Through scaffold modification to 3,5-diaryl-4,5-dihydroisoxazoles: new potent and selective inhibitors of monoamine oxidase B

Figures & data

Figure 1. Scaffold evolution of MAO B inhibitors: from dihydro-pyrazoles to dihydro-isoxazoles.

Table 1. Chemical, analytical, and physical data of derivatives EMAC II (i–m).

			C–H–N
Compound	R	Cryst. solvent	Calc.	Found	M.P. (°C)	Yield %	E.I. 70 eV mass spectra m/z (rel. ab.)
EMAC II i	4-Cl	Ethanol	70.72, 5.19, 5.15	70.67, 5.17, 5.12	134-5	42	271 (100); 240 (78); 139 (17)
EMAC II j	4-CH3	Ethanol	81.24, 6.82, 5.57	81.27, 6.83, 5.54	103-4	46	251 (100); 220 (83); 119 (13)
EMAC II k	4-F	Ethanol	75.28, 5.53, 5.49	75.31, 5.54, 5.43	132-4	41	255 (100); 224 (75); 123 (19)
EMAC II l	3,4-Cl	Water/ethanol	62.76, 4.28, 4.57	62.72, 4.31, 4.54	134-5	65	305 (100); 274 (81); 173 (22)
EMAC II m	4-OCH3	ethanol	76.38, 6.41, 5.24	76.33, 6.40, 5.22	131-3	44	267 (100); 236 (71); 135 (9)

Table 2. ¹H NMR data and main fragments in E.I. mass spectrometry of derivatives EMAC II (i–m).

Compound	^1H NMR δ (ppm)
EMAC II i	^1H-NMR: (300 MHz, CDCl3) δH 2.38 (s, 3H, CH3), 3.23 (dd, 1H, Jab 16.5, Jax 7.8, CH2, isoxazole), 3.76 (dd, 1H, Jab 16.5, Jbx 10.1, CH2, isoxazole), 5.59 (dd, 1H, Jbx 10.8, Jax 8.1, CH, isoxazole), 7.21 (d, 2H, J 7.8, CH, arom.), 7.33 (m, 4H, CH, arom.), 7.57 (2H, d, J 8.1, CH, arom.).
EMAC II j	^1H-NMR: (300 MHz, CDCl3) δH 2.37 (s, 3H, CH3), 2.43 (s, 3H, CH3), 3.30 (dd, 1H, Jab 16.5, Jax 8.3, CH2, isoxazole), 3.72 (dd, 1H, Jab 16.5, Jbx 10.8, CH2, isoxazole), 5.66 (dd, 1H, Jbx 10.8, Jax 8.3, CH, isoxazole), 7.16–7.29 (m, 6H, CH, arom.), 7.57 (d, 2H, J 8.3, CH, arom.).
EMAC II k	^1H-NMR: (300 MHz, CDCl3) δH 2.35 (s, 3H, CH3), 3.27 (dd, 1H, Jab 16.7, Jax 8.0, CH2, isoxazole), 3.74 (dd, 1H, Jab 16.5, Jbx 10.7, CH2, isoxazole), 5.69 (dd, 1H, Jbx 10.7, Jax 8.3, CH, isoxazole), 7.04 (t, 2H, J 8.3, CH, arom.), 7.21 (d, 2H, J 8.1, CH, 4-CH3-phenyl), 7.36 (dd, 2H, CH, J 8.3/5.3, arom.), 7.56 (d, 2H, J 8.2, CH, 4-CH3-phenyl).
EMAC II l	^1H-NMR: (300 MHz, CDCl3) δH 2.38 (s, 3H, CH3), 3.27 (dd, 1H, Jab 16.7, Jax 7.8, CH2, isoxazole), 3.78 (dd, 1H, Jab 16.4, Jbx 10.8, CH2, isoxazole), 5.67 (dd, 1H, Jbx 10.8, Jax 7.8, CH, isoxazole), 7.21 (m, 3H, CH, arom.), 7.44 (d, 1H, J 8.2, CH, arom.), m, 7.49 (d, 1H, J 1.2, CH, arom.), 7.56 (d, 2H, J 8.1, CH, arom.).
EMAC II m	^1H-NMR: (300 MHz, CDCl3) δH 2.38 (s, 3H, CH3), 3.30 (dd, 1H, Jab 16.7, Jax 7.8, CH2, isoxazole), 3.71 (dd, 1H, Jab 16.5, Jbx 10.6, CH2, isoxazole), 3.80 (s, 3H, OCH3), 5.66 (dd, 1H, Jbx 10.5, Jax 8.6, CH, isoxazole), 6.89 (d, 2H, J 8.3, CH, arom.), 7.21 (d, 2H, J 8.2, CH, 4-CH3-phenyl), 7.31 (d, 2H, J 8.7, CH, arom.), 7.58 (d, 2H, J 8.2, CH, 4-CH3-phenyl).

