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We performed a microdialysis study to examine the effects of local perfusion of COA‑Cl on the extracellular levels of dopamine (DA) and its metabolites in the dorsal striatum of mice in vivo. The mice were perfused with Ringer's solution (control) and COA‑Cl (0.05, 0.1, or 0.5 mM) into the dorsal striatum. Dialysate samples were collected every 30 min and then analyzed using high‑performance liquid chromatography coupled with an electrochemical detector. We found that local perfusion of COA‑Cl (0.1 or 0.5 mM) into the dorsal striatum of living mice produced a significant and dose‑dependent increase in extracellular levels of DA, 3‑methoxytyramine (3‑MT), and homovanillic acid (HVA), where only 0.5 mM COA‑Cl increased dihydroxyphenylacetic acid (DOPAC) levels. However, 0.05 mM of COA‑Cl did not significantly affect either DA levels or its metabolites. Then, we administered the monoamine oxidase (MAO) inhibitor clorgyline alone or in combination with COA‑Cl (0.1 mM) to test whether COA‑Cl‑induced increases in DOPAC and HVA are mediated by increased MAO activity. Clorgyline alone increased 3‑MT levels and decreased DOPAC and HVA levels but not DA levels. When combined with COA‑Cl, clorgyline increased 3‑MT levels and reversed the decrease in DOPAC and HVA levels caused by clorgyline. The increase in DA metabolism induced by COA‑Cl suggests that some DA was further metabolized into DOPAC, 3‑MT, and HVA. This indicates that COA‑Cl plays a role in DA metabolism via increased DA release and/or activation of MAO, offering new insights into the effects of COA‑Cl on DA metabolism in the brain.

Introduction: To investigate the inhibition properties and structure-activity relationship between monoamine oxidase (MAO) and selected monoamine oxidase inhibitors (MAOIs, including selegiline, rasagiline and clorgiline)., Methods: The inhibition effect and molecular mechanism between MAO and MAOIs were identified via the half maximal inhibitory concentration (IC 50 ) and molecular docking technology., Results: It was indicated that selegiline and rasagiline were MAO B inhibitors, but clorgiline was MAO-A inhibitor based on the selectivity index (SI) of MAOIs (0.000264, 0.0197 and 14607.143 for selegiline, rasagiline and clorgiline, respectively). The high-frequency amino acid residues of the MAOIs and MAO were Ser24, Arg51, Tyr69 and Tyr407 for MAO-A and Arg42 and Tyr435 for MAO B. The MAOIs and MAO A/B pharmacophores included the aromatic core, hydrogen bond acceptor, hydrogen bond donor-acceptor and hydrophobic core., Conclusion: This study shows the inhibition effect and molecular mechanism between MAO and MAOIs and provides valuable findings on the design and treatment of Alzheimer's and Parkinson's diseases., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Monoamine oxidases (MAOA/MAOB) are enzymes known for their role in neurotransmitter regulation in the central nervous system (CNS). Irreversible and non-selective MAO inhibitors (MAOi's) were the first class of antidepressants, thus subsequent work on drugs such as the selective MAOA inhibitor clorgyline has focussed on selectivity and increased CNS penetration. MAOA is highly expressed in high grade and metastatic prostate cancer with a proposed effect on prostate cancer growth, recurrence, and drug resistance. A Phase II Clinical Trial has demonstrated the therapeutic effects of the irreversible nonselective MAOi phenelzine for prostate cancer. However, neurologic adverse effects led to early withdrawal in 25% of the enrolled patient-population. In this work, we revised the clorgyline scaffold with the goal of decreasing CNS penetration to minimize CNS-related side effects while retaining or enhancing MAOA inhibition potency and selectivity. Using the known co-crystal structure of clorgyline bound with FAD co-factor in the hMAOA active site as a reference, we designed and synthesized a series of compounds predicted to have lower CNS penetration (logBB). All synthesized derivatives displayed favorable drug-like characteristics such as predicted Caco-2 permeability and human oral absorption, and exhibited highly selective hMAOA binding interactions. Introduction of an HBD group (NH 2 or OH) at position 5 of the phenyl ring clorgyline resulted in 3x more potent hMAOA inhibition with equivalent or better hMAOB selectivity, and similar prostate cancer cell cytotoxicity. In contrast, introduction of larger substituents at this position or at the terminal amine significantly reduced the hMAOA inhibition potency, attributed in part to a steric clash within the binding pocket of the MAOA active site. Replacement of the N-methyl group by a more polar, but larger 2-hydroxyethyl group did not enhance potency. However, introduction of a polar 2-hydroxy in the propyl chain retained the highly selective MAOA inhibition and cancer cell cytotoxicity of clorgyline while reducing its CNS score from 2 to 0. We believe that these results identify a new class of peripherally directed MAOIs that may allow safer therapeutic targeting of MAOA for a variety of anti-cancer and anti-inflammatory indications., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Monoamine oxidase A (MAO A) and heat shock protein 90 (HSP90) inhibitors have been shown to decrease the progression of glioblastoma (GBM) and other cancers. In this study, a series of MAO A/HSP90 dual inhibitors were designed and synthesized in the hope to develop more effective treatment of GBM. Compounds 4-b and 4-c are conjugates of isopropylresorcinol (pharmacophore of HSP90 inhibitor) with the phenyl group of clorgyline (MAO A inhibitor) by a tertiary amide bond substituted with methyl (4-b) or ethyl (4-c) group, respectively. They inhibited MAO A activity, HSP90 binding, and the growth of both TMZ-sensitive and -resistant GBM cells. Western blots showed that they increased HSP70 expression indicating reduced function of HSP90, reduced HER2 and phospho-Akt expression similar to MAO A or HSP90 inhibitor itself. Both compounds decreased IFN-γ induced PD-L1 expression in GL26 cells, suggesting they can act as immune checkpoint inhibitor. Further, they reduced tumor growth in GL26 mouse model. NCI-60 analysis showed they also inhibited the growth of colon cancer, leukemia, non-small cell lung and other cancers. Taken together, this study demonstrates MAO A/HSP90 dual inhibitors 4-b and 4-c reduced the growth of GBM and other cancers, and they have potential to inhibit tumor immune escape., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Masson SAS. All rights reserved.)

Monoamine oxidase (MAO) is an enzyme located on the outer mitochondrial membrane that metabolizes amine substrates like serotonin, norepinephrine and dopamine. MAO inhibitors (MAOIs) are frequently utilized to treat disorders such as major depression or Parkinson's disease (PD), though their effects on brain mitochondrial bioenergetics are unclear. These studies measured bioenergetic activity in mitochondria isolated from the mouse cortex in the presence of inhibitors of either MAO-A, MAO-B, or both isoforms. We found that only 10 μM clorgyline, the selective inhibitor of MAO-A and not MAO-B, increased mitochondrial oxygen consumption rate in State V(CI) respiration compared to vehicle treatment. We then assessed mitochondrial bioenergetics, reactive oxygen species (ROS) production, and Electron Transport Chain (ETC) complex function in the presence of 0, 5, 10, 20, 40, or 80 μM of clorgyline to determine if this change was dose-dependent. The results showed increased oxygen consumption rates across the majority of respiration states in mitochondria treated with 5, 10, or 20 μM with significant bioenergetic inhibition at 80 μM clorgyline. Next, we assessed mitochondrial ROS production in the presence of the same concentrations of clorgyline in two different states: high mitochondrial membrane potential (ΔΨ m ) induced by oligomycin and low ΔΨ m induced by carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone (FCCP). There were no changes in ROS production in the presence of 5, 10, 20, or 40 μM clorgyline compared to vehicle after the addition of oligomycin or FCCP. There was a significant increase in mitochondrial ROS in the presence of 80 μM clorgyline after FCCP addition, as well as reduced Complex I and Complex II activities, which are consistent with inhibition of bioenergetics seen at this dose. There were no changes in Complex I, II, or IV activities in mitochondria treated with low doses of clorgyline. These studies shed light on the direct effect of MAO-A inhibition on brain mitochondrial bioenergetic function, which may be a beneficial outcome for those taking these medications., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interest., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Aim: Previously, we revealed a crucial role of 5-HT degradation system (5DS), consisting of 5-HT2A receptor (5-HT2AR), 5-HT synthases and monoamine oxidase A (MAO-A), in ischemia-reperfusion (IR)-caused organ injury. Whereas, platelet activating factor receptor (PAFR) also mediates myocardial ischemia-reperfusion injury (MIRI). Here, we try to clarify the relationship between 5DS and PAFR in mediating MIRI., Methods: H9c2 cell injury and rat MIRI were caused by hypoxia/reoxygenation (H/R) or PAF, and by ligating the left anterior descending coronary artery then untying, respectively. 5-HT 2A R and PAFR antagonists [sarpogrelate hydrochloride (SH) and BN52021], MAO-A, AKT, mTOR and 5-HT synthase inhibitors (clorgyline, perifosine, rapamycin and carbidopa), and gene-silencing PKCε were used in experiments RESULTS: The mitochondrial ROS production, respiratory chain damage, inflammation, apoptosis and myocardial infarction were significantly prevented by BN52021, SH and clorgyline in H/R and PAF-treated cells and in IR myocardium. BN52021 also significantly suppressed the upregulation of PAFR, 5-HT 2A R, 5-HT synthases and MAO-A expression (mRNA and protein), and Gα q and PKCε (in plasmalemma) expression induced by H/R, PAF or IR; the effects of SH were similar to that of BN52021 except for no affecting the expression of PAFR and 5-HT 2A R. Gene-silencing PKCε suppressed H/R and PAF-induced upregulation of 5-HT synthases and MAO-A expression in cells; perifosine and rapamycin had not such effects; however, clorgyline suppressed H/R and PAF-induced phosphorylation of AKT and mTOR., Conclusion: MIRI is probably due to PAFR-mediated 5-HT 2A R activation, which further activates PKCε-mediated 5-HT synthesis and degradation, leading to mitochondrial ROS production., Competing Interests: Declaration of Competing Interest The authors declare that there are no conflicts of interest., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

