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  • About
  • The Global ETD Search service is a free service for researchers to find electronic theses and dissertations. This service is provided by the Networked Digital Library of Theses and Dissertations.
    Our metadata is collected from universities around the world. If you manage a university/consortium/country archive and want to be added, details can be found on the NDLTD website.
1

Novel sulfanyl- and sulfinylcaffeine analogues as inhibitors of monoamine oxidase / Wayne Mentz

Mentz, Wayne January 2013 (has links)
Parkinson’s disease (PD) is a neurodegenerative disorder, which is progressive in nature and usually associated with the elderly. It is the second most common age-related neurodegenerative disorder after Alzheimer’s disease (AD). PD occurs when there is a dramatic loss of dopamine (DA) in the striatum, a substructure of the basal ganglia, of the brain due to the degeneration of the nigrostriatal pathway that contains the dopaminergic neurons. Motor symptoms of PD include bradykinesia, muscular rigidity and resting tremors. Non-motor symptoms include speech and sleep problems, hallucinations and depression. Diverse treatment options are available to treat the symptoms of PD, including levodopa (L-Dopa), DA agonists and monoamine oxidase B (MAO-B) inhibitors. The MAOs are flavoproteins that are bound to the outer membrane of the mitochondria and catalyze the oxidative deamination of neurotransmitters such as serotonin (5-HT), noradrenaline (NA) and DA. Two isoforms occur, namely MAO-A and –B, which share a 70% sequence identity. MAO-A catalyzes the oxidation of 5-HT and MAO-B has a substrate specificity towards benzylamine and 2-phenylethylamine. DA, NA, adrenaline and tryptamine are oxidized by both forms. MAO-A plays an important role in depression while MAO-B plays an important role in PD. The two isoforms are not evenly distributed in the brain. Of particular relevance to PD is the observation that, in the basal ganglia, MAO-B is the predominant form and the oxidation of DA in this region is largely due to MAO-B activity. Also, with an increase in age, there is an up to fourfold increase in MAO-B activity in the brain. In the aged parkinsonian brain, MAO-B is therefore a major DA metabolizing enzyme and MAO-B inhibitors have an important role in the therapy of PD. MAO-B inhibitors may potentially reduce the metabolic destruction of DA and thereby provide relief from the symptoms of PD. MAO-B inhibitors may also exert a neuroprotective effect in PD. In the catalytic cycle of MAO-B, one mole each of an aldehyde, hydrogen peroxide and ammonia are formed for each mole of primary amine substrate oxidized. Ferrous iron, which is abundant in the basal ganglia, may react with the hydrogen peroxide to form hydroxyl radicals in the Fenton reaction. The hydroxyl radical damages virtually all types of biomolecules including proteins, DNA, lipids, carbohydrates and amino acids. The aldehyde, in turn, may react with amino groups of proteins, and thus lead to cell injury. Inhibitors of MAO-B may reduce the MAO-catalyzed formation of hydrogen peroxide and aldehydes in the basal ganglia, and thus act as neuroprotective agents. MAO-B inhibitors that are currently being used in the treatment of PD are selegiline and rasagiline. Both are irreversible inhibitors of MAO-B. While irreversible inhibitors of MAO have been used extensively as drugs, irreversible inhibition has a number of disadvantages. These include the loss of selectivity as a result of repeated drug administration and a slow and variable rate of enzyme recovery following termination of drug treatment. The turnover rate for the biosynthesis of MAO-B in the human brain may require as much as 40 days while with reversible inhibition, enzyme activity is recovered when the inhibitor is eliminated from the tissues. For these reasons the discovery of novel MAO-B inhibitors, which interact reversibly with the enzymes are of value in the therapy of PD. The goal of this study was to design novel and reversible inhibitors of MAO-B, which may find application in the therapy of PD. In the current study, caffeine