Identification of Antipneumococcal Molecules Effective Against Different Streptococcus pneumoniae Serotypes Using a Resazurin-Based High-Throughput Screen

Assay Drug Dev Technol. 2017 Jul;15(5):198-209. doi: 10.1089/adt.2017.789.

Abstract

Streptococcus pneumoniae is a major human pathogen, causing around 1.6 million deaths worldwide each year. By optimizing a resazurin-based assay to detect S. pneumoniae growth in 384-well microplates, we developed a new high-throughput screening (HTS) system for the discovery of antipneumococcal molecules, which was unsuccessful using conventional absorbance measurements. Before applying our protocol to a large-scale screen, we validated the system through a pilot screen targeting about 7,800 bioactive molecules using three different S. pneumoniae serotypes. Primary screenings of a further 27,000 synthetic small molecules facilitated the identification of 3-acyl-2-phenylamino-1,4-dihydropquinolin-4-one (APDQ) derivatives that inhibited growth of S. pneumoniae with MIC90 values <1 μM (0.03-0.81 μM). Five selected APDQ derivatives were also active against Staphylococcus aureus but neither Klebsiella pneumoniae nor Pseudomonas aeruginosa, suggesting that APDQ may act specifically against Gram-positive bacteria. Our results both validated and demonstrated the utility of the resazurin-based HTS system for the identification of new antipneumococcal molecules. Moreover, the identified new antipneumococcal molecules in this study may have potential to be further developed as new antibiotics.

Keywords: Streptococcus pneumoniae; antipneumococcals; high-throughput screening; resazurin.

MeSH terms

  • Anti-Bacterial Agents / analysis*
  • Anti-Bacterial Agents / pharmacology
  • Dose-Response Relationship, Drug
  • Drug Evaluation, Preclinical / methods
  • High-Throughput Screening Assays / methods*
  • Humans
  • Indicators and Reagents / analysis*
  • Oxazines / analysis*
  • Serogroup*
  • Streptococcus pneumoniae / drug effects
  • Streptococcus pneumoniae / isolation & purification*
  • Xanthenes / analysis*

Substances

  • Anti-Bacterial Agents
  • Indicators and Reagents
  • Oxazines
  • Xanthenes
  • resazurin