New pyrazolyl-thiazolidinone/thiazole derivatives as celecoxib/dasatinib analogues with selective COX-2, HER-2 and EGFR inhibitory effects: design, synthesis, anti-inflammatory/anti-proliferative activities, apoptosis, molecular modelling and ADME studies

Figures & data

Figure 1. Structure of reversible EGFR inhibitor Erlotinib (1), irreversible EGFR inhibitor Dacamitinib (2), Dasatinib (3), Dabrafenib (4) EGFR inhibitor Pyrazolyl-thiazole derivatives (5a), (5b), EGFR inhibitor hydrazonoyl-thiazole derivative (6), EGFR inhibitor pyrazolyl-thiazolidinone derivative (7), HER2 and EGFR inhibitor Pyrazolyl-thiazole (8), selective COX-2 inhibitors Celecoxib (9), thiazole containing NSAID Fentiazac (10a), thiazole containing NSAID Sudoxicam (10b) and Meloxicam (10c).

Figure 2. Structural hyperdization of designed compounds from EGFR inhibitor Dasatinib (3), reported EGFR inhibitor Pyrazolyl-thiazole (5a,b), reported EGFR inhibitor Hydrazono-thiazole (6), reported EGFR inhibitor pyrazolyl-thiazolidinone, reported HER2 and EGFR inhibitor Pyrazolyl-thiazole (8), COX-2 inhibitors Celecoxib (9) and Meloxicam (10c).

Table 1. Cell cycle analysis of compounds 18d and control for breast MCF7 cells.

Cell cycle phase	%G0–G1	%S	%G2-M
18d	62.43	33.18	4.39
Cont. MCF7	59.61	27.28	13.11

Table 2. Apoptosis of compounds 18d and control against breast MCF7 cell line.

	Total	Early	Late	Necrosis
18d	46.55	12.58	27.45	6.52
Cont. MCF7	2.47	0.55	0.16	1.76

Scheme 1. Reagents and conditions: (a) ethanol 95% reflux 12 h, (b) POCl₃/DMF reflux 24 h, (c) thiosemicarbazide, ethanol 95%, reflux 10 h, (d) ethanol 95%, sod. acetate, reflux, 24 h, (e) ethanol 95%, pyridine, reflux, 24 h.

Table 3. In vitro COX-1 and COX-2 inhibitory activity of thiazole 16a,b, 18a–j and reference drugs indomethacin, celecoxib.

Compound	COX-1 IC50 (μM)^a ± S.E.	COX-2 IC50 (μM)^a ± S.E.	COX-2 S.I.^b
16a	46.98 ± 2.01	0.349 ± 0.01	134.6
16b	19.72 ± 0.84	0.756 ± 0.03	26.08
18a	34.55 ± 1.48	7.667 ± 0.3	4.506
18b	49.64 ± 2.12	11.34 ± 0.45	4.377
18c	62.68 ± 2.68	18.64 ± 0.74	3.363
18d	23.21 ± 0.99	8.828 ± 0.35	2.629
18e	27.47 ± 1.18	3.006 ± 0.12	9.138
18f	34.42 ± 1.47	0.817 ± 0.03	42.13
18g	68.2 ± 2.92	27.76 ± 1.1	2.457
18h	38.52 ± 1.65	15.52 ± 0.61	2.482
18i	9.417 ± 0.4	5.824 ± 0.23	1.617
18j	24.5 ± 1.05	11.46 ± 0.45	2.138
Celecoxib	16.5 ± 0.71	0.685 ± 0.03	24.09
Indomethacin	0.196 ± 0.01	0.399 ± 0.02	0.491

^aThe concentration of test compound produce 50% inhibition of COX-1, COX-2 enzyme.

^bThe in vitro COX-2 selectivity index (COX-1/COX-2).

Figure 3. (a) 2D NOESY H¹NMR of compounds 18a showing correlation between the pyrazole H-5 and olefinic proton of each geometrical isomer, (b) 2D NOESY H¹NMR of compounds 18d showing correlation between the pyrazole H-5 and olefinic proton of each geometrical isomer, (c) 2D NOESY H¹NMR of compounds 18g showing correlation between the pyrazole H-5 and olefinic proton of each geometrical isomer.

Table 4. In vitro MCF-7, A549, F180 inhibitory activity of thiazole containing derivatives 16a,b & 18a–j and reference drugs dasatinib & doxorubicin.

Compound	MCF7^a	A549^a	F180^a	S.I. (MCF7)	S.I. (A549)
16a	0.73 ± 0.03	1.64 ± 0.06	24.2 ± 0.89	33.15	14.75
16b	6.25 ± 0.23	3.44 ± 0.12	44.6 ± 1.63	7.13	12.96
18a	12.7 ± 0.47	23.2 ± 0.81	48.7 ± 1.78	3.83	2.09
18b	36 ± 1.32	57.6 ± 2.01	99.5 ± 3.64	2.76	1.72
18c	3.18 ± 0.12	14.3 ± 0.5	59.6 ± 2.18	18.72	4.16
18d	2.15 ± 0.08	4.03 ± 0.14	28.5 ± 1.04	13.25	7.07
18e	8.26 ± 0.3	15.3 ± 0.53	73.7 ± 2.7	8.92	4.81
18f	2.85 ± 0.1	1.25 ± 0.04	23.6 ± 0.86	8.28	18.88
18g	19.9 ± 0.73	28.4 ± 0.99	36.6 ± 1.34	1.83	1.28
18h	10.1 ± 0.39	22.2 ± 0.77	70.6 ± 2.59	6.99	3.18
18i	16.7 ± 0.61	9.41 ± 0.33	59.8 ± 2.19	3.58	6.35
18j	35.7 ± 1.31	68.7 ± 2.4	71.8 ± 2.63	2.01	1.04
Dasatinib	7.99 ± 0.29	11.8 ± 0.41	32.2 ± 1.18	4.03	2.72
Doxorubicin	3.1 ± 0.11	2.42 ± 0.08	9.39 ± 0.34	3.02	3.88

