In vitro and in silico inhibition performance of choline against CYP1A2, CYP2D6 and CYP3A4

Figures & data

Table 1. Predicted CYP1A2, CYP2D6, and CYP3A4 inhibition potency of choline and the positive controls with the DL-CYP Prediction Server.

Compounds	Predicted inhibition values
	CYP1A2	CYP2D6	CYP3A4
Choline	0	0	0
Ketoconazole	0.08	0.81	1.0
alfa-Naphtoflavone	0.99	0	0.06
Quinidine	0.02	0.98	0.04

Figure 1. Effect of choline on CYP1A2 activity in vitro. Five concentrations of choline and α-naphthoflavone (as a selective inhibitor) were assayed: 0.150, 0.250, 0.500, 0.750, and 1 µmol/L. *p < 0.05; **p < 0.01 vs. control (pure CYP1A2).

Figure 2. Effect of choline on CYP2D6 in vitro. Five concentrations of choline and quinidine (as a selective inhibitor) were assayed: 0.150, 0.250, 0.500, 0.750, and 1 µmol/L. *p < 0.05; **p < 0.01 vs. control (pure CYP2D6).

Figure 3. Effect of choline on CYP3A4 activity in vitro. Five concentrations of choline and ketoconazole (as a selective inhibitor) were assayed: 0.150, 0.250, 0.500, 0.750, and 1 µmol/L. *p < 0.05; **p < 0.01 vs. control (pure CYP3A4).

Table 2. Scoring method, docking scores, participating active residues and distance from the heme structure in docking simulations of choline in the active site of CYP1A2, CYP2D6, and CYP3A4 employing XP, IFD, and MM/GBSA recalculations (Schrödinger Inc.).

CYP isoform	Scoring method	Docking scores		Residues participating in stabilization	Distance from the heme group
		Co-crystallized ligand	Choline
CYP1A2	XP Glide	−10.82	−3.69	Thr124, Phe125, Asp313, Ala317, Asp320, Thr321, Leu382, Ile386, Leu497, Thr498, Hem900	3.29 Å
	IFD	−11.24	−4.60	Thr124, Phe125, Ala317, Thr321, Leu382, Thr385, Ile386, Leu497, Thr498, Hem900	3.05 Å
	MM/GBSA	−89.43	−38.71	Thr124, Phe125, Ala317, Asp320, Thr321, Leu382, Thr385, Ile386, Leu497, Thr498, Hem900	3.53 Å
CYP2D6	XP Glide	−9.89	−5.04	Phe120, Leu121, Glu216, Gln244, Ser304, Ala305, Phe483	8.70 Å
	IFD	−10.29	−5.24	Phe120, Leu121, Glu216, Ser304, Ala305, Val374, Phe483	6.01 Å
	MM/GBSA	−101.94	−58.10	Phe120, Leu121, Glu216, Ser304, Asp301, Ala305, Val374, Phe483	5.76 Å
CYP3A4	XP Glide	−9.11	−4.38	Leu216, Phe304, Ala305, Thr309, Ile369, Ala370, Leu482, Hem601	3.15 Å
	IFD	−10.12	−5.75	Leu216, Ala305, Gly306, Thr309, Ile369, Ala370, Leu482, Hem601	2.64 Å
	MM/GBSA	−115.79	−63.82	Leu216, Ala305, Gly306, Thr309, Ile369, Ala370, Leu482, Hem601	2.81 Å

Figure 4. Visualized major intermolecular interactions of choline in the active sites of CYP1A2 (A, B), CYP2D6 (C, D), and CYP3A4 (E, F) after employing IFD and MM/GBSA recalculations. The interactions are provided in both 2D and 3 D forms. The enzyme structures are depicted in green, and the heme group is given in red colored sticks.