Unbiased Metabolite Profiling Indicates That a Diminished Thymidine Pool Is the Underlying Mechanism of Colon Cancer Chemoprevention by Alpha-Difluoromethylornithine

Witherspoon, Mavee; Chen, Qiuying; Kopelovich, Levy; Gross, Steven S.; Lipkin, Steven M.

doi:10.1158/2159-8290.cd-12-0305

Cited by 43 publications

(51 citation statements)

References 40 publications

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“…Thymidine depletion is the basis for anti-proliferative therapies that range from antibiotics (Hawser et al, 2006; Kwon et al, 2010) to cancer chemotherapy (Gangjee et al, 2007; Witherspoon et al, 2013). The success of drugs that target thymidine pools highlights the importance of generating this nucleotide in proliferating cells.…”

Section: Discussionmentioning

confidence: 99%

Pyruvate Kinase Isoform Expression Alters Nucleotide Synthesis to Impact Cell Proliferation

et al. 2015

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SUMMARY Metabolic regulation influences cell proliferation. The influence of pyruvate kinase isoforms on tumor cells has been extensively studied, but whether PKM2 is required for normal cell proliferation is unknown. We examine how PKM2-deletion affects proliferation and metabolism in non-transformed, non-immortalized PKM2-expressing primary cells. We find that deletion of PKM2 in primary cells results in PKM1 expression and proliferation arrest. PKM1 expression, rather than PKM2 loss, is responsible for this effect, and proliferation arrest cannot be explained by cell differentiation, senescence, death, changes in gene expression, or prevention of cell growth. Instead, PKM1 expression impairs nucleotide production and the ability to synthesize DNA and progress through the cell cycle. Nucleotide biosynthesis is limiting, as proliferation arrest is characterized by severe thymidine depletion, and supplying exogenous thymine rescues both nucleotide levels and cell proliferation. Thus, PKM1 expression promotes a metabolic state that is unable to support DNA synthesis.

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Section: Discussionmentioning

confidence: 99%

Pyruvate Kinase Isoform Expression Alters Nucleotide Synthesis to Impact Cell Proliferation

et al. 2015

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“…A metabolomic approach applied to colorectal cancer models led to the hypothesis that reduced cellular thymidine pools contribute to DFMO-induced cytostatic activity (52). DFMO-mediated polyamine depletion leads to compensatory hyperactivation of Amd1 with futile S-adenosylmethionine consumption, followed by depletion of folate-dependent metabolites and thymidine.…”

Section: Discussionmentioning

confidence: 99%

Polyamine Antagonist Therapies Inhibit Neuroblastoma Initiation and Progression

Evageliou

Haber

et al. 2016

Clinical Cancer Research

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Purpose: Deregulated MYC drives oncogenesis in many tissues yet direct pharmacologic inhibition has proven difficult. MYC coordinately regulates polyamine homeostasis as these essential cations support MYC functions, and drugs that antagonize polyamine sufficiency have synthetic-lethal interactions with MYC. Neuroblastoma is a lethal tumor in which the MYC homologue MYCN, and ODC1, the rate-limiting enzyme in polyamine synthesis, are frequently deregulated so we tested optimized polyamine depletion regimens for activity against neuroblastoma.Experimental Design: We used complementary transgenic and xenograft-bearing neuroblastoma models to assess polyamine antagonists. We investigated difluoromethylornithine (DFMO; an inhibitor of Odc, the rate-limiting enzyme in polyamine synthesis), SAM486 (an inhibitor of Amd1, the second ratelimiting enzyme), and celecoxib (an inducer of Sat1 and polyamine catabolism) in both the preemptive setting and in the treatment of established tumors. In vitro assays were performed to identify mechanisms of activity.Results: An optimized polyamine antagonist regimen using DFMO and SAM486 to inhibit both rate-limiting enzymes in polyamine synthesis potently blocked neuroblastoma initiation in transgenic mice, underscoring the requirement for polyamines in MYC-driven oncogenesis. Furthermore, the combination of DFMO with celecoxib was found to be highly active, alone, and combined with numerous chemotherapy regimens, in regressing established tumors in both models, including tumors harboring highest risk genetic lesions such as MYCN amplification, ALK mutation, and TP53 mutation with multidrug resistance.Conclusions: Given the broad preclinical activity demonstrated by polyamine antagonist regimens across diverse in vivo models, clinical investigation of such approaches in neuroblastoma and potentially other MYC-driven tumors is warranted.

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“…Metabolomics has shed light on the changes in metabolism that accompany diseases and drug treatment [2], identified changes in metabolism of human diseases such as cancer [3, 4], and provide vital biological insights through the study of a wide variety of organisms [5, 6]. …”

Section: Introductionmentioning

confidence: 99%

Extraction parameters for metabolomics from cultured cells

Ser

Liu

Tang

et al. 2015

Analytical Biochemistry

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The successful extraction of metabolites is a critical step in metabolite profiling. By optimizing metabolite extraction, the range and quantitative capacity of metabolomics studies can be improved. We considered eight separate extraction protocols for the preparation of a metabolite extract from cultured mammalian cells. Parameters considered included temperature, pH, and cell washing before extraction. The effects on metabolite recovery were studied using a high resolution liquid chromatography mass spectrometry (LC-HRMS) platform that measures metabolites of diverse chemical classes including among others amino acids, lipids, and sugar derivatives. The temperature considered during the extraction or the presence of formic acid, a commonly used additive, was shown to have minimal effects on the measured ion intensities of metabolites. However, washing of samples before metabolite extraction whether with water or PBS (both commonly considered practices) exhibited dramatic effects on measured intensities of both intra- and extra-cellular metabolites. Together these findings present a systematic assessment of extraction conditions for metabolite profiling.

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Unbiased Metabolite Profiling Indicates That a Diminished Thymidine Pool Is the Underlying Mechanism of Colon Cancer Chemoprevention by Alpha-Difluoromethylornithine

Cited by 43 publications

References 40 publications

Pyruvate Kinase Isoform Expression Alters Nucleotide Synthesis to Impact Cell Proliferation

Pyruvate Kinase Isoform Expression Alters Nucleotide Synthesis to Impact Cell Proliferation

Polyamine Antagonist Therapies Inhibit Neuroblastoma Initiation and Progression

Extraction parameters for metabolomics from cultured cells

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