First, we aimed to investigate ex vivo the effects of ethanol (EtOH) on levels of norepinephrine (NE), dopamine (DA), serotonin (5-HT), and their metabolites in the frontal cortex, hippocampus, and striatum of Aldh2-knockout (Aldh2-KO) and wild-type (WT) mice. Animals were treated intraperitoneally with saline (control) or EtOH (1.0, 2.0, or 3.0 g/kg). Brain samples were collected 60 and 120 min after EtOH injection, and monoamines and their metabolites were measured by HPLC-ECD. We found in both WT and Aldh2-KO mice that 3.0 g/kg EtOH increased the levels of 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) and decreased the level of 3-methoxytyramine (3-MT). A 2.0 g/kg dose of EtOH also increased HVA, but there was not a consistent effect within the brain regions of Aldh2-KO and WT mice. There were inconsistent findings of genotype differences in the levels of DA, 5-HT, and their metabolites in the brain regions tested. None of the EtOH doses altered NE, DA, 5-HT, or 5-hydroxyindoleacetic acid contents in any of the brain regions studied. Second, we tested whether EtOH-induced increases in DOPAC and HVA are mediated by increased monoamine oxidase (MAO) or catechol-O-methyltransferase (COMT) activity. To test this, we used the MAO blocker clorgyline (2.0 and 4.0 mg/kg) and the COMT blocker tolcapone (15 and 30 mg/kg) alone or in combination with EtOH (3.0 g/kg). Clorgyline alone increased 3-MT and decreased DOPAC and HVA levels, whereas tolcapone alone increased DOPAC and decreased 3-MT and HVA levels. Surprisingly, the combination of EtOH with clorgyline (4.0 mg/kg) or tolcapone (30 mg/kg) further decreased 3-MT and increased DOPAC and HVA levels, an effect that reversed the inhibitor-induced decreases in HVA. These results suggest that a high concentration of EtOH can accelerate DA metabolism, as evidenced by the increase in DOPAC and HVA, and this effect is likely a consequence of increased degradation of DA by MAO., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Monoamine oxidase (MAO) is believed to mediate the degradation of monoamine neurotransmitters, including dopamine, in the brain. Between the two types of MAO, MAO-B has been believed to be involved in dopamine degradation, which supports the idea that the therapeutic efficacy of MAO-B inhibitors in Parkinson's disease can be attributed to an increase in extracellular dopamine concentration. However, this belief has been controversial. Here, by utilizing in vivo phasic and basal electrochemical monitoring of extracellular dopamine with fast-scan cyclic voltammetry and multiple-cyclic square wave voltammetry and ex vivo fluorescence imaging of dopamine with GRAB DA2m , we demonstrate that MAO-A, but not MAO-B, mainly contributes to striatal dopamine degradation. In contrast, our whole-cell patch-clamp results demonstrated that MAO-B, but not MAO-A, was responsible for astrocytic GABA-mediated tonic inhibitory currents in the rat striatum. We conclude that, in contrast to the traditional belief, MAO-A and MAO-B have profoundly different roles: MAO-A regulates dopamine levels, whereas MAO-B controls tonic GABA levels., (© 2021. The Author(s).)

Monoamine oxidases (MAOs), a class of enzymes bound to the outer mitochondrial membrane, are important sources of reactive oxygen species. Increased MAO-A activity in endothelial cells and cardiomyocytes contributes to vascular dysfunction and progression of left heart failure. We hypothesized that inhibition of MAO-A can be used to treat pulmonary arterial hypertension (PAH) and right ventricular (RV) failure. MAO-A levels in lung and RV samples from patients with PAH were compared with levels in samples from donors without PAH. Experimental PAH was induced in male Sprague-Dawley rats by using Sugen 5416 and hypoxia (SuHx), and RV failure was induced in male Wistar rats by using pulmonary trunk banding (PTB). Animals were randomized to receive either saline or the MAO-A inhibitor clorgyline at 10 mg/kg. Echocardiography and RV catheterization were performed, and heart and lung tissues were collected for further analysis. We found increased MAO-A expression in the pulmonary vasculature of patients with PAH and in experimental experimental PAH induced by SuHx. Cardiac MAO-A expression and activity was increased in SuHx- and PTB-induced RV failure. Clorgyline treatment reduced RV afterload and pulmonary vascular remodeling in SuHx rats through reduced pulmonary vascular proliferation and oxidative stress. Moreover, clorgyline improved RV stiffness and relaxation and reversed RV hypertrophy in SuHx rats. In PTB rats, clorgyline had no direct clorgyline had no direct effect on the right ventricle effect. Our study reveals the role of MAO-A in the progression of PAH. Collectively, these findings indicated that MAO-A may be involved in pulmonary vascular remodeling and consecutive RV failure.

A series of rasagiline-clorgyline hybrids was designed, synthesized and investigated in vitro for their inhibition of monoamine oxidase and amyloid-β aggregation. Most of compounds were found to be selective and highly potent hMAO-B inhibitors showing IC 50 values in the nanomolar, and exhibited a moderate inhibition of amyloid-β aggregation. 7-((5-(methyl(prop-2-yn-1-yl)amino) pentyl)oxy)chroman-4-one (6j) was the most interesting compound identified in this research, endowed with higher hMAO-B potency (IC 50  = 4 nM) and selectivity (SI > 25000) compared to the reference selective inhibitor rasagiline (IC 50  = 141 nM, SI > 355), and exhibited good inhibitory activity against Aβ 1-42 aggregation (40.78%, 25 μM). Kinetic and molecular modeling studies revealed that 6j was a competitive reversible inhibitor for hMAO-B. Moreover, compound 6j displayed low toxicity and good neuroprotective effects in SH-SY5Y cell assay, and could penetrate the blood-brain barrier according to the parallel artificial membrane permeability assay. Pharmacokinetics assay revealed that compound 6j possessed good pharmacokinetic profiles after intravenous and oral administrations. Overall, these results highlighted that compound 6j was an effective and promising multitarget agent against Alzheimer's disease., Competing Interests: Declaration of competing interest The authors declare no competing financial interest., (Copyright © 2020 Elsevier Masson SAS. All rights reserved.)

The antiandrogen enzalutamide (Enz) has improved survival in castration resistant prostate cancer (CRPC) patients. However, most patients eventually develop Enz resistance that may involve inducing the androgen receptor (AR) splicing variant 7 (ARv7). Here we report that high expression of monoamine oxidase-A (MAO-A) is associated with positive ARv7 detection in CRPC patients following Enz treatment. Targeting MAO-A with phenelzine or clorgyline, the FDA-approved drugs for antidepression, resensitize the Enz resistant (EnzR) cells to Enz treatment and further suppress EnzR cell growth in vitro and in vivo. Our findings suggest that Enz-increased ARv7 expression can transcriptionally enhance MAO-A expression resulting in Enz resistance via altering the hypoxia HIF-1α signals. Together, our results show that targeting the Enz/ARv7/MAO-A signaling with the antidepressants phenelzine or clorgyline can restore Enz sensitivity to suppress EnzR cell growth, which may indicate that these antidepression drugs can overcome the Enz resistance to further suppress the EnzR CRPC.

