Enzyme Inhibition Assays for Monoamine Oxidase.

Monoamine oxidase (MAO) catalyzes the oxidative deamination of monoamines with two isoforms, namely, MAO-A and MAO-B, in mitochondrial outer membranes. These two types of MAO-A and MAO-B participate in changes in levels of neurotransmitter such as serotonin (5-hydroxytryptamine) and dopamine. Selective MAO-A inhibitors have been targeted for anti-depression treatment, while selective MAO-B inhibitors are targets of therapeutic agents for Alzheimer's disease and Parkinson's disease. For this reason, study on the development of MAO inhibitors has recently become important. Here, we describe methods of MAO activity assay, especially continuous spectrophotometric methods, which give relatively high accuracy. MAO-A and MAO-B can be assayed using kynuramine and benzylamine as substrates, respectively, at 316 nm and 250 nm, respectively, to measure their respective products, 4-hydroxyquinoline and benzaldehyde. Inhibition degree and pattern can be analyzed by using the Lineweaver-Burk and secondary plots in the presence of inhibitor, and reversibility of inhibitor can be determined by using the dialysis method.

Inhibiting Monoamine Oxidase in CNS and CVS would be a Promising Approach to Mitigating Cardiovascular Complications in Neurodegenerative Disorders.

The flavoenzyme monoamine oxidases (MAOs) are present in the mitochondrial outer membrane and are responsible for the metabolism of biogenic amines. MAO deamination of biological amines produces toxic byproducts such as amines, aldehydes, and hydrogen peroxide, which are significant in the pathophysiology of multiple neurodegenerative illnesses. In the cardiovascular system (CVS), these by-products target the mitochondria of cardiac cells leading to their dysfunction and producing redox imbalance in the endothelium of the blood vessels. This brings up the biological relationship between the susceptibility of getting cardiovascular disorders in neural patients. In the current scenario, MAO inhibitors are highly recommended by physicians worldwide for the therapy and management of various neurodegenerative disorders. Many interventional studies reveal the benefit of MAO inhibitors in CVS. Drug candidates who can target both the central and peripheral MAO could be a better to compensate for the cardiovascular comorbidities observed in neurodegenerative patients.

Flavonoid and Chalcone Scaffolds as Inhibitors of BACE1: Recent Updates.

Flavonoids and chalcones are two major classes of chemical moieties that have a vast background of pharmacological activities. Chalcone is a subclass of flavonoids whose therapeutic potential has been implicated due to an array of bioactivities. A lot of research works have shown interest in investigating the neuroprotective effect of these molecules, and have revealed them to be much more potent molecules that can be used to treat neurodegenerative disorders. Beta-site APP cleaving enzyme (BACE1), which is majorly found in the brain, is one of the reasons behind the development of Alzheimer's disease (AD). Flavonoids and chalcones have proven clinical data that they inhibit the production of Aß plaques that are involved in the progression of AD. In this article, we have provided a detailed chronological review of the research work on the BACE1 inhibiting potency of both flavonoids and chalcones. Almost all the flavonoids and chalcones mentioned in this article have shown very good in vitro and in vivo BACE1 inhibiting activity. The docking studies and the structural importance of some BACE1-inhibiting flavonoids, as well as chalcones, are also mentioned here.

Halogenated class of oximes as a new class of monoamine oxidase-B inhibitors for the treatment of Parkinson's disease: Synthesis, biochemistry, and molecular dynamics study.