was used as scaffold for the design of new MAO inhibitors. Caffeine is reported to be a weak inhibitor of MAO-B, with an IC50 value of 5084 μM. Substitution at C-8 of the caffeine moiety, however, yields compounds with potent MAO-B inhibitory properties. Of particular importance to this study is a recent report that a series of 8-sulfanylcaffeine analogues acts as selective inhibitors of human MAO-B. Among the compounds examined, 8-[(phenylethyl)sulfanyl]caffeine was found to be a particularly potent MAO-B inhibitor with an IC50 value of 0.223 μM. In an attempt to further enhance the MAO-B inhibition potency of 8-[(phenylethyl)sulfanyl]caffeine, and possibly to discover highly potent MAO-B inhibitors, a series of five 8-[(phenylethyl)sulfanyl]caffeine analogues was synthesized and evaluated as inhibitors of human MAO-A and –B. For the purpose of this study 8- [(phenylethyl)sulfanyl]caffeine homologues containing C-3 alkyl (CF3, CH3 and OCH3) and halogen (Cl and Br) substituents on the phenyl ring were considered. Furthermore, a series of two 8-sulfinylcaffeine analogues and one 8-sulfonylcaffeine were synthesized and their MAO inhibitory potencies were measured. The purpose with these compounds was to compare the MAO inhibitory properties of the 8-sulfinylcaffeine analogues and 8-sulfonylcaffeine with those of the 8-sulfanylcaffeine analogues. This study also investigates the MAO inhibition properties of three selected 8-[(phenylpropyl)sulfanyl]caffeine and two 8-(benzylsulfanyl)caffeine analogues. Chemistry: The target 8-sulfanylcaffeine analogues were synthesized according to the literature procedure. 8-Chlorocaffeine was reacted with an appropriate mercaptan in the presence of NaOH, to yield the target 8-sulfanylcaffeine analogues in yields of 6.4–50.7%. 8-Chlorocaffeine, in turn, was conveniently synthesized in high yield by reacting chlorine with caffeine in chloroform. In certain instances, the mercaptan starting materials were not commercially available and were thus synthesized according to the literature procedure by reacting an appropriate alkylbromide with thiourea. The resulting thiouronium salt was hydrolyzed in the presence of NaOH to yield the target mercaptan. The 8-sulfinylcaffeine analogues and 8- sulfonylcaffeine were synthesized by reacting the 8-sulfanylcaffeines with H2O2 in the presence of glacial acetic acid and acetic anhydride. The structures and the purities of the inhibitors were verified by NMR, MS and HPLC analyses. MAO inhibition studies: The MAO inhibitory properties of the test compounds were examined using the recombinant human enzymes. The mixed MAO-A/B substrate, kynuramine, was employed as substrate for both enzymes and the inhibition potencies were expressed as the IC50 values. The 8-[(phenylethyl)sulfanyl]caffeine analogues were found to be highly potent inhibitors of MAO-B. The IC50 values recorded for these homologues ranged from 0.017–0.125 μM, making them twofold to 13-fold more potent MAO-B inhibitors than the lead compound, 8- [(phenylethyl)sulfanyl]caffeine (IC50 = 0.223 μM). For comparison, the reversible MAO-B selective inhibitor, lazabemide, exhibits an IC50 value of 0.091 μM under the same conditions (unpublished data from our laboratory). Interestingly, both alkyl (CF3, CH3 and OCH3) and halogen (Cl and Br) substitution lead to highly potent MAO-B inhibition. It may therefore be concluded that substitution on C-3 is a general strategy to enhance the MAO-B inhibition potency of 8-[(phenylethyl)sulfanyl]caffeine. The results of the MAO inhibitory studies with the 8- [(phenylpropyl)sulfanyl]caffeine analogues showed that these compounds are also inhibitors of MAO-B with IC50 values of 0.061–0.500 μM. Those homologues substituted with chlorine on the para and meta positions of the phenyl ring were found to be exceptionally potent inhibitors with IC50 values of 0.061 μM and 0.062 μM, respectively. For the series of 8- (benzylsulfanyl)caffeines, meta substitution with chlorine (IC50 = 0.227 μM) and bromine (IC50 = 0.199 μM) was also found to enhance the MAO-B inhibition potency of 8- (benzylsulfanyl)caffeine (IC50 = 1.86 μM). The results document that the 8-sulfinylcaffeines