The maladaptive form of aggressive behavior confers risk for violence and criminal incidences with profound impact on society. Although considerable research has been devoted to elucidate the etiology of aggression, molecular correlates of sex differences remains largely unexplored. Also, little attention has been given to whether males and females respond differently to similar causal factor of aggression. Here, we show the possible association of brain region specific neural activity (c-Fos expression) and monoamine oxidase A (MAOA) epigenetic state with sexual dimorphism in peripubertal stress (PPS) induced adulthood aggression. While PPS adult males exhibited escalated aggression, females spent maximal time in social exploration. c-Fos expression was brain region and sex specific. In the PPS adult cohort, only males showed elevated c-Fos expression in the prefrontal cortex, indicative of their hyper-responsive behavior. MAOA expression and enzyme activity was reduced in hypothalamus and increased in prefrontal cortex of hyper-aggressive male mice. Investigation into the underlying mechanisms revealed hypomethylation in prefrontal cortex and hypermethylation in hypothalamus of MAOA promoter negatively correlating with the expression pattern. On the other hand, binding of Sirt1 to MAOA promoter was diametrically opposite being increased in prefrontal cortex and reduced in hypothalamus. In females, neither expression nor epigenetic state of MAOA gene was significantly altered between control and PPS adult mice. Our study revealed novel epigenetic correlates of sexual dimorphism in stress induced aggressive psychopathology. However, given the multi-factorial nature with environmental influences, further studies are warranted to uncover the biological hub., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Background: Monoamine oxidase A (MAOA) is best known for its role in neuro-transmitter regulation. Monoamine oxidase inhibitors are used to treat atypical depression. MAOA is highly expressed in high grade prostate cancer and modulates tumorigenesis and progression in prostate cancer. Here, we investigated the potential role of MAOA inhibitors (MAOAIs) in relation to the androgen receptor (AR) pathway and resistance to antiandrogen treatment in prostate cancer., Methods: We examined MAOA expression and the effect of MAOI treatment in relation to AR-targeted treatments using the LNCaP, C4-2B, and 22Rv1 human prostate cancer cell lines. MAOA, AR-full length (AR-FL), AR splice variant 7 (AR-V7), and PSA expression was evaluated in the presence of MAOAIs (clorgyline, phenelzine), androgenic ligand (R1881), and antiandrogen (enzalutamide) treatments. An enzalutamide resistance cell line was generated to test the effect of MAOAI treatment in this model., Results: We observed that MAOAIs, particularly clorgyline and phenelzine, were effective at decreasing MAOA activity in human prostate cancer cells. MAOAIs significantly decreased growth of LNCaP, C4-2B, and 22Rv1 cells and produced additive growth inhibitory effects when combined with enzalutamide. Clorgyline decreased expression of AR-FL and AR-V7 in 22Rv1 cells and was effective at decreasing growth of an enzalutamide-resistant C4-2B cell line with increased AR-V7 expression., Conclusions: MAOAIs decrease growth and proliferation of androgen-sensitive and castration-resistant prostate cancer cells. Clorgyline, in particular, decreases expression of AR-FL and AR-V7 expression and decreases growth of an enzalutamide-resistant cell line. These findings provide preclinical validation of MAOA inhibitors either alone or in combination with antiandrogens for therapeutic intent in patients with advanced forms of prostate cancer., (© 2019 Wiley Periodicals, Inc.)

Inhibition of MAO-B has been an effective strategy for the treatment of Parkinson's disease. To find more potent and selective MAO-B inhibitors with novel chemical scaffold, we designed and synthesized a series of new 2,3-dihydro-1H-inden-1-amine derivatives on basis of our previous study. Furthermore, the corresponding structure-activity relationship (SAR) of these compounds is detailedly discussed. Compounds L4 (IC 50  = 0.11 μM), L8 (IC 50  = 0.18 μM), L16 (IC 50  = 0.27 μM) and L17 (IC 50  = 0.48 μM) showed similar MAO-B inhibitory activity as Selegiline. Moreover, L4, L16 and L17 also exhibited comparable selectivity with Selegiline, indicating that L4, L16 and L17 could be promising selective MAO-B inhibitors for further study., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Monoamine oxidase A (MAOA) is a mitochondrial enzyme that catalyzes oxidative deamination of neurotransmitters and dietary amines and produces H 2 O 2 . It facilitates the progression of gliomas and prostate cancer, but its expression and functional relevance have not been studied in lymphoma. Here, we evaluated MAOA in 427 cases of Hodgkin and non-Hodgkin lymphoma and in a spectrum of reactive lymphoid tissues by immunohistochemistry on formalin-fixed, paraffin-embedded specimens. MAOA was expressed by Hodgkin Reed-Sternberg (HRS) cells in the majority of classical Hodgkin lymphomas (cHLs) (181/241; 75%), with 34.8% showing strong expression. Weak MAOA was also noted in a minority of primary mediastinal large B-cell lymphomas (8/47; 17%) and in a mediastinal gray-zone lymphoma. In contrast, no MAOA was found in non-neoplastic lymphoid tissues, nodular lymphocyte-predominant Hodgkin lymphoma (NLPHL; 0/8) or any other non-Hodgkin lymphomas studied (0/123). MAOA was more common in Epstein-Barr virus (EBV)-negative compared to EBV-positive cHL (p < 0.0001) and was especially prevalent in the EBV-negative nodular sclerosing subtype. Similar to primary human lymphoma specimens, most cHL-derived cell lines displayed MAOA activity, whereas non-Hodgkin-lymphoma-derived cell lines did not. The MAOA inhibitor clorgyline reduced the growth of L1236 cells and U-HO1 cells, and shRNA knockdown of MAOA reduced the growth of L1236 cells. Conversely, ectopic overexpression of MAOA increased the growth of MAOA-negative HDLM2 cells. Combined treatment with clorgyline and ABVD (doxorubicin, bleomycin, vinblastine, dacarbazine) was more effective in reducing cell growth than either regimen alone. In summary, MAOA is highly expressed in cHL and may reflect the distinct biology of this lymphoma. Further studies on the potential utility of MAOA as a diagnostic marker and therapeutic target are warranted. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd., (Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.)

Because of the lack of significant disease-modifying drugs for neurodegenerative disorders, a pressing need for new chemical entities endowed with IMAO-B still exists. Within this framework, and for the first time, a study was performed to compare coumarin- and chomone-3-phenylcarboxamide scaffolds. Compounds 10a and 10b were the most potent, selective, and reversible noncompetitive IMAO-B. The benzopyrone sp 2 oxygen atom was found to be position independent and a productive contributor for the ligand-enzyme complex stability.

The regulation of brain monoamine levels is paramount for cognitive functions, and the monoamine oxidase (MAO A and B) enzymes play a central role in these processes. The aim of this study was to evaluate whether the procognitive properties exerted by propargylamine N-(furan-2-ylmethyl)-N-methylprop-2-yn-1-amine (F2MPA) are related to changes in monoamine content via MAO inhibition. In vivo microdialysis and ex vivo amine metabolite measurement demonstrated region-specific alterations in monoamine metabolism that differ from both of the classic MAO A and MAO B inhibitors, clorgyline and l-deprenyl, respectively. Although all the inhibitors (1 and 4 mg/kg) increased cortical serotonin tissue content, only F2MPA increased the levels of cortical noradrenaline. In the striatum, clorgyline (1 mg/kg), but not F2MPA (1 mg/kg), reduced extracellular levels of dopamine metabolites at rest or stimulated by the intrastriatal application of the MAO substrate 3-methoxytyramine. In vitro, F2MPA exhibited a low affinity toward MAO B and MAO A. Nonetheless, it modified the B form of MAO, forming a flavin adduct structurally similar to that with deprenyl. F2MPA was rapidly metabolized in the presence of rat but not human microsomes, producing a hydroxylated derivative. In conclusion, the effect of F2MPA on cognition may arise from monoaminergic changes in the cortex, but the role of MAO in this process is likely to be negligible, consistent with the poor affinity of F2MPA for MAO.

3,4-methylenedioxymethamphetamine (MDMA; ecstasy) is a commonly abused recreational drug that causes neurotoxic effects in both humans and animals. The mechanism behind MDMA-induced neurotoxicity is suggested to be species-dependent and needs to be further investigated on the cellular level. In this study, the effects of MDMA in neuronally differentiated P19 mouse embryonal carcinoma cells have been examined. MDMA produces a concentration-, time- and temperature-dependent toxicity in differentiated P19 neurons, as measured by intracellular MTT reduction and extracellular LDH activity assays. The P19-derived neurons express both the serotonin reuptake transporter (SERT), that is functionally active, and the serotonin metabolizing enzyme monoamine oxidase A (MAO-A). The involvement of these proteins in the MDMA-induced toxicity was investigated by a pharmacological approach. The MAO inhibitors clorgyline and deprenyl, and the SERT inhibitor fluoxetine, per se or in combination, were not able to mimic the toxic effects of MDMA in the P19-derived neurons or block the MDMA-induced cell toxicity. Oxidative stress has been implicated in MDMA-induced neurotoxicity, but pre-treatment with the antioxidants α-tocopherol or N-acetylcysteine did not reveal any protective effects in the P19 neurons. Involvement of mitochondria in the MDMA-induced cytotoxicity was also examined, but MDMA did not alter the mitochondrial membrane potential (ΔΨm) in the P19 neurons. We conclude that MDMA produce a concentration-, time- and temperature-dependent neurotoxicity and our results suggest that the mechanism behind MDMA-induced toxicity in mouse-derived neurons do not involve the serotonergic system, oxidative stress or mitochondrial dysfunction., Competing Interests: The authors have declared that no competing interests exist.

