Substrate specificity and inhibitor sensitivity of monoamine oxidase in rat kidney mitochondria.

The deamination of the substrates 5-hydroxytryptamine (5-HT), tyramine, dopamine, β-phenylethylamine and benzylamine by rat kidney mitochondrial monoamine oxidase (MAO) was studied, and kinetic constants are reported for each substrate. By the use of the selective MAO inhibitors, clorgyline and deprenyl, 5-HT and benzylamine were found to be substrates for types A and B MAO, respectively, in this tissue, whereas the other substrates were metabolized by both forms of MAO. No evidence for any significant metabolism of 5-HT or benzylamine by other amine oxidases was obtained. However, some conditions under which the carbonyl reagents semicarbazide, isoniazid and aminoguanidine may interfere with assays for MAO, without actually affecting enzyme activity directly, are described. Preincubation of kidney mitochondria with histamine resulted in a time- and oxygen-dependent irreversible inhibition of both type A and type B MAO activity; the exact nature of the inhibitory agent and its mode of action remain to be determined.     

Inhibition of monoamine oxidases A and B by simple isoquinoline alkaloids: racemic and optically active 1,2,3,4-tetrahydro-, 3,4-dihydro-, and fully aromatic isoquinolines

A series of 1,2,3,4-tetrahydro-, 3,4-dihydro-, and fully aromatic isoquinolines were tested as substrates and/or inactivators of highly purified human monoamine oxidase A and B (MAO A and B). None were found to be a substrate for either enzyme, but many of these isoquinolines could selectively inhibit either MAO A or B. Stereoselective competitive inhibition of MAO A was found with the R enantiomer of all the stereoisomers tested, including salsolinol (Ki = 31 microM), salsoline (Ki = 77 microM), salsolidine (Ki = 6 microM), and carnegine (Ki = 2 microM). As a class, the 3,4-dihydro-isoquinolines were the most potent inhibitors tested (Ki = 2-130 microM), and the fully aromatic isoquinolines had intermediate activity (Ki = 17-130 microM) against MAO A. In contrast, only a few of these compounds markedly inhibited MAO B. 1,2,3,4-Tetrahydroisoquinoline, its 2-methyl derivative, and o-methylcorypalline gave apparent Ki values of 15, 1, and 29 microM, respectively, and two 3,4-dihydroisoquinolines (compounds 22 and 25) showed substantial inhibition of MAO B (Ki = 76 and 15 microM, respectively). These results support the concept that the topography of the inhibitor binding site differs in MAO A and B.     

Rasagiline [N-propargyl-1R(+)-aminoindan], a selective and potent inhibitor of mitochondrial monoamine oxidase B

1. Rasagiline [N-propargyl-1R(+)-aminoindan], was examined for its monoamine oxidase (MAO) A and B inhibitor activities in rats together with its S(-)-enantiomer (TVP 1022) and the racemic compound (AGN-1135) and compared to selegiline (1-deprenyl). The tissues that were studied for MAO inhibition were the brain, liver and small intestine. 2. While rasagiline and AGN1135 are highly potent selective irreversible inhibitors of MAO in vitro and in vivo, the S(-) enantiomer is relatively inactive in the tissues examined. 3. The in vitro IC(50) values for inhibition of rat brain MAO activity by rasagiline are 4.43+/-0.92 nM (type B), and 412+/-123 nM (type A). The ED(50) values for ex vivo inhibition of MAO in the brain and liver by a single dose of rasagiline are 0.1+/-0.01, 0.042+/-0.0045 mg kg(-1) respectively for MAO-B, and 6.48+/-0.81, 2.38+/-0.35 mg kg(-1) respectively for MAO-A. 4. Selective MAO-B inhibition in the liver and brain was maintained on chronic (21 days) oral dosage with ED(50) values of 0.014+/-0.002 and 0.013+/-0.001 mg kg(-1) respectively. 5. The degree of selectivity of rasagiline for inhibition of MAO-B as opposed to MAO-A was similar to that of selegiline. Rasagiline was three to 15 times more potent than selegiline for inhibition of MAO-B in rat brain and liver in vivo on acute and chronic administration, but had similar potency in vitro. 6. These data together with lack of tyramine sympathomimetic potentiation by rasagiline, at selective MAO-B inhibitory dosage, indicate that this inhibitor like selegiline may be a useful agent in the treatment of Parkinson's disease in either symptomatic or L-DOPA adjunct therapy, but lack of amphetamine-like metabolites could present a therapeutic advantage for rasagiline.     

