by ecancer reporter Clare Sansom

Cancer cells exhibit significant differences in basic metabolism from their normal equivalents; these differences are driven by mutations and are necessary to maintain their characteristic, constantly dividing phenotype.

Differences in metabolism can cause cellular vulnerabilities that may be exploited in the design of novel cancer therapies.

One essential metabolic pathway that has been targeted by carcinogenic mutations is the tricarboxylic acid (TCA) pathway, which generates energy in the form of ATP and the precursors of a number of essential molecules through the oxidation of acetate derived from carbohydrates.

Mutations in three enzymes in this pathway – succinate dehydrolase (SDH), fumarate hydratase (FH) and isocitrate dehydrogenase – have recently been discovered in tumour cells.

Succinate dehydrolase is a multi-subunit enzyme that catalyses the oxidation of succinate to fumarate and generates electrons for the production of ATP via mitochondrial respiration.

Deactivating mutations in subunits of this complex have been associated with several tumour types, including neuroendocrine tumours and renal cell carcinoma (RCC).

Loss of this protein, which has been classed as a tumour suppressor, causes an accumulation of succinate and the development of a hypoxic phenotype.

However, the metabolic re-wiring that allows tumour cells to grow and divide without this important enzyme is not yet completely understood.

A group of researchers led by Eyal Gottlieb of the Cancer Research UK Beatson Institute, Glasgow, UK has now demonstrated the metabolic adaptations to complete loss of this complex using a mouse cell model.

Gottlieb and his co-workers generated a strain of transgenic mice in which the gene for the B subunit of SDH, Sdhb, could be deleted by adenovirus infection, isolated and immortalised primary kidney epithelial cells (Sdhb fl/fl cells) from these mice, and then selectively deleted Sdhb to produce a Sdhb Δ/Δ cell line,

All activity of the SDH complex was lost in the Sdhb Δ/Δ cells, leading to a build-up of succinate and a consequent drop in the level of fumarate.

Culturing both cell lines in media containing uniformly labelled 13C-glucose or 13C-glutamine showed that glutamine was a major source of carbon for the TCA cycle in both Sdhb fl/fl and Sdhb Δ/Δ cells.

Succinate processing continued beyond the step catalysed by SDH in the Sdhb fl/fl cells but not the Sdhb Δ/Δ cells, indicating that loss of this enzyme was sufficient to block the cycle.

The researchers next investigated the effect of SDH loss on the rate of cellular oxygen consumption and mitochondrial respiration.

The SDH deficient cells were found to consume oxygen at a lower rate than the cells with active SDH, although this reduced rate was close to their bioenergetic limit.

This decline in oxygen consumption was associated with a specific loss of respiratory complex I in the mitochondria that has also been observed in SDH null neuroendocrine tumours.

These findings indicate that SDH null cells must generate their ATP through glycolysis.

The Sdhb Δ/Δ cells were found to consume extracellular pyruvate at approximately 2.5 times the rate of their Sdhb fl/fl counterparts, and this pyruvate consumption was seen to be necessary for glycolysis.

Metabolic profiling of Sdhb Δ/Δ cells in the presence and absence of pyruvate showed that the concentration of the amino acid aspartate was significantly decreased in these cells, and that aspartate depletion increased when pyruvate was removed from the medium.

Further experiments showed that the enzyme pyruvate carboxylase is necessary for the formation of oxaloacetate, a precursor of acetate, in Sdhb Δ/Δ cells.

This enzyme was also shown to be over-expressed in several SDH-deficient human tumour cell lines.

Silencing the pyruvate carboxylase gene Pcx using short hairpin RNA in SDH-null cells reduced the incorporation of radiolabelled carbon atoms from glucose into aspartarte, and also reduced the capacity of these cells to form tumours.

Taken together, these results indicate that pyruvate carboxylase is an essential enzyme in SDH deficient tumour cells, that it defines a metabolic vulnerability and that it might, therefore, be a useful target for drugs against these tumours.

Reference

Cardaci, S., Zheng, L., MacKay, G. and 14 others (2015). Pyruvate carboxylation enables growth of SDH-deficient cells by supporting aspartate biosynthesis. Nature Cell Biology, published online ahead of print 24 August 2015.