CCG-1423 The data from drug screening in P

The data from drug screening in P. berghei (Fig. 3D) provide proof of concept that chemogenomic profiling can be accomplished in P. berghei in the same manner as P. falciparum, in that significant increases in the EC50 ratio were found only for antifolate drugs in the parasite with attenuated TgDHFR-TS expression. That increased antifolate sensitivity under this condition was observed despite the presence of endogenous PbDHFR-TS indicates that chemogenomic profiling is not limited to parasites with modification of endogenous drug target only. The specificity of the approach is highlighted by the insignificant EC50 ratio for SDX, a drug targeting dihydropteroate synthase that acts immediately upstream of DHFR-TS in the folate/pyrimidine metabolic pathway (Triglia et al., 1998). The significant EC50 ratio for PGN suggests that this CCG-1423 targets DHFR-TS, perhaps because conversion to the antifolate drug CYC is manifest in the P. berghei in vitro system, possibly by leukocytes. The low EC50 ratio of PGN compared with antifolate drugs, however, suggests that conversion to CYC occurs at a low level, or PGN has other significant targets. The insignificant EC50 ratio for PGN in P. falciparum suggests that this compound does not target PfDHFR-TS, in agreement with an earlier study using a different transgenic parasite approach (Fidock et al., 1998). In conclusion, our study opens the door for chemogenomic profiling in Plasmodium spp. The method could be developed further to the level in yeast where compounds are tested in parallel against a whole-genome collection (Giaever et al., 1999, Giaever et al., 2004). Given that there are so few examples of Plasmodium genes that have been successfully modified and their expressions controlled using reverse genetic tools, it is not known if any of the current tools, including the glmS ribozyme tool, are sufficiently robust such that they could be applied on a genome-wide scale. The low efficiency of transgenic DNA integration is a common obstacle for application of reverse genetic tools, especially in P. falciparum. The recent CRISPR/Cas9 genome editing tools described for Plasmodium that give much improved integration efficiency may be useful in this regard (Ghorbal et al., 2014, Wagner et al., 2014, Zhang et al., 2014).
Acknowledgements We thank Dr. Chris Janse for providing the pL0017 plasmid and the PbGFPko230p-SMCON (exp 507 clone1) parasite, Pongpisid Koonyosying for animal husbandry assistance and the Medicines for Malaria Venture (Switzerland) for providing the Malaria Box compound library. We acknowledge the following sources of funding: the Thailand Research Fund grant nos. RSA5780007 and RSA5880064 to PJS and CU, respectively; National Science and Technology Development Agency/Cluster and Program Management (Thailand) project nos. P-14-50752, P-13-00832 and P-14-50883 to PJS, CU and SK, respectively; the United Nations Children’s Fund/the United Nations Development Programme/World Bank/ the World Health Organization (WHO) Special Programme for Research on Diseases of Poverty (TDR) grant no. A50098 to SK; Howard Hughes Medical Institute, USA, grant nos. 55005512 and 55005915 to SK; the Grant (UK) for the Institute for Infectious Diseases of Poverty (IIDP) ID WT087536MA to GOG and the IIDP scholarship award ID Number 2011/01 (from 2011–2014) to OA. OA was also co-sponsored by WHO/TDR and the Thailand National Center for Genetic Engineering and Biotechnology (BIOTEC) for research travel fellowship at BIOTEC during 2010–2011. The funders had no role in study design, the collection, analysis and interpretation of data, the writing of the report, or in the decision to submit the article for publication.
Introduction From antiquity, diseases like malaria have been a challenge for human civilization. Last decade has witnessed millions of deaths due to this devastating disease, especially in tropic and sub-tropics (World Malaria Report, 2013, Machado et al., 2009). It imposes a great socio-economic burden on the poor economies of third world and is a major contributor of mortality and morbidity (Machado et al., 2009).