Discovery and Evaluation of Enantiopure 9H-pyrimido[4,5-b]indoles as Nanomolar GSK-3β Inhibitors with Improved Metabolic Stability

Abstract

Glycogen synthase kinase-3β (GSK-3β) is a potential target in the field of Alzheimer’s disease drug discovery. We recently reported a new class of 9H-pyrimido[4,5-b]indole-based GSK-3β inhibitors, of which 3-(3-((7-chloro-9H-pyrimido[4,5-b]indol-4-yl)(methyl)amino)piperidin-1-yl)propanenitrile (1) demonstrated promising inhibitory potency. However, this compound underwent rapid degradation by human liver microsomes. Starting from 1, we prepared a series of amide-based derivatives and studied their structure–activity relationships against GSK-3β supported by 1 µs molecular dynamics simulations. The biological potency of this series was substantially enhanced by identifying the eutomer configuration at the stereocenter. Moreover, the introduction of an amide bond proved to be an effective strategy to eliminate the metabolic hotspot. The most potent compounds, (R)-3-(3-((7-chloro-9H-pyrimido[4,5-b]indol-4-yl)(methyl)amino)piperidin-1-yl)-3-oxopropanenitrile ((R)-2) and (R)-1-(3-((7-bromo-9Hpyrimido[4,5-b]indol-4-yl)(methyl)amino)piperidin-1-yl)propan-1-one ((R)-28), exhibited IC50 values of 480 nM and 360 nM, respectively, and displayed improved metabolic stability. Their favorable biological profile is complemented by minimal cytotoxicity and neuroprotective properties.

Addressing a Trapped High-Energy Water: Design and Synthesis of Highly Potent Pyrimidoindole-Based Glycogen Synthase Kinase-3β Inhibitors

“…We recently reported the synthetic preparation of piperidines 4a – j and the enantiopure analogs ( R ) -4c , ( R )-4d , ( S ) -4c , and ( S )-4d from 9-tosyl-9 H -pyrimido­[4,5- b ]­indoles 3a – j (Scheme ). These piperidine intermediates display diverse substitution patterns of the tricyclic core and served as precursors to prepare the final compounds 5a – j , ( R ) -5c , ( R )-5d , ( S ) -5c , and ( S )-5d within the herein reported study. As previously described in the synthesis of inhibitor 1 , the cyanoethyl substituent was installed on the piperidine nitrogen by a Michael reaction with acrylonitrile …”

“…Furthermore, the introduction of a methyl group in the pyrimidine 2-position of parent compound 1 afforded an inactive derivative (5j, IC 50 value >10 μM). As we recently observed a strong influence of the stereoconfiguration on the bioactivity of structurally related GSK-3β inhibitors, 35 we also decided to evaluate the activity of enantiopure compounds within this series. Here, however, this effect was less pronounced.…”

“…12 Moreover, our established synthetic methodologies were successfully applied to the corresponding 7-iodine derivatives (Scheme 3). 35 In detail, previously reported 7-iodo-9-tosyl-9Hpyrimido[4,5-b]indole (6) was reacted with N-Boc-3-(methylamino)piperidine in a nucleophilic aromatic substitution (S N Ar) to yield intermediate 8. In the case of the enantiopure analog (S)-8, commercially available (S)-N-Boc-3-aminopiperidine was used in the S N Ar resulting in (S)-7, which was subsequently methylated with iodomethane under strictly anhydrous basic conditions.…”

(S)-3-(3-((7-Ethynyl-9H-pyrimido[4,5-b]indol-4-yl)amino)piperidin-1-yl)propanenitrile

“…The title compound 2 was prepared starting from the previously reported 4-chloro-7-iodo-9H-pyrimido [4,5-b]indole (3) [5], which was treated with an excess of commercially available tert-butyl (S)-3-aminopiperidine-1-carboxylate and DIPEA in n-BuOH [6] (Scheme 1). Substitution product 4 underwent Boc deprotection under acidic conditions, resulting in free piperidine 5, which was reacted with acrylonitrile in MeOH to introduce the cyanoethyl substituent (6).…”