Rationale: Fas-associated death domain (FADD) is an adaptor of death receptors that can also induce anti-apoptotic actions through its phosphorylated form (p-FADD). Activation of monoamine receptors, indirect targets of classic anti-depressant drugs (ADs), reduced FADD and increased p-FADD and p-FADD/FADD ratio in brain., Objectives: To ascertain whether ADs, which indirectly regulate monoamine receptors, modulate FADD protein forms to promote anti-apoptotic actions., Methods: The effects of selected norepinephrine transporter (NET), serotonin transporter (SERT), monoamine oxidase (MAO) inhibitors, atypical ADs, and electroconvulsive shock (ECS) or behavioral procedures (forced swim test, FST) on FADD forms and pro-survival FADD-like interleukin-1β-converting enzyme-inhibitory protein (FLIP-L) and phosphoprotein enriched in astrocytes of 15 kDa (p-PEA-15) contents were assessed in rat brain cortex by western blot analysis., Results: Acute NET (e.g., nisoxetine) but not SERT (e.g., fluoxetine) inhibitors decreased cortical FADD (up to 37 %) and increased p-FADD/FADD ratio (up to 1.9-fold). Nisoxetine effects were prevented by α2-antagonist RX-821002, suggesting the involvement of presynaptic α2-autoreceptors. Immobility time in the FST correlated with increases of pro-apoptotic FADD and decreases of anti-apoptotic p-FADD. The MAO-A/B inhibitor phenelzine decreased FADD (up to 33 %) and increased p-FADD (up to 65 %) and p-FADD/FADD (up to 2.4-fold). Other MAO inhibitors (clorgyline, Ro 41-1049, rasagiline), atypical ADs (ketamine and mirtazapine), or ECS did not modulate cortical FADD. Chronic (14 days) desipramine and fluoxetine, but not phenelzine, increased p-FADD (up to 59 %), p-FADD/FADD ratio (up to 1.8-fold), and pro-survival p-PEA-15 (up to 46 %) in rat brain cortex., Conclusions: Multifunctional FADD protein, through an increased p-FADD/FADD ratio, could participate in the mechanisms of anti-apoptotic actions induced by ADs.

A series of 2-amino-6-nitrobenzothiazole-derived extended hydrazones were designed, synthesized, and investigated for their ability to inhibit monoamine oxidase A and B (MAO-A/MAO-B). The compounds were found to exhibit inhibitory activities in the nanomolar to micromolar range. Some of the compounds showed excellent potency and selectivity against the MAO-B isoform. N'-(5-Chloro-2-oxoindolin-3-ylidene)-2-(6-nitrobenzothiazol-2-ylamino)acetohydrazide (compound 31) showed the highest MAO-B inhibitory activity (IC50 =1.8±0.3 nm, selectivity index [SI]=766.67), whereas compound 6 [N'-(1-(4-bromophenyl)ethylidene)-2-(6-nitrobenzothiazol-2-ylamino)acetohydrazide] was found to be the most active MAO-A inhibitor (IC50 =0.42±0.003 μm). Kinetic studies revealed that compounds 6 and 31 exhibit competitive-type reversible inhibition against both MAO-A and MAO-B, respectively. Structure-activity relationship (SAR) studies disclosed several structural aspects significant for potency and the contribution of the methylene spacer toward MAO-B inhibitory potency, with minimal or no neurotoxicity. Molecular modeling studies yielded a good correlation between experimental and theoretical inhibitory data. Binding pose analysis revealed the significance of cumulative effects of π-π stacking and hydrogen bond interactions for effective stabilization of virtual ligand-protein complexes. Further optimization studies of compound 31, including co-crystallization of inhibitor-MAO-B complexes, are essential to develop these compounds as potential therapeutic agents for MAO-B-associated neurodegenerative diseases., (© 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The 3-substituted oxindole derivatives were designed, synthesized, and evaluated for antidepressant activity by employing forced swimming test, tail suspension test, and MAO-A inhibition assay. Results of biological studies revealed that the majority of compounds exhibited potent to moderately potent activity and among them, 12 displayed potency comparable to that of the imipramine with %DID of 37.95 and 44.84 in the FST and TST, respectively. At the same time, imipramine showed %DID of 43.62 and 50.64 in the FST and TST, correspondingly. In the MAO-A inhibition assay, 12 showed an IC50 of 18.27 μmol, whereas the reference drug moclobemide displayed an IC50 of 13.1 μmol. The SAR study disclosed that the presence of bromo atom at the phenyl/furanyl or thienyl moiety in the oxindole derivatives was critical for the antidepressant activity., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Three series of 4,6-dimethoxy-, 4,6-dipiperidino- and 4,6-dimorpholino-1,3,5-triazin-2-yl) amino acid derivatives were synthesized and characterized. A preliminary study for their monoamine oxidase inhibitory activity showed that compounds 7, 18, and 25 had MAO-A inhibition activity comparable to that of the standard clorgyline, with apparently more selective inhibitory activity toward MAO-A than MAO-B and no significant acute toxicity.

Monoamine oxidases (MAOs) have recently emerged as important mitochondrial sources of oxidative stress in the cardiovascular system. Generation of reactive oxygen species during the brief episodes of ischemic preconditioning (IPC) is responsible for the cardioprotection at reperfusion. The aim of this study was to assess the effects of two MAO inhibitors (clorgyline and pargyline) on the IPC-related protection in isolated rat hearts. Animals subjected to 30 min global ischemia and 120 min reperfusion were assigned to the following groups: (i) Control, no additional intervention; (ii) IPC, 3 cycles of 5 min ischemia and 5 min reperfusion before the index ischemia; (iii) IPC-clorgyline, IPC protocol bracketed for 5 min with clorgyline (50 μmol/L); (iv) IPC-pargyline, IPC protocol bracketed for 5 min with pargyline (0.5 mmol/L). The postischemic functional recovery was assessed by the left ventricular developed pressure (LVDP) and the indices of contractility (+dLVP/dt max) and relaxation (-dLVP/dt max). Infarct size (IS) was quantified by TTC staining. In both genders, IPC significantly improved functional recovery that was further enhanced in the presence of either clorgyline or pargyline. IS reduction was comparable among all the preconditioned groups, regardless of the presence of MAO inhibitors. In isolated rat hearts, acute inhibition of MAOs potentiates the IPC-induced postischemic functional recovery without interfering with the anti-necrotic protection.

A series of quinazolinone amino acid ester and quinazolinone amino acid hydrazides were prepared under microwave irradiation as well as conventional condition. The microwave irradiation afforded the product in less reaction time, higher yield and purity. The structures of the synthesized compounds were confirmed by IR, NMR, and elemental analysis. The new synthesized compounds were studied for their monoamine oxidase inhibitory activity. They showed more selective inhibitory activity toward MAO-A than MAO-B. Compounds 7, 10, and 15 showed MAO-A inhibition activity (IC50=3.6×10(-9), 2.8×10(-9), 2.1×10(-9) M, respectively) comparable to that of the standard clorgyline (IC50=2.9×10(-9)M). 2-(2-(Benzo[d][1,3]dioxol-5-yl)-4-oxo-1,2-dihydroquinazolin-3(4H)-yl)acetohydrazide 15 showed selective MAO-A inhibition activity (SI=39524) superior to that of the standard clorgyline (SI=33793). The acute toxicity of the synthesized compounds was determined. In addition, computer-assisted simulated docking experiments were performed to rationalize the biological activity., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

The objective of the study was to evaluate the metabolism dependent inhibition of CYP2B6 catalyzed bupropion hydroxylation in human liver microsomes by monoamine oxidase (MAO) inhibitors and to predict the drug-drug interaction potential of monoamine oxidase inhibitors as perpetrators of drug interaction. Human liver microsomal CYP2B6 activities were investigated using bupropion hydroxylation as probe substrate marker. The results from single point time dependent inhibition and shift assays suggest that clorgyline, pargyline, phenelzine, and selegiline were metabolism based inhibitors of CYP2B6. In IC50 shift assays, clorgyline, pargyline, phenelzine and selegiline are metabolism based inhibitors of CYP2B6 with fold shit of 3.0-, 3.7-, 2.9-, and 11.4-fold respectively. The inactivation of clorgyline was characterized by KI value of 2.5 ± 0.3 and k(inact) value of 0.045 ± 0.001 min(-1). Phenelzine inactivated CYP2B6 with KI and k(inact) values of 44.9 ± 6.9 μM and 0.085 ± 0.003 min(-1) respectively. Inactivation of selegiline was characterized with KI and k(inact) values of 22.0 ± 3.3 and 0.074 ± 0.002 min(-1) respectively. The inactivation caused by these inhibitors was not reversed by dialysis indicating irreversible inhibition. Based on the mechanistic static model, selegiline showed an increase in the area under the curve (AUC) of efavirenz and bupropion by 1.01-fold. Phenelzine predicted to cause an increase in the AUC of efavirenz and bupropion by 9.4- and 2.4-fold respectively considering unbound hepatic inlet concentrations of phenelzine. In conclusion, the results from this study demonstrated that MAO inhibitors can inactivate human liver microsomal CYP2B6. The likelihood of drug interaction when selegiline co-administered with CYP2B6 substrates is remote. Caution is required while co-administering phenelzine with substrates that are exclusively metabolized by CYP2B6 enzyme and substrates that have narrow therapeutic index., (Copyright © 2015. Published by Elsevier Ireland Ltd.)

