Transcriptional Responses to Bumped Kinase Inhibitor BKI-1708 in Toxoplasma gondii and Human Fibroblasts

Bumped kinase inhibitors are safe with promising efficacy against apicomplexan parasites. The 5-aminopyrazole-4-carboxamide BKI-1708 effectively inhibited vertical transmission of Toxoplasma gondii and significantly reduced the cerebral parasite loads in experimentally infected pregnant mice. In vitro experiments revealed that exposure of T. gondii tachyzoites to BKI-1708 induces the formation of intracellular multinucleated complexes called "baryzoites", exhibiting increased expression of bradyzoite-stage proteins while still displaying classical tachyzoite markers. Differential affinity chromatography of T. gondii extracts identified numerous BKI-1708-binding proteins involved in invasion/egress, redox homeostasis, and RNA processing. To understand the transcriptional implications of BKI-1708 treatment on T. gondii tachyzoites and human foreskin fibroblast host cells, T. gondii-infected host cells, either treated with BKI-1708 or untreated, were subjected to dual RNA-seq analysis. BKI-1708 induced a significant transcriptional remodeling in the parasite, with an enrichment in pathways related to translation, RNA metabolism, and stress responses. In contrast, host-cell transcriptional changes were more limited, with transcripts related to metabolic and detoxification programs upregulated in uninfected fibroblasts, and increased transcription of immune, lysosomal, and glycan degradation pathways in infected fibroblasts. These findings suggest that BKI-1708 modulates the transcriptome in a predominantly parasite-specific manner, disrupting essential biological processes in T. gondii while largely preserving host cell function.