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Befloxatone is a reversible, selective, and competitive monoamine oxidase (MAO-A) inhibitor of the phenyloxazolidinone family group that was studied in the past for its treatment of depression, social phobia, and panic disorders. Befloxatone causes an increase in extracellular striatal dopamine and cortical norepinephrine levels, but does not affect cortical serotonin levels. It also inhibits the firing rate of serotonergic neurons, partially decreases the firing of noradrenergic neurons and has no effect on the firing of dopaminergic neurons. Currently befloxatone is no longer being developed as a therapeutic drug. Sanofi-Synthelabo, the manufacturers of the drug, stated that there was no significant beneficial effect achieved in their phase III studes.

Synthesis


The first reversible MOA-A inhibitor of the phenoxyloxazolidinone family discovered was toloxatone, and after its discovery efforts were made to synthesize a more potent and selective MOA-A inhibitor. Thus, befloxatone was created, which made use of the aromatic system, phenyl ring substitution, and the substitution of the hydroxymethyl chain on to the phenyloxazolidinone moiety. Befloxatone is typically synthesized by the condensation of 1,1,1,-trifuloro-4-(toxyloxy)-2(R)-butanol with 3-(4-hydroxyphenyl)-5(R)-(methoxymethyl)oxazoldin-2-one by means of K2Co3 in hot dimethylformamide (DMF). However, recently befloxatone has been used as a positron emission tomography (PET) tracer for the in vivo imaging of MAO-A density, and therefore methods of efficiently synthesizing befloxatone using radiosynthesis have been developed.

Pharmacodynamics
Befloxatone is a competitive MAO-A antagonist useful due to its selectivity in inhibiting MAO-A as well as its reversibility, which minimizes side effects that have commonly been associated with MAOIs. Befloxatone is hydrophobic, with two surrounding hydrophilic regions that interact with the polar residues flavin adenine dinucleotide-cofactor of the MAO-A as well as with the MAO-A protein moiety. These chemical interaction properties confer befloxatone with high selectivity for the MAO-A isoform.

Befloxatone can also act at the metabotropic glutamate 2 receptors and its binding is highly concentrated in the locus coeruleus and interpeduncular nucleus, while less concentrated in the dorsal raphe, cerebral cortex and limbic system. In monoaminergic neurons, the physiological release of monoamine transmitters due to befloxatone is subject to feedback from autoreceptors. Befloxatone transport is modulated by human breast cancer resistance protein.

The potency of befloxatone for MAO-A (IC50 = 4nM) is superior to that of other reversible MAO-A inhibitors because it is a fully competitive, potent and selective MOA-A inhibitor. . Befloxatone has a Kd of 1.3nM, a Bmax of 7.5 pmol/mg, and it inhibits rat brain MAO-A activity with an ED50 of 0.02mg/kg for MAO-A. Also, Ki values for befloxatone range from 2.1 to 3.6nM for MAO-A and 269 to 920nM for MAO-B. The high efficacy and potency of befloxatone can be shown through preclinical models of depression which include the learned helplessness paradigm, acknowledged as one of the most valid ways of measuring animal depression, and the forced swimming test. Befloxatone has a pharmaco-EEG profile similar to that of non-sedative antidepressants currently in the market.

Pharmacokinetics
Befloxatone has a half-life of around 11 hours in males and is rapidly absorbed with a Tmax of approximately 2.5 hours. Using deaminated metabolite 3-4dihydroxyphenylglycol (DHPG) of plasma levels as an index of MAO inhibition it was found that a dose of 10mg of oral befloxatone administration not only produced a significant decrease in plasma DHPG levels with a peak after 2 hours, but the inhibition effect also persisted over 24 hours. According to the phase I and II trials conducted thus far it has been shown that a daily 10 mg oral dose of befloxatone seems to induce the optimal effects, although up to 20 mg per day orally has been considered safe. Befloxatone has no significant effect on skilled performance, memory, or sleep patterns, and rather has the effect of improving long-term memory in subjects.

O-demethylated befloxatone is the major befloxatone metabolite in the human body and has a half life of 22 hours. It is a 5 to 10-fold less potent MAO-A inhibitor compared to befloxatone. Befloxatone and its metabolites are glucuronidated, after which they are excreted in the urine.

Treatment
Befloxatone was studied for its potential treatment of depression, social phobias and panic disorders by preventing the catalyzation of monoamines. In the past, irreversible MAOIs have been used in the treatment of various emotional diseases such as depression, anxiety disorders, hysterical traits, phobic disorders, obsessive-compulsive disorders, bulimia, and fatigue. However, due to side effects such as hepatotoxic effects, insomnia, orthostatic hypotension, hypomania, and the negative interaction they have with tyramine, which can cause the hypertensive 'cheese effect', these drugs have been largely replaced in studies by a new class of reversible inhibitors. These inhibitors include moclobemide, brofaromine, toloxatone and befloxatone. In mice, befloxatone is shown to be 10~500-fold more potent than reference monoamine oxidase inhibitors including clorgyline, nialamide, phenelzine and tranycypromine and monoamine reuptake inhibitors such as fluoxetine. It is especially potent compared to the reference inhibitors in the learned helplessness paradigm and in the forced swimming test for mice. Unlike irreversible monoamine oxidase inhibitors, befloxatone has no potentiating pressor effect when administered with tyramine. This can be attributed to the fact that befloxatone potently blocks MAO-A activity and has the ability to enhance catecholamine release.

Side Effects
Befloxatone has relatively little known side effects, with the clinical trials suggesting that the drug lacks sedative, convulsant, anticholinergic, cardiovascular and (negligible) tyramine effects. When administered with a 10 mg dose, patients were seen to exhibit significant long-term memory improvement with no significant side effect in skilled performance or sleep patterns. In mice, befloxatone had no sedative or stimulant activity up to doses of 200 mg/kg orally and likewise showed no anticholinergic activity. MAO inhibitors have been restricted because of their side effects including hepatotoxicity, orthostatic hypotension and hypertension caused when they interact with tyramine, but befloxatone does not exhibit any of these side effects. However, when administered at 1mg/kg orally befloxatone, like all other MAOIs, caused hypotension with a 10% decrease in blood pressure. This shows that hypotension may be a dose-limiting clinical side effect for befloxatone. Regardless, the propensity for a lack of side effects coupled with high potency and wide range of active doses in antidepressant models gives befloxatone a good therapeutic index.

Other Uses
Befloxatone has good imaging characteristics and a low nonsaturable uptake. Also, its volume of distribution correlates to the protein and mRNA levels of MAO-A in the human brain. Therefore befloxatone is used as a positron emission tomography (PET) radioligand to image MAO-A density in the brain.

The most prominent use of befloxatone as a PET radioligand has been to confirm the inhibition of cerebral monoamine oxidase A in tobacco smokers. It was previously known that cigarette smoke was a potent inhibitor of MAO-A and –B isozymes in humans, which accounted for the addictive properties and mood-modulating effects of tobacco. However, the extent to which the smoke inhibited the monoamine oxidases was unknown. Using befloxatone as a selective radioligand it was found that smokers had a reduction of befloxatone binding potential in cortical areas by 60% and in the caudate and thalamus by 40%, indicating a widespread inhibition of cerebral MAO-A in smokers. In monkeys, befloxatone confirmed that cigarette smoke was seen to affect cardiac monoamine oxidase A as well. In fact, the inhalation of tobacco smoke was seen to decrease the Bmax of befloxatone within the myocardium from 208 pmol/ml to 150 pmol/ml, showing that for monkeys, a single cigarette could affect the cardiac turnover of catecholamines.