are also inhibitors of MAO-B with IC50 values of 11.8–131 μM. The 8-sulfonylcaffeine was also found to be a MAO-B inhibitor. Compared to the 8-sulfanylcaffeines, these homologues are, however, weaker inhibitors. It may, therefore, be concluded that 8-sulfinylcaffeines and 8-sulfonylcaffeines are comparatively weak MAO-B inhibitors and less suited for the design of high potency MAO-B inhibitors. The results also document that the 8-[(phenylethyl)sulfanyl]caffeines are relatively weak MAO-A inhibitors with IC50 values of 5.66–141 μM, with one homologue exhibiting no inhibition under the experimental conditions. As evident from the selectivity indices (SI values), the 8- [(phenylethyl)sulfanyl]caffeines were all selective inhibitors of the MAO-B isoform. Two compounds exhibited SI values in excess of 1000. Since these compounds are also highly potent MAO-B inhibitors, they represent suitable leads for the design of potent and selective MAO-B inhibitors. The 8-sulfinylcaffeines and 8-sulfonylcaffeine were found to be weak MAO-A inhibitors with IC50 values of 166–250 μM. The SI values demonstrate that these compounds are MAO-B selective inhibitors, although to a lesser degree than the 8- [(phenylethyl)sulfanyl]caffeines. The 8-[(phenylpropyl)sulfanyl]caffeines are also MAO-A inhibitors with IC50 values of 0.708–6.48 μM. It is noteworthy that these homologues are the most potent MAO-A inhibitors among the compounds evaluated in this study. In fact, one of the 8-[(phenylpropyl)sulfanyl]caffeines, 8-{[3-(4-chlorophenyl)propyl]sulfanyl}caffeine (IC50 = 0.708 μM), is the only compound with an IC50 value for the inhibition of MAO-A in the submicromolar range. The 8-[(phenylpropyl)sulfanyl]caffeines display, in general, lower degrees of selectivity for MAO-B than the corresponding 8-[(phenylethyl)sulfanyl]caffeines. Reversibility studies: The reversibility of the interaction of a representative inhibitor, 8-{[2-(3- (trifluoromethyl)phenyl)ethyl]sulfanyl}caffeine, with MAO-B was investigated by evaluating the recovery of the enzymatic activity after dilution of the enzyme-inhibitor complex. For this purpose, MAO-B was preincubated with the test compound at concentrations of 10 × IC50 and 100 × IC50 for 30 min. The reactions were subsequently diluted 100-fold to 0.1 × IC50 and 1 × IC50, respectively. The results show that, after dilution to 0.1 × IC50 and 1 × IC50, the MAO-B catalytic activities are recovered to 35% and 22%, respectively, of the control value. For reversible enzyme inhibition, the enzyme activities are expected to recover to levels of approximately 90% and 50%, respectively, after 100-fold dilution of the preincubations containing inhibitor concentrations of 10 × IC50 and 100 × IC50. After preincubation of MAO-B with the irreversible inhibitor (R)-deprenyl (at 10 × IC50), and dilution of the resulting complex to 0.1 × IC50, MAO-B activity is not recovered (3.0% of control). These data indicate that the test compound does indeed react reversibly with MAO-B but because enzyme activities are not recovered to the expected 90% and 50% respectively, it may suggest that the test compound possess a quasi-reversible or tight-binding component. Hansch-type structure activity relationship studies: A limited Hansch-type QSAR study was performed for the inhibition of MAO by the 8-[(phenylethyl)sulfanyl]caffeines. For this purpose, five parameters were used to describe the physicochemical properties of the C-3 substituents on the phenyl rings of the inhibitors. The Van der Waals volume (Vw) and Taft steric parameter (Es) served as descriptors of the bulkiness of the substituents, while the lipophilicities were described by the Hansch constant (π). The electronic properties were described by the classical Hammett constant (σm) and the Swain-Lupton constant (F). A one-parameter fit with the Taft steric parameter versus the inhibition potency (logIC50) yielded the best correlation with a correlation coefficient (R2) of 0.912 and a statistical F value of 41.27 (Fmax = 35). The positive sign of the Es (+0.47) parameter coefficient indicated that the inhibition potencies of the 8- [(phenylethyl)sulfanyl]caffeines towards MAO-B may be enhanced by substitution with sterically large groups at C-3 of the phenyl rings of the inhibitors. / Thesis (MSc (Pharmaceutical Chemistry))--North-West University, Potchefstroom Campus, 2013