Synthesis of novel N-substituted imidazolecarboxylic acid hydrazides as monoamine oxidase inhibitors

Novel 2-alkylsulfanyl-1-benzyl-5-imidazolecarboxylic acid hydrazides (15a,b) were synthesized as analogues of isocarboxazide, which is a known nonselective irreversible monoamine oxidase inhibitor and tested for monoamine oxidase A and B inhibitory activity. Neither of the compounds showed any inhibition of MAO B activity up to a high concentration of 100 microM. An MAO A activity was only slowly inhibited at this high concentration after prolonged incubation with either compound. This suggests any observed inhibition is not very specific.     

Enzyme-activated irreversible inhibitors of monoamine oxidase: phenylallylamine structure-activity relationships

Seventeen 2-aryl-3-haloallylamine derivatives were prepared and evaluated as inhibitors of monoamine oxidase (MAO, EC 1.4.3.4). The synthesis of these compounds was achieved from either alpha-methylstyrene or ring-substituted phenylacetic acid derivatives. With one exception, these 2-arylallylamines were found to be enzyme-activated, irreversible inhibitors of MAO. The most potent inhibitors were ring-substituted derivatives of (E)-2-phenyl-3-fluoroallylamine with IC50 values ranging from 10(-6) to 10(-8) M. Selectivity for the A and B form of MAO was found to depend on the nature of aromatic ring substitution. In general, hydroxyl substitution favored the inactivation of the A form of MAO, while very selective B inhibitors were obtained when the aromatic ring was substituted with a 4-methoxy group. (E)-2-(4-Methoxyphenyl)-3-fluoroallylamine and (E)-2-(3,4-dimethoxyphenyl)-3-fluoroallylamine proved to be in vitro as selective for the B form of MAO as deprenyl.     

The kinetics of monoamine oxidase inhibition by three 2-indolylmethylamine derivatives

The inhibition of bovine brain mitochondrial MAO-A and MAO-B by three acetylenic and non-acetylenic derivatives of 2-indolylmethylamine, chosen among more than 100 new compounds, were studied. The non-acetylenic derivative N-methyl-2(5-hydroxy-1-methylindolyl)methylamine (1) was a weak non-selective inhibitor which was shown to act in a reversible and competitive manner towards the deamination of tyramine. The two acetylenic derivatives N-methyl-N-(2-propynyl)-2-(5-benzyloxy-1-methylindolyl)methylamine (2) and N-methyl-N-(2-propynyl)-2-(5-hydroxy-1-methylindolyl)methylamine (3) were potent MAO inhibitors, one of them non-selective (compound 2) and the other MAO-A selective inhibitor (compound 3). Both of them were irreversible and competitive inhibitors, compound 2 towards the deamination of tyramine and compound 3 towards the deamination of serotonin and beta-phenylethylamine. A mechanism for the inhibition of the enzyme by both irreversible inhibitors is proposed and the inhibition parameters are determined.     

Kinetic Studies of N-Allenic Analogues of Tryptamine as Monoamine Oxidase Inhibitors

A series of N-allenic analogues of tryptamine in which the side chain is located at the 2 position of the indole ring, but which differed in the ring and side-chain nitrogen substituents, were assayed kinetically as MAO A and MAO B inhibitors. All the compounds studied were mechanism-based inhibitors. The kinetic constants of each inhibition step K_i and k_i, were determined for both MAO A and B. The data obtained indicated that these allenic derivatives show a greater selectivity and potency towards MAO A as inhibitors than the corresponding acetylenic derivatives.     

Rapid synthesis of flavone‐based monoamine oxidase (MAO) inhibitors targeting two active sites using click chemistry

A new library of flavone derivatives targeting two active sites of monoamine oxidases (“aromatic cage” and substrate cavity) were designed and synthesized using click chemistry (CuAAC reaction) between 6‐N₃‐2‐phenyl chromones ( Az1–Az2 ) and a series of alkynes ( k1–k20 ). Their inhibitory activities against MAO isoforms (MAO‐A and MAO‐B) are evaluated. Compounds with fluorine, amide bonds, or amino bonds have shown better inhibition. The most potent flavone MAO inhibitor studied is Az2k19 (1.6 μm for MAO‐A, 2.1 μm for MAO‐B), while Az1k15 and Az2k15 displayed better selectivity toward MAO‐B (SI > 10). Docking studies are in accordance with our hypothesis that these inhibitors are most likely located at both the substrate cavity and the “aromatic cage”. Our results show that it is considerable to develop new MAO inhibitors from C6 substitution of flavone derivatives and that these compounds are also potential for the treatment of diseases associated with MAOs.     

