Chromone-based monoamine oxidase B inhibitor with potential iron-chelating activity for the treatment of Alzheimer’s disease

Figures & data

Figure 1. Representative dual-targeting anti-AD agents.

Figure 2. Representative dual-targeting anti-AD agents.

Scheme 1. Reagents and conditions: (i) 4-Methoxybenzyl chloride, K₂CO₃, acetone, reflux, 3–8 h. (ii) Ethane-1,2-diamine, Ethanol: H₂O = 1:1, 70 °C, 1.5 h.

Scheme 2. Reagents and conditions: (i) POCl₃, DMF, −10 °C, 15 h. (ii) NH₂SO₃H, NaClO₂, 0 °C, 12 h. (iii) POCl₃, DMF, 3, r.t., 10 h. (iv) BCl₃, anhydrous DCM, −48 °C to r.t., 12 h.

Scheme 3. Reagents and conditions: (i) POCl₃, DMF, −10 °C 15 h. (ii) Acetyl chloride, Triethylamine, anhydrous DCM, 0 °C to r.t., 3 h. (iii) NH₂SO₃H, NaClO₂, 0 °C, 12 h. (iv) Oxalyl chloride, DMF, anhydrous DCM, 3, r.t., 3 h. (v) K₂CO₃, acetonitrile: H₂O = 1:1, reflux, 0.5 h. (vi) corresponding substituted benzyl bromide, K₂CO₃, DMF, 12 h (vi) BCl₃, anhydrous DCM, −48 °C to r.t., 12 h.

Scheme 4. Reagents and conditions: (i) 4-Methoxybenzyl chloride, K₂CO₃, acetone, reflux, 3–8 h. (ii) Ethane-1,2-diamine, Ethanol: H₂O = 1:1, 70 °C, 1.5 h.

Table 1. Physicochemical parameters for compounds 8a–e, 15a–g, and 17a–l.

Comp.	pKa1^a	pKa2^a	Log β1^c	Log β2^c	Log β3^c	pFe^3^+
8a	3.29	9.67	13.67	25.22	34.95	18.68
8b	3.32	9.58	13.74	25.04	34.36	18.35
8c	3.26	9.69	13.86	25.07	33.48	17.19
8d	3.20	9.60	13.82	25.31	34.23	18.18
8e	3.18	9.67	13.67	25.38	35.37	19.08
15a	3.30	9.59	13.77	24.72	32.17	16.45
15b	3.06	9.73	13.64	25.57	34.21	17.80
15c	3.19	9.34	13.40	23.16	31.01	15.87
15d	3.36	9.52	13.50	25.31	34.85	19.01
15e	2.89	9.57	13.24	24.65	34.50	18.52
15f	3.35	9.52	13.79	25.03	33.91	18.09
15g	3.25	9.56	13.82	24.02	31.70	16.00
17a	2.81	9.80	13.46	25.19	35.28	18.62
17b	2.95	9.82	13.58	25.38	35.34	18.61
17c	2.75	9.76	13.31	25.00	34.07	17.55
17d	2.86	9.78	13.50	25.21	35.13	18.52
17e	2.76	9.83	13.32	25.08	34.89	18.12
17f	2.73	9.69	13.20	24.70	33.77	17.45
17g	2.68	9.62	13.02	24.75	33.78	17.66
17h	2.56	9.59	12.87	24.73	33.36	17.34
17i	2.58	9.68	12.93	24.99	34.10	17.81
17j	2.76	9.53	13.13	24.78	33.73	17.89
17k	2.53	9.43	12.99	24.76	33.88	18.31
17l	2.61	9.47	13.02	24.99	34.39	18.70
DFP	3.65	9.78	13.96	26.43	36.36	19.75
DFP^b	3.61	9.78	15.03	27.42	37.35	20.74
DFP^a	3.65	9.78	14.67	27.58	36.90	20.29

^aThe compounds were measured in 0.1 M KCl.

^bThe data from the reference tested in a 0.1 M KCl solution³¹.

^cThe compounds were tested in DMSO: KCl (0.1 M) = 1:1.5 (v/v) to address the solubility issue.

Figure 3. Rational design of chromone-pyridinone hybrids as potential anti-AD agents.