2

Novel sulfanyl- and sulfinylcaffeine analogues as inhibitors of monoamine oxidase / Wayne Mentz

Mentz, Wayne January 2013 (has links)
Parkinson’s disease (PD) is a neurodegenerative disorder, which is progressive in nature and usually associated with the elderly. It is the second most common age-related neurodegenerative disorder after Alzheimer’s disease (AD). PD occurs when there is a dramatic loss of dopamine (DA) in the striatum, a substructure of the basal ganglia, of the brain due to the degeneration of the nigrostriatal pathway that contains the dopaminergic neurons. Motor symptoms of PD include bradykinesia, muscular rigidity and resting tremors. Non-motor symptoms include speech and sleep problems, hallucinations and depression. Diverse treatment options are available to treat the symptoms of PD, including levodopa (L-Dopa), DA agonists and monoamine oxidase B (MAO-B) inhibitors. The MAOs are flavoproteins that are bound to the outer membrane of the mitochondria and catalyze the oxidative deamination of neurotransmitters such as serotonin (5-HT), noradrenaline (NA) and DA. Two isoforms occur, namely MAO-A and –B, which share a 70% sequence identity. MAO-A catalyzes the oxidation of 5-HT and MAO-B has a substrate specificity towards benzylamine and 2-phenylethylamine. DA, NA, adrenaline and tryptamine are oxidized by both forms. MAO-A plays an important role in depression while MAO-B plays an important role in PD. The two isoforms are not evenly distributed in the brain. Of particular relevance to PD is the observation that, in the basal ganglia, MAO-B is the predominant form and the oxidation of DA in this region is largely due to MAO-B activity. Also, with an increase in age, there is an up to fourfold increase in MAO-B activity in the brain. In the aged parkinsonian brain, MAO-B is therefore a major DA metabolizing enzyme and MAO-B inhibitors have an important role in the therapy of PD. MAO-B inhibitors may potentially reduce the metabolic destruction of DA and thereby provide relief from the symptoms of PD. MAO-B inhibitors may also exert a neuroprotective effect in PD. In the catalytic cycle of MAO-B, one mole each of an aldehyde, hydrogen peroxide and ammonia are formed for each mole of primary amine substrate oxidized. Ferrous iron, which is abundant in the basal ganglia, may react with the hydrogen peroxide to form hydroxyl radicals in the Fenton reaction. The hydroxyl radical damages virtually all types of biomolecules including proteins, DNA, lipids, carbohydrates and amino acids. The aldehyde, in turn, may react with amino groups of proteins, and thus lead to cell injury. Inhibitors of MAO-B may reduce the MAO-catalyzed formation of hydrogen peroxide and aldehydes in the basal ganglia, and thus act as neuroprotective agents. MAO-B inhibitors that are currently being used in the treatment of PD are selegiline and rasagiline. Both are irreversible inhibitors of MAO-B. While irreversible inhibitors of MAO have been used extensively as drugs, irreversible inhibition has a number of disadvantages. These include the loss of selectivity as a result of repeated drug administration and a slow and variable rate of enzyme recovery following termination of drug treatment. The turnover rate for the biosynthesis of MAO-B in the human brain may require as much as 40 days while with reversible inhibition, enzyme activity is recovered when the inhibitor is eliminated from the tissues. For these reasons the discovery of novel MAO-B inhibitors, which interact reversibly with the enzymes are of value in the therapy of PD. The goal of this study was to design novel and reversible inhibitors of MAO-B, which may find application in the therapy of PD. In the current study, caffeine was used as scaffold for the design of new MAO inhibitors. Caffeine is reported to be a weak inhibitor of MAO-B, with an IC50 value of 5084 μM. Substitution at C-8 of the caffeine moiety, however, yields compounds with potent MAO-B inhibitory