Coumarins derivatives as dual inhibitors of acetylcholinesterase and monoamine oxidase.

A set of 17 coumarin and 2 chromone derivatives with known inhibitory activity toward monoamine oxidase (MAO) A and B were tested as acetylcholinesterase (AChE) inhibitors. All compounds inhibited AChE with values in the micromolar range (3-100 microM). A kinetic study showed that most compounds acted as noncompetitive AChE inhibitors. This finding may be of interest in the context of Alzheimer's disease because recent observations suggest that MAO and AChE inhibition might decrease beta-amyloid deposition.     

4-Fluoro-3-nitrophenyl azide, a selective photoaffinity label for type B monoamine oxidase

The effects of 4-fluoro-3-nitrophenyl azide (FNPA) on types A and B monoamine oxidase in rat brain cortex were studied using serotonin and phenylethylamine as substrates respectively. FNPA competitively inhibited the oxidative deamination of both serotonin (Ki = 3 microM) and phenylethylamine (Ki = 0.78 microM) in the dark. Upon photoirradiation in the presence of FNPA, a photodependent inhibition of type B MAO activity resulted. This photodependent inhibition was apparently irreversible since there was no recovery of activity upon washing of the photolyzed FNPA-enzyme mixture. Additional evidence for the photoinduced covalent binding of FNPA to type B MAO is that non-competitive inhibition kinetics resulted after photolysis. The specificity of the photodependent incorporation of FNPA to type B MAO was shown by the protective effect of phenylethylamine and by decreased [3H]pargyline labeling after the enzyme was photolyzed with FNPA. Under the same experimental conditions, only minimal photodependent inhibition of type A MAO by FNPA was found. The observed difference in the efficiencies of the photodependent inactivation of the two types of MAO by FNPA suggests that there is a conformational or a structural difference in the active sites of the two types of MAO. The active site of type B MAO could be characterized by utilizing FNPA as a photoaffinity labeling probe.     

串果藤中甘草素和异甘草素对大鼠单胺氧化酶的体外抑制作用(英文)

目的:研究串果藤中甘草素和异甘草素体外对单胺氧化酶A型和B型的抑制作用.方法:根据不同的离心速度制备大鼠全脑粗线粒体作为单胺氧化酶的酶源;分别以5-羟基[侧链-2-~(14)C]色胺肌酸硫酸盐([~(14)C]5-HT)和2-苯基[1-~(14)C]乙基胺盐酸盐([~(14)C]β-PEA)为单胺氧化酶A型和B型放射性底物,用液体闪烁技术,研究甘草素和异甘草素酶抑制作用和抑制类型.结果:甘草素和异甘草素对单胺氧化酶A型和B型均具有抑制作用,呈良好的量效关系,对单胺氧化酶A型的IC_(50)(95％的可信限)分别为32(26-36)μmol/L和13.9(12.8-15.6)μmol/L,对单胺氧化酶B型的IC_(50)值分别为104.6(89.0-118.9)μmol/L和47.2(39.5-54.5)μmol/L.酶抑制特征曲线显示甘草素和异甘草素对单胺氧化酶A型呈非竞争性抑制,K_i值分别为31.5μmol/L和14.3μmol/L,而对单胺氧化酶B型呈混合竞争性抑制,K_i值分别为164.7μmol/L和62.2μmol/L,K_I值分别为15.2μmol/L和9.3μmol/L.结论:甘草素和异甘草素体外对单胺氧化酶A型呈非竞争性抑制作用,对单胺氧化酶B型呈混合竞争性抑制作用.     

Inhibition of monoamine oxidase in 5-hydroxytryptaminergic neurones by substituted p-aminophenylalkylamines.

