Tissue transglutaminase (TG2) is a calcium-dependent enzyme that natively catalyses the formation of isopeptidic bonds between protein- or peptide-bound glutamine and lysine residues. Physiologically, it is ubiquitously expressed in tissues, with roles in cellular differentiation, extracellular matrix stabilisation, and apoptosis, among others. However, its unregulated activity has been associated with various pathologies including fibrosis, cancer and celiac disease. Since most pathologies are associated with an increased transamidation activity, efforts have been directed towards the development of TG2 inhibitors. In this context, the work described in this thesis is centred on reversible inhibitors, building on recent work done within the Keillor group in two directions, namely localisation and potency. In a localisation-driven approach, we developed a photolabile derivative of a known reversible inhibitor, in order to form a covalent bond with the enzyme and determine the inhibitor’s binding site. In tandem, we optimised a protocol for the expression of TG2 incorporating ArgΔ10 and LysΔ8, amino acids that are 13C- and 15N-labelled to provide a mass shift of 10 and 8 Da, respectively, compared to the corresponding unlabelled amino acids. This “heavy” TG2 was developed as a tool for reference in the analysis of the tryptic digest of labelled protein. In a potency-driven approach, based on the observation that previous trans cinnamoyl inhibitor scaffolds were susceptible to nucleophilic attack by glutathione, we developed a bis(triazole) scaffold with reduced electrophilicity. The preparation of a small library of compounds showed that this scaffold demonstrates a preference for electron-withdrawing substituents, such as nitro groups. Continuing in a potency-driven approach, and inspired by work done in the identification of glutathione-resistant scaffolds, we studied a new alkynyl scaffold. While still susceptible to glutathione addition, these compounds showed a marked improvement in potency, with the lead compound having an IC50 of 930 nM and being established as a competitive inhibitor with a Ki of 420 nM, our most potent reversible inhibitor to date. Furthermore, this scaffold also produced an inhibitor lacking nitro groups (to limit eventual cellular toxicity), but maintaining good potency, with an IC50 value of 3.03 μM.

The shortcomings of current antidepressant agents prompts the design of novel multimodal antidepressants and the identification of new antidepressant targets, especially those located at sub-cellular level. Such antidepressants should possess improved response rates as well as safety profiles. Methylene blue (MB) is reported to possess diverse pharmacological actions and is attracting increasing attention for the treatment of a variety of disorders including Alzheimer’s disease, bipolar disorder, anxiety and depression. MB acts on both monoamine oxidase (MAO) and the nitric oxide (NO)-cGMP pathway, and possesses antidepressant activity in rodents. The principal goal of this study was to design a close structural analogue of MB and to evaluate the effects of these structural changes on MAO inhibition, a well-known antidepressant target. Furthermore, MAO inhibition is also responsible for cardiovascular toxicity in clinically used MAOI inhibitors. For this purpose we investigated the antidepressant properties of the synthetic MB analogue (ethyl-thioniniumchloride; ETC) as well as azure B, the major metabolite of MB, in the forced swim test (FST). ETC was synthesized with a high degree of purity from diethyl-p-phenylenediamine with 6% yield. ETC was firstly evaluated as a potential inhibitor of recombinant human MAO-A and MAO-B. Azure B and ETC were evaluated over a dosage range of 4-30 mg/kg for antidepressant-like activity in the acute FST in rats, and the results were compared to those obtained with saline, imipramine (15 mg/kg) and MB (15 mg/kg) treated rats. Locomotor activity was evaluated to ensure that changes in swim motivation are based on antidepressant response and not due to an indirect effect of the drug on locomotor activity. The results document that ETC inhibits MAO-A and MAO-B with IC50 values of 0.51 μM and 0.592 μM, respectively. Furthermore, ETC inhibits MAO-A and MAO-B reversibly, while the mode of inhibition is most likely competitive. In the acute FST, azure B and ETC were more effective than imipramine and MB in reversing immobility, without inducing locomotor effects. Azure B and ETC increased swimming behaviour during acute treatment, which is indicative of enhanced serotonergic neurotransmission. Azure B and ETC did not affect noradrenergicmediated climbing behaviour. These results suggest that azure B may be a contributor to the antidepressant effect of MB, and acts via increasing serotonergic transmission. Secondly, small structural changes made to MB do not abolish its antidepressant effect even though ETC is a less potent MAO-A inhibitor than MB. / MSc (Pharmaceutical Chemistry), North-West University, Potchefstroom Campus, 2014

