Hypoxic human cancer cells are sensitized to BH-3 mimetic–induced apoptosis via downregulation of the Bcl-2 protein Mcl-1
Luke R.E. Harrison, … , Guy Makin, Caroline Dive
Luke R.E. Harrison, … , Guy Makin, Caroline Dive
Published February 14, 2011
Citation Information: J Clin Invest. 2011;121(3):1075-1087. https://doi.org/10.1172/JCI43505.
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Research Article Oncology

Hypoxic human cancer cells are sensitized to BH-3 mimetic–induced apoptosis via downregulation of the Bcl-2 protein Mcl-1

Abstract

Solid tumors contain hypoxic regions in which cancer cells are often resistant to chemotherapy-induced apoptotic cell death. Therapeutic strategies that specifically target hypoxic cells and promote apoptosis are particularly appealing, as few normal tissues experience hypoxia. We have found that the compound ABT-737, a Bcl-2 homology domain 3 (BH-3) mimetic, promotes apoptotic cell death in human colorectal carcinoma and small cell lung cancer cell lines exposed to hypoxia. This hypoxic induction of apoptosis was mediated through downregulation of myeloid cell leukemia sequence 1 (Mcl-1), a Bcl-2 family protein that serves as a biomarker for ABT-737 resistance. Downregulation of Mcl-1 in hypoxia was independent of hypoxia-inducible factor 1 (HIF-1) activity and was consistent with decreased global protein translation. In addition, ABT-737 induced apoptosis deep within tumor spheroids, consistent with an optimal hypoxic oxygen tension being necessary to promote ABT-737–induced cell death. Tumor xenografts in ABT-737–treated mice also displayed significantly more apoptotic cells within hypoxic regions relative to normoxic regions. Synergies between ABT-737 and other cytotoxic drugs were maintained in hypoxia, suggesting that this drug may be useful in combination with chemotherapeutic agents. Taken together, these findings suggest that Mcl-1–sparing BH-3 mimetics may induce apoptosis in hypoxic tumor cells that are resistant to other chemotherapeutic agents and may have a role in combinatorial chemotherapeutic regimens for treatment of solid tumors.

Authors

Luke R.E. Harrison, Dimitra Micha, Martin Brandenburg, Kathryn L. Simpson, Christopher J. Morrow, Olive Denneny, Cassandra Hodgkinson, Zaira Yunus, Clare Dempsey, Darren Roberts, Fiona Blackhall, Guy Makin, Caroline Dive

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Figure 4

Effect of hypoxia and HIF-1 on Mcl-1 protein expression levels.

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Effect of hypoxia and HIF-1 on Mcl-1 protein expression levels. (A) Effe...
(A) Effect of hypoxia on Mcl-1 and HIF-1α protein levels after 18, 24, or 48 hours hypoxia (1% O2) or normoxia. (B) Validation of HCT116 cell line expressing DN HIF-1α protein. HCT116 EV control and HCT116 DN HIF-1α cells were incubated in hypoxia (1% O2) or normoxia for 18 hours, after which the fold induction of firefly luciferase in hypoxia was calculated over that of Renilla luciferase (see Methods). (C) HCT116 EV or HCT116 DN HIF-1α cells were incubated in normoxia or hypoxia (1% O2) for 18 hours, after which they were exposed to a range of ABT-737 concentrations under continuous normoxia or hypoxia for 72 hours prior to determination of IC50 values using the SRB assay. (D) Western blot analysis of Mcl-1 expression level in HCT116 EV and HCT116 DN cells after 18, 24, or 48 hours hypoxia (1% O2) or normoxia. (E) HCT116 cells were treated with HIF-1α or HIF-2α or NT siRNA for 24 hours, and then siRNA was removed and cells were incubated in normoxia or hypoxia for 24 hours, after which cells were harvested and levels of HIF-1α, HIF-2α, Mcl-1, and GAPDH were determined by Western blot. Data are mean ± SEM of 3 independent experiments.