Mechanism of thymidylate synthase inhibition by methotrexate in human neoplastic cell lines and normal human myeloid progenitor cells.

Chu, Edward; Drake, James C.; Boarman, Donna; Baram, J; Allegra, Carmen J.

doi:10.1016/s0021-9258(19)38912-4

Cited by 50 publications

(8 citation statements)

References 28 publications

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“…And indeed, polyglutamated MTX inhibits phosphoribosylaminoimidazolecarboxamide (AICAR) transformylase (ATIC) ( 39 , 40 ). MTX, when polyglutamated, also inhibits thymidylate synthetase (TYMS) directly ( 41 , 42 ), which, along with the depletion of 5,10 methylene THF, further diminishes the synthesis of deoxythymidine monophosphate ( 43 ). This then reduces the availability of deoxythymidine triphosphate, which is required for the synthesis of DNA but not RNA.…”

Section: Resultsmentioning

confidence: 99%

The folate antagonist methotrexate diminishes replication of the coronavirus SARS-CoV-2 and enhances the antiviral efficacy of remdesivir in cell culture models

et al. 2021

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The search for successful therapies of infections with the coronavirus SARS-CoV-2 is ongoing. We tested inhibition of host cell nucleotide synthesis as a promising strategy to decrease the replication of SARS-CoV-2-RNA, thus diminishing the formation of virus progeny. Methotrexate (MTX) is an established drug for cancer therapy and to induce immunosuppression. The drug inhibits dihydrofolate reductase and other enzymes required for the synthesis of nucleotides. Strikingly, the replication of SARS-CoV-2 was inhibited by MTX in therapeutic concentrations around 1 µM, leading to more than 1000-fold reductions in virus progeny in Vero C1008 (Vero E6) and ∼100-fold reductions in Calu-3 cells. Virus replication was more sensitive to equivalent concentrations of MTX than of the established antiviral agent remdesivir. MTX strongly diminished the synthesis of viral structural proteins and the amount of released virus RNA. Virus replication and protein synthesis were rescued by folinic acid (leucovorin) and also by inosine, indicating that purine depletion is the principal mechanism that allows MTX to reduce virus RNA synthesis. The combination of MTX with remdesivir led to synergistic impairment of virus replication, even at 100 nM MTX. The use of MTX in treating SARS-CoV-2 infections still awaits further evaluation regarding toxicity and efficacy in infected organisms, rather than cultured cells. Within the frame of these caveats, however, our results raise the perspective of a two-fold benefit from repurposing MTX for treating COVID-19. Firstly, its previously known ability to reduce aberrant inflammatory responses might dampen respiratory distress. In addition, its direct antiviral activity described here would limit the dissemination of the virus.

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Section: Resultsmentioning

confidence: 99%

The folate antagonist methotrexate diminishes replication of the coronavirus SARS-CoV-2 and enhances the antiviral efficacy of remdesivir in cell culture models

et al. 2021

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“…Methotrexate is a classical DHFR inhibitor characterized by an intact glutamate side chain that can undergo polyglutamylation by folylpolyglutamate synthase ( 32 ). These methotrexate polyglutamates can not only demonstrate prolonged intracellular retention times but also inhibit other folate metabolism enzymes, including TS ( 33 , 34 , 35 ). In contrast, pyrimethamine contains a lipophilic terminal group that cannot be polyglutamylated.…”

Section: Discussionmentioning

confidence: 99%

The antimicrobial drug pyrimethamine inhibits STAT3 transcriptional activity by targeting the enzyme dihydrofolate reductase

Heppler

Attarha

Persaud

et al. 2022

Journal of Biological Chemistry

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Cancer is often characterized by aberrant gene expression patterns caused by the inappropriate activation of transcription factors. Signal transducer and activator of transcription 3 (STAT3) is a key transcriptional regulator of many protumorigenic processes and is persistently activated in many types of human cancer. However, like many transcription factors, STAT3 has proven difficult to target clinically. To address this unmet clinical need, we previously developed a cell-based assay of STAT3 transcriptional activity and performed an unbiased and high-throughput screen of small molecules known to be biologically active in humans. We identified the antimicrobial drug pyrimethamine as a novel and specific inhibitor of STAT3 transcriptional activity. Here, we show that pyrimethamine does not significantly affect STAT3 phosphorylation, nuclear translocation, or DNA binding at concentrations sufficient to inhibit STAT3 transcriptional activity, suggesting a potentially novel mechanism of inhibition. To identify the direct molecular target of pyrimethamine and further elucidate the mechanism of action, we used a new quantitative proteome profiling approach called proteome integral solubility alteration coupled with a metabolomic analysis. We identified human dihydrofolate reductase as a target of pyrimethamine and demonstrated that the STAT3-inhibitory effects of pyrimethamine are the result of a deficiency in reduced folate downstream of dihydrofolate reductase inhibition, implicating folate metabolism in the regulation of STAT3 transcriptional activity. This study reveals a previously unknown regulatory node of the STAT3 pathway that may be important for the development of novel strategies to treat STAT3-driven cancers.

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“…5M). Furthermore, treatment of IDH3-deficient cells with methotrexate (MTX), an inhibitor of TYMS and DHFR (29,30), which promotes apoptosis through ROS generation (31), reduced cell viability in IDH3 KO NHAs compared to WT control cells (fig. S7A), while IDH3-overexpressing NHAs were protected against MTX cytotoxic effect (fig.…”

Section: Idh3 Regulates Nucleotide Biosynthesis and Dna Methylationmentioning

confidence: 99%

IDH3α regulates one-carbon metabolism in glioblastoma

et al. 2019

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Mechanism of thymidylate synthase inhibition by methotrexate in human neoplastic cell lines and normal human myeloid progenitor cells.

Cited by 50 publications

References 28 publications

The folate antagonist methotrexate diminishes replication of the coronavirus SARS-CoV-2 and enhances the antiviral efficacy of remdesivir in cell culture models

The folate antagonist methotrexate diminishes replication of the coronavirus SARS-CoV-2 and enhances the antiviral efficacy of remdesivir in cell culture models

The antimicrobial drug pyrimethamine inhibits STAT3 transcriptional activity by targeting the enzyme dihydrofolate reductase

IDH3α regulates one-carbon metabolism in glioblastoma

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