Article
A novel high throughput screening approach targeting the vitamin B6 biosynthesis in Plasmodium falciparum
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Authors
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Markus Wolf
- European ScreeningPort GmbH, Hamburg, Germany
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Ingrid B. Müller
- Biochemical Parasitology, Bernhard Nocht Institute for Tropical Medicine, Hamburg, Germany
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Bernhard Ellinger
- European ScreeningPort GmbH, Hamburg, Germany
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Oliver Keminer
- European ScreeningPort GmbH, Hamburg, Germany
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Philip Gribbon
- European ScreeningPort GmbH, Hamburg, Germany
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Carsten Wrenger
- Unit for Drug Discovery, Department of Parasitology, Institute of Biomedical Science, University of São Paulo, São Paulo, Brasil
| Published: | January 29, 2014 |
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Outline
Text
Pyridoxal phosphate (PLP), the respective active molecule of vitamin B6, is de novo synthesised in Plasmodium falciparum by the PLP-synthase which consists of the enzymes PfPdx1 and PfPdx2 (Wrenger et al., 2005). Due to the lack of this synthase complex in humans the enzymes provide an ideal starting point for medicinal chemistry programs. In a fluorescent medium throughput screen different lead series were identified and their activity was confirmed on cellular level utilising a 96-well 3H-hypoxanthine incorporation assay established at the Wrenger group using P. falciparum cultured in human red blood cells.
These promising data encouraged us to develop a high throughput assay in 384-well plate format. After analysing the assay in terms of stability, variability and DMSO tolerance 256,000 compounds were screened on 9 consecutive days using 945 384-well plates. The median z‘ of the primary screen was 0.72 and the assay yielded 3607 Hits corresponding to a Hit rate of 1.4% (50% activity cut off). Hits were analysed in two consecutive counter screenings leading to active molecules which are now analysed in vivo. The data presented will cover some aspects of the high throughput screening performed at European ScreeningPort.