Oximes are the promising structural scaffold for inhibiting monoamine oxidase (MAO)-B. Eight chalcone-based oxime derivatives were synthesized by microwave-assisted technique, and their ability to inhibit human MAO (hMAO) enzymes were tested. All compounds showed higher inhibitory activity of hMAO-B than hMAO-A. In the CHBO subseries, CHBO4 most potently inhibited hMAO-B with an IC50 value of 0.031 µM, followed by CHBO3 (IC50 = 0.075 µM). In the CHFO subseries, CHFO4 showed the highest inhibition of hMAO-B with an IC50 value of 0.147 µM. Compound CHBO4 had the highest selectivity index (SI) value of 1290.3. However, CHBO3 and CHFO4 showed relatively low SI values of 27.7 and 19.2, respectively. The -Br substituent in the CHBO subseries at the para-position in the B-ring showed higher hMAO-B inhibition than the -F substituent in the CHFO subseries. In both series, hMAO-B inhibition increased with the substituents at para-position in A-ring (-F > -Br > -Cl > -H in order). Compound CHBO4 (-F in A-ring and -Br in B-ring) was 12.6-times potent than the substituents-reversed compound CHFO3 (-Br in A-ring and -F in B-ring; IC50 = 0.391 µM). In the kinetic study, Ki values of CHBO4 and CHFO4 for hMAO-B were 0.010 ± 0.005 and 0.040 ± 0.007 µM, respectively, with competitive inhibitions. Reversibility experiments showed that CHBO4 and CHFO4 were reversible hMAO-B inhibitors. In the cytotoxicity test using the Vero cells by the MTT technique, CHBO4 had low toxicity with an IC50 value of 128.8 µg/mL. In H2O2-induced cells, CHBO4 significantly reduced cell damage by scavenging reactive oxygen species (ROS). Molecular docking and dynamics showed the stable binding mode of the lead molecule CHBO4 on the active site of hMAO-B. These results suggest that CHBO4 is a potent reversible, competitive, and selective hMAO-B inhibitor and can be used as a treatment agent for neurological disorders.

KIF1A-Associated Neurological Disorder: An Overview of a Rare Mutational Disease.

KIF1A-associated neurological diseases (KANDs) are a group of inherited conditions caused by changes in the microtubule (MT) motor protein KIF1A as a result of KIF1A gene mutations. Anterograde transport of membrane organelles is facilitated by the kinesin family protein encoded by the MT-based motor gene KIF1A. Variations in the KIF1A gene, which primarily affect the motor domain, disrupt its ability to transport synaptic vesicles containing synaptophysin and synaptotagmin leading to various neurological pathologies such as hereditary sensory neuropathy, autosomal dominant and recessive forms of spastic paraplegia, and different neurological conditions. These mutations are frequently misdiagnosed because they result from spontaneous, non-inherited genomic alterations. Whole-exome sequencing (WES), a cutting-edge method, assists neurologists in diagnosing the illness and in planning and choosing the best course of action. These conditions are simple to be identified in pediatric and have a life expectancy of 5-7 years. There is presently no permanent treatment for these illnesses, and researchers have not yet discovered a medicine to treat them. Scientists have more hope in gene therapy since it can be used to cure diseases brought on by mutations. In this review article, we discussed some of the experimental gene therapy methods, including gene replacement, gene knockdown, symptomatic gene therapy, and cell suicide gene therapy. It also covered its clinical symptoms, pathogenesis, current diagnostics, therapy, and research advances currently occurring in the field of KAND-related disorders. This review also explained the impact that gene therapy can be designed in this direction and afford the remarkable benefits to the patients and society.

A comprehensive review of the docking studies of chalcone for the development of selective MAO-B inhibitors.

Monoamine oxidase B is a crucial therapeutic target for neurodegenerative disorders like Alzheimer's and Parkinson's since they assist in disintegrating neurotransmitters such as dopamine in the brain. Pursuing efficacious monoamine oxidase B inhibitors is a hot topic, as contemporary therapeutic interventions have many shortcomings. Currently available FDA-approved monoamine oxidase inhibitors like safinamide, selegiline and rasagiline also have a variety of side effects like depression and insomnia. In the quest for a potent monoamine oxidase B inhibitor, sizeable, diverse chemical entities have been uncovered, including chalcones. Chalcone is a renowned structural framework that has been intensively explored for its monoamine oxidase B inhibitory activity.The structural resemblance of chalcone (1,3-diphenyl-2-propen-1-one) based compounds and 1,4-diphenyl-2-butene, a recognized MAO-B inhibitor, accounts for their MAO-B inhibitory activity. Therefore, multiple revisions to the chalcone scaffold have been attempted by the researchers to scrutinize the implications of substitutions onthe molecule's potency. In this work, we outline the docking investigation results of various chalcone analogues with monoamine oxidase B available in the literature until now to understand the interaction modes and influence of substituents. Here we focused on the interactions between reported chalcone derivatives and the active site of monoamine oxidase B and the influence of substitutions on those interactions. Detailed images illustrating the interactions and impact of the substituents or structural modifications on these interactions were used to support the docking results.