The shortcomings of current antidepressant agents prompts the design of novel multimodal antidepressants and the identification of new antidepressant targets, especially those located at sub-cellular level. Such antidepressants should possess improved response rates as well as safety profiles. Methylene blue (MB) is reported to possess diverse pharmacological actions and is attracting increasing attention for the treatment of a variety of disorders including Alzheimer’s disease, bipolar disorder, anxiety and depression. MB acts on both monoamine oxidase (MAO) and the nitric oxide (NO)-cGMP pathway, and possesses antidepressant activity in rodents. The principal goal of this study was to design a close structural analogue of MB and to evaluate the effects of these structural changes on MAO inhibition, a well-known antidepressant target. Furthermore, MAO inhibition is also responsible for cardiovascular toxicity in clinically used MAOI inhibitors. For this purpose we investigated the antidepressant properties of the synthetic MB analogue (ethyl-thioniniumchloride; ETC) as well as azure B, the major metabolite of MB, in the forced swim test (FST). ETC was synthesized with a high degree of purity from diethyl-p-phenylenediamine with 6% yield. ETC was firstly evaluated as a potential inhibitor of recombinant human MAO-A and MAO-B. Azure B and ETC were evaluated over a dosage range of 4-30 mg/kg for antidepressant-like activity in the acute FST in rats, and the results were compared to those obtained with saline, imipramine (15 mg/kg) and MB (15 mg/kg) treated rats. Locomotor activity was evaluated to ensure that changes in swim motivation are based on antidepressant response and not due to an indirect effect of the drug on locomotor activity. The results document that ETC inhibits MAO-A and MAO-B with IC50 values of 0.51 μM and 0.592 μM, respectively. Furthermore, ETC inhibits MAO-A and MAO-B reversibly, while the mode of inhibition is most likely competitive. In the acute FST, azure B and ETC were more effective than imipramine and MB in reversing immobility, without inducing locomotor effects. Azure B and ETC increased swimming behaviour during acute treatment, which is indicative of enhanced serotonergic neurotransmission. Azure B and ETC did not affect noradrenergicmediated climbing behaviour. These results suggest that azure B may be a contributor to the antidepressant effect of MB, and acts via increasing serotonergic transmission. Secondly, small structural changes made to MB do not abolish its antidepressant effect even though ETC is a less potent MAO-A inhibitor than MB. / MSc (Pharmaceutical Chemistry), North-West University, Potchefstroom Campus, 2014

Monoamine oxidase (MAO) is responsible for dopamine catabolism in the brain and therefore is especially important in the treatment of Parkinson's disease (PD). MAO–B inhibition provides symptomatic relief by indirectly elevating dopamine levels in the PD brain. PD is caused by the loss of dopaminergic neurons in the substantia nigra and the formation of proteinaceous structures in the brain. The cause of idiopathic PD is unknown, but one theory states that reactive oxygen species (ROS), partly derived from the catalytic cycle of MAO, may be to blame for damaging dopaminergic neurons. Since MAO inhibitors may reduce the MAO–catalyzed production of ROS, these compounds may protect dopaminergic neurons against degeneration in PD. It is commonly accepted that by the time PD symptoms manifest, about 80% of striatal dopamine has been lost. MAO is present as two subtypes in the human brain, namely MAO–A and MAO–B. MAOs are found mainly attached to the mitochondrial membrane and is responsible for the oxidative deamination of various monoamines, including dopamine. MAO is a dimeric enzyme which operates in conjunction with a co–factor, flavin adenine dinucleotide (FAD), to which it is covalently bound. The flavin is in a bent conformation, which assists the catalytic activity of MAO. As mentioned above, the catalytic action of MAO also produces harmful substances such as