^aThe concentration of test compound produce 50% inhibition of MCF-7, A549, F180 cell lines, the result is the mean of six values.

Figure 4. IC₅₀ of compounds 16a, b & 18a–j against of MCF-7, A549, F180 cell lines and reference drugs dasatinib & doxorubicin.

Figure 5. IC₅₀ values of compounds 16a, 18c, 18d and 18f compared to dasatinib on EGFR-WT, EGFR (L858R), EGFR (T790M/L858R).

Figure 6. IC₅₀ values of compounds 16a, 18c, 18d, and 18f compared to that of dasatinib on HER-2.

Figure 7. SAR of in vitro anti-inflammatory and cytotoxic activities of compounds 16a,b, and 18a–j.

Figure 8. Compounds 18d effect on DNA-ploidy flow cytometric analysis of breast MCF-7 cells compared to negative control.

Figure 9. The percentage of Annexin-V-FITC-positive staining in breast MCF-7 cells when treated with compounds 18d compared to negative control.

Figure 10. (A) Visual representation (2D) of celecoxib docked with 3LN1 active site (B) Visual representation (2D) of compound 16a docked with 3LN1active site, (C) Visual representation (2D) of compound 16b docked with 3LN1 active site (D) Visual representation (2D) of compound 18c docked with 3LN1 active site (E) Visual representation (2D) of compound 18f docked with 3LN1 active site (F) Visual representation (2D) of compound 18d docked with 3LN1 active site.

Figure 11. (A) Visual representation (2D) of compound 16a docked with 1M17 active site, (B) Visual representation (2D) of compound 16b docked with 1M17 active site (C) Visual representation (2D) of compound 18c docked with 1M17 active site. (D) Visual representation (2D) of compound 18f docked with 1M17 active site (E) Visual representation (2D) of compound 18d docked with 1M17 active site (F) Visual representation (2D) of dasatinib docked with 1M17 active site.

Figure 12. (A) Visual representation (2D) of 16a docked with HER-2 active site (3PP0) (B) Visual representation (2D) of compound 16b docked with HER-2 active site (3PP0), (C) Visual representation (2D) of compound 18c docked with HER-2 active site (3PP0), (D)Visual representation (2D) of compound 18d docked with HER-2 active site (3PP0),(E) Visual representation (2D) of compound 18f docked with HER-2 active site (3PP0) (E) Visual representation (2D) of compound dasatinib docked with HER-2 active site (3PP0).

Table 8. Physicochemical parameters of compounds 16a, 16b, 18c, 18d, 18f and dsastinib.

Compound No	M.wt	No of H-bond acceptors	No of H-bond donors	No of rotoable bonds	Molar refractivity	TPSA
16a	453.54	6	1	5	126.21	139.46
16b	495.02	6	1	7	140.63	139.46
18c	608.53	6	1	8	158.36	138.32
18d	574.63	8	1	8	154.67	184.14
18f	600.71	7	1	9	167.57	174.91
Dasatinib	491.97	7	3	8	133.62	134.75

Table 9. Pharmacokinetics of compounds 16a, 16b, 18c, 18d, 18f and dsastinib.

Compound No	GI absorption	BBB permeant	CYP1A2 inhibitor	CYP2C19 inhibitor	CYP2C9 inhibitor
16a	High	No	No	Yes	Yes
16b	Low	No	No	Yes	Yes
18c	Low	No	No	No	No
18d	Low	No	No	No	No
18f	Low	No	No	No	No
Dasatinib	High	No	No	Yes	Yes

Table 10. Lipophilicity and water solubility of compounds 16a, 16b, 18c, 18d, 18f and dsastinib.

Compound No	Logpo/w (ilogP)	Log S (ESOl)
16a	3.15	−4.84
16b	3.39	−5.81
18c	4.47	−7.03
18d	3.39	−6.40
18f	3.39	−6.40
Dasatinib	3.15	−4.84

Table 11. Drug likeness of compounds 16a, 16b, 18c, 18d, 18f and dsastinib.

Compound No	Lipinski	Bioavailability score
16a	0 violations	0.55
16b	0 violations	0.55
18c	1 violations	0.55
18d	2 violations	0.17
18f	2 violations	0.17
Dasatinib	0 violations	0.55

Figure 14. Structure activity relationship of synthesised compounds 16a, b and 18a–j.

Figure 13. The physicochemical diagram for the oral bioavailiability of compounds 16a, 16b, 18c, 18d, 18f and dasatinib.