Design, synthesis and molecular docking of new fused 1H-pyrroles, pyrrolo[3,2-d]pyrimidines and pyrrolo[3,2-e][1, 4]diazepine derivatives as potent EGFR/CDK2 inhibitors

Figures & data

Figure 1. FDA approved and reported kinases inhibitors with their essential pharmacophoric features.

Figure 2. ATP binding site of EGFR cavity composed of five main parts^47–49.

Figure 3. Design strategies of Scaffolds A, B and C.

Scheme 1. Construction of compounds 5a–c. Reagents and conditions: (a) (CH₃)₂SO₄, benzene, CH₂(CN)₂, reflux, 6 h; (b) ClCH₂COCl, glacial acetic acid, CH₂COONa, 30–40 °C, 2 h; (c) acetone, K₂CO₃, reflux, 24 h.

Scheme 2. Synthesis of compounds 6a–c, 7a–c and 8a–c. Reagents and conditions: (a) 4-Nitrobenzaldehyde, absolute ethanol, glacial acetic acid; (b) ClCH₂COCl, benzene; r.t.; 48 h; (c) piperidine, NaHCO₃, absolute ethanol, reflux, 8 h.

Scheme 3. Synthesis of compounds 9a–c, 10a–c and 11a–c. Reagents and conditions: (a) Excess CH₃C(OC₂H₅)₃, reflux, 12 h; (b) K₂CO₃, DMF, r.t., 48 h; (c) ClCH₂COOC₂H₅, K₂CO₃, acetone, reflux, 6 h.

Table 1. IC₅₀ values of the new compounds (6a–c, 8a–c, 9a–c, and 11a–c) against Hep3B, HCT116 and MCF-7 cell lines.

Comp.	Scaffold	R	Spacer	R1	IC50 (µM)
					Hep3B	HCT116	MCF-7
6a	A	H	N = CH-		0.219	0.213	0.422
6b	A	CH3	N = CH-		1.956	2.069	0.336
6c	A	Cl	N = CH-		1.470	0.955	2.195
8a	A	H	-NHCOCH2-		0.313	0.408	0.315
8b	A	CH3	-NHCOCH2-		0.049	0.031	0.043
8c	A	Cl	-NHCOCH2-		0.144	0.180	0.294
9a	B	H	–	–	0.487	0.011	0.137
9b	B	CH3	–	–	0.265	0.820	0.644
9c	B	Cl	–	–	0.072	0.009	1.479
11a	C	H	–	–	0.218	0.840	0.154
11b	C	CH3	–	–	0.271	0.303	0.354
11c	C	Cl	–	–	0.179	0.286	0.364
DOX	-	-	-	–	0.005	0.008	0.008

Hep3B, Liver cancer cell line; HCT116, colon cancer cell line; MCF-7, breast cancer cell line; DOX, doxorubicin.

Table 2. Inhibitory activity of compound 8b and imatinib against 20 kinases at 10 µM⁶⁶.

Kinase	% Inhibition		Kinase	% Inhibition
	Imatinib^a	8b^b		Imatinib^a	8b^b
AMPK (A1/B1/G1)	−14%	−18%	EPHA1	−22%	−46%
ALK1	−18%	−21%	FLT1	10%	−8%
ASK1	−5%	−23%	GRK1	−2%	−1%
Aurora A	−38%	−6%	GSK3 alpha	1%	10%
BLK	−3%	−36%	MSK1	−49%	−48%
BRAF	−13%	−77%	NEK1	2%	−8%
CDK2/Cyclin A1	2%	15%	p38 Alpha	−17%	−23%
CK1 Alpha 1	0%	−10%	PDK1	−34%	−28%
DYRK3	10%	23%	PRKG1	−4%	−4%
EGFR	−13%	−70%	SGK1	−3%	−10%

The + ve values indicate the inhibition % and − ve values indicate increase in enzyme activity.

Figure 4. Cell cycle distribution of MCF-7 treated with compound 6a (µM, 72 h: x axis); % cell (y axis).

Figure 5. Cell cycle distribution of MCF-7 treated with compound 8b (µM, 72 h: x axis); % cell (y axis).

Table 3. The binding free energies of the synthesised compounds against EGFR and CDK-2.