hydrogen peroxide, ammonia, aldehydes and may also increase the levels of hydroxyl radicals. In the healthy brain, these substances are metabolized rapidly, but the PD brain may exhibit reduced clearance of these species. Thus the inhibition of MAOs may be beneficial to the PD sufferer as it indirectly increases dopamine levels in the brain and may also slow the formation of harmful substances. MAO inhibitors, of the MAO–A type, were first used as anti–depressants. It was these drugs that first prompted researchers to explore MAO inhibitors as novel anti–parkinsonian drugs, as MAO–A inhibition slows the degradation of dopamine. Two types of inhibition modes exist, irreversible and reversible inhibition. Irreversible inhibitors do not allow for competition with the substrate and inactivate the enzyme permanently. Selegiline, a propargyl amine derivative, is an example of an irreversible MAO–B selective inhibitor. The major disadvantage of irreversible inhibitors is that after terminating treatment, recovery of the enzyme activity may require several weeks, since the turnover rate for the biosynthesis of MAO in the human brain may be as much as 40 days. Reversible inhibitors have better safety profiles since they allow for competition with the substrate. (E)–8–(3–Chlorostyryl)caffeine (CSC) is an example of a reversible inhibitor of MAO–B and is also an antagonist of the adenosine A2A receptor. Since antagonism of A2A receptors also produces an antiparkinsonian effect, dual acting compounds such as CSC, which block both the A2A receptors and MAO–B, may have an enhanced therapeutic potential in PD therapy. Current PD therapy available only treats the symptoms of PD and do not halt or slow the progression of the neurodegenerative processes. There therefore exists the need for the development of antiparkinsonian drugs with neuroprotective effects. Since both MAO–B inhibitors and A2A receptor antagonists are reported to possess protective effects in PD and PD animal models, dual acting drugs, that antagonize A2A receptors and inhibit MAO–B, may be candidates for neuroprotection. Using the structure of CSC as lead, we investigate in the current study, the possibility that aminocaffeines may also possess potent MAO–B inhibitory properties. The structures of the aminocaffeine derivatives that were investigated bear close structural resemblance to CSC as well as to a series of alkyloxycaffeine analogues that was recently found to be potent MAO inhibitors. This study therefore further explores the structural requirements of caffeine derivatives to act as MAO inhibitors by examining the possibility that aminocaffeine derivatives may be MAO inhibitors. Such compounds may act as lead compounds for the development of improved PD therapy. In this study, a series of 8–aminocaffeine derivatives were synthesized and evaluated as inhibitors of human MAO–A and B. For this purpose, 8–chlorocaffeine was reacted with the appropriate amine at high temperatures to produce the desired 8–aminocaffeine derivatives. The inhibitory activities of the compounds were determined towards recombinant human MAO–A and B and expressed as IC50 values. The results showed that human MAO–B was most potently inhibited by 8–[methyl(4–phenylbutyl)amino]caffeine with an IC50 value of 2.97 ?M. Human MAO–A was most potently inhibited by 8–[2–(3–chlorophenyl)–ethylamino]caffeine with an IC50 value of 5.78 ?M. It was found that methylation of the amine group at C8 of the caffeine ring increases inhibition but also selectivity towards MAO–B inhibition. For example, 8–[4–(phenylbutylamino)]caffeine inhibits MAO–B with an IC50 value of 7.56 ?M whereas 8–[methyl(4–phenylbutyl)amino]–caffeine has an increased inhibition potency of 2.97 ?M. The selectivity for MAO–B inhibition also increases over MAO–A when the C8 amine is methylated. It was found that the aminocaffeine derivatives bind reversibly to both enzyme isoforms and the mode of inhibition is competitive for MAO–B. From these results it can be concluded that although the 8–aminocaffeine derivatives are only moderately potent MAO–B inhibitors, they may act as lead compounds for the design of more potent reversible MAO inhibitors. Docking studies revealed that the 8–aminocaffeine and 8–[(methyl)amino]caffeine derivatives traverse both the entrance and substrate cavities of the MAO–B enzyme, with the caffeinyl moiety oriented towards the FAD co–factor while the amino–side chain protrudes into the entrance cavity. / Thesis (M.Sc. (Pharmaceutical Chemistry))--North-West University, Potchefstroom Campus, 2012.

