Pyrimidine salvage in<i>Toxoplasma gondii</i>as a target for new treatment
Abstract Toxoplasmosis is a common protozoan infection that can have severe outcomes in the immunocompromised and during pregnancy, but treatment options are limited. Recently, nucleotide metabolism has received much attention as a target for new antiprotozoal agents and here we focus on pyrimidine salvage byToxoplasma gondiias a drug target. Whereas uptake of [3H]-cytidine and particularly [3H]-thymidine was at most marginal, [3H]-uracil and [3H]-uridine were readily taken up. Kinetic analysis of uridine uptake was consistent with a single transporter with a Kmof 3.3 ± 0.8 µM, which was inhibited by uracil with high affinity (Ki= 1.15 ± 0.07 µM) but not by thymidine or 5-methyluridine, showing that the 5-Me group is incompatible with uptake byT. gondii. Conversely, [3H]-uracil transport displayed a Kmof 2.05 ± 0.40 µM, not significantly different from the uracil Kion uridine transport, and was inhibited by uridine with a Ki2.44 ± 0.59 µM, also not significantly different from the experimental uridine Km. The reciprocal, complete inhibition, displaying Hill slopes of approximately ∼1, strongly suggest that uridine and uracil share a single transporter with similarly high affinity for both, and we designate it uridine/uracil transporter 1 (TgUUT1). While TgUUT1 excludes 5-methyl substitutions, the smaller 5F substitution was tolerated as 5F-uracil inhibited uptake of [3H]-uracil with a Kiof 6.80 ± 2.12 µM (P> 0.05 compared to uracil Km). Indeed, we found that 5F-Uridine, 5F-uracil and 5F,2’-deoxyuridine were all potent antimetabolites againstT. gondiiwith EC50values well below that of the current first line treatment, sulfadiazine.In vivoevaluation also showed that 5F-uracil and 5F,2’-deoxyuridine were similarly effective as sulfadiazine against acute toxoplasmosis. Our preliminary conclusion is that TgUUT1 mediates potential new anti-toxoplasmosis drugs with activity superior to the current treatment..
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Preprint| Erscheinungsjahr: |
2023 |
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| Erschienen: |
2023 |
| Enthalten in: |
bioRxiv.org - (2023) vom: 06. Nov. Zur Gesamtaufnahme - year:2023 |
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| Sprache: |
Englisch |
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| Beteiligte Personen: |
Elati, Hamza A. A. [VerfasserIn] |
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| Links: |
Volltext [kostenfrei] |
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| doi: |
10.1101/2023.11.01.565095 |
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| PPN (Katalog-ID): |
XBI041415760 |
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| 520 | |a Abstract Toxoplasmosis is a common protozoan infection that can have severe outcomes in the immunocompromised and during pregnancy, but treatment options are limited. Recently, nucleotide metabolism has received much attention as a target for new antiprotozoal agents and here we focus on pyrimidine salvage byToxoplasma gondiias a drug target. Whereas uptake of [3H]-cytidine and particularly [3H]-thymidine was at most marginal, [3H]-uracil and [3H]-uridine were readily taken up. Kinetic analysis of uridine uptake was consistent with a single transporter with a Kmof 3.3 ± 0.8 µM, which was inhibited by uracil with high affinity (Ki= 1.15 ± 0.07 µM) but not by thymidine or 5-methyluridine, showing that the 5-Me group is incompatible with uptake byT. gondii. Conversely, [3H]-uracil transport displayed a Kmof 2.05 ± 0.40 µM, not significantly different from the uracil Kion uridine transport, and was inhibited by uridine with a Ki2.44 ± 0.59 µM, also not significantly different from the experimental uridine Km. The reciprocal, complete inhibition, displaying Hill slopes of approximately ∼1, strongly suggest that uridine and uracil share a single transporter with similarly high affinity for both, and we designate it uridine/uracil transporter 1 (TgUUT1). While TgUUT1 excludes 5-methyl substitutions, the smaller 5F substitution was tolerated as 5F-uracil inhibited uptake of [3H]-uracil with a Kiof 6.80 ± 2.12 µM (P> 0.05 compared to uracil Km). Indeed, we found that 5F-Uridine, 5F-uracil and 5F,2’-deoxyuridine were all potent antimetabolites againstT. gondiiwith EC50values well below that of the current first line treatment, sulfadiazine.In vivoevaluation also showed that 5F-uracil and 5F,2’-deoxyuridine were similarly effective as sulfadiazine against acute toxoplasmosis. Our preliminary conclusion is that TgUUT1 mediates potential new anti-toxoplasmosis drugs with activity superior to the current treatment. | ||
| 650 | 4 | |a Biology |7 (dpeaa)DE-84 | |
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| 700 | 1 | |a Goerner, Amber L. |4 aut | |
| 700 | 1 | |a Di Genova, Bruno Martorelli |4 aut | |
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| 700 | 1 | |a de Koning, Harry P. |0 (orcid)0000-0002-9963-1827 |4 aut | |
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