Hypoxia, or reduced oxygen supply, is a common attribute of solid tumours and is caused by excessive cellular proliferation.
Adaptation to this microenvironmental stress is mediated by the activation of the hypoxia-inducible factor (HIF) pathway promoting metabolic alterations, invasion, metastasis and the induction of angiogenesis.
The transcription factor HIF-1, itself composed of HIF-1a and HIF-1b, is the major player within this pathway. HIF-1 is degraded under normoxic conditions and accumulates under oxygen deprivation. In contrast, HIF-1b is regarded as constitutively expressed. HIF signalling contributes to the development of malignant melanoma, the most aggressive form of skin cancer. There is evidence of elevated HIF activity even under normoxia within this cell type. The HIF pathway thus represents an attractive drug target for cancer therapy. However, currently consequences of HIF inhibition are poorly defined.
Therefore the objective of this project was to investigate cellular effects elicited by HIF inhibition under hypoxia and exposure to the hypoxia-mimetic cobalt chloride. Suppression of HIF signalling was achieved by siRNA mediated knockdown of HIF subunits, ectopic expression of a dominant-negative form of HIF-1a, as well as by pharmacological inhibition. A Hens egg test - chorioallantoic membrane (Het-CAM) model was used to study the HIF-dependent processes angiogenesis and invasion.
The data obtained demonstrates unique angiogenic and invasive characteristics of different human melanoma cell lines in the Het-CAM assay. Secretion of the pro-angiogenic cytokine Interleukin (IL)-8 by melanoma cells was elevated via HIF inhibition under cobalt chloride-induced hypoxia. In addition, the results indicate that HIF-1b obviously is a hypoxia-responsive protein within certain melanoma cell lines. A series of experiments revealed the existence of a novel, cell specific mechanism by which HIF-1a upregulates HIF-1b expression under hypoxic conditions.
In conclusion, the results indicate an IL-8-mediated compensatory response due to HIF inhibition and demonstrate that melanoma cells can acquire the capability to upregulate HIF-1b under hypoxia, which might prevent this subunit to become a limiting factor.