Monoamine oxidase (MAO) is responsible for dopamine catabolism in the brain and therefore is especially important in the treatment of Parkinson's disease (PD). MAO–B inhibition provides symptomatic relief by indirectly elevating dopamine levels in the PD brain. PD is caused by the loss of dopaminergic neurons in the substantia nigra and the formation of proteinaceous structures in the brain. The cause of idiopathic PD is unknown, but one theory states that reactive oxygen species (ROS), partly derived from the catalytic cycle of MAO, may be to blame for damaging dopaminergic neurons. Since MAO inhibitors may reduce the MAO–catalyzed production of ROS, these compounds may protect dopaminergic neurons against degeneration in PD. It is commonly accepted that by the time PD symptoms manifest, about 80% of striatal dopamine has been lost. MAO is present as two subtypes in the human brain, namely MAO–A and MAO–B. MAOs are found mainly attached to the mitochondrial membrane and is responsible for the oxidative deamination of various monoamines, including dopamine. MAO is a dimeric enzyme which operates in conjunction with a co–factor, flavin adenine dinucleotide (FAD), to which it is covalently bound. The flavin is in a bent conformation, which assists the catalytic activity of MAO. As mentioned above, the catalytic action of MAO also produces harmful substances such as hydrogen peroxide, ammonia, aldehydes and may also increase the levels of hydroxyl radicals. In the healthy brain, these substances are metabolized rapidly, but the PD brain may exhibit reduced clearance of these species. Thus the inhibition of MAOs may be beneficial to the PD sufferer as it indirectly increases dopamine levels in the brain and may also slow the formation of harmful substances. MAO inhibitors, of the MAO–A type, were first used as anti–depressants. It was these drugs that first prompted researchers to explore MAO inhibitors as novel anti–parkinsonian drugs, as MAO–A inhibition slows the degradation of dopamine. Two types of inhibition modes exist, irreversible and reversible inhibition. Irreversible inhibitors do not allow for competition with the substrate and inactivate the enzyme permanently. Selegiline, a propargyl amine derivative, is an example of an irreversible MAO–B selective inhibitor. The major disadvantage of irreversible inhibitors is that after terminating treatment, recovery of the enzyme activity may require several weeks, since the turnover rate for the biosynthesis of MAO in the human brain may be as much as 40 days. Reversible inhibitors have better safety profiles since they allow for competition with the substrate. (E)–8–(3–Chlorostyryl)caffeine (CSC) is an example of a reversible inhibitor of MAO–B and is also an antagonist of the adenosine A2A receptor. Since antagonism of A2A receptors also produces an antiparkinsonian effect, dual acting compounds such as CSC, which block both the A2A receptors and MAO–B, may have an enhanced therapeutic potential in PD therapy. Current PD therapy available only treats the symptoms of PD and do not halt or slow the progression of the neurodegenerative processes. There therefore exists the need for the development of antiparkinsonian drugs with neuroprotective effects. Since both MAO–B inhibitors and A2A receptor antagonists are reported to possess protective effects in PD and PD animal models, dual acting drugs, that antagonize A2A receptors and inhibit MAO–B, may be candidates for neuroprotection. Using the structure of CSC as lead, we investigate in the current study, the possibility that aminocaffeines may also possess potent MAO–B inhibitory properties. The structures of the aminocaffeine derivatives that