Radiation and inhibition of angiogenesis by canstatin synergize to induce HIF-1α–mediated tumor apoptotic switch
Claire Magnon, Paule Opolon, Marcel Ricard, Elisabeth Connault, Patrice Ardouin, Ariane Galaup, Didier Métivier, Jean-Michel Bidart, Stéphane Germain, Michel Perricaudet, Martin Schlumberger
Claire Magnon, Paule Opolon, Marcel Ricard, Elisabeth Connault, Patrice Ardouin, Ariane Galaup, Didier Métivier, Jean-Michel Bidart, Stéphane Germain, Michel Perricaudet, Martin Schlumberger
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Research Article Oncology

Radiation and inhibition of angiogenesis by canstatin synergize to induce HIF-1α–mediated tumor apoptotic switch

Abstract

Tumor radioresponsiveness depends on endothelial cell death, which leads in turn to tumor hypoxia. Radiation-induced hypoxia was recently shown to trigger tumor radioresistance by activating angiogenesis through hypoxia-inducible factor 1–regulated (HIF-1–regulated) cytokines. We show here that combining targeted radioiodide therapy with angiogenic inhibitors, such as canstatin, enhances direct tumor cell apoptosis, thereby overcoming radio-induced HIF-1–dependent tumor survival pathways in vitro and in vivo. We found that following dual therapy, HIF-1α increases the activity of the canstatin-induced αvβ5 signaling tumor apoptotic pathway and concomitantly abrogates mitotic checkpoint and tetraploidy triggered by radiation. Apoptosis in conjunction with mitotic catastrophe leads to lethal tumor damage. We discovered that HIF-1 displays a radiosensitizing activity that is highly dependent on treatment modalities by regulating key apoptotic molecular pathways. Our findings therefore support a crucial role for angiogenesis inhibitors in shifting the fate of radiation-induced HIF-1α activity from hypoxia-induced tumor radioresistance to hypoxia-induced tumor apoptosis. This study provides a basis for developing new biology-based clinically relevant strategies to improve the efficacy of radiation oncology, using HIF-1 as an ally for cancer therapy.

Authors

Claire Magnon, Paule Opolon, Marcel Ricard, Elisabeth Connault, Patrice Ardouin, Ariane Galaup, Didier Métivier, Jean-Michel Bidart, Stéphane Germain, Michel Perricaudet, Martin Schlumberger

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Figure 3

In vivo, detection of hypoxic, apoptotic, and endothelial markers in MDA-MB-231 xenografted tumors following AdNIS-131I therapy combined with AdCanHSA.

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In vivo, detection of hypoxic, apoptotic, and endothelial markers in MDA...
(A) The number of CD34-stained intratumor microvessels per square millimeter was quantified by digital analysis of tumor tissue sections at day 30 after infection. Original magnification, ×100. (B) Representative images of CD34 endothelial immunostaining. (C) Quantification of the area fraction of hypoxic CAIX-stained cells within digitized sections from AdCO1-, AdNIS-, AdCanHSA-, and AdCanHSA-AdNIS–injected tumors at days 15 and 30 (i.e., at days 8 and 20 after injection of 131I). (D) Representative CAIX-stained digitized sections for each group. (E) Quantification of HIF-1 mRNA levels by in situ hybridization analysis. The number of cells per field producing mRNA HIF-1α was quantified within tumors sections (×40) at day 30 following injections of appropriate adenoviruses listed above. (F) Western blotting analysis of HIF-1α protein levels within nuclear (N) and cytoplasmic (C) extracts of adenovirus-infected xenografted tumors. A whole-cell lysate of RCC (VHL mutated) cells was considered as positive control. Note that HIF-1α and CAIX have a similar expression pattern at day 30. (G) The proportion of apoptotic cells per field was quantified within sections (×200) (TUNEL method) at days 15 and 30 after infection. Quantification of the area fraction of caspase-9– (H) and caspase-3–stained cells (I) within digitized sections from treated tumors at day 30 as described in Figure 1A. (J) Representative illustrations of both TUNEL and caspase-3 staining. Columns, mean ± SEM of 9 mice for each treated group. All digitized paraffin-embedded tumor sections were obtained from the experiment described in Figure 1A. Original magnification, ×200.