Molecular Response to Hypoxia; from C. elegans to cancer

Abstract

Oncogenesis is governed by genetic and epigenetic events that co-opt to malignant progression. The role of the microenvironment in tumorigenesis and maintenance is increasingly appreciated. Oxygen supply is one of the rate limiting microenvironmental factors. Like healthy cells, cancer cells rely on oxygen. Hypoxia is the condition in which the partial oxygen pressure has dropped to levels that are no longer sufficient to sustain normal cellular function. Hypoxic cells alter metabolism and growth rate and produce angiogenic factors. This ultimately promotes tumor progression. The hypoxia-inducible factor (HIF) transcription factors account for a major part of this response. The HIF transcription factor is a heterodimer composed of a constitutively expressed beta subunit and a hypoxia-regulated alpha subunit. There are three distinct alpha subunits, HIF-1alpha, -2alpha and -3alpha. At physiological oxygen tensions (normoxia), the alpha subunits are continuously degraded upon synthesis, which is dependent on the Von Hippel Lindau tumorsuppressor protein. In the absense of oxygen (hypoxia), the alpha subunits stabilize and constitute the HIF transcription factor after dimerisation with the beta subunit. The HIF trancription factor is involved in the regulation of genes involved in angiogenesis, energy metabolism, cell survival and metastasis. In multiple solid cancer types, expression of HIFalpha subunits has been associated with poor prognosis. Understanding the molecular pathways triggered by hypoxia will enhance insight into the pathological mechanism that underlies hypoxia-associated poor prognosis of cancer. This thesis addresses two main questions: i. how is HIF-1alpha regulated, and ii. how does hypoxia, and particularly HIFalpha overexpression, result in a poor prognosis? Studies concerning the first research question have provided insights into regulation of the HIF pathway and these are presented in this thesis. First of all, experiments in a cell culture model mimicking the serum- and oxygen-deprived microenvironment of solid cancers indicate a crosstalk between oncogenic pathways like the PI 3-kinase pathway and the HIF pathway. This was confirmed in specimen from breast cancer patients. Furthermore, genetic screening has resulted in the identification of multiple genes potentially involved in regulation of the HIF pathway. The second research question has been addressed in multiple chapters of this thesis. In endometrium cancer the HIF pathway induces expression of the cell cycle regulator p27. Subsequent induction of cell cycle arrest might promote hypoxic cancer cell survival. Importantly, this thesis describes the identification of TWIST1 as a HIF-2alpha target gene by genetic screening in the nematode C. elegans. The HIF pathway has been conserved in the evolution, which allows usage of these model organisms. The TWIST1 gene has previously been shown to be a key molecule in metastasis formation in breast cancer. Therefore, the expression profiles of TWIST1 in breast cancer and relation to promotor methylation are also presented. Hypoxic regulation of TWIST1 might explain the association between hypoxia and metastasis formation in human cancer. In addition to the novel results on hypoxic regulation of TWIST1, this thesis also includes a current review on the latest insights in HIF-mediated regulation of metastasis.