Figure 4. The pH-dependent UV spectra of compound 17d. (A) The pH-dependence of the spectrum of compound 17d in the presence of Fe³⁺ over the pH range 0.8–2.1 in 0.1 M KCl at 25 °C, [Fe³⁺] = 1.0 μM, [17d] = 1.1 μM. (B) The pH-dependence of the spectrum of compound 17d in the presence of Fe³⁺ over the pH range 2.1 and 9.0 in 0.1 M KCl: DMSO = 3:2 (v/v) at 25 °C, [Fe³⁺] = 1.0 μM, [17d] = 5.0 μM. C. Speciation plot of Fe³⁺/17d as measured by the percentage formation relative to [Fe³⁺]_total as a function of pH. This plot was calculated from the affinity constants reported in Table 1 and the Fe³⁺ hydrolysis constants were as follows: FeOH = –2.563, Fe (OH)₂ = –6.205, Fe (OH)₃ = –15.100, Fe₂ (OH)₂ = –2.843, Fe₃ (OH)₄ = –6.059, and Fe (OH)₄ = –21.883.

Figure 5. Kinetic study and cytotoxicity assay of compound 17d. Mode of MAO-B inhibition saturation curves (A) and Lineweaver − Burk plot (B) of the inhibition of MAO-B enzyme by different concentrations of 17d (0, 33.3, 100, and 300 nM) in the presence of p-tyramine (0.05, 0.1, 0.25, 0.5, 1.0, and 1.25 mM) as a substrate. The Cytotoxicity effect of compound 17d on PC-12 cells (C).

Table 2. The hMAO-B inhibitory activities of compounds.

Comp.	R	Inhibitory rate (%, 100 nM)	Comp.	R	Inhibitory rate (%, 100 nM)
8a	H-	42.20 ± 3.7	17a	H-	53.11 ± 1.6
8b	7-methyl-	35.79 ± 0.2	17b	7-methyl-	49.50 ± 0.3
8c	6-methyl-	42.10 ± 0.8	17c	6-methyl-	44.23 ± 0.1
8d	7-methoxy-	41.87 ± 1.4	17d	7-methoxy-	48.88 ± 1.4
8e	6-methoxy-	34.28 ± 2.2	17e	6-methoxy-	46.40 ± 1.3
15a	H-	38.95 ± 1.1	17f	H-	49.00 ± 0.6
15b	3-methyl-	39.13 ± 1.1	17g	3-methyl-	55.75 ± 2.7
15c	4-methyl-	38.70 ± 0.5	17h	4-methyl-	30.10 ± 3.7
15d	3-fluoro-	38.48 ± 1.4	17i	3-fluoro-	39.48 ± 0.3
15e	4-fluoro-	42.41 ± 2.1	17j	4-fluoro-	45.95 ± 0.7
15f	3-chloro-	41.19 ± 2.8	17k	3-chloro-	43.06 ± 2.0
15 g	4-chloro-	39.07 ± 0.5	17l	4-chloro-	61.43 ± 3.4
Pargyline		48.93 ± 1.7

Table 3. IC₅₀ and SI for the chromone derivatives on the enzyme activity of MAO isoforms.

Comp.	IC50 (nM)
	hMAO-B	hMAO-A	SI^a
8a	112.00 ± 1.3	–^b	>89
8c	111.75 ± 5.7	–^b	>89
8d	107.25 ± 5.6	–^b	>93
15e	106.35 ± 0.2	–^b	>94
15f	129.50 ± 0.8	–^b	>80
17a	67.27 ± 1.9	2063 ± 35.4	30.7
17b	82.78 ± 0.7	–^b	>120
17c	86.30 ± 0.7	–^b	>116
17d	67.02 ± 4.3	739 ± 21.9	11.0
17e	88.66 ± 5.3	–^b	>112
17f	76.45 ± 3.8	795 ± 0.7	10.4
17g	78.00 ± 4.3	1666 ± 5.7	21.4
17j	101.66 ± 3.2	–^b	>99
17k	113.90 ± 3.8	–^b	>88
17l	75.56 ± 3.1	1312 ± 13.4	17.4
Pargyline	111.30 ± 0.6	4184 ± 7.1	37.6

^aSI: hMAO-B selectivity index = IC₅₀ (hMAO-A)/IC₅₀ (hMAO-B). ^bInhibitory rate less than 20% at 10 μM (highest concentration tested).