properties. Of particular importance to this study is a recent report that a series of 8-sulfanylcaffeine analogues acts as selective inhibitors of human MAO-B. Among the compounds examined, 8-[(phenylethyl)sulfanyl]caffeine was found to be a particularly potent MAO-B inhibitor with an IC50 value of 0.223 μM. In an attempt to further enhance the MAO-B inhibition potency of 8-[(phenylethyl)sulfanyl]caffeine, and possibly to discover highly potent MAO-B inhibitors, a series of five 8-[(phenylethyl)sulfanyl]caffeine analogues was synthesized and evaluated as inhibitors of human MAO-A and –B. For the purpose of this study 8- [(phenylethyl)sulfanyl]caffeine homologues containing C-3 alkyl (CF3, CH3 and OCH3) and halogen (Cl and Br) substituents on the phenyl ring were considered. Furthermore, a series of two 8-sulfinylcaffeine analogues and one 8-sulfonylcaffeine were synthesized and their MAO inhibitory potencies were measured. The purpose with these compounds was to compare the MAO inhibitory properties of the 8-sulfinylcaffeine analogues and 8-sulfonylcaffeine with those of the 8-sulfanylcaffeine analogues. This study also investigates the MAO inhibition properties of three selected 8-[(phenylpropyl)sulfanyl]caffeine and two 8-(benzylsulfanyl)caffeine analogues. Chemistry: The target 8-sulfanylcaffeine analogues were synthesized according to the literature procedure. 8-Chlorocaffeine was reacted with an appropriate mercaptan in the presence of NaOH, to yield the target 8-sulfanylcaffeine analogues in yields of 6.4–50.7%. 8-Chlorocaffeine, in turn, was conveniently synthesized in high yield by reacting chlorine with caffeine in chloroform. In certain instances, the mercaptan starting materials were not commercially available and were thus synthesized according to the literature procedure by reacting an appropriate alkylbromide with thiourea. The resulting thiouronium salt was hydrolyzed in the presence of NaOH to yield the target mercaptan. The 8-sulfinylcaffeine analogues and 8- sulfonylcaffeine were synthesized by reacting the 8-sulfanylcaffeines with H2O2 in the presence of glacial acetic acid and acetic anhydride. The structures and the purities of the inhibitors were verified by NMR, MS and HPLC analyses. MAO inhibition studies: The MAO inhibitory properties of the test compounds were examined using the recombinant human enzymes. The mixed MAO-A/B substrate, kynuramine, was employed as substrate for both enzymes and the inhibition potencies were expressed as the IC50 values. The 8-[(phenylethyl)sulfanyl]caffeine analogues were found to be highly potent inhibitors of MAO-B. The IC50 values recorded for these homologues ranged from 0.017–0.125 μM, making them twofold to 13-fold more potent MAO-B inhibitors than the lead compound, 8- [(phenylethyl)sulfanyl]caffeine (IC50 = 0.223 μM). For comparison, the reversible MAO-B selective inhibitor, lazabemide, exhibits an IC50 value of 0.091 μM under the same conditions (unpublished data from our laboratory). Interestingly, both alkyl (CF3, CH3 and OCH3) and halogen (Cl and Br) substitution lead to highly potent MAO-B inhibition. It may therefore be concluded that substitution on C-3 is a general strategy to enhance the MAO-B inhibition potency of 8-[(phenylethyl)sulfanyl]caffeine. The results of the MAO inhibitory studies with the 8- [(phenylpropyl)sulfanyl]caffeine analogues showed that these compounds are also inhibitors of MAO-B with IC50 values of 0.061–0.500 μM. Those homologues substituted with chlorine on the para and meta positions of the phenyl ring were found to be exceptionally potent inhibitors with IC50 values of 0.061 μM and 0.062 μM, respectively. For the series of 8- (benzylsulfanyl)caffeines, meta substitution with chlorine (IC50 = 0.227 μM) and bromine (IC50 = 0.199 μM) was also found to enhance the MAO-B inhibition potency of 8- (benzylsulfanyl)caffeine (IC50 = 1.86 μM). The results document that the 8-sulfinylcaffeines are also inhibitors of MAO-B with IC50 values of 11.8–131 μM. The 8-sulfonylcaffeine was also found to be a MAO-B inhibitor. Compared to the 8-sulfanylcaffeines, these homologues are, however, weaker inhibitors. It may, therefore, be concluded that 