A screening pipeline for antiparasitic agents targeting cryptosporidium inosine monophosphate dehydrogenase.
BACKGROUND
The protozoan parasite Cryptosporidium parvum is responsible for significant disease burden among children in developing countries. In addition Cryptosporidiosis can result in chronic and life-threatening enteritis in AIDS patients, and the currently available drugs lack efficacy in treating these severe conditions. The discovery and development of novel anti-cryptosporidial therapeutics has been hampered by the poor experimental tractability of this pathogen. While the genome sequencing effort has identified several intriguing new targets including a unique inosine monophosphate dehydrogenase (IMPDH), pursuing these targets and testing inhibitors has been frustratingly difficult.


RESULTS
Here we have developed a pipeline of tools to accelerate the in vivo screening of inhibitors of C. parvum IMPDH. We have genetically engineered the related parasite Toxoplasma gondii to serve as a model of C. parvum infection as the first screen. This assay provides crucial target validation and a large signal window that is currently not possible in assays involving C. parvum. To further develop compounds that pass this first filter, we established a fluorescence-based assay of host cell proliferation, and a C. parvum growth assay that utilizes automated high-content imaging analysis for enhanced throughput.


CONCLUSIONS
We have used these assays to evaluate C. parvum IMPDH inhibitors emerging from our ongoing medicinal chemistry effort and have identified a subset of 1,2,3-triazole ethers that exhibit excellent in vivo selectivity in the T. gondii model and improved anti-cryptosporidial activity.
Sharling
Lisa
L
Center for Tropical and Emerging Global Diseases, University of Georgia, Athens, Georgia, United States of America.
Liu
Xiaoping
X
Gollapalli
Deviprasad R
DR
Maurya
Sushil K
SK
Hedstrom
Lizbeth
L
Striepen
Boris
B
D000981:Antiprotozoal Agents / Q000302:isolation & purification / Q000494:pharmacology*; D001331:Automation; D016785:Cryptosporidium parvum / Q000187:drug effects* / Q000201:enzymology*; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D004791:Enzyme Inhibitors / Q000302:isolation & purification / Q000494:pharmacology*; D057166:High-Throughput Screening Assays / Q000379:methods; D006801:Humans; D007168:IMP Dehydrogenase / Q000037:antagonists & inhibitors*; D007091:Image Processing, Computer-Assisted; D013194:Staining and Labeling; D014122:Toxoplasma / Q000201:enzymology / Q000235:genetics; D014230:Triazoles / Q000302:isolation & purification / Q000494:pharmacology*
D000981:Antiprotozoal Agents; D004791:Enzyme Inhibitors; D014230:Triazoles; D007168:IMP Dehydrogenase