A series of substituted p-aminophenethylamines and some related compounds were examined with regards to the inhibition of monoamine oxidase (MAO) in vivo inside and outside 5-hydroxytryptaminergic neurones in the rat hypothalamus. This was recorded as the protection against the irreversible inhibition of MAO produced by phenelzine by determining the remaining deaminating activity in the absence and presence of citalopram using a low (0.1 microM) concentration of [14C]-5-hydroxytryptamine (5-HT) as substrate. Some of the phenethylamines were much more potent inside than outside the 5-hydroxytryptaminergic neurones. This neuronal selectivity was antagonized by pretreatment of the rats with norzimeldine, a 5-HT uptake inhibitor, which indicates that these compounds are accumulated in the 5-HT nerve terminals by the 5-HT pump. Selectivity was obtained for compounds with dimethyl, monomethyl or unsubstituted p-amino groups. An isopropyl group appears to substitute for the dimethylamino group but with considerably lower potency. Compounds with 2-substitution showed selectivity for aminergic neurones and this effect decreased with increased size of the substituent. The 2,6-dichloro derivative FLA 365 had, however, no neuronal selective action but was a potent MAO inhibitor. Substitutions in the 3- and 5-positions decreased both potency and selectivity. Prolongation of the side chain with one methylene group abolished the preference for the MAO in 5-hydroxytryptaminergic neurones although the MAO inhibitory potency remained. The selectivity disappeared by increasing the alpha-substituent to an ethyl group but remained for the alpha,alpha-dimethyl substituted derivatives. It is concluded that compounds which are (1) transported by the 5-HT pump and (2) potent reversible MAO-A inhibitors produce pronounced inhibition of MAO in 5-hydroxytryptaminergic neurones.     

Synthesis,biological evaluation,and molecular modeling of nitrile-containing compounds: Exploring multiple activities as anti-Alzheimer agents

Based on the monoamine oxidase (MAO) inhibition properties of aminoheterocycles with a carbonitrile group we have carried out a systematic exploration to discover new classes of carbonitriles endowed with dual MAO and AChE inhibitory activities, and Aβ anti-aggregating properties. Eighty-three nitrile-containing compounds, 13 of which are new, were synthesized and evaluated. in vitro screening revealed that 31 , a new compound, presented the best lead for trifunctional inhibition against MAO A (0.34 μM), MAO B (0.26 μM), and AChE (52 μM), while 32 exhibited a lead for selective MAO A (0.12 μM) inhibition coupled to AChE (48 μM) inhibition. Computational analysis revealed that the malononitrile group can find an advantageous position with the aromatic cleft and FAD of MAO A or MAO B. However, the total binding energy can be handicapped by an internal penalty caused by twisting of the ligand molecule and subsequent disruption of the conjugation ( 32 in MAO B compared to the conjugated 31 ). Conjugation is also important for AChE as well as the hydrophilic character of malononitrile that allows this group to be in close contact with the aqueous environment as seen for 83 . Although the effect of 31 and 32 against Aβ_1–42, was very weak, the effect of 63 and 65 , and of the new compound 75 , indicated that these compounds were able to disaggregate Aβ_1–42 fibrils. The most effective was 63 , a (phenylhydrazinylidene)propanedinitrile derivative that also inhibited MAO A (1.65 μM), making it a potential lead for Alzheimer's disease application.     

Novel dual inhibitors of AChE and MAO derived from hydroxy aminoindan and phenethylamine as potential treatment for Alzheimer's disease

Carbamate derivatives of N-propargylaminoindans (Series I) and N-propargylphenethylamines (Series II) were synthesized via multistep procedures from the corresponding hydroxy precursors. The respective rasagiline- and selegiline-related series were designed to combine inhibitory activities of both acetylcholine esterase (AChE) and monoamine oxidase (MAO) by virtue of their carbamoyl and propargylamine pharmacophores. Each compound was tested for these activities in vitro in order to find molecules with similar potencies against each enzyme. Compounds with such dual AChE and MAO inhibitory activities are expected to have potential for the treatment of Alzheimer's disease. The observed SAR also offers insight into the requirements of the active sites on these enzymes. A carbamate moiety was found to be essential for AChE inhibition, which was absent in the corresponding hydroxy precursors. The propargyl group caused 2-70-fold decrease in AChE inhibitory activity (depending on the position of the carbamoyl group) of Series I, but had little or no effect in Series II. Thus, the 6- and 7-carbamyloxyphenyls in Series I were either equipotent to, or slightly (2- to 5-fold) less active as AChE inhibitors than, the corresponding compounds in Series II, while the 4-carbamyloxyphenyls were more potent. The presence of the carbamate moiety in 6- and 7-carbamyloxyphenyls of Series I, considerably decreased MAO-A and -B inhibitory activity, compared to that of the parent hydroxy analogues, while the opposite was true for Series II. Thus, the 6- and 7-carbamyloxyphenyls in Series I were 2-3 orders of magnitude weaker MAO inhibitors while the 4- carbamyloxyphenyls were equipotent with the corresponding compounds in Series II. In both series, N-methylation of the propargylamine enhanced the MAO (A and B equally) inhibitory activities and decreased the AChE inhibitory activity. Two candidates belonging to the indan and tetralin ring systems (24c, 27b) and one phenethylamine (53d) were identified as possible leads for further development based on the following criteria: (a) comparable AChE and MAO-B inhibitory activities, (b) good to moderate AChE inhibitory activity, and (c) lack of strong MAO-A selectivity. However, it is likely that these compounds will be metabolized to the corresponding phenols, with inhibitory activities against AChE and/or MAO-A or -B, different from those of the parent carbamates. Thus, the apparent enzyme inhibition will be a result of the combined inhibition of all of these individual metabolites. The results of our ongoing in vivo screening programs will be published elsewhere.     