were investigated bear close structural resemblance to CSC as well as to a series of alkyloxycaffeine analogues that was recently found to be potent MAO inhibitors. This study therefore further explores the structural requirements of caffeine derivatives to act as MAO inhibitors by examining the possibility that aminocaffeine derivatives may be MAO inhibitors. Such compounds may act as lead compounds for the development of improved PD therapy. In this study, a series of 8–aminocaffeine derivatives were synthesized and evaluated as inhibitors of human MAO–A and B. For this purpose, 8–chlorocaffeine was reacted with the appropriate amine at high temperatures to produce the desired 8–aminocaffeine derivatives. The inhibitory activities of the compounds were determined towards recombinant human MAO–A and B and expressed as IC50 values. The results showed that human MAO–B was most potently inhibited by 8–[methyl(4–phenylbutyl)amino]caffeine with an IC50 value of 2.97 ?M. Human MAO–A was most potently inhibited by 8–[2–(3–chlorophenyl)–ethylamino]caffeine with an IC50 value of 5.78 ?M. It was found that methylation of the amine group at C8 of the caffeine ring increases inhibition but also selectivity towards MAO–B inhibition. For example, 8–[4–(phenylbutylamino)]caffeine inhibits MAO–B with an IC50 value of 7.56 ?M whereas 8–[methyl(4–phenylbutyl)amino]–caffeine has an increased inhibition potency of 2.97 ?M. The selectivity for MAO–B inhibition also increases over MAO–A when the C8 amine is methylated. It was found that the aminocaffeine derivatives bind reversibly to both enzyme isoforms and the mode of inhibition is competitive for MAO–B. From these results it can be concluded that although the 8–aminocaffeine derivatives are only moderately potent MAO–B inhibitors, they may act as lead compounds for the design of more potent reversible MAO inhibitors. Docking studies revealed that the 8–aminocaffeine and 8–[(methyl)amino]caffeine derivatives traverse both the entrance and substrate cavities of the MAO–B enzyme, with the caffeinyl moiety oriented towards the FAD co–factor while the amino–side chain protrudes into the entrance cavity. / Thesis (M.Sc. (Pharmaceutical Chemistry))--North-West University, Potchefstroom Campus, 2012.

Parkinson’s disease (PD) is an age-related neurodegenerative disorder. The degeneration of the neurons of the substantia nigra in the midbrain leads to the loss of dopamine from the striatum, which is responsible for the motor symptoms of PD. In the brain, the enzyme, monoamine oxidase B (MAOB), An analysis of the Lineweaver-Burk plots indicated that 7-(3-bromobenzyloxy)-3,4-dihydro-2(1H)- quinolinone inhibits MAO-B with a Ki value of 2.7 nM. An analysis of the structure-activity relationships for MAO-B inhibition shows that substitution on the C7 position of the 3,4-dihydro- 2(1H)-quinolinone moiety leads to significantly more potent inhibition compared to substitution on C6. In this regard, a benzyloxy substituent on C7 is more favourable than phenylethoxy and phenylpropoxy substitution on this position. In spite of this, C6-substituted 3,4-dihydro-2(1H)-quinolinone with potent MAO-B inhibitory activities were also identified. An analyses of selected properties of the 3,4-dihydro-2(1H)- quinolinones showed that the compounds are highly lipophilic with logP values in the range of 3.03- 4.55. LogP values between 1 and 3 are, however, in the ideal range for bioavailability. The compounds synthesised have logP values higher than 3, which may lead to lower bioavailability. Laboratory data further showed that none of the 3,4-dihydro-2(1H)-quinolinones are highly toxic to cultured cells at the concentrations, 1 μM and 10 μM, tested. For