Identification and Mechanistic Understanding of Dihydroorotate Dehydrogenase Point Mutations in Plasmodium falciparum that Confer in Vitro Resistance to the Clinical Candidate DSM265

John White, Satish K. Dhingra, Xiaoyi Deng, Farah El Mazouni, Marcus C.S. Lee, Gustavo A. Afanador, Aloysus Lawong, Diana R. Tomchick, Caroline L. Ng, Jade Bath, Pradipsinh K. Rathod, David A. Fidock, Margaret A. Phillips

Research output: Contribution to journalArticle

7 Scopus citations

Abstract

Malaria is one of the most challenging human infectious diseases, and both prevention and control have been hindered by the development of Plasmodium falciparum resistance to existing therapies. Several new compounds with novel mechanisms are in clinical development for the treatment of malaria, including DSM265, an inhibitor of Plasmodium dihydroorotate dehydrogenase. To explore the mechanisms by which resistance might develop to DSM265 in the field, we selected for DSM265-resistant P. falciparum parasites in vitro. Any of five different amino acid changes led to reduced efficacy on the parasite and to decreased DSM265 binding to P. falciparum DHODH. The DSM265-resistant parasites retained full sensitivity to atovaquone. All but one of the observed mutations were in the DSM265 binding site, and the remaining C276F was in the adjacent flavin cofactor site. The C276F mutation was previously identified in a recrudescent parasite during a Phase IIa clinical study. We confirmed that this mutation (and the related C276Y) accounted for the full level of observed DSM265 resistance by regenerating the mutation using CRISPR/Cas9 genome editing. X-ray structure analysis of the C276F mutant enzyme showed that conformational changes of nearby residues were required to accommodate the larger F276 residue, which in turn led to a restriction in the size of the DSM265 binding pocket. These findings underscore the importance of developing DSM265 as part of a combination therapy with other agents for successful use against malaria.

Original languageEnglish (US)
Pages (from-to)90-101
Number of pages12
JournalACS infectious diseases
Volume5
Issue number1
DOIs
StatePublished - Nov 1 2019

Keywords

  • Plasmodium
  • dihydroorotate dehydrogenase
  • drug resistance
  • gene editing
  • malaria

ASJC Scopus subject areas

  • Infectious Diseases

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    White, J., Dhingra, S. K., Deng, X., El Mazouni, F., Lee, M. C. S., Afanador, G. A., Lawong, A., Tomchick, D. R., Ng, C. L., Bath, J., Rathod, P. K., Fidock, D. A., & Phillips, M. A. (2019). Identification and Mechanistic Understanding of Dihydroorotate Dehydrogenase Point Mutations in Plasmodium falciparum that Confer in Vitro Resistance to the Clinical Candidate DSM265. ACS infectious diseases, 5(1), 90-101. https://doi.org/10.1021/acsinfecdis.8b00211