Characterization of eight biogenic indoleamines as substrates for type A and type B monoamine oxidase

Tryptamine, N-methyltryptamine, N,N-dimethyltryptamine, 5-hydroxytryptamine (5-HT), 5-hydroxy-N-methyltryptamine, bufotenine, 5-methoxytryptamine, and 5-methoxy-N,N-dimethyltryptamine were characterized as substrates for type A and type B monoamine oxidase (MAO) in rat liver mitochondria. Experiments on sensitivity to clorgyline and to deprenyl, using two substrate concentrations, showed that tryptamine and its N-methylated and N,N-dimethylated derivatives were common substrates for both types of MAO at a substrate concentration of 20.0 μM; at 1000 μM, tryptamine and N-methyltryptamine were common, but N,N-dimethyltryptamine became specific for type B MAO. All the 5-hydroxy- or 5-methoxy-indole derivatives were almost completely specific for type A MAO at a substrate concentration of 20.0 μM; when the concentration was 1000 μM, some of the MAO activity was due to type B MAO for 5-HT, bufotenine and 5-methoxytryptamine. The rat liver mitochondrial enzyme was pretreated with 10^?7M clorgyline and 10^?7M deprenyl to obtain, respectively, the type B-rich and the type A-rich enzyme. These enzyme preparations were subjected to kinetic analyses for the eight amines. From the kinetic analyses, together with data on inhibitor sensitivity, the following phenomena can be described. N-Methylation of tryptamine or of 5-HT did not cause appreciable changes in the specificity of the substrates toward each type of MAO, but it elevated the K_mvalue of type B MAO when the values for tryptamine and N-methyltryptamine were compared. N,N-Dimethylation of tryptamine and 5-HT tended to increase the specificity for type B MAO. All the dimethylated compounds had very low activities with either type A or type B MAO. Either the 5-hydroxy- or the 5-methoxy-group contributed to the specificity of the substrates for type A MAO.     

Irreversible Inhibition of Rat Liver Mitochondrial MAO A and MAO B by Enantiomers of Deprenyl and α-Methylpargyline

Using rat liver mitochondrial monoamine oxidase (MAO) A and MAO B, the possible influence of stereochemical factors upon the irreversible inhibition by propargylamine derivatives has been studied using the enantiomers of deprenyl and of α-methylpargyline. Whether studying the inhibition of MAO A or MAO B, little difference was found among enantiomeric pairs in the first-order rate constant (k₂) for formation of the enzyme inhibitor adduct. Similarly, and with the exception of (S)-d -(+)-deprenyl (k₂ = 0 or an extremely low value at MAO A), the computed value of k₂ for the individual enantiomers showed little variation between MAO A and MAO B. These results suggest that inhibitor selectivity towards a particular form of the enzyme is determined predominantly at the competitive phase of the inhibition.     