Table 4. Predicted drug-like properties and PAMPA-BBB results for 17a and 17d.

Compound	MW^b	HBD^b	HBA^b	LogP^b	Pe ± SEM (*10^−6 cm/s)^a	CNS^c	BBB permeability^b
17a	326.35	2	6	1.76	1.22 ± 0.22	–	+
17d	356.38	2	7	1.77	2.73 ± 0.07	±	+

^aData are the mean ± SEM of eight independent experiments.

^bThese results were predicted via online programs (https://admet.scbdd.com).

^cCNS (+) (high BBB permeation predicted); P_e (× 10⁻⁶cm s^{− 1}) > 4.0; CNS (±) (BBB permeation uncertain); P_e (× 10⁻⁶cm s⁻¹) from 2.0 to 4.0; CNS (−) (low BBB permeation predicted); P_e (× 10⁻⁶cm s⁻¹) <2.0.

Figure 6. Best docking poses of compound 17d at the active site of MAO-B. The most relevant interacting residues are presented in green carbons polytube, FAD cofactor is shown as grey carbons polytube and ligands as yellow carbons polytube.

Figure 7. Pargyline (15 mg/kg), Memantine (15 mg/kg), and compound 17d (15 mg/kg) were evaluated for scopolamine-induced (15 mg/kg) memory impairment in ICR mice in the Morris water maze. The mouse trajectories of the mice are shown as the control (D–1), model (D–2), pargyline (D–3), Memantine (D–4), and 17d (D–5) groups. Data are presented as the mean ± SEM (n = 15; ^#p < 0.05, ^##p < 0.01, ^###p < 0.001, and ^####p < 0.0001, Control group vs. scopolamine model group; *p < 0.05, **p < 0.01, ***p < 0.001, and ^****p < 0.0001, Pargyline group, Memantine group, 17d vs. scopolamine model group).

Table 5. Effects of compound 17d (15 mg/kg) on scopolamine-induced memory impairment in ICR mice evaluated by the Morris water maze test.

Group	Entries to target	Latency to first entry	Distance to first entry
Control	6.1 ± 0.6	8 ± 1	2.0 ± 0.3
Model	2.1 ± 0.3^####	42 ± 6^####	8.9 ± 1.2^####
17d	3.3 ± 0.3*	14 ± 1***	3.3 ± 0.3***
Pargyline	5.3 ± 0.9**	11 ± 1****	2.6 ± 0.3****
Memantine	4.0 ± 0.3****	13 ± 1***	3.2 ± 0 .2***

Pargyline (15 mg/kg) and Memantine (15 mg/kg) were used as reference.

Data are presented as the mean ± SEM (n = 15; ^####p < 0.0001, Control group vs. scopolamine model group; *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001, Pargyline group, Memantine group, 17d vs. scopolamine model group).

Figure 8. The pH-dependent UV spectra of compound 17d. (A) The pH-dependence of the titration spectrum of compound 17d. (B) The pK_a values of compound 17d over the pH range of 1.3–11.0 in 0.1 M KCl at 25 °C.

Table 6. Pharmacokinetic parameters of 17d after its administration^a.

Administration	i.v. (2.34 mg/kg)	i.g. (15 mg/kg)
AUC0–24 (ng/ml*h)	745.23 ± 88.38	439.36 ± 59.60
AUC0–∞ (n /ml*h)	748.74 ± 87.10	443.82 ± 65.33
MRT0–∞ (h)	0.25 ± 0.08	0.81 ± 0.09
t1/2 (h)	0.32 ± 0.00	0.51 ± 0.18
CL (L/h/kg)	3.15 ± 0.37	0.03 ± 0.01
V (L/kg)	1.46 ± 0.19	0.02 ± 0.01
Cmax (ng/ml)	2153.83 ± 636.73	389.51 ± 50.09
Tmax (h)	0.083 ± 0	0.25 ± 0.00
F (%)	–	9.25%
Brain 17d content (ng/g)^b	–	6.02 ± 0.08

^aValues are expressed as the mean ± SD (n = 3).

^bData collected from C_max time point.

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