Monoamine oxidase-A (MAO-A) dysfunction has been historically associated with depression. Recently, depression as well as altered MAO-A expression have both been associated with a poor prognosis in cancers, although the mechanism involved remains ambiguous. For example, MAO-A mRNA is repressed across cancers, yet MAO-A protein and levels of serotonin, a substrate of MAO-A implicated in depression, are paradoxically increased in malignancies, including breast cancer. The effect of clorgyline (CLG), a selective inhibitor of MAO-A, on malignant behaviour, expression of transitional markers, and biochemical correlates was examined in two human breast carcinoma cell lines, i.e. the epithelial, oestrogen receptor (ER)-positive MCF-7 cell line and the post-EMT (mesenchymal), ER-negative MDA-MB-231 cell line. CLG exerted little effect on malignant behaviour in MCF-7 cells, but inhibited proliferation and anchorage-independent growth, and increased invasiveness and active migration of MDA-MB-231 cells. CLG induced the expression of the mesenchymal marker vimentin in MCF-7 cells, but not in MDA-MB-231 cells. In contrast, CLG induced the epithelial protein marker E-cadherin in both cell lines, with a more robust effect in MDA-MB-231 cells (where a nuclear E-cadherin signal was also detected). This effect appears to be independent of any canonical Snai1-mediated regulation of E-cadherin mRNA expression. CLG interfered with the β-catenin/[phospho]GSK-3β complex as well as the E-cadherin/β-catenin complex in both cell lines cells, but, again, the effect was more robust in MDA-MB-231 cells. Parallel studies revealed a general lack of effect of CLG on the ER-negative, epithelial Au565 breast cancer cell line. Thus, any effect of CLG on metastatic behaviours appears to rely on the cell's EMT status rather than on the cell's ER status. These data suggest that inactivation of MAO-A triggers a mesenchymal-to-epithelial transition in MDA-MB-231 cells via a non-canonical mechanism. This potentially implicates an MAO-A-sensitive step in advanced breast cancer and should be borne in mind when considering pharmacological treatment options for co-morbid depression in breast cancer patients., (Copyright © 2014 Elsevier Inc. All rights reserved.)

ABCB5, an ATP-binding cassette (ABC) transporter, is highly expressed in melanoma cells, and may contribute to the extreme resistance of melanomas to chemotherapy by efflux of anti-cancer drugs. Our goal was to determine whether we could functionally express human ABCB5 in the model yeast Saccharomyces cerevisiae, in order to demonstrate an efflux function for ABCB5 in the absence of background pump activity from other human transporters. Heterologous expression would also facilitate drug discovery for this important target. DNAs encoding ABCB5 sequences were cloned into the chromosomal PDR5 locus of a S. cerevisiae strain in which seven endogenous ABC transporters have been deleted. Protein expression in the yeast cells was monitored by immunodetection using both a specific anti-ABCB5 antibody and a cross-reactive anti-ABCB1 antibody. ABCB5 function in recombinant yeast cells was measured by determining whether the cells possessed increased resistance to known pump substrates, compared to the host yeast strain, in assays of yeast growth. Three ABCB5 constructs were made in yeast. One was derived from the ABCB5-β mRNA, which is highly expressed in human tissues but is a truncation of a canonical full-size ABC transporter. Two constructs contained full-length ABCB5 sequences: either a native sequence from cDNA or a synthetic sequence codon-harmonized for S. cerevisiae. Expression of all three constructs in yeast was confirmed by immunodetection. Expression of the codon-harmonized full-length ABCB5 DNA conferred increased resistance, relative to the host yeast strain, to the putative substrates rhodamine 123, daunorubicin, tetramethylrhodamine, FK506, or clorgyline. We conclude that full-length ABCB5 can be functionally expressed in S. cerevisiae and confers drug resistance.

Serotonin (5-hydroxytryptamine; 5-HT) plays important roles in the cardiovascular system; however, the relationship between 5-HT-induced vasocontraction and the arterial 5-HT system including metabolism and signal transduction, in the presence of chronic type 2 diabetes (T2D) remains unclear. Therefore, we investigated 5-HT-induced contraction and associated mechanisms in carotid arteries from chronic T2D Goto-Kakizaki (GK) rats. Contractions in response to 5-HT were examined in carotid arteries from GK rats (42-46 weeks old). To investigate the response mechanisms of arterial smooth muscle, we constructed concentration-response curves for TCB2 (5-HT2A-receptor agonist), BW723C86 (5-HT2B-receptor agonist), and 5-HT in the presence of various inhibitors using endothelium-denuded preparations. Carotid arterial expressions of monoamine oxidase-A (MAO-A), serotonin transporter (SERT), and 5-HT2A were detected by immunoblotting. 5-HT-induced contraction was increased in carotid arteries from GK compared to control Wistar rats in both endothelium-intact and -denuded preparations. In denuded preparations, we found that: (1) TCB2-induced contraction was increased in GK rat arteries (vs. Wistar); (2) MAO-A inhibitor did not affect 5-HT-induced contraction, whereas SERT inhibitor augmented such contractions in both groups; and (3) differences in 5-HT-induced contractions were abolished by p38 MAPK, PI3K, and Rho kinase inhibitors. Carotid arterial expressions of MAO-A, SERT, and 5-HT2A remained unchanged in the groups. The results suggest that 5-HT-induced contraction is augmented in T2D GK rat carotid arteries. This augmentation is due to smooth muscle activation partly mediated by p38 MAPK, PI3K, and Rho kinases, and may also be partly due to arterial SERT activity., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

The efficiency (work/oxygen consumption) of isolated papillary muscles from failing hearts is reduced. We investigated whether this can be due to an increase of intrinsic cardiac adrenergic (ICA) cell density. The number of ICA cells in the septum and both ventricular walls was determined by tyrosine hydroxylase immunohistochemistry in rats with monocrotaline-induced pulmonary hypertension. We found that the number of ICA cells is about 200,000 per rat heart. ICA cell density was significantly lower in right ventricular myocardium of hypertrophied hearts (P < 0.01). MAO-A enzyme histochemistry and inhibition experiments with clorgyline in papillary muscles were performed to localize the enzyme and to determine its oxygen consumption. Upregulation of MAO-A was found in the right ventricular wall and papillary muscles of failing hearts (P = 0.018). A positive correlation between ICA cell density and MAO-A activity was absent. Clorgyline (2 μM) decreased the basal rate of oxygen consumption of right ventricular papillary muscles by 65 μM O(2)/s (P = 0.027). This rate can only be maintained for several seconds judging from the catecholamine content of the preparations reported previously. High ICA cell activity rather than density and/or recycling of oxidized catecholamines are discussed as alternative explanations for the low myocardial efficiency in experimental pulmonary hypertension.

Hydrogen peroxide is the co-substrate used by the enzyme catalase to form Compound I (the catalase-H2O2 system), which is the major pathway for the conversion of ethanol (EtOH) into acetaldehyde in the brain. This acetaldehyde has been involved in many of the effects of EtOH. Previous research demonstrated that treatments that change the levels of cerebral H2O2 available to catalase modulate the locomotor-stimulating effects of EtOH and its volitional intake in rodents. However, the source of H2O2 which is used by catalase to form Compound I and mediates the psychoactive actions of EtOH is unknown. One cause of the generation of H2O2 in the brain comes from the deamination of biogenic amines by the activity of MAO-A. Here we explore the consequences of the administration of the MAO-A inhibitor clorgyline on EtOH-induced locomotion and voluntary EtOH drinking. For the locomotor activity tests, we injected Swiss (RjOrl) mice intraperitoneally (IP) with clorgyline (0-10mg/kg) and later (0.5-8h) with EtOH (0-3.75 g/kg; IP). Following these treatments, mice were placed in locomotor activity chambers to measure their locomotion. For the drinking experiments, mice of the C57BL/6J strain were injected IP with clorgyline prior to offering them an EtOH (20%) solution following a drinking-in-the-dark procedure. Additional experiments were performed to assess the selectivity of this compound in altering EtOH-stimulated locomotion and EtOH intake. Moreover, we indirectly tested the ability of clorgyline to reduce brain H2O2 levels. We showed that this treatment selectively reduced EtOH-induced locomotion and its self-administration. Moreover, this compound decreased central H2O2 levels available to catalase. We suggest that H2O2 derived from the deamination of biogenic amines by the activity of MAO-A could determine the formation of brain EtOH-derived acetaldehyde. This centrally-formed acetaldehyde within the neurons of the aminergic system could play a role in the neurobehavioral properties of EtOH., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Background and Purpose: Selective MAO type B (MAO-B) inhibitors are effective in potentiation of the clinical effect of L-DOPA in Parkinson's disease, but dopamine (DA) is deaminated mainly by MAO type A (MAO-A) in rat brain. We sought to clarify the roles of MAO-A and MAO-B in deamination of DA formed from exogenous L-DOPA in rat striatum depleted of dopaminergic, or both dopaminergic and serotonergic innervations. We also studied the effect of organic cation transporter-3 (OCT-3) inhibition by decinium-22 on extracellular DA levels following L-DOPA., Experimental Approach: Striatal dopaminergic and/or serotonergic neuronal innervations were lesioned by 6-hydroxydopamine or 5,7-dihydroxytryptamine respectively. Microdialysate DA levels after systemic L-DOPA were measured after inhibition of MAO-A or MAO-B by clorgyline or rasagiline respectively. MAO subtype localization in the striatum was determined by immunofluorescence., Key Results: Rasagiline increased DA extracellular levels following L-DOPA to a greater extent in double- than in single-lesioned rats (2.8- and 1.8-fold increase, respectively, relative to saline treatment); however, clorgyline elevated DA levels in both models over 10-fold. MAO-A was strongly expressed in medium spiny neurons (MSNs) in intact and lesioned striata, while MAO-B was localized in glia and to a small extent in MSNs. Inhibition of OCT-3 increased DA levels in the double- more than the single-lesion animals., Conclusions and Implications: In striatum devoid of dopaminergic and serotonergic inputs, most deamination of L-DOPA-derived DA is mediated by MAO-A in MSN and a smaller amount by MAO-B in both MSN and glia. OCT-3 plays a significant role in uptake of DA from extracellular space. Inhibitors of OCT-3 are potential future targets for anti-Parkinsonian treatments., (© 2013 The British Pharmacological Society.)

