CANCER CELLS AND THE REQUIRED ENERGY
Since cancer cells are growing and multiplying, thus using much greater amounts of ATP, one might suppose that they would use the highly efficient aerobic pathway whenever possible, but this is not the case.
The first person to spot that there was something odd about metabolism in cancer cells was the biochemist Otto Warburg and he eventually demonstrated that they obtain most of their energy from glucose using the glycolytic pathway, rather than the second, oxygen- using stage. Typically, the glycolytic flux in rapidly growing tumour cells is a hundred-fold greater than in the normal cells from which a tumour originated. This conversion of glucose into lactate, even in the presence of abundant oxygen, is known as the Warburg effect or aerobic glycolysis. Presumably, it reflects the fact that tumour cells have adapted to being fed by the disorganised blood supply they create, which gives rise to regions of low oxygen pressure. One consequence of this switch is that tumour cells mirror what happens in normal cells that are dividing in that carbon skeletons are diverted into the pentose phosphate pathway for macromolecular synthesis. Warburg suggested, incorrectly, that cancers actually occurred because mitochondria developed faults and it was disrupted metabolism that drove tumour formation. We now know that it is mutational changes in sets of genes that act as ‘drivers’ and metabolic perturbation is just one of the consequences. A critical contributor in this context is hyperactive MYC, which regulates many glycolytic genes in its transcription control portfolio and thereby contributes to the Warburg effect.
Warburg’s discovery, reported in the 1920s, languished largely ignored in the literature for some 70 years until technical advances began to reveal the extraordinary behaviour that lies behind his observation. Notwithstanding, there were some significant observations in the intervening period. In the 1940s and 1950s, it was shown that the lethal effect on normal cells of radiation used to treat cancers can largely be blocked if the tissue has previously been exposed to low levels of oxygen (anoxia). Conversely, tumour cells exposed to high levels of oxygen become sensitive to radiation treatment. In an ingenious experiment in which the two halves of a tumour were irradiated separately, one half when the patient was breathing normal air, the other when he was inhaling hyperbaric oxygen (three times atmospheric oxygen levels), the effect of radiation was shown to be much greater on the half irradiated under high oxygen conditions. To understand why this is we must first answer a critical question: how do cells detect changes in oxygen levels.
Various cellular responses, including angiogenesis, are affected by oxygen levels. Indeed the transcription of many genes is activated when cells are exposed to low levels of oxygen, for example, VEGFA. In the 1990s a number of transcription factors – regulatory proteins that control gene expression – were found to respond to hypoxia and named, therefore, hypoxia-inducible factors (HIFs). It was subsequently shown that the regulatory regions of oxygen-responsive genes contain specific sequence motifs to which HIFs bind – and hence control gene expression. Hypoxia-inducible factors are heterodimeric transcription factors composed of alpha subunits (HIF1A, HIF2A (for which the approved gene name is EPAS1) or HIF3A), and a beta subunit (the aryl hydrocarbon nuclear translocator (ARNT)). The levels of HIF1A and HIF2A are frequently up-regulated in tumour cells and, in addition to hypoxia, they can respond independently to activated-oncoproteins (e.g. RAS). Among the genes they regulate are VEGFA, glucose transporters (SLC2A1/GLUT1 and SLC2A5/GLUT3), and enzymes of the glycolytic pathway and regulators of apoptosis. HIFA proteins do not actually ‘sense’ oxygen but their very existence depends on proteins that do. Enzymes that use oxygen directly are oxygenases, and a group of these control the amount of HIF proteins in cells and consequently the cell’s response to oxygen (Fig. 1).
1. Regulation of HIF activity.
When normal levels of oxygen are present (normoxia) HIFA proteins are made but rapidly tagged with an –OH (hydroxyl) group: this is recognised by the cellular machinery responsible for breaking down proteins so that normally HIFA proteins have a very short half-life. The enzyme that does the tagging is an oxygenase (prolyl hydroxylase) and it acts as a sensor of oxygen concentration in the cell. As cells become hypoxic the activity of the oxygenase declines, levels of HIFA proteins rise and their target genes are switched on.
In oxygenated cells, the rapid degradation of HIFA proteins occurs via the ubiquitin-proteasome pathway, mediated by the von Hippel–Lindau (VHL) protein as part of a multimeric ubiquitin ligase complex. VHL is thus a critical regulator of cellular responses to hypoxia and is a ‘tumour suppressor’. Loss of VHL function means that proteins that would normally be degraded by the proteasome continue to function, and some of these contribute to tumour development.