Discovery of a New DNA Gyrase A Inhibitor, 4-[(1-methyl-6-nitroquinolin-1-ium-4-yl)amino]-N-[4- [(1-methylpyridin-1-ium-4-yl)amino]phenyl]benzamide

Incheol Seo; Seong-Il Suh; Min-Ho Suh; Won-Ki Baek

doi:10.4167/jbv.2017.47.4.179

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Seo, Suh, Suh, and Baek: Discovery of a New DNA Gyrase A Inhibitor, 4-[(1-methyl-6-nitroquinolin-1-ium-4-yl)amino]-N-[4- [(1-methylpyridin-1-ium-4-yl)amino]phenyl]benzamide

Original Article

J Bacteriol Virol 2017;47(4):179-188.

Published online: 9 February 2017

DOI: https://doi.org/10.4167/jbv.2017.47.4.179

Discovery of a New DNA Gyrase A Inhibitor, 4-[(1-methyl-6-nitroquinolin-1-ium-4-yl)amino]-N-[4- [(1-methylpyridin-1-ium-4-yl)amino]phenyl]benzamide

Incheol Seo¹, Seong-Il Suh², Min-Ho Suh², Won-Ki Baek^2,^∗

¹Public Health & Welfare Bureau, Daegu Metropolitan City, Daegu, Korea

²Department of Microbiology, School of Medicine, Keimyung University, Daegu, Korea

^∗Corresponding author: Won-Ki Baek, M.D., Ph.D. Department of Microbiology, Keimyung University School of Medicine, 1095 Dalgubeol-daero, Dalseo-Gu, Daegu, 42601, Korea. Phone: +82-53-580-3843, Fax: +82-53-580-3788, e-mail: wonki@dsmc.or.kr

^∗∗ We thank the Developmental Therapeutics Program, National Cancer Institute, USA, for providing us with vialed samples, and the Caenorhabditis Genetics Center for providing a C. elegans strain.

Received 25 September 201725 October 2017 Accepted 27 November 2017

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Escherichia coli (E. coli) is a clinically important causative organism that can lead to urinary tract infections. Quinolone antibiotics are among the first-line treatments for urinary tract infections. However, the frequency of resistance to quinolone in E. coli has been increasing. Therefore, new antimicrobial agents that can be used for treatment in lieu of quinolone antibiotics are needed. In this study, thirty-six compounds with higher scores in a virtual screening based on the three-dimensional structure of E. coli DNA gyrase were selected for in vitro antimicrobial activity testing. An in vitro test confirmed the antimicrobial activity of 4-[(1-methyl-6-nitroquinolin-1-ium-4-yl)amino]-N-[4-[(1-methylpyridin-1-ium-4-yl)amino]phenyl]benzamide (ZINC18057104) against E. coli among the 36 compounds. The minimum inhibitory concentration (MIC) of ZINC18057104 against E. coli ATCC^® 25922 ^TM was 2 μg/ml, and the MIC₅₀ and MIC₉₀ for the 72 quinolone-resistant E. coli clinical isolates were 4 and 64 μg/ml, respectively. ZINC18057104, which has a quinoline structure which is similar to the quinolone antibiotics, is predicted to exhibit antimicrobial activity in quinolone-resistant E. coli because it has different molecular interactions with the DNA gyrase than that of existing quinolone antibiotics.

Keywords: Escherichia coli, DNA gyrase, Anti-bacterial agents, Drug discovery, ZINC18057104

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Figure 1.

The molecular structures of Escherichia coli DNA gyrase. (A) Secondary and tertiary structures of the GyrA homodimer. (B) DNA-gate region of the GyrA homodimer. (C) Hydrophobicity of the DNA-gate region (hydrophobic, red; hydrophilic, white). (D) Cubic box for the virtual screening centered at the DNA-gate region. Green, red, blue, white, and yellow spheres represent carbon, oxygen, nitrogen, hydrogen, and sulfur atoms, respectively.

Figure 2.

