A Novel Benzodioxole-Containing Inhibitor of Toxoplasma gondii Growth Alters the Parasite Cell Cycle

Abstract

Toxoplasma gondii is an obligate intracellular parasite that can cause disease in the developing fetus and in immunocompromised humans. Infections can last for the life of the individual, and to date there are no drugs that eliminate the chronic cyst stages that are characteristic of this parasite. In an effort to identify new chemical scaffolds that could form the basis for new therapeutics, we carried out a chemoinformatic screen for compounds that had the potential to interact with members of a superfamily of parasite-secreted kinases and assayed them for growth inhibition in vitro. Of 17 candidate compounds, we identified one with potent antiparasitic activity. The compound has a 50% inhibitory concentration (IC 50 ) of ϳ2 nM, and structurefunction analyses implicate the benzodioxole moiety in its action. The compound does not appear to be cytotoxic to host cells. Using microarray analyses of both parasites and host cells treated with the compound, we found that the levels of very few host cell transcripts are altered by the compound, while a large number of parasite transcripts have a different abundance after compound treatment. Gene ontology analyses of parasite transcripts with a different abundance revealed an enrichment of cell cycle-related genes, suggesting that the compound alters progression of the parasite through the cell cycle. Assaying the nuclear content of treated parasites demonstrated that compound treatment significantly increased the percentage of parasites in the S/M phase of the cell cycle compared to controls. This compound and its analogs represent a novel scaffold with antiparasitic activity.

Hammondia hammondi Harbors Functional Orthologs of the Host-Modulating Effectors GRA15 and ROP16 but Is Distinguished from Toxoplasma gondii by a Unique Transcriptional Profile

“…Genes were deemed to be significantly different in abundance if the adjusted P value was Յ0.01. Gene ontology (GO) analyses of differentially expressed transcripts were conducted with GeneMerge (45) as described previously (46). GO categories were deemed significantly enriched if they had adjusted P values of Յ0.05.…”

Review of Experimental Compounds Demonstrating Anti-Toxoplasma Activity

“…Cheminformaticsbased virtual screens are being performed to identify novel antiToxoplasma compounds (87,124,125). The availability of crystal structures for some of the T. gondii proteins and the improvement in computational molecular modeling approaches facilitate this approach.…”

A Focused Small-Molecule Screen Identifies 14 Compounds with Distinct Effects on Toxoplasma gondii

“…Electron microscopy studies were done as previously described (14), with the following minor changes. The monolayers were infected at multiplicities of infection (MOIs) of 5 and 10.…”

“…The monolayers were infected at multiplicities of infection (MOIs) of 5 and 10. After 4 h, monolayers infected at an MOI of 5 were thoroughly washed to remove uninvading parasites and then treated with 10 M compounds C3 and C5 or vehicle for 40 h. Monolayers infected at an MOI of 10 were grown overnight, washed, and then treated with 10 M compounds C2 and C1 or vehicle for 24 h. Monolayers were then fixed and processed for electron microscopy (14).…”

“…Host cell viability in the presence of the compounds was determined using both a colorimetric assay and microscopy. The colorimetric assay was done as described previously (14) after 48 h compound (10 M) exposure. Staurosporine was used as a positive control for cytotoxicity, and vehicle was used as the negative control.…”