Epigenetic gene silencing, mediated by aberrant promoter DNA hypermethylation and repressive histone modifications, is a hallmark of cancer. Although heritable, the dynamic nature and potential reversibility through pharmacological interventions make such aberrations attractive targets. Since cancers contain multiple epigenetic abnormalities, combining therapies that target different defects could potentially enhance their individual efficacies. 5-Aza-2'-deoxycytidine (5-Aza-CdR), FDA-approved drug for the treatment of myelodysplastic syndrome, can inhibit DNA methyltransferases (DNMTs) upon incorporation into the DNA of dividing cells, resulting in global demethylation. More recently, the first histone demethylase, lysine specific demethylase 1 (LSD1), which demethylates both histone and non-histone substrates, has become a new target for epigenetic therapy. Using, clorgyline, an LSD1 inhibitor (LSD1i) to treat cancer cell lines, we show that clorgyline employs two mechanisms of action depending on the cell type: it can either induce global DNA demethylation or inhibit LSD1-driven H3K4me2 and H3K4me1 demethylation to establish an active chromatin configuration. We also investigate the therapeutic efficacy of combining 5-Aza-CdR with clorgyline and determine that this combinatorial treatment has synergistic effects on reactivating aberrantly silenced genes by enriching H3K4me2 and H3K4me1. Many of the reactivated genes are categorized as cancer testis antigens or belong to the interferon-signaling pathway, suggesting potential implications for immunotherapy. Together, our results demonstrate that combinatorial treatment consisting of a DNMT inhibitor (DNMTi) and an LSD1i have enhanced therapeutic values and could improve the efficacy of epigenetic therapy.

Density functional theory calculations were employed to investigate the nature of chemical bond formation between the flavin co-factor of the enzyme monoamine oxidase (MAO) and its irreversible acetylenic inhibitor clorgyline in its terminally deprotonated anionic form. Since MAOs regulate the level of neurotransmitters in living cells, this reaction is pharmacologically relevant for treating depression and other mood disorders. The results revealed that this pathway is associated with the activation free energy of ΔG act (#) = 17.4 kcal mol(-1), which, together with our previous results, suggests that clorgyline is intrinsically a more effective MAO inhibitor than antiparkinsonian drugs rasagiline and selegiline considering the preferred MAO isoforms in each case, thus displaying a trend in agreement with experimental data. The reaction is facilitated by the pronounced electrophilic character of the flavin moiety, due to its ability to efficiently accommodate excess negative charge from the approaching anionic inhibitor through resonance effect. The investigated mechanism was additionally validated by the inspection of the geometry of the flavin moiety in the formed adduct, which exhibit distortion from planarity consistent with experimental observations. These results offer valuable insight for mechanistic studies on other flavoenzymes and for the design of new antidepressants and antiparkinsonian drugs.

Introduction: Depression is a mental disease causing large personal and socio-economic problems, and new improved drugs are therefore needed. Selective monoamine oxidase A (MAO-A) inhibitors are potential anti-depressants, but discovering new MAO-A inhibitors from natural sources by bioassay-guided approaches are a lengthy and time-consuming process. New analytical technologies that allow simultaneously chemical and biological screening of extracts are therefore urgently needed., Method: In the present study we describe coupling of a photometric microplate-based high-resolution MAO-A inhibitor assay with a hyphenated system consisting of high-performance liquid chromatography, solid-phase extraction and tube transfer nuclear magnetic resonance (HPLC-SPE-ttNMR). The standard compound clorgyline, and an extract of black pepper (Piper nigrum L.), representing a complex plant matrix, were used for proof-of-concept., Results: The work with clorgyline showed that the microplate-based high-resolution assay produced MAO-A inhibition profiles that easily allowed detection of submicrogram amounts of this selective MAO-A inhibitor. Furthermore, the HPLC-SPE-ttNMR/high-resolution MAO-A inhibition assay platform allowed identification of piperine and two piperine analogues as the main MAO-A inhibitors in the black pepper petroleum ether extract., Conclusion: The HPLC-SPE-ttNMR/high-resolution MAO-A inhibition assay platform is a powerful tool for fast and efficient identification of new MAO-A inhibitors from complex extracts, and promise future advancement in the search for new anti-depressants from natural sources., (Copyright © 2012 John Wiley & Sons, Ltd.)

Aim: Monoamine oxidases (MAOs) are mitochondrial enzymes, with 2 isoforms, A and B that convert biogenic amines to their corresponding aldehydes via a reaction that produces hydrogen peroxide. Since MAO-A is the predominant form at vascular level we hypothesized that MAO-A-dependent H2O2 production may contribute to the development of endothelial dysfunction and, MAOs inhibition could improve the vascular function, respectively., Material and Methods: To this aim aortic rings were isolated from female adult spontaneously hypertensive rats (SHR) and their corresponding (Wistar-Kyoto) controls. The effect of MAO-A inhibitor, clorgyline (10 micromol/l) on endothelium-dependent relaxation (EDR) in response to acetylcholine and endothelium-independent relaxation in response to sodium nitroprusside, was studied in isolated phenylephrine-preconstricted aortic segments in the presence of indometacine (10 micromol/l)., Results: In hypertensive group EDR was significantly decreased - maximal relaxation (% of KCI, mean +/- SD) being 37 +/- 3.5 in SHR vs. 3.7 +/- 1.8 in controls. If experiments were done in the presence of clorgyline, EDR in control segments was unaffected. However, the compound restored normal EDR in aortic segments from hypertensive rats (maximal relaxation % of KCI, 13.7 +/- 2.3)., Conclusions: Inhibition of the MAO-A isoform might be useful in restoring endothelium-dependent relaxation in this experimental model of hypertension in rat.

In this study we assessed the involvement of monoamine oxidase B (MAO-B), a key enzyme implicated in monoamine metabolism, on postoperative (plantar incision) and neuropathic (partial sciatic nerve ligation) pain models in mice. Paw incision submitted mice showed a significant decrease in mechanical threshold compared with the sham-operated mice, characterizing the development of mechanical allodynia. The selective and irreversible MAO-B inhibitor selegiline, at a dose sufficient to selectively inhibit MAO-B activity (10 mg/kg), showed an anti-allodynic effect from 0.5 to 6 h after incision. Likewise, partial sciatic nerve ligation submitted mice also developed mechanical allodynia, which was reversed by selegiline (10 mg/kg) from 2 to 6 h after treatment. In addition, a significant increase on striatal MAO-B activity was observed in neuropathic mice compared with the sham-operated animals, which was reversed by selegiline treatment. Taken together, our results showed that MAO-B seems to exert a critical role in the development of postoperative and neuropathic pain., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Cardiovascular baroreceptor responsiveness of conscious rats treated with selective inhibitors of monoamine oxidase (MAO) types A and B was determined by measurement of blood pressure (BP) and heart rate (HR) responses to intravenous injection of phenylephrine and sodium nitroprusside. Treatment with selegiline (1 or 5 mg/kg p.o. daily for 7 days) did not significantly modify resting levels of BP and HR, lower or upper HR plateau levels, or HR/BP gain. Treatment with clorgyline (2 mg/kg p.o. daily for 7 days) increased HR/BP gain but also did not modify resting BP or HR, or lower and upper plateau levels of HR. The results are compatible with an effect of MAO-A inhibition to modify monoamine levels in medullary areas participating in CNS control of blood pressure., (Copyright © 2012 Elsevier B.V. All rights reserved.)

BACKGROUND AND PURPOSE Preclinical pharmacology of 3-iodothyronamine (T1AM), an endogenous derivative of thyroid hormones, indicates that it is a rapid modulator of rodent metabolism and behaviour. Since T1AM undergoes rapid enzymatic degradation, particularly by MAO, we hypothesized that the effects of T1AM might be altered by inhibition of MAO. EXPERIMENTAL APPROACH We investigated the effects of injecting T1AM (i.c.v.) on (i) feeding behaviour, hyperglycaemia and plasma levels of thyroid hormones and (ii) T1AM systemic bioavailability, in overnight fasted mice, under control conditions and after pretreatment with the MAO inhibitor clorgyline. T1AM (1.3, 6.6, 13, 20 and 26 µg·kg(-1) ) or vehicle were injected i.c.v. in fasted male mice not pretreated or pretreated i.p. with clorgyline (2.5 mg·kg(-1) ). Glycaemia was measured by a glucorefractometer, plasma triiodothyronine (fT3) by a chemiluminescent immunometric assay, c-fos activation immunohistochemically and plasma T1AM by HPLC coupled to tandem-MS. KEY RESULTS T1AM, 1.3 µg·kg(-1) , produced a hypophagic effect (-24% vs. control) and reduced c-fos activation. This dose showed systemic bioavailability (0.12% of injected dose), raised plasma glucose levels and reduced peripheral insulin sensitivity (-33% vs. control) and plasma fT3 levels. These effects were not linearly related to the dose injected. Clorgyline pretreatment strongly increased the systemic bioavailability of T1AM and prevented the hyperglycaemia and reduction in fT3 induced by T1AM. CONCLUSIONS AND IMPLICATIONS T1AM induces central and peripheral effects including hyperglycaemia and a reduction in plasma fT3 levels in fasted mice. These effects critically depend on the concentration of T1AM or its metabolites in target organs., (© 2012 The Authors. British Journal of Pharmacology © 2012 The British Pharmacological Society.)