example, the most potent MAO-B inhibitor, 7-(3-bromobenzyloxy)-3,4-dihydro-2(1H)-quinolinone, reduced cell viability to 88.11% and 86.10% at concentrations of 1 μM and 10 μM, respectively. These concentrations are well above its IC50 value for the inhibition of MAO-B. At concentrations required for MAO-B inhibition, the more potent 3,4-dihydro-2(1H)-quinolinones are thus unlikely to be cytotoxic. It may thus be concluded that C7-substituted 3,4-dihydro-2(1H)-quinolinones are promising highly potent and selective MAO-B inhibitors, and thus leads for the therapy of Parkinson’s disease. represents a major catabolic pathway of dopamine. Inhibitors of MAO-B conserve the depleted supply of dopamine and are thus used in the therapy of PD. In the present study, a series of 3,4- dihydro-2(1H)-quinolinone derivatives were synthesized and evaluated as inhibitors of recombinant human MAO-A and MAO-B. These quinolinone derivatives are structurally related to a series of coumarin (1-benzopyran-2-one) derivatives, which has been reported to act as MAO-B inhibitors. C6- and C7-substituted 3,4-dihydro-2(1H)-quinolinone derivatives were synthesized by reacting 6- or 7- hydroxy-3,4-dihydro-2(1H)-quinolinone with an appropriately substituted alkyl bromide in the presence of base. To evaluate the MAO inhibitory properties (IC50 values) of the quinolinone derivatives the recombinant human MAO-A and MAO-B enzymes were used. The reversibility of inhibition of a representative 3,4-dihydro-2(1H)-quinolinone derivative was examined by measuring the recovery of enzyme activity after the dilution of the enzyme-inhibitor complexes, while the mode of MAO inhibition was determined by constructing Lineweaver-Burk plots. To determine the lipophilicity of the 3,4-dihydro-2(1H)-quinolinone derivatives, the logP values were measured. The toxicity of the 3,4-dihydro-2(1H)-quinolinone derivatives towards cultured cells (cytotoxicity) was also measured. The results document that the 3,4-dihydro-2(1H)-quinolinone derivatives are highly potent and selective MAO-B inhibitors with most homologues exhibiting IC50 values in the nanomolar range. The most potent MAO-B inhibitor, 7-(3-bromobenzyloxy)-3,4-dihydro-2(1H)-quinolinone, exhibits an IC50 value of 2.9 nM with a 2750-fold selectivity for MAO-B over the MAO-A isoform. As a MAO-B inhibitor, this compound is approximately equipotent to the most potent coumarin derivative (IC50 = 1.14 nM) reported in literature. Since MAO-B activity could be recovered after dilution of enzyme-inhibitor mixtures, it may be concluded that 7-(3-bromobenzyloxy)-3,4-dihydro-2(1H)- quinolinone is a reversible MAO-B inhibitor. The Lineweaver-Burk plots constructed for the inhibition of MAO-B by 7-(3-bromobenzyloxy)-3,4-dihydro-2(1H)-quinolinone were linear and intersected on the y-axis. These data indicated that this compound also is a competitive MAO-B inhibitor. / MSc (Pharmaceutical Chemistry), North-West University, Potchefstroom Campus, 2014

Parkinson’s disease (PD) is an age-related neurodegenerative disorder. The degeneration of the neurons of the substantia nigra in the midbrain leads to the loss of dopamine from the striatum, which is responsible for the motor symptoms of PD. In the brain, the enzyme, monoamine oxidase B (MAOB), An analysis of the Lineweaver-Burk plots indicated that 7-(3-bromobenzyloxy)-3,4-dihydro-2(1H)- quinolinone inhibits MAO-B with a Ki value of 2.7 nM. An analysis of the structure-activity relationships for MAO-B inhibition shows that substitution on the C7 position of the 3,4-dihydro- 2(1H)-quinolinone moiety leads to significantly more potent inhibition compared to substitution on C6. In this regard, a benzyloxy substituent