L-Deprenyl as an inhibitor of menadione-induced permeability transition in liver mitochondria

L-Deprenyl, an inhibitor of mitochondrial monoamine oxidase B (MAO B), inhibits the swelling of liver mitochondria induced by the pro-oxidant 2-methyl-1,4-naphtoquinone with a K(i) dependent on quinone concentration. L-Deprenyl also inhibits the collapse of membrane potential, cation efflux, pyridine nucleotide oxidation and cytochrome c release, all events which accompany the osmotic change and are typical of membrane permeability transition induction, thus emphasizing the inhibitory effect of the drug on this phenomenon. Results show that this inhibition is not due to the effect of L-deprenyl on monoamine oxidase activity but is most likely due to a direct interaction of the drug with the pore forming structures. It is here proposed that L-deprenyl, being a propargylamine, at physiological pH has a protonated amino group able to interact with critical aromatic or anionic amino acidic residues. As a consequence, the opening of the transition pore is prevented. These results indicate a more generalized protective effect of L-deprenyl on mitochondrial functions, involving the inhibition of membrane permeability transition induced not only by the oxidation of substrates of MAO B, but also by pro-oxidant agents such as 2-methyl-1,4-naphtoquinone, which does not involve MAO B activity.     

Orientation of oxazolidinones in the active site of monoamine oxidase

Oxazolidinone inhibitors of monoamine oxidase (MAO) and oxazolidinone antibacterials are two distinct classes of drug, often with linear structures and overlapping activities for some derivatives. By synthesizing novel dimerised derivatives with identical substitution of the two C-5 side chains, we have obtained experimental evidence for the orientation of oxazolidinones in the active site of MAO A. Two types of spectral changes, either increasing the absorbance at 510 nm or decreasing it at 495 nm depending on the group nearest to the flavin cofactor, were seen on ligand binding to MAO A. Side chain derivatives with amine substituents are very poor substrates so that it was possible to examine the spectral change due to binding of a substrate before reduction of the flavin occurred. Binding of these amino derivative substrates to MAO A induced a spectral change characterized by a strong decrease in absorbance at 495 nm. These substrates reduced the enzyme fully without any trace of a semiquinone intermediate. Only oxazolidinone inhibitors with a bromo-imidazole substituent increased the yield of semiquinone intermediate obtained during chemical reduction. In accord with the experimental data, results of docking experiments showed that binding of the oxazolidinone ring in the aromatic cage close to the flavin was favored and that the nitrogen of the derivatives that were substrates was within van der Waals distance of N-5 of the flavin.     

Observations on the inhibition of rat liver monoamine oxidase by clorgyline

Two forms of monoamine oxidase (MAO) denned as MAO A and B by others differ in their specificities to substrates and their sensitivities to the irreversible inhibitor clorgyline. From studies using the substrates 5-HT, tyramine and benzylamine, the presence of both MAO forms in rat liver mitochondria has been confirmed and some characteristics of their inhibition by varying concentrations of clorgyline investigated. Although both MAO forms showed time-dependent inhibition, this process occurred, in general, at a qualitatively slower rate for MAO B, despite the fact that this enzyme form requires higher concentrations of clorgyline than MAO A for inhibition of its activity. However, factors such as the concentration of enzyme, the concentration of clorgyline and the enzyme: drug ratio employed in the assay all influence the resultant time-course and the final degree of the inhibition observed. The possible importance of the lipid environment of the outer mitochondrial membrane in generating multiple MAO forms and in regulating the inhibition kinetics of these forms is discussed. The results indicate that the effects of pre-incubation time and the enzyme: drug ratio on inhibition of MAO by clorgyline should be fully recognized when using the drug to indicate multiple forms in animal tissues.     

Synthesis and monoamine oxidase inhibitory activity of new pyridazine-, pyrimidine- and 1,2,4-triazine-containing tricyclic derivatives

A number of condensed azines, mostly belonging to the families of indeno-fused pyridazines (1), pyrimidines (2, 3), and 1,2,4-triazines (4, 5), were synthesized and evaluated in vitro as monoamine oxidase (MAO) A and B inhibitors. Most of them showed higher inhibition potency toward MAO-B, the most effective one being 3-(3-nitrophenyl)-9H-indeno[1,2-e] [1,2,4]triazin-9-one (4c), which displayed an IC50 value of 80 nM and proved to be 10-fold more potent than its [2,1-e] fusion isomer 5. Replacing the 3-phenyl group of the known indeno[1,2-c]pyridazin-5-one MAO-B inhibitors with a flexible phenoxymethyl group enhanced the inhibitory potency. The inhibition data highlighted the importance of the aza-heterocyclic scaffold in affecting the MAO isoform selectivity. The 3-phenyl derivatives with type 1, 4, and 5 scaffolds were inhibitors of MAO-B with little or no MAO-A effect, whereas 2- or 3-phenyl derivatives of type 2 and 3 pyrimidine-containing fusion isomers inhibited both isoenzymes with a structure-dependent preference toward MAO-A.