Large-scale High-throughput Screening Revealed 5'-(carbonylamino)-2,3'- bithiophene-4'-carboxylate as Novel Template for Antibacterial Agents
Authors: Ivanenkov Yan A. , Yamidanov Renat S. , Osterman Ilya A. , Sergiev Petr V. , Aladinskiy Vladimir A. , Aladinskaya Anastasia V. , Terentiev Victor A. , Veselov Mark S. , Ayginin Andrey A. , Skvortsov Dmitry A. , Komarova Katerina S. , Zagribelnyy Bogdan A. , Baimiev Alexey Kh. , Shvetc Kseniya Yu. , Baimiev Andrey Kh. , Sofronova Alina A. , Machulkin Alexey E. , Petrov Rostislav A. , Zainullina Liana F. , Maximova Marina A. , Zileeva Zulfiya R. , Vakhitova Yulia V. , Bezrukov Dmitry S. , Puchinina Maria M. , Dontsova Olga A. 
Published: June 2019

Background:
The key issue in the development of novel antimicrobials is a rapid expansion of
new bacterial strains resistant to current antibiotics. Indeed, World Health Organization has reported that
bacteria commonly causing infections in hospitals and in the community, e.g. E. Coli, K. pneumoniae and
S. aureus, have high resistance vs the last generations of cephalosporins, carbapenems and fluoroquinolones.
During the past decades, only few successful efforts to develop and launch new antibacterial medications
have been performed. This study aims to identify new class of antibacterial agents using novel
high-throughput screening technique.

Methods:
We have designed library containing 125K compounds not similar in structure (Tanimoto coeff.<
0.7) to that published previously as antibiotics. The HTS platform based on double reporter system
pDualrep2 was used to distinguish between molecules able to block translational machinery or induce
SOS-response in a model E. coli system. MICs for most active chemicals in LB and M9 medium were determined
using broth microdilution assay.

Results:
In an attempt to discover novel classes of antibacterials, we performed HTS of a large-scale small
molecule library using our unique screening platform. This approach permitted us to quickly and robustly
evaluate a lot of compounds as well as to determine the mechanism of action in the case of compounds being
either translational machinery inhibitors or DNA-damaging agents/replication blockers. HTS has resulted
in several new structural classes of molecules exhibiting an attractive antibacterial activity. Herein,
we report as promising antibacterials. Two most active compounds from this series showed MIC value of
1.2 (5) and 1.8 μg/mL (6) and good selectivity index. Compound 6 caused RFP induction and low SOS response.
In vitro luciferase assay has revealed that it is able to slightly inhibit protein biosynthesis. Compound
5 was tested on several archival strains and exhibited slight activity against gram-negative bacteria
and outstanding activity against S. aureus. The key structural requirements for antibacterial potency were
also explored. We found, that the unsubstituted carboxylic group is crucial for antibacterial activity as well
as the presence of bulky hydrophobic substituents at phenyl fragment.

Conclusion:
The obtained results provide a solid background for further characterization of the 5'-
(carbonylamino)-2,3'-bithiophene-4'-carboxylate derivatives discussed herein as new class of antibacterials
and their optimization campaign.