Reversing hypoxic cell chemoresistance in vitro using genetic and small molecule approaches targeting hypoxia inducible factor-1.

Abstract

The resistance of hypoxic cells to conventional chemotherapy is well documented. Using both adenovirus-mediated gene delivery and small molecules targeting hypoxia-inducible factor-1 (HIF-1), we evaluated the impact of HIF-1 inhibition on the sensitivity of hypoxic tumor cells to etoposide. The genetic therapy exploited a truncated HIF-1alpha protein that acts as a dominant-negative HIF-1alpha (HIF-1alpha-no-TAD). Its functionality was validated in six human tumor cell lines using HIF-1 reporter assays. An EGFP-fused protein demonstrated that the dominant-negative HIF-1alpha was nucleus-localized and constitutively expressed irrespective of oxygen tension. The small molecules studied were quinocarmycin monocitrate (KW2152), its analog 7-cyanoquinocarcinol (DX-52-1), and topotecan. DX-52-1 and topotecan have been previously established as HIF-1 inhibitors. HT1080 and HCT116 cells were treated with either AdHIF-1alpha-no-TAD or nontoxic concentrations (0.1 microM; <IC(10)) of KW2152 and DX-52-1 and exposed to etoposide in air or anoxia (<0.01% oxygen). Topotecan inhibited HIF-1 activity only at cytotoxic concentrations and was not used in the combination study. Etoposide IC(50) values in anoxia were 3-fold higher than those in air for HT1080 (2.2 +/- 0.3 versus 0.7 +/- 0.2 microM) and HCT116 (9 +/- 4 versus 3 +/- 2 microM) cells. KW2152 and DX-52-1 significantly reduced the anoxic etoposide IC(50) in HT1080 cells, whereas only KW2152 yielded sensitization in HCT116 cells. In contrast, AdHIF-1alpha-no-TAD (multiplicity of infection 50) ablated the anoxic resistance in both cell lines (IC(50) values: HT1080, 0.7 +/- 0.04 microM; HCT116, 3 +/- 1 microM). HIF-1alpha-no-TAD expression inhibited HIF-1-mediated down-regulation of the proapoptotic protein Bid under anoxia. These data support the potential development of HIF-1 targeted approaches in combination with chemotherapy, where hypoxic cell resistance contributes to treatment failure.

• Authors: Louisa M Brown, Rachel L Cowen, Camille Debray, Amanda Eustace, Janine T Erler, Freda C D Sheppard, Catriona A Parker, Ian J Stratford, Kaye J Williams
• Journal: Molecular pharmacology (Mol Pharmacol) Vol. 69 Issue 2 Pg. 411-8 (Feb 2006) ISSN: 0026-895X [Print] United States
• PMID: 16254058 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Antineoplastic Agents, Phytogenic
• Hypoxia-Inducible Factor 1
• Hypoxia-Inducible Factor 1, alpha Subunit
• Isoquinolines
• enhanced green fluorescent protein
• Green Fluorescent Proteins
• DX 52-1
• Etoposide
• Topotecan
• quinocarcin
• Oxygen

Topics

• Adenoviridae (genetics)
• Antineoplastic Agents, Phytogenic (therapeutic use)
• Cell Hypoxia
• Cell Line, Tumor
• Cell Nucleus (chemistry)
• Drug Resistance, Neoplasm (drug effects)
• Etoposide (therapeutic use)
• Genetic Vectors (genetics)
• Green Fluorescent Proteins (analysis, genetics)
• Humans
• Hypoxia-Inducible Factor 1 (antagonists & inhibitors, genetics)
• Hypoxia-Inducible Factor 1, alpha Subunit (analysis, genetics)
• Isoquinolines (pharmacology)
• Neoplasms (drug therapy)
• Oxygen (metabolism)
• Sequence Deletion
• Topotecan (pharmacology)
• Transcriptional Activation