Exploration of a new class of monoamine oxidase B inhibitors by assembling benzyloxy pharmacophore on halogenated chalcones.

Eight derivatives of benzyloxy-derived halogenated chalcones (BB1-BB8) were synthesized and tested for their ability to inhibit monoamine oxidases (MAOs). MAO-A was less efficiently inhibited by all compounds than MAO-B. Additionally, the majority of the compounds displayed significant MAO-B inhibitory activities at 1 µM with residual activities of less than 50%. With an IC50 value of 0.062 µM, compound BB4 was the most effective in inhibiting MAO-B, followed by compound BB2 (IC50 = 0.093 µM). The lead molecules showed good activity than the reference MAO-B inhibitors (Lazabemide IC50 = 0.11 µM and Pargyline Pargyline IC50 = 0.14). The high selectivity index (SI) values for MAO-B were observed in compounds BB2 and BB4 (430.108 and 645.161, respectively). Kinetics and reversibility experiments revealed that BB2 and BB4 were reversible competitive MAO-B inhibitors with Ki values of 0.030 ± 0.014 and 0.011 ± 0.005 µM, respectively. Swiss target prediction confirmed the high probability in the targets of MAO-B for both compounds. Hypothetical binding mode revealed that the BB2 or BB4 is similarly oriented to the binding cavity of MAO-B. Based on the modelling results, BB4 showed a stable confirmation during the dynamic simulation. From these results, it was concluded that BB2 and BB4 were potent selective reversible MAO-B inhibitors and they can be considered drug candidates for treating related neurodegenerative diseases such as Parkinson's disease.

Current and Future Nano-Carrier-Based Approaches in the Treatment of Alzheimer's Disease.

It is a very alarming situation for the globe because 55 million humans are estimated to be affected by Alzheimer's disease (AD) worldwide, and still it is increasing at the rapid speed of 10 million cases per year worldwide. This is an urgent reminder for better research and treatment due to the unavailability of a permanent medication for neurodegenerative disorders like AD. The lack of drugs for neurodegenerative disorder treatment is due to the complexity of the structure of the brain, mainly due to blood-brain barrier, because blood-brain drug molecules must enter the brain compartment. There are several novel and conventional formulation approaches that can be employed for the transportation of drug molecules to the target site in the brain, such as oral, intravenous, gene delivery, surgically implanted intraventricular catheter, nasal and liposomal hydrogels, and repurposing old drugs. A drug's lipophilicity influences metabolic activity in addition to membrane permeability because lipophilic substances have a higher affinity for metabolic enzymes. As a result, the higher a drug's lipophilicity is, the higher its permeability and metabolic clearance. AD is currently incurable, and the medicines available merely cure the symptoms or slow the illness's progression. In the next 20 years, the World Health Organization (WHO) predicts that neurodegenerative illnesses affecting motor function will become the second-leading cause of mortality. The current article provides a brief overview of recent advances in brain drug delivery for AD therapy.

Introduction of benzyloxy pharmacophore into aryl/heteroaryl chalcone motifs as a new class of monoamine oxidase B inhibitors.