Minimum inhibitory concentration (MIC) of ZINC18057104 against Escherichia coli. A broth microdilution test was done according to Clinical and Laboratory Standards Institute's recommendations, using the cation-adjusted Müller-Hinton broth. E. coli ATCC^®25922 ^TM was used for the test. Turbidity was measured after 20 hours of treatment of ZINC18057104, and then the viability of the E. coli was confirmed again by a resazurin reduction assay. Conc.: concentration. Ctrl: control without inoculation.

Figure 3.

ZINC18057104 and its molecular interaction with E. coli DNA gyrase A.(A-B) Chemical structure of ZINC18057104 (C-D) Three-dimensional structure of the E. coli DNA gyrase ZINC18057104 complex. E Hydrophobicity of the ZINC18057104 binding site (hydrophobic, red; hydrophilic, white). (F) Molecular interaction (yellow dotted lines) between the E. coli DNA gyrase (purple) and ZINC18057104.

Figure 4.

Toxicity analyses of ZINC18057104 against Caenorhabditis elegans and human glioblastoma cells. (A) Cox proportional hazards survival analysis of C. elegans treated with ZIN18057104 at different concentrations. Conc^a: concentration (μg/ml). HR^b: hazard ratio (95% confidence interval). (B) Percent viable human cells [measured as average fluorescence intensity ratio (test agent relative to Dimethyl sulfoxide)] for cytotoxicity analysis of ZINC18057104 at 0.0625, 0.125, 0.25, 0.5, 1, 2, 4, 8, 16, and 32 μg/ml against U-87MG and U-251MG cells using the resazurin reduction assay. Dimethyl sulfoxide was used as a negative control to determine a baseline measurement for the cytotoxic impact of each concentration. CC50 is the concentration at which 50% cell survival is obtained. DMSO: dimethyl sulfoxide.

Table 1.

Virtual screening hit list against Escherichia coli DNA gyrase

ZINC ID^a	NSC ID^b	Binding energy (kcal/mol)
04824645	97920	-19.6
13152284	84100	-10.6
18055497	84100	-10.6
01736227	84100	-10.5
01690699	61610	-10.4
04896472	80313	-10.3
01855333	670283	-10.1
01568793	308835	-10.0
18057104	260594	-9.7
12671898	59620	-9.7
13154306	60785	-9.7
29590275	309874	-9.7
00990239	345845	-9.6
01726776	142335	-9.5
04707806	71097	-9.5
17465958	158413	-9.5
01687247	5856	-9.4
01568966	309892	-9.3
01635676	163443	-9.3
05664697	328101	-9.3
01045530	319435	-9.2
01573829	327702	-9.2
01627329	81750	-9.2
01701460	109451	-9.2
02476372	6844	-9.2
04822288	102742	-9.2
01568035	305798	-9.1
01590864	379555	-9.1
04628938	305798	-9.1
04773602	217697	-9.1
04783229	37553	-9.1
01559756	268487	-9.0
01608817	24032	-9.0
01665801	33575	-9.0
05124960	156565	-9.0
08627502	109747	-9.0

^a ZINC ID: the ZINC database identifier.

^b NSC ID: an identifier for substances submitted to the Developmental Therapeutics Program repository at the National Cancer Institute.

Table 2.

The antimicrobial effect of ZINC18057104 on the quinolone-resistant Escherichia coli clinical isolates

No.	MIC	No.	MIC	No.	MIC
1	4	26	4	51	4
2	4	27	4	52	32
3	16	28	4	53	2
4	4	29	4	54	4
5	32	30	16	55	4
6	4	31	8	56	4
7	4	32	16	57	8
8	16	33	4	58	4
9	4	34	4	59	4
10	32	35	4	60	4
11	4	36	4	61	32
12	> 64	37	4	62	4
13	16	38	4	63	4
14	4	39	8	64	2
15	4	40	> 64	65	4
16	64	41	8	66	16
17	4	42	8	67	4
18	> 64	43	16	68	2
19	4	44	4	69	64
20	> 64	45	64	70	4
21	4	46	4	71	2
22	> 64	47	4	72	4
23	4	48	4	MIC₅₀	4
24	2	49	> 64	MIC₉₀	64
25	32	50	4

MIC: minimum inhibitory concentration (μg/ml). MIC₅₀, MIC₉₀: Minimum inhibitory concentration at which 90% and 50% of the isolates were inhibited by ZINC18057104, respectively.

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