical properties, metabolic stability, and mechanism of inhibition.

Spiro-oxindole dihydroquinazolinone based IRAP inhibitors were synthesized via fast and simple microwave-promoted reactions, either in batch or in a continuous flow approach. The most potent compounds displayed sub-µM affinity, and interestingly an uncompetitive mode of inhibition with the synthetic substrate used in the assay. Molecular modeling confirmed the possibility of simultaneous binding of the compounds and the substrate. Furthermore, the molecular modeling suggested that the S-enantiomer accounts for the inhibitory effect observed with this compound series. The compounds also proved inactive on the closely related enzyme aminopeptidase N. Unfortunately, the spiro-oxindole based inhibitors suffered from poor solubility and metabolic stability.

Imidazo[1,5-α]pyridine based IRAP inhibitors were synthesized via a five step procedure, providing inhibitors in the low-µM range. The stereospecificity of a methyl group proved important for inhibition. The compound series displayed no inhibitory activity on aminopeptidase N. Intriguing, these compounds exhibit a noncompetitive inhibition mechanism with the model substrate. As observed for the spiro-compounds, the imidazopyridines suffered from both poor solubility and metabolic stability.  

In summary, the work presented in this thesis provide synthetic procedures, initial structure-activity relationship, and pharmacological evaluation of two distinct inhibitors classes. The compounds are among the first non-peptidic IRAP inhibitors presented, serving as interesting starting points in the development of research tools for use in models of cognition.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2020. p. 86
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Pharmacy, ISSN 1651-6192 ; 285
Keywords
compound screening, insulin-regulated aminopeptidase, IRAP, inhibitors, cognitive disorders, spiro-oxindole, quinazolinone, imidazopyridine, medicinal chemistry, structure-activity relationship, microwave heating
National Category
Medicinal Chemistry
Research subject
Medicinal Chemistry
Identifiers
urn:nbn:se:uu:diva-406417 (URN)978-91-513-0894-4 (ISBN)
Public defence
2020-04-24, Room B41, BMC, Husargatan 3, Uppsala, 13:15 (Swedish)
Opponent
Supervisors
Available from: 2020-04-03 Created: 2020-03-07 Last updated: 2020-05-19

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Rosenström, UlrikaGising, JohanHallberg, MathiasLarhed, Mats

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Engen, KarinLundbäck, ThomasRosenström, UlrikaGising, JohanHallberg, MathiasLarhed, Mats
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