The inhibitory action of fifteen benzyloxy ortho/para-substituted chalcones (B1-B15) was evaluated against human monoamine oxidases (hMAOs). All the molecules inhibited hMAO-B isoform more potently than hMAO-A. Furthermore, the majority of the molecules showed strong inhibitory actions against hMAO-B at 10 µM level with residual activities of less than 50%. Compound B10 has an IC50 value of 0.067 µM, making it the most potent inhibitor of hMAO-B, trailed by compound B15 (IC50 = 0.12 µM). The thiophene substituent (B10) in the A-ring exhibited the strongest hMAO-B inhibition structurally, however, increased residue synthesis did not result in a rise in hMAO-B inhibition. In contrast, the benzyl group at the para position of the B-ring displayed more hMAO-B inhibition than the other positions. Compounds B10 and B15 had relatively high selectivity index (SI) values for hMAO-B (504.791 and 287.600, respectively). Ki values of B10 and B15 were 0.030 ± 0.001 and 0.033 ± 0.001 µM, respectively. The reversibility study showed that B10 and B15 were reversible inhibitors of hMAO-B. PAMPA assay manifested that the benzyloxy chalcones (B10 and B15) had a significant permeability and CNS bioavailability with Pe value higher than 4.0 × 10-6 cm/s. Both compounds were stabilized in protein-ligand complexes by the π-π stacking, which enabled them to bind to the hMAO-B enzyme's active site incredibly effectively. The hMAO-B was stabilized by B10- and B15-hMAO-B complexes, with binding energies of - 74.57 and - 87.72 kcal/mol, respectively. Using a genetic algorithm and multiple linear regression, the QSAR model was created. Based on the best 2D and 3D descriptor-based QSAR model, the following statistics were displayed: R2 = 0.9125, Q2loo = 0.8347. These findings imply that B10 and B15 are effective, selective, and reversible hMAO-B inhibitors.

Conjugated Dienones from Differently Substituted Cinnamaldehyde as Highly Potent Monoamine Oxidase-B Inhibitors: Synthesis, Biochemistry, and Computational Chemistry.

Fifteen multiconjugated dienones (MK1-MK15) were synthesized and evaluated to determine their inhibitory activities against monoamine oxidases (MAOs) A and B. All derivatives were found to be potent and highly selective MAO-B inhibitors. Compound MK6, with an IC50 value of 2.82 nM, most effectively inhibited MAO-B, like MK12 (IC50 = 3.22 nM), followed by MK5, MK13, and MK14 (IC50 = 4.02, 4.24, and 4.89 nM, respectively). The selectivity index values of MK6 and MK12 for MAO-B over MAO-A were 7361.5 and 1780.5, respectively. Compounds MK6 and MK12 were competitive reversible inhibitors of MAO-B, with K i values of 1.10 ± 0.20 and 3.0 ± 0.27 nM, respectively. Cytotoxic studies showed that MK5, MK6, MK12, and MK14 exhibited low toxicities on Vero cells, with IC50 values of 218.4, 149.1, 99.96, and 162.3 µg/mL, respectively, which were much higher than those for their effective nanomolar-level concentrations. Also, MK5, MK6, MK12, and MK14 decreased cell damage in H2O2-induced cells via a significant scavenging effect of reactive oxygen species. Molecular modeling was performed to rationalize the potential inhibitory activities of MK5, MK6, MK12, and MK14 toward MAO-B and their possible binding mechanisms, showing high-affinity binding pocket interactions and conformation perturbations of the compounds with MAO-B, which were interpreted as the conformational dynamics of MAO-B. This study concluded that all the compounds tested were more potent MAO-B inhibitors than the reference drugs, and leading compounds could be further explored for their effectiveness in various kinds of neurodegenerative disorders.

Navigating into the Chemical Space of Monoamine Oxidase Inhibitors by Artificial Intelligence and Cheminformatics Approach.

