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Updated: Aug 26, 2025
In Vitro Drug Screening Against All Life Cycle Stages of Trypanosoma cruzi Using Parasites Expressing β-galactosidase
Published on: November 5, 2021
Kinetic Characterization and Inhibition of Trypanosoma cruzi Hypoxanthine-Guanine Phosphoribosyltransferases
Kayla Glockzin1, Demetrios Kostomiris1, Yacoba V T Minnow2
1Department of Biochemistry and Biophysics, Texas A&M University, 300 Olsen Boulevard, College Station, Texas 77843-2128, United States.
Insights
New inhibitors targeting hypoxanthine-guanine phosphoribosyltransferases (HGPRTs) show promise for treating Chagas disease. These compounds, repurposed from malaria research, effectively inhibit key enzymes essential for parasite survival.
Area of Science:
- Parasitology
- Biochemistry
- Drug Discovery
Background:
- Chagas disease affects millions globally, with current treatments lacking efficacy and exhibiting toxicity, especially in chronic stages.
- The parasite *Trypanosoma cruzi* relies on host purines due to its inability to synthesize them *de novo*.
- Hypoxanthine-guanine phosphoribosyltransferases (HGPRTs) are crucial for parasite purine salvage and represent potential therapeutic targets.
Purpose of the Study:
- To characterize *Trypanosoma cruzi* HGPRT enzymes (TcA and TcC) and identify potential drug targets.
- To evaluate the efficacy of transition-state analogue inhibitors (TSAIs) against *T. cruzi* HGPRTs.
- To validate the repurposing of TSAIs for Chagas disease treatment.
Main Methods:
- Characterization of two *T. cruzi* HGPRT gene products (TcA and TcC).
- Kinetic characterization of the TcC isozyme to determine catalytic mechanisms.
- Screening and characterization of TSAIs originally developed for *Plasmodium falciparum* HGXPRT against *T. cruzi* HGPRTs.
Main Results:
- Two *T. cruzi* gene products, TcA and TcC, demonstrated HGPRT activity *in vitro*.
- The TcC isozyme's catalytic steps were elucidated through kinetic analysis.
- A potent TSAI, originally developed for *Pf*HGXPRT, exhibited low nanomolar affinity for *T. cruzi* HGPRT.
Conclusions:
- *T. cruzi* HGPRTs are validated as promising targets for novel Chagas disease therapies.
- Repurposing of TSAIs is an effective strategy for developing selective inhibitors against conserved molecular targets.
- The identified TSAIs offer a potential new avenue for treating Chagas disease, particularly in advanced stages.
Abstract:
Chagas disease, caused by the parasitic protozoan Trypanosoma cruzi, affects over 8 million people worldwide. Current antiparasitic treatments for Chagas disease are ineffective in treating advanced, chronic stages of the disease, and are noted for their toxicity. Like most parasitic protozoa, T. cruzi is unable to synthesize purines de novo, and relies on the salvage of preformed purines from the host. Hypoxanthine-guanine phosphoribosyltransferases (HGPRTs) are enzymes that are critical for the salvage of preformed purines, catalyzing the formation of inosine monophosphate (IMP) and guanosine monophosphate (GMP) from the nucleobases hypoxanthine and guanine, respectively. Due to the central role of HGPRTs in purine salvage, these enzymes are promising targets for the development of new treatment methods for Chagas disease. In this study, we characterized two gene products in the T. cruzi CL Brener strain that encodes enzymes with functionally identical HGPRT activities in vitro: TcA (TcCLB.509693.70) and TcC (TcCLB.506457.30). The TcC isozyme was kinetically characterized to reveal mechanistic details on catalysis, including identification of the rate-limiting step(s) of catalysis. Furthermore, we identified and characterized inhibitors of T. cruzi HGPRTs originally developed as transition-state analogue inhibitors (TSAIs) of Plasmodium falciparum hypoxanthine-guanine-xanthine phosphoribosyltransferase (PfHGXPRT), where the most potent compound bound to T. cruzi HGPRT with low nanomolar affinity. Our results validated the repurposing of TSAIs to serve as selective inhibitors for orthologous molecular targets, where primary and secondary structures as well as putatively common chemical mechanisms are conserved.
