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 7

Hypoxic sensitization to ABT-737 in H526 SCLC cells in vitro and in vivo.

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Hypoxic sensitization to ABT-737 in H526 SCLC cells in vitro and in vivo...
(A–C) Hypoxic sensitization of H526 cells to ABT-737 in vitro. (A) H256 cell population growth (by resazurin assay) under continuous normoxia or hypoxia as in Figure 1A. (B) Apoptotic cell death (by DAPI staining and nuclear morphology) after 24 hours incubation in normoxia or hypoxia with or without ABT-737 (1 μM) or (C) as assessed by CC3 levels after 0, 4, or 8 hours incubation with ABT-737. CC3 and GAPDH protein level data shown are for noncontiguous lanes run on the same gel. (D) Expression levels of Mcl-1 in untreated H526 cells exposed to normoxia or hypoxia for up to 8 hours. GAPDH was used as a loading control. Data in A and B are mean ± SEM from 3 independent experiments. Data in C and D are representative of at least 3 independent experiments. (E–G) Hypoxic sensitization of H526 cells to ABT-737 in vivo. (E) Effect of ABT-737 (100 mg/kg/d) on tumor xenograft growth in SCID-bg mice (size-matched and dosed from day 17). Data represent the average of 6 mice per group. (F) Representative images of serial tumor sections showing staining for pimonidazole (hypoxia) and CC3 (apoptosis) from tumors harvested after 72 hours of ABT-737 treatment. Images are of identical magnification. Scale bar: 100 μm. (G) Percentage area of CC3-positive staining in 4 hypoxic or 4 normoxic tumor regions. Data are the average of 4 mice per time point and per treatment. *P < 0.05 (Student’s 2-tailed t test).