Evaluation of N-alkyl isatins and indoles as acetylcholinesterase and butyrylcholinesterase inhibitors

Figures & data

Scheme 1. Synthesis of N-alkyl isatins 4a-j and N-alkyl indoles 5a-j.

Figure 1. Structures of isatin, indole, and previously investigated isatin/indole-based AChEI and BChEI.

Table 1. Inhibition of AChE and BChE by N-alkyl isatins 4a-j and indoles 5a-j.

Compound	n	AChE IC50 (µM)^a	BChE IC50 (µM)^a	Selectivity^b	BBB permeability^c
4a	0	647 ± 57	> 1000	< 0.6	1.16 (moderate)
4b	1	297 ± 35	193 ± 8	1.5	1.48 (moderate)
4c	2	519 ± 11	257 ± 8	2.0	2.00 (moderate)
4d	3	540 ± 23^c	73.7 ± 1.3	7.3	2.38 (high)
4e	4	197 ± 51	83.8 ± 3.4	2.4	2.55 (high)
4f	5	> 125	90.8 ± 2.3	> 1.4	0.25 (moderate)
4g	6	67.8 ± 7.8	20.7 ± 3.3	3.3	0.39 (moderate)
4h	7	> 125	17.9 ± 1.8	> 7.0	0.54 (moderate)
4i	8	82.1 ± 8.1	3.77 ± 0.45	21.8	0.49 (moderate)
4j	9	62.2 ± 8.5	5.16 ± 0.66	12.1	0.39 (moderate)
5a	0	86.7 ± 4.5	> 125	< 0.7	1.65 (moderate)
5b	1	101 ± 6	> 125	< 0.8	1.67 (moderate)
5c	2	78.6 ± 2.3	> 125	< 0.6	1.96 (moderate)
5d	3	> 125	> 125	–	2.56 (high)
5e	4	95.0 ± 5.9	> 125	< 0.8	3.58 (high)
5f	5	76.2 ± 1.6	> 125	< 0.6	6.60 (high)
5g	6	35.0 ± 10.5	> 62.5	< 0.6	9.04 (high)
5h	7	49.5 ± 1.8	> 62.5	< 0.8	11.00 (high)
5i	8	44.6 ± 1.7	> 62.5	< 0.7	13.01 (high)
5j	9	48.6 ± 1.4	> 62.5	< 0.8	19.26 (high)
isatin	–	> 1000^d	> 1000^d	–	0.67 (moderate)
rivastigmine	–	275 ± 25^d	1.14 ± 0.13^d	241	1.16 (moderate)
tacrine	–	0.00187 ± 0.00002^d	0.0128 ± 0.0012^d	0.14	0.87 (moderate)

^a IC₅₀ values are reported as the mean of three independent experiments ± SEM. AChE was from Electrophorus electricus. BChE was from equine serum. For compounds with limited solubility under the assay conditions, the IC₅₀ is given as > X, where X = the highest soluble concentration tested.

^b Selectivity was calculated as AChE IC₅₀/BChE IC₅₀.

^c Predicted using the PreADMET application (https://preadmet.qsarhub.com/.) where low BBB permeability < 0.1, moderate BBB permeability 0.1–2.0, high BBB permeability > 2.0.

^d Values previously determined^18,19.

Figure 2. Lineweaver-Burk plot showing non-competitive inhibition of AChE with respect to acetylthiocholine (ATC) for compound 4i.

Figure 3. Lineweaver-Burk plot showing mixed inhibition of BChE with respect to butyrylthiocholine (BTC) for compound 4i.

Figure 4. Docking of TcAChE (PDB: 1ACJ; A and C) and hBChE (PDB: 4BDS; B and D) with 1-nonylindoline-2,3-dione (4i, cyan) and 1-nonyl-1H-indole (5i, yellow). Active site residues of TcAChE include the catalytic triad (Ser200, His440, Glu327; gold), anionic site (Trp84, Tyr130, Phe330, and Phe331; green), oxyanion hole (Gly118, Gly119, Ala201; purple), acyl pocket (Phe288, Phe290; blue), and PAS (Tyr70, Asp72, Tyr121, Ser122, Trp279, Tyr334; magenta). Active site residues of hBChE include the catalytic triad (Ser198, His438, Glu325; gold), anionic site (Trp82, Tyr128, Phe329; green), oxyanion hole (Gly116, Gly117, Ala199; purple), acyl pocket (Trp231, Leu286, Val288; blue), and PAS (Asp70, Tyr332; magenta). Orange dashes indicate π-π stacking interactions, and yellow dashes indicate H-bonding interactions. All distances shown are in Å. O, N, and H atoms are shown in red, blue, and white, respectively.

Reiland KM, Eckroat TJ. Selective butyrylcholinesterase inhibition by isatin dimers and 3-indolyl-3-hydroxy-2-oxindole dimers. Bioorg Med Chem Lett. 2022;77:129037.

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Davis SM, Eckroat TJ. Isatin-linked 4,4-dimethyl-5-methylene-4,5-dihydrothiazole-2-thiols for inhibition of acetylcholinesterase. Med Chem Res. 2021;30(12):2289–2300.

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Data availability statement

The data that support the findings of this study are available from the corresponding author, TJE, upon reasonable request.