Comp.	Binding free energy (kcal/mol)
	EGFR	CDK-2
6a	−18.33	−23.67
6b	−18.97	−23.84
6c	−19.14	−23.60
8a	−20.67	−24.89
8b	−22.46	−24.52
8c	−20.08	−24.20
9a	−17.69	−17.77
9b	−18.90	−17.74
9c	−17.72	−16.71
11a	−21.54	−24.08
11b	−23.30	−24.18
11c	−18.93	−23.98
Erlotinib	−23.49	–
AZD5438	–	−21.66

Figure 6. (A) superimposition of the docked ligand of erlotinib (turquoise) and the original ligand (green) with an RMSD value of 0.88 Å. (B) superimposition of the docked ligand of AZD5438 (pink) and the original ligand (green) with RMSD value of 0.54 Å.

Figure 7. Erlotinib docked into the active site of EGFR.

Figure 8. Compound 8b docked into the active site of EGFR.

Figure 9. Co-crystallised ligand (AZD5438) docked into the active site of CDK-2.

Figure 10. (A) Binding of compound 8b with CDK-2.

Figure 11. (A) Binding of compound 8b with DYRK3.

Figure 12. (A) Binding of compound 8b with GSK3 alpha.

Table 4. Toxicity properties of the synthesised compounds.

Comp.	FDA rodent carcinogenicity (mouse-female)	Carcinogenic potency TD50 (Rat)^a	Rat maximum tolerated dose (feed)^b	Rat oral LD50^b	Rat chronic LOAEL^b	Ocular irritancy	Skin irritancy
6a	Single-Carcinogen	18.143	0.060	0.800	0.157	Irritant	Non-Irritant
6b	Non-Carcinogen	2.356	0.049	2.050	0.096	Irritant	Non-Irritant
6c	Non-Carcinogen	1.839	0.073	0.838	0.063	Irritant	Non-Irritant
8a	Non-Carcinogen	71.541	0.087	1.162	0.071	Irritant	Non-Irritant
8b	Non-Carcinogen	11.813	0.072	3.451	0.054	Irritant	Non-Irritant
8c	Non-Carcinogen	7.264	0.107	1.219	0.029	Irritant	Non-Irritant
9a	Non-Carcinogen	11.774	0.045	0.433	0.163	Irritant	Non-Irritant
9b	Non-Carcinogen	3.458	0.037	0.829	0.137	Irritant	Non-Irritant
9c	Non-Carcinogen	2.160	0.056	0.531	0.073	Irritant	Non-Irritant
11a	Non-Carcinogen	22.882	0.025	1.344	0.080	Irritant	Non-Irritant
11b	Non-Carcinogen	6.639	0.021	2.522	0.067	Irritant	Non-Irritant
11c	Non-Carcinogen	4.081	0.031	1.603	0.035	Irritant	Non-Irritant
Erlotinib	Non-Carcinogen	8.057	0.083	0.662	0.036	Irritant	Non-Irritant

^aUnit: mg/kg body weight/day.

^bUnit: g/kg body weight.

Figure 13. The expected ADMET study.

Table 5. Predicted ADMET profile for the synthesised compounds

Comp.	BBB level^a	Solubility level^b	Absorption level^c	CYP2D6 prediction^d	PPB prediction^e
6a	4	2	1	False	True
6b	4	2	1	False	True
6c	4	2	2	False	True
8a	3	3	0	False	True
8b	3	2	0	False	True
8c	3	2	0	False	True
9a	2	2	0	False	True
9b	2	2	0	False	True
9c	2	2	0	False	True
11a	3	3	0	False	True
11b	3	3	0	False	True
11c	3	2	0	False	True
Erlotinib	1	2	0	False	True

^aBBB level, blood brain barrier level, 0 = very high, 1 = high, 2 = medium, 3 = low, 4 = very low.

^bSolubility level, 1 = very low, 2 = low, 3 = good, 4 = optimal.

^cAbsorption level, 0 = good, 1 = moderate, 2 = poor, 3 = very poor.

^dCYP2D6, cytochrome P2D6, TRUE = inhibitor, FALSE = non inhibitor.

^ePBB, plasma protein binding, FALSE means less than 90%, TRUE means more than 90%.

Elsayed MS, El-Araby ME, Serya RA, et al. Structure-based design and synthesis of novel pseudosaccharine derivatives as antiproliferative agents and kinase inhibitors. Eur J Med Chem 2013;61:122–31.

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