Oxidative deamination of the GABA(A) partial agonist CP-409,092 and sumatriptan represents a major metabolic pathway and seems to play an important role for the clearance of these two compounds. Similar to sumatriptan, human mitochondrial incubations with deprenyl and clorgyline, probe inhibitors of monoamine oxidase B and monoamine oxidase A (MAO-B and MAO-A), respectively, showed that CP-409,092 was metabolized to a large extent by the enzyme MAO-A. The metabolism of CP-409,092 and sumatriptan was therefore studied in human liver mitochondria and in vitro intrinsic clearance (CL(int)) values were determined and compared to the corresponding in vivo oral clearance (CL(PO)) values. The overall objective was to determine whether an in vitro-in vivo correlation (IVIVC) could be described for compounds cleared by MAO-A. The intrinsic clearance, CL(int), of CP-409,092 was approximately 4-fold greater than that of sumatriptan (CL(int), values were calculated as 0.008 and 0.002 ml/mg/min for CP-409,092 and sumatriptan, respectively). A similar correlation was observed from the in vivo metabolic data where the unbound oral clearance, CL(u)(PO), values in humans were calculated as 724 and 178 ml/min/kg for CP-409,092 and sumatriptan, respectively. The present work demonstrates that it is possible to predict in vivo metabolic clearance from in vitro metabolic data for drugs metabolized by the enzyme monoamine oxidase.

Resistance to the commonly used azole antifungal fluconazole (FLC) can develop due to overexpression of ATP-binding cassette (ABC) and major facilitator superfamily (MFS) plasma membrane transporters. An approach to overcoming this resistance is to identify inhibitors of these efflux pumps. We have developed a pump assay suitable for high-throughput screening (HTS) that uses recombinant Saccharomyces cerevisiae strains hyperexpressing individual transporters from the opportunistic fungal pathogen Candida albicans. The recombinant strains possess greater resistance to azoles and other pump substrates than the parental host strain. A flow cytometry-based HTS, which measured increased intracellular retention of the fluorescent pump substrate rhodamine 6G (R6G) within yeast cells, was used to screen the Prestwick Chemical Library (PCL) of 1,200 marketed drugs. Nine compounds were identified as hits, and the monoamine oxidase A inhibitor (MAOI) clorgyline was identified as an inhibitor of two C. albicans ABC efflux pumps, CaCdr1p and CaCdr2p. Secondary in vitro assays confirmed inhibition of pump-mediated efflux by clorgyline. Clorgyline also reversed the FLC resistance of S. cerevisiae strains expressing other individual fungal ABC transporters (Candida glabrata Cdr1p or Candida krusei Abc1p) or the C. albicans MFS transporter Mdr1p. Recombinant strains were also chemosensitized by clorgyline to other azoles (itraconazole and miconazole). Importantly, clorgyline showed synergy with FLC against FLC-resistant C. albicans clinical isolates and a C. glabrata strain and inhibited R6G efflux from a FLC-resistant C. albicans clinical isolate. Clorgyline is a novel broad-spectrum inhibitor of two classes of fungal efflux pumps that acts synergistically with azoles against azole-resistant C. albicans and C. glabrata strains.

The concentration of presenilin-1 (PS-1) protein at the mitochondrial-associated aspect of the endoplasmic reticulum supports the potential for a mitochondrial influence of PS-1. Given that carriers of certain Alzheimer's disease (AD)-related PS-1 variants are predisposed to clinical depression and that depression has been historically associated with the mitochondrial enzyme, monoamine oxidase-A (MAO-A), we investigated cortical MAO-A function in the AD-related PS-1(M146V) knock-in mouse. The MAO-A system was clearly altered in the PS-1(M146V) mouse as revealed by (a) a mismatch between MAO-A protein expression and MAO-A activity; (b) changes in MAO-A-mediated monoaminergic neurotransmitter metabolism; (c) changes in non-cognitive behavior following treatment with the irreversible MAO-A inhibitor clorgyline; and (d) an increase in the potency of clorgyline in these same mice. We next investigated whether PS-1(M146V) could be influencing MAO-A directly. We observed (a) an enhanced MAO-A activity in necropsied PS-1(M146V) mouse cortical extracts incubated with DAPT (a PS-1 substrate-competitor); (b) the proximity of PS-1 with MAO-A and mitochondrial markers in cortical sections and in primary cortical neurons; (c) the co-segregation and co-immunoprecipitation of PS-1 and MAO-A within the mitochondrial fraction; and (d) the co-immunoprecipitation of overexpressed PS-1(M146V) and MAO-A proteins from N2a lysates. The PS-1(ΔEx9) and PS-1(D257A) variants, known to have low substrate-binding capacity, co-immunoprecipitated weakly with MAO-A. These combined data support a physical interaction between PS-1 and MAO-A that could influence MAO-A activity and contribute to the monoaminergic disruptions common to disorders as seemingly diverse as depression and AD.

Epigenetic changes including histone methylation, histone acetylation, and DNA methylation are thought to play important roles in the onset and progression of cancer in numerous tumor types. Recent evidence shows that dysregulated epigenetic modifications are as significant as genetic mutations and can act as oncogenic driver lesions causing autonomous growth of cancer cells. Here, we investigated the role of lysine-specific demethylase 1 in mesenchymal tumors. Lysine-specific demethylase 1 is the first discovered histone lysine demethylase and can demethylate both H3K4me2/1 and H3K9me2/1. By analyzing a total of 468 tumors, we describe for the first time high lysine-specific demethylase 1 expression in several highly malignant sarcomas, including synovial sarcomas, rhabdomyosarcomas, desmoplastic small round cell tumors and malignant peripheral nerve sheath tumors. Among the intermediate tumors only solitary fibrous tumors were found to be highly lysine-specific demethylase 1 positive, whereas lysine-specific demethylase 1 expression was low or absent in benign tumors. Lysine-specific demethylase 1 inhibition with small molecule inhibitors resulted in growth inhibition of synovial sarcoma cells in vitro and an increase in global H3K4me2 methylation. Sarcomas continue to remain a clinical challenge and therefore the identification of both diagnostic markers and novel drug targets for the development of new therapeutic options are needed. Our results suggest that dysregulation of lysine-specific demethylase 1 is associated with highly malignant sarcomas proposing them as molecular tumor markers as well as targets for the treatment of these tumor types., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Monoamine oxidases (MAO-A, MAO-B) metabolize biogenic amines and have been implicated in neuronal apoptosis. Although apoptosis is an important process in embryo development, the role of MAO isoenzymes has not been investigated in detail. We found that expression of MAO-A and MAO-B can be detected early on during embryo development. Expression levels remained constant until around midgestation but then dropped to almost undetectable levels toward birth. Similar expression kinetics were observed in the brain. Isoform-specific expression silencing of MAO-A mediated by siRNA during in vitro embryogenesis induced developmental defects, as indicated by a reduction of the crown rump length and impaired cerebral development. These alterations were paralleled by elevated serotonin levels. Similar abnormalities were observed when embryos were cultured in the presence of the MAO-A inhibitor clorgyline or when the transcriptional inhibitor of MAO-A expression R1 was overexpressed. In contrast, no such alterations were detected when expression of MAO-B was knocked down. To explore the underlying mechanisms for the developmental abnormalities in MAO-A knockdown embryos, we quantified the degree of developmental apoptosis in the developing brain. MAO-A knockdown reduced the number of apoptotic cells in the neuroepithelium, which coincided with impaired activation of caspases 3 and 9. Moreover, we observed reduced cyclin D1 levels as an indicator of impaired cell proliferation in MAO-A knockdown embryos. This data highlights MAO-A as a vital regulator of embryonic brain development.