on C7 is more favourable than phenylethoxy and phenylpropoxy substitution on this position. In spite of this, C6-substituted 3,4-dihydro-2(1H)-quinolinone with potent MAO-B inhibitory activities were also identified. An analyses of selected properties of the 3,4-dihydro-2(1H)- quinolinones showed that the compounds are highly lipophilic with logP values in the range of 3.03- 4.55. LogP values between 1 and 3 are, however, in the ideal range for bioavailability. The compounds synthesised have logP values higher than 3, which may lead to lower bioavailability. Laboratory data further showed that none of the 3,4-dihydro-2(1H)-quinolinones are highly toxic to cultured cells at the concentrations, 1 μM and 10 μM, tested. For example, the most potent MAO-B inhibitor, 7-(3-bromobenzyloxy)-3,4-dihydro-2(1H)-quinolinone, reduced cell viability to 88.11% and 86.10% at concentrations of 1 μM and 10 μM, respectively. These concentrations are well above its IC50 value for the inhibition of MAO-B. At concentrations required for MAO-B inhibition, the more potent 3,4-dihydro-2(1H)-quinolinones are thus unlikely to be cytotoxic. It may thus be concluded that C7-substituted 3,4-dihydro-2(1H)-quinolinones are promising highly potent and selective MAO-B inhibitors, and thus leads for the therapy of Parkinson’s disease. represents a major catabolic pathway of dopamine. Inhibitors of MAO-B conserve the depleted supply of dopamine and are thus used in the therapy of PD. In the present study, a series of 3,4- dihydro-2(1H)-quinolinone derivatives were synthesized and evaluated as inhibitors of recombinant human MAO-A and MAO-B. These quinolinone derivatives are structurally related to a series of coumarin (1-benzopyran-2-one) derivatives, which has been reported to act as MAO-B inhibitors. C6- and C7-substituted 3,4-dihydro-2(1H)-quinolinone derivatives were synthesized by reacting 6- or 7- hydroxy-3,4-dihydro-2(1H)-quinolinone with an appropriately substituted alkyl bromide in the presence of base. To evaluate the MAO inhibitory properties (IC50 values) of the quinolinone derivatives the recombinant human MAO-A and MAO-B enzymes were used. The reversibility of inhibition of a representative 3,4-dihydro-2(1H)-quinolinone derivative was examined by measuring the recovery of enzyme activity after the dilution of the enzyme-inhibitor complexes, while the mode of MAO inhibition was determined by constructing Lineweaver-Burk plots. To determine the lipophilicity of the 3,4-dihydro-2(1H)-quinolinone derivatives, the logP values were measured. The toxicity of the 3,4-dihydro-2(1H)-quinolinone derivatives towards cultured cells (cytotoxicity) was also measured. The results document that the 3,4-dihydro-2(1H)-quinolinone derivatives are highly potent and selective MAO-B inhibitors with most homologues exhibiting IC50 values in the nanomolar range. The most potent MAO-B inhibitor, 7-(3-bromobenzyloxy)-3,4-dihydro-2(1H)-quinolinone, exhibits an IC50 value of 2.9 nM with a 2750-fold selectivity for MAO-B over the MAO-A isoform. As a MAO-B inhibitor, this compound is approximately equipotent to the most potent coumarin derivative (IC50 = 1.14 nM) reported in literature. Since MAO-B activity could be recovered after dilution of enzyme-inhibitor mixtures, it may be concluded that 7-(3-bromobenzyloxy)-3,4-dihydro-2(1H)- quinolinone is a reversible MAO-B inhibitor. The Lineweaver-Burk plots constructed for the inhibition of MAO-B by 7-(3-bromobenzyloxy)-3,4-dihydro-2(1H)-quinolinone were linear and intersected on the y-axis. These data indicated that this compound also is a competitive MAO-B inhibitor. / MSc (Pharmaceutical Chemistry), North-West University, Potchefstroom Campus, 2014