8-sulfinylcaffeines and 8-sulfonylcaffeines are comparatively weak MAO-B inhibitors and less suited for the design of high potency MAO-B inhibitors. The results also document that the 8-[(phenylethyl)sulfanyl]caffeines are relatively weak MAO-A inhibitors with IC50 values of 5.66–141 μM, with one homologue exhibiting no inhibition under the experimental conditions. As evident from the selectivity indices (SI values), the 8- [(phenylethyl)sulfanyl]caffeines were all selective inhibitors of the MAO-B isoform. Two compounds exhibited SI values in excess of 1000. Since these compounds are also highly potent MAO-B inhibitors, they represent suitable leads for the design of potent and selective MAO-B inhibitors. The 8-sulfinylcaffeines and 8-sulfonylcaffeine were found to be weak MAO-A inhibitors with IC50 values of 166–250 μM. The SI values demonstrate that these compounds are MAO-B selective inhibitors, although to a lesser degree than the 8- [(phenylethyl)sulfanyl]caffeines. The 8-[(phenylpropyl)sulfanyl]caffeines are also MAO-A inhibitors with IC50 values of 0.708–6.48 μM. It is noteworthy that these homologues are the most potent MAO-A inhibitors among the compounds evaluated in this study. In fact, one of the 8-[(phenylpropyl)sulfanyl]caffeines, 8-{[3-(4-chlorophenyl)propyl]sulfanyl}caffeine (IC50 = 0.708 μM), is the only compound with an IC50 value for the inhibition of MAO-A in the submicromolar range. The 8-[(phenylpropyl)sulfanyl]caffeines display, in general, lower degrees of selectivity for MAO-B than the corresponding 8-[(phenylethyl)sulfanyl]caffeines. Reversibility studies: The reversibility of the interaction of a representative inhibitor, 8-{[2-(3- (trifluoromethyl)phenyl)ethyl]sulfanyl}caffeine, with MAO-B was investigated by evaluating the recovery of the enzymatic activity after dilution of the enzyme-inhibitor complex. For this purpose, MAO-B was preincubated with the test compound at concentrations of 10 × IC50 and 100 × IC50 for 30 min. The reactions were subsequently diluted 100-fold to 0.1 × IC50 and 1 × IC50, respectively. The results show that, after dilution to 0.1 × IC50 and 1 × IC50, the MAO-B catalytic activities are recovered to 35% and 22%, respectively, of the control value. For reversible enzyme inhibition, the enzyme activities are expected to recover to levels of approximately 90% and 50%, respectively, after 100-fold dilution of the preincubations containing inhibitor concentrations of 10 × IC50 and 100 × IC50. After preincubation of MAO-B with the irreversible inhibitor (R)-deprenyl (at 10 × IC50), and dilution of the resulting complex to 0.1 × IC50, MAO-B activity is not recovered (3.0% of control). These data indicate that the test compound does indeed react reversibly with MAO-B but because enzyme activities are not recovered to the expected 90% and 50% respectively, it may suggest that the test compound possess a quasi-reversible or tight-binding component. Hansch-type structure activity relationship studies: A limited Hansch-type QSAR study was performed for the inhibition of MAO by the 8-[(phenylethyl)sulfanyl]caffeines. For this purpose, five parameters were used to describe the physicochemical properties of the C-3 substituents on the phenyl rings of the inhibitors. The Van der Waals volume (Vw) and Taft steric parameter (Es) served as descriptors of the bulkiness of the substituents, while the lipophilicities were described by the Hansch constant (π). The electronic properties were described by the classical Hammett constant (σm) and the Swain-Lupton constant (F). A one-parameter fit with the Taft steric parameter versus the inhibition potency (logIC50) yielded the best correlation with a correlation coefficient (R2) of 0.912 and a statistical F value of 41.27 (Fmax = 35). The positive sign of the Es (+0.47) parameter coefficient indicated that the inhibition potencies of the 8- [(phenylethyl)sulfanyl]caffeines towards MAO-B may be enhanced by substitution with sterically large groups at C-3 of the phenyl rings of the inhibitors. / Thesis (MSc (Pharmaceutical Chemistry))--North-West University, Potchefstroom Campus, 2013

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