Monoamine oxidase (MAO) is an essential enzyme in the catabolism of monoamines, and implicated in the immune response of vertebrates. In the present study, the full-length cDNA encoding monoamine oxidase (designated CfMAO) was cloned from Chlamys farreri by using rapid amplification of cDNA ends (RACE) approaches and expression sequence tag (EST) analysis. The open reading frame of CfMAO cDNA encoded 519 amino acids, which shared 73.9% similarity with that from oyster Crassostrea gigas, and 64.5-66.3% similarity with those from vertebrates. A conserved Amino&#95;oxidase domain and a transmembrane domain were identified in the deduced CfMAO protein. The mRNA transcripts of CfMAO could be detected in all the tested tissues, including haemocytes, hepatopancreas, kidney, adductor muscle, mantle, gill and gonad. The mRNA expression of CfMAO was up-regulated significantly in haemocytes of scallops during 6-48 h after bacteria Vibrio anguillarum challenge, and it reached the peak (25.9-fold, P < 0.05) at 12h. The cDNA fragment encoding the mature peptide of CfMAO was expressed in the prokaryotic expression system, and 1mg of the recombinant protein (rCfMAO) could catalyze the deamination of 3665.59 nmol serotonin, 2061.89 nmol norepinephrine, 2104.85 nmol epinephrine or 3040.34 nmol dopamine within 1 min (nmol min⁻¹ mg⁻¹) in vitro. When the reaction mixture was coincubated with 0.1 mmol L⁻¹ MAO inhibitor clorgyline, its catalyzing activity to deaminize serotonin and dopamine was decreased significantly to 1603.69 and 955.39 nmol min⁻¹ mg⁻¹ (P < 0.05) respectively. These results indicated that CfMAO, as the homologue of monoamine oxidase in scallop C. farreri, could modulate the immune response of scallops through the deamination of monoamines., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

We previously demonstrated that intracerebroventricular (ICV) injection of orexin-A induces arousal and increased metabolic turnover rates of norepinephrine, dopamine, and serotonin in layer (egg-type) chicks. Because monoamine oxidase-A (MAO-A) is a potent degrading enzyme of these monoamines, we hypothesized that orexin-A may mediate its arousal-inducing effects through MAO-A. Therefore, we simultaneously injected clorgyline, a specific inhibitor of MAO-A, with orexin-A and examined behavior of chicks. Behaviors associated with arousal were attenuated in the group of chicks that received clorgyline and orexin-A compared with those that received orexin-A alone. For the monoamine turnover rate, enhancement of the turnover rate of serotonin by orexin-A was attenuated by clorgyline. Therefore, we conclude that orexin-A-induced arousal is dependent upon monoamine neural activities stimulated by MAO-A in chicks.

Objective: We have evaluated the impact of chronic administration of clorgyline, a potent monoamine oxidase A inhibitor and a former antidepressant, on the preimplantation embryo development in Wistar rats., Material and Methods: Females were injected intraperitoneally daily for 30 days with saline (control animals, C), or with a low dose of clorgyline (0.1 mg/kg/day, LDC) or with a high dose of clorgyline (1 mg/kg/day, HDC). Embryos were isolated on day 5 of pregnancy and their urine was collected., Results: The number of embryos per female did not differ between experimental groups and control, but we have recorded a decreased number of embryos in HDC group compared to LDC (p < 0.05). We have found that LDC significantly reduced the presence of healthy embryos and increased the presence of the degenerated embryos (p < 0.001). The administration of the LDC resulted in the lowest cell number in blastocysts (p < 0.01). Concerning monoamines in urine, we have observed significantly increased serotonin levels in HDC group compared to control (p < 0.05) and LDC animals (p < 0.01). Norepinephrine levels in both experimental groups were significantly elevated compared to controls, too (p < 0.001 LDC vs C; p < 0.01 HDC vs C)., Conclusions: We assume that the impaired embryo development recorded in the LDC group after clorgyline administration was a consequence of the higher levels of the norepinephrine. We speculate that lesser negative effect of HDC compared to LDC on the preimplantation embryo development could be the consequence of the lesser norepinephrine levels and/or elevated serotonin levels (Tab. 2, Fig. 2, Ref. 37). Full Text in free PDF www.bmj.sk.

Since a substantial proportion of smokers have comorbid mood disorders, the smoking cessation aid varenicline might occasionally be prescribed to patients who are simultaneously treated with antidepressants. Given that varenicline is a selective nicotinic acetylcholine receptor partial agonist and not a substrate or inhibitor of drug metabolizing enzymes, pharmacokinetic interactions with various classes of antidepressants are highly unlikely. It is, however, conceivable that varenicline may have a pharmacodynamic effect on antidepressant-evoked increases in central monoamine release. Interactions resulting in excessive transmitter release could cause adverse events such as serotonin syndrome, while attenuation of monoamine release could impact the clinical efficacy of antidepressants. To investigate this we examined whether varenicline administration modulates the effects of the selective serotonin reuptake inhibitor sertraline and the monoamine oxidase inhibitor clorgyline, given alone and combined, on extracellular concentrations of the monoamines serotonin, dopamine, and norepinephrine in rat brain by microdialysis. Given the important role attributed to cortical monoamine release in serotonin syndrome as well as antidepressant activity, the effects on extracellular monoamine concentrations were measured in the medial prefrontal cortex. Responses to maximally effective doses of sertraline or clorgyline and of sertraline plus clorgyline were the same in the absence as in the presence of a relatively high dose of varenicline, which by itself had no significant effect on cortical monoamine release. This is consistent with the binding profile of varenicline that has insufficient affinity for receptors, enzymes, or transporters to inhibit or potentiate the pharmacologic effects of antidepressants. Since varenicline neither diminished nor potentiated sertraline- or clorgyline-induced increases in neurotransmitter levels, combining varenicline with serotonergic antidepressants is unlikely to cause excessive serotonin release or to attenuate antidepressant efficacy via effects on cortical serotonin, dopamine or norepinephrine release., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

Purpose: Inhibitors of monoamine oxidase A (MAOA), a mitochondrial enzyme that degrades neurotransmitters including serotonin and norepinephrine, are commonly used to treat neurological conditions including depression. Recently, we and others identified high expression of MAOA in normal basal prostatic epithelium and high-grade primary prostate cancer (PCa). In contrast, MAOA is low in normal secretory prostatic epithelium and low-grade PCa. An irreversible inhibitor of MAOA, clorgyline, induced secretory differentiation in primary cultures of normal basal epithelial cells and high-grade PCa. Furthermore, clorgyline inhibited several oncogenic pathways in PCa cells, suggesting clinical value of MAOA inhibitors as a pro-differentiation and anti-oncogenic therapy for high-risk PCa. Here, we extended our studies to a model of advanced PCa, VCaP cells, which were derived from castration-resistant metastatic PCa and express a high level of MAOA., Methods: Growth of VCaP cells in the presence or absence of clorgyline was evaluated in vitro and in vivo. Gene expression changes in response to clorgyline were determined by microarray and validated by quantitative real-time polymerase chain reaction., Results: Treatment with clorgyline in vitro inhibited growth and altered the transcriptional pattern of VCaP cells in a manner consistent with the pro-differentiation and anti-oncogenic effects seen in treated primary PCa cells. Src, beta-catenin, and MAPK oncogenic pathways, implicated in androgen-independent growth and metastasis, were significantly downregulated. Clorgyline treatment of mice bearing VCaP xenografts slowed tumor growth and induced transcriptome changes similar to those noted in vitro., Conclusion: Our results support the possibility that anti-depressant drugs that target MAOA might find a new application in treating PCa.

Previous studies have demonstrated that serotonin (5-HT) syndromes, particularly for the malignant cases, can be alleviated by ice water mists, cooling blankets and many other external cooling measures. In this study, we tested the hypothesis that external cooling measures reduce the responsivity of 5-HT(2A) receptors to excessive 5-HT efflux, which may be a possible mechanism underlying the treatment of serotonin syndrome. To test this, rat experiments were carried out in the standard and cool ambient temperature (T(amb) ) by administration of the 5-HT precursor 5-hydroxy-L-tryptophan combined with the monoamine oxidase inhibitor clorgyline. The first set of experiments was to assess severity of the syndromes by measuring body temperature responses. Consistent with the hypothesis, we found that the syndrome was malignant at the standard T(amb) of 22°C but alleviated at 12 or 6°C, these results being similar to those in rats pre-treated with the 5-HT(2A) receptor antagonist ketanserin. The second set of experiments was to utilize microdialysis to determine the relationship between the syndrome severity and 5-HT levels at the above-mentioned T(amb) . We found that excessive 5-HT efflux consisted of primary and secondary components through two distinct mechanisms. Furthermore, the secondary component efflux, which can be ascribed to 5-HT(2A) receptor activation, was proportionally reduced at the cool T(amb) of 12 and 6°C. In conclusion, results of this study support the hypothesis that cooling T(amb) reduces the functional activity of 5-HT(2A) receptors, thus alleviating the malignant syndrome., (© 2010 The Authors. © 2010 Nordic Pharmacological Society.)

We have evaluated the impact of chronic administration of clorgyline, a potent monoamine oxidase A inhibitor and a former antidepressant, on the preimplantation embryo development in Wistar rats. Females were injected intraperitoneally daily for 30 days with saline (control animals), or with a low-dose clorgyline (LDC, 0.1 mg/kg per d) or with a high-dose clorgyline (HDC, 1 mg/kg per d). Embryos were isolated on day 5 of pregnancy and urine was collected by puncture of the urinary bladder. The number of embryos per female did not differ between experimental groups and control, but we have recorded a decreased number of embryos in HDC group compared to LDC (P < .05). We have found that LDC significantly reduced the presence of healthy embryos and increased the presence of the degenerated embryos (P < .001). The administration of the LDC resulted in the lowest cell number in blastocysts. We have observed significantly increased serotonin levels in HDC group compared to both control (P < .05) and LDC animals (P < .01). Norepinephrine (NE) levels in both experimental groups were significantly elevated compared to controls. Dopamine levels did not differ between groups (P > .05). We speculate that lesser negative effect of HDC compared to LDC on the preimplantation embryo development could be the consequence of the lower NE levels and/or elevated serotonin levels. Potential mechanisms mediating clorgyline-induced impaired preimplantation embryo development are proposed.