Novel chemical scaffolds for inhibition of rifamycin-resistant RNA polymerase discovered from high-throughput screening

Nathan T. Scharf, Vadim Molodtsov, Arrin Kontos, Katsuhiko S. Murakami, George A. Garcia

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

Rifampin has been a cornerstone of tuberculosis (TB) treatment since its introduction. The rise of multidrug-resistant and extensively drug-resistant TB makes the development of novel therapeutics effective against these strains an urgent need. Site-specific mutations in the target enzyme of rifampin, RNA polymerase (RNAP) comprises the majority (~97%) of rifamycin-resistant (RifR) strains of Mycobacterium tuberculosis (MTB). To identify novel inhibitors of bacterial RNAP, an in vitro plasmid-based transcription assay that uses malachite green (MG) to detect transcribed RNA containing MG aptamers was developed. This assay was optimized in a 384-well plate format and used to screen 150,000 compounds against an Escherichia coli homolog of the most clinically relevant RifR RNAP (βS531L) containing a mutation (β‘V408G) that compensates for the fitness defect of this RifR mutant. Following confirmation and concentration-response studies, 10 compounds were identified with similar in vitro inhibition values across a panel of wild-type and RifR E. coli and MTB RNAPs. Four compounds identified from the screen are active against MTB in culture at concentrations below 200 µ\M. Initial follow-up has resulted in the elimination of one scaffold due to potential pan-assay interference.

Original languageEnglish (US)
Pages (from-to)287-297
Number of pages11
JournalSLAS Discovery
Volume22
Issue number3
DOIs
StatePublished - Jan 1 2017

Fingerprint

DNA-Directed RNA Polymerases
Scaffolds
Screening
Mycobacterium tuberculosis
Throughput
Assays
Rifampin
Escherichia coli
Extensively Drug-Resistant Tuberculosis
Bacterial RNA
Mutation
Transcription
Tuberculosis
Plasmids
RNA
Defects
rifamycin SV
Enzymes
Pharmaceutical Preparations
malachite green

All Science Journal Classification (ASJC) codes

  • Biotechnology
  • Analytical Chemistry
  • Biochemistry
  • Molecular Medicine

Cite this

Scharf, Nathan T. ; Molodtsov, Vadim ; Kontos, Arrin ; Murakami, Katsuhiko S. ; Garcia, George A. / Novel chemical scaffolds for inhibition of rifamycin-resistant RNA polymerase discovered from high-throughput screening. In: SLAS Discovery. 2017 ; Vol. 22, No. 3. pp. 287-297.
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Novel chemical scaffolds for inhibition of rifamycin-resistant RNA polymerase discovered from high-throughput screening. / Scharf, Nathan T.; Molodtsov, Vadim; Kontos, Arrin; Murakami, Katsuhiko S.; Garcia, George A.

In: SLAS Discovery, Vol. 22, No. 3, 01.01.2017, p. 287-297.

Research output: Contribution to journalArticle

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