The monoamine oxidase (MAO) enzyme class is a prevalent target for many neurodegenerative and depressive disorders. Even though scrutinization of many promising drugs for the treatment of MAO inhibition has been carried out in recent times, a conclusive structural requirement for potent activity needs to be developed. Numerous approaches have been examined for the identification of structural features for potent MAO inhibitors (MAOIs) that mainly involve an array of computational studies, synthetic approaches, and biological evaluation. In this paper, we have analyzed ∼2200 well-known MAOIs to expand perceptions in the chemical space of MAOIs. The physicochemical properties of the MAOIs disclosed a discernible hydrophobic feature making a bunch discrete from the central nervous system (CNS) acting drugs, as exposed using the principal component analysis (PCA). The Murcko scaffold structure study revealed unfavorable and favorable scaffold structures, in both data sets, with the highest biological activity shown by the 3-phenyl-2H-chromen-2-one scaffold. This scaffold showed a polypharmacological effect. R-group disintegration and automatic structure-activity relationship (SAR) study resulted in identification of substructures responsible for the inhibitory bioactivity of the MAO-A and MAO-B enzymes. Moreover, with activity cliff analysis, significant biological activity was detected by simple molecular conversion in the chemical compound structure. In addition, we used the machine learning tool to generate a hypothesis wherein pyrazole, benzene ring, and amide containing structural functionalities can exhibit potential biological activities. This hypothesis revealed that CNS target drugs, C4155, C13390, C21265, C43862, C31524, C24810, C37100, C42075, and C43644, could be repurposed as valuable candidates for the MAO-B enzyme. For researchers, this study will bring new perceptions in the discovery and development of MAOIs and direct lead and hit optimization for the progress of small molecules beneficial for MAO-targeting associated diseases.

Deciphering the detailed structure-activity relationship of coumarins as Monoamine oxidase enzyme inhibitors-An updated review.

In the last few years, Monoamine oxidase (MAO) have emerged as a target for the treatment of many neurodegenerative diseases including anxiety, depression, Alzheimer's, and Parkinson's diseases. The MAO inhibitors especially selective and reversible inhibitors of either of the isoenzymes (MAO-A & MAO-B) have been given more attention as both the form have different therapeutic properties and hence can be used for different neurological disorders. The lack of selective and reversible inhibitors available for both the enzymes and severity of the neuronal disorder in society have opened a new door to the researchers to carry out large and dedicated researches in this field. Among the several classes of the molecule as the inhibitors, coumarins hold a rank as a potent scaffold with its ease of synthesis, high therapeutic potential, and reversibility in inhibiting MAOs. The current review is an update of the research in the field that covers the works during the last six years (2014-2020) with a major focus on the SAR of the coumarin derivatives including synthetic, natural, and hybrids of coumarins with FDA-approved drugs.

Development of Halogenated Pyrazolines as Selective Monoamine Oxidase-B Inhibitors: Deciphering via Molecular Dynamics Approach.

Halogens have been reported to play a major role in the inhibition of monoamine oxidase (MAO), relating to diverse cognitive functions of the central nervous system. Pyrazoline/halogenated pyrazolines were investigated for their inhibitory activities against human monoamine oxidase-A and -B. Halogen substitutions on the phenyl ring located at the fifth position of pyrazoline showed potent MAO-B inhibition. Compound 3-(4-ethoxyphenyl)-5-(4-fluorophenyl)-4,5-dihydro-1H-pyrazole (EH7) showed the highest potency against MAO-B with an IC50 value of 0.063 µM. The potencies against MAO-B were increased in the order of -F (in EH7) > -Cl (EH6) > -Br (EH8) > -H (EH1). The residual activities of most compounds for MAO-A were > 50% at 10 µM, except for EH7 and EH8 (IC50 = 8.38 and 4.31 µM, respectively). EH7 showed the highest selectivity index (SI) value of 133.0 for MAO-B, followed by EH6 at > 55.8. EH7 was a reversible and competitive inhibitor of MAO-B in kinetic and reversibility experiments with a Ki value of 0.034 ± 0.0067 µM. The molecular dynamics study documented that EH7 had a good binding affinity and motional movement within the active site with high stability. It was observed by MM-PBSA that the chirality had little effect on the overall binding of EH7 to MAO-B. Thus, EH7 can be employed for the development of lead molecules for the treatment of various neurodegenerative disorders.