Scheme 1. Synthesis of EMAC II (i–m) compounds. Reagents and conditions: (i) ethanol, NaOH; (ii) ethanol, hydroxylamine hydrochloride, potassium hydroxide, reflux.

Figure 2. An ORTEP view of the molecular structure of (R)-(−)-EMAC II i enantiomer.

Figure 3. Effect of Fe²⁺ or Fe³⁺ ions on the spectrum of absorbance of EMAC II l. UV/vis spectrum was measured with 100 μM of compound alone (unbroken line) or in the presence of 10 mM FeCl₃ or FeSO₄ (dotted line).

Table 3. Inhibitory activities towards hMAO-A and hMAO-B of EMAC II (i–m) derivatives.

Compound	Structure	MAO-A (IC50)	MAO-B (IC50)	Ratio^e
EMAC II i		^b	104.04 ± 3.69 nM	>961^d
EMAC II j		^b	41.05 ± 1.52 nM	>2436^d
EMAC II k		^b	320.22 ± 13.61 nM	>312^d
EMAC II l		^b	11.97 ± 0.37 nM	>8354^d
EMAC II m		^b	449.57 ± 18.02 nM	>222^d
Clorgiline		4.46 ± 0.32 nM^c	61.35 ± 1.13 μM	0.000073
l-Deprenyl		67.25 ± 1.02 μM^c	19.60 ± 0.86 nM	3431.12
Iproniazide		6.56 ± 0.76 μM	7.54 ± 0.36 μM	0.87
Moclobemide		361.38 ± 19.37 μM	^a	<0.36^e

^a Inactive at 1 mM (highest concentration tested).

^b Inactive at 100 μM (highest concentration tested). At higher concentration, the compounds precipitate.

^c Results are mean ± SEM from five experiments. Level of statistical significance: ^cP <0.01 versus the corresponding IC₅₀ values obtained against MAO-B, as determined by ANOVA/Dunnett’s.

^d Values obtained under the assumption that the corresponding IC₅₀ against MAO-A is the highest concentration tested (100 μM).

^e Selectivity ratios [IC₅₀(MAO-A)]/[IC₅₀(MAO-B)].

Table 4. Inhibitory activities towards hMAO-A and hMAO-B of EMAC II (i–m) single enantiomers.

Compound	MAO-A (IC50)	MAO-B (IC50)	Ratio
R-(−) EMAC II i	^a	15.27 ± 0.95 nM	6549^b
S-(+) EMAC II i	^a	^a
R-(−) EMAC II j	^a	2.55 ± 0.11 nM	39 216^b
S-(+) EMAC II j	^a	31.37 ± 1.67 μM	3188^b
R-(−) EMAC II k	^a	299.86 ± 12.04 nM	333.5^b
S-(+) EMAC II k	^a	33.82 ± 1.12 μM	2956^b
R-(−) EMAC II l	^a	1.89 ± 0.16 nM	52 910^b
S-(+) EMAC II l	^a	10.96 ± 0.83 μM	9124
R-(−) EMAC II m	^a	126.16 ± 8.44 nM	792.6^b
S-(+) EMAC II m	^a	291.82 ± 13.45 nM	342.6

IC₅₀ values are the mean ± SEM from five experiments.

^a Inactive at 100 μM (highest concentration tested).

^bValues obtained under the assumption that the corresponding IC₅₀ against MAO-B is the highest concentration tested (100 μM).

Figure 4. Putative binding mode of compound EMAC II l. (a) R-(−)-EMAC II l/MAO-B complex; (c) S-(+)-EMAC II l/MAO-B complex; (b, d) compound 2D representation and binding pocket interacting residues: green, hydrophobic; cyan, polar residues. Green arrows indicate π−π interactions.

Figure 5. Binding mode of previously published compounds and approved inhibitors. (a) 4f-(R) MAO-B complex21; (c) safinamide MAO-B complex (pdb code 4v5z)25; (b, d) compound 2D representation and binding pocket interacting residues: in green, hydrophobic; cyan, polar residues. Green arrows indicate π−π interactions, magenta arrow indicates H bonds.

Distinto S, Meleddu R, Yanez M, et al. Drug design, synthesis, in vitro and in silico evaluation of selective monoaminoxidase B inhibitors based on 3-acetyl-2-dichlorophenyl-5-aryl-2,3-dihydro-1,3,4-oxadiazole chemical scaffold. Eur J Med Chem 2016;108:542–52.

 PubMed Web of Science ®Google Scholar

Binda C, Wang J, Pisani L, et al. Structures of human monoamine oxidase B complexes with selective noncovalent inhibitors: safinamide and coumarin analogs. J Med Chem 2007;50:5848–52.

 PubMed Web of Science ®Google Scholar