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HIF prolyl hydroxylase-3 mediates alpha-ketoglutarate-induced apoptosis and tumor suppression

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Abstract

Many solid tumors consist of large regions of poorly perfused cells, resulting in areas of low oxygen (hypoxia) throughout the cell mass. Cells subjected to hypoxia turn on a complex set of responses that alter their metabolism, rebalance their survival mechanisms, increase their invasive capacity, and stimulate angiogenesis. This allows them to at least temporarily escape the nutrient starvation and cell death resulting from this hostile environment. Accordingly, the hypoxic regions of tumors are often sources of the most aggressive and therapy-resistant cells, and therefore those cells that drive tumorigenesis. The hypoxia inducible factor (HIF) prolyl hydroxylases (PHDs) are enzymes that are functionally inactivated in hypoxia, as they use both oxygen and α-ketoglutarate as substrates to hydroxylate target prolyl residues. Although HIF1α, the most highly characterized PHD target, orchestrates many of the cellular responses to hypoxia observed in tumors, PHDs themselves have previously been shown to regulate some hypoxia responses, including apoptosis, in a HIF-independent mechanism. We have previously shown that PHDs can be reactivated under hypoxia and that this results in a metabolic defect, both in vitro and in vivo. This led us to investigate whether chronic reactivation of these enzymes may inhibit tumor progression. We show here that esterified α-ketoglutarate given daily will induce apoptosis and inhibit tumor growth, in vivo. The effects are independent of HIF1α but dependent on the presence of PHD3. These data suggest that PHD3 may be a valid target in vivo for anti-tumor therapy.

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Acknowledgments

This work was supported by Cancer Research UK. We would like to thank Tom Hamilton and Derek Miller for their invaluable assistance with the in vivo work, Colin Nixon and Mairi Macdonald for helping with the histology, Margaret O’Prey for help with the microscopy, and Celeste Simon for generously providing us with the HIF1α-expressing plasmid.

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The authors declare they have no conflicts of interest.

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Authors and Affiliations

  1. Cancer Research UK, The Beatson Institute for Cancer Research, Glasgow, UK

    Daniel A. Tennant & Eyal Gottlieb

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  1. Daniel A. Tennant
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  2. Eyal Gottlieb
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Correspondence to Eyal Gottlieb.

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Figure S1

Two examples of tumors treated with a single dose of ETaKG. Serial sections were stained with pimonidazole and cleaved caspase-3. Images captured at ×20 magnification, scale bar indicates 200 μm. “N” marks a necrotic zone, and “H”, the hypoxic ring surrounding this area. (PDF 990 kb)

Figure S2

Proliferation and tumor growth for individual shPHD clones, as shown. (PDF 297 kb)

Figure S3

Immunoblots of xenograft tumor lysates showing PHD levels for each knockdown, after vehicle or ETaKG treatment. (PDF 102 kb)

Figure S4

Immunoblot of HIF1α after knockdown of PHD1, 2, or 3. (PDF 23 kb)

Figure S5

Negative control for anti-myc tag staining using a xenograft tumor not expressing any myc-tagged protein. Scale bar indicates 200 μm. (PDF 193 kb)

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Tennant, D.A., Gottlieb, E. HIF prolyl hydroxylase-3 mediates alpha-ketoglutarate-induced apoptosis and tumor suppression. J Mol Med 88, 839–849 (2010). https://doi.org/10.1007/s00109-010-0627-0

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  • DOI: https://doi.org/10.1007/s00109-010-0627-0

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