Tumours require the non-essential amino acids serine and glycine for growth
Feeding a diet lacking serine and glycine results in reduced growth of a subset of mouse tumours, but some oncogenic mutations upregulate the endogenous serine synthesis pathway, making dietary restriction ineffective
Inhibition of the serine synthesis pathway combined with a diet free of serine and glycine blocks tumour growth
Stratification of tumours based on their nutritional requirements, and precise manipulation of nutrient availability may augment the efficacy of existing cancer treatments
In order to proliferate and survive, cancer cells must acquire distinct metabolic adaptations. Dependence on these novel metabolic requirements may make tumours vulnerable to therapies limiting the amounts of key nutrients, and the possibility that this might be accomplished by dietary modification has been explored for many years.
As well as studies assessing the effects of making large-scale changes such as caloric restriction or feeding ketogenic diets, more nuanced approaches involving selective removal of defined nutrients have been tested. Karen Vousden’s laboratory has been at the forefront of this latter field, showing in the last quinquennium that limiting the availability of the non-essential amino acids serine and glycine has profound effects on tumour growth, and that how this is accomplished must be tailored to individual cancer genotypes.
Serine plays critical roles in a number of metabolic pathways, including nucleotide and lipid synthesis, methylation reactions, and antioxidant defence through glutathione synthesis. In conditions of dietary restriction, the body can manufacture serine either by serine-glycine interconversion, or from glucose via the serine synthesis pathway, which converts the glycolytic intermediate 3-phosphoglycerate to serine by successive action of the enzymes phosphoglycerate dehydrogenase (PHGDH), phosphoserine aminotransferase 1 (PSAT1), and phosphoserine phosphatase (PSPH).
In a landmark paper in 2013 (Maddocks et al, 2013), Vousden and co-workers showed that the human colon cancer cell line HCT116 was extremely dependent on exogenous serine, and in its absence, despite cells switching on the serine synthesis pathway, growth was impaired. The ability of the cells to survive serine deprivation was dependent on p53, as p53 null cells exhibited oxidative stress, reduced viability and even more severely impaired proliferation. The importance of exogenous serine was confirmed in vivo: if HCT116 cells were injected subcutaneously into mice they rapidly formed tumours, but tumour growth and lethality were both significantly slowed if the animals were fed a diet lacking serine and glycine, which they tolerated well.
This paper was notable not just for its demonstration of a tumour’s metabolic vulnerability upon p53 loss, but also because it overturned previous assumptions that diets lacking non-essential amino acids would be ineffective in altering endogenous levels, due to the body’s ability to compensate by synthesising more. In subsequent years, diets depleted in similar ways were shown by a number of groups to have an impact on survival of animals carrying xenografts and allografts (reviewed in Tajan and Vousden, 2020).
In 2017, Maddocks and colleagues began to explore oncogene-specific differences in responses to dietary deprivation of serine and glycine, using genetically engineered autochthonous mouse models of cancer, which allow more clinically relevant investigation of whole-body metabolic effects.
The group used two rapid onset tumour models: intestinal adenomas in mice heterozygous for the Apc gene (Apc+/-), and lymphomas driven by constitutive Myc expression (Eμ-Myc). Feeding a diet lacking serine and glycine (the -SG diet) to both strains reduced tumour burden, and this held true even if the switch to the -SG diet was made after tumour induction: to mimic a therapeutic intervention, in an inducible Apc+/- model, introduction of the -SG diet after tumour induction significantly increased survival, and this was also the case for HCT116 xenografts and allografts of Eμ-Myc-driven lymphomas.
If cells are starved of exogenous serine, they switch on the endogenous serine synthesis pathway by diverting glycolytic intermediates away from energy production. To maintain ATP production, oxidative phosphorylation is increased, with a concomitant build up in reactive oxygen species (ROS). To prevent unacceptable levels of damage due to oxidative stress, cells must then switch on their antioxidant response, suggesting that antagonising this process could further improve survival and negatively affect tumour growth. This proved to be the case: treating serine and glycine-starved intestinal tumour organoids with a ROS inducer decreased growth and conversely, limiting ROS helped cells starved of serine and glycine to survive.
These data were confirmed in a mouse model in which the Tigar gene, which can support tumour development by limiting ROS, was deleted. Fed normal diets, double Tigar-/- Eμ-Myc mutants rapidly died of lymphoma, but on the -SG diet, there was a significant additive increase in survival. As many chemotherapeutics and radiotherapy induce ROS, these results suggest there may be a benefit to combining the -SG diet with standard anti-cancer treatments.
The -SG diet can clearly inhibit tumour growth in these two mouse models, but it has little effect on tumours carrying Kras mutations: in two models of Kras-driven pancreatic ductal adenocarcinoma there was no significant change in survival in mice fed the -SG diet, suggesting that Kras driven tumours are less reliant on exogenous serine. This was indeed the case: Kras can upregulate the enzymes of the serine synthesis pathway, and conferred resistance to the -SG diet when co-expressed in the APC-driven model of intestinal adenoma.
Taken together, these data demonstrate that in mouse models, dietary limitation of serine and glycine can be effective in the treatment of some cancers, and could be a useful adjunct to therapies which induce ROS. However, Kras-driven tumours, by dint of upregulation of endogenous serine synthesis, are resistant to serine and glycine starvation. Subsequent to this paper, other oncogenic drivers have also been shown to increase expression of the enzymes of the serine synthesis pathway.
These differences in response to the -SG diet are reminiscent of the wide variation in responses to targeted drug therapies, and underline the necessity of developing a similar precision approach to dietary intervention, based on a thorough understanding of the genetic landscape of a tumour, and most likely, its tissue of origin and cellular milieu. This was further emphasised in a recent paper from Vousden and coworkers, where they analysed two of the commonest hotspot mutations in human p53-driven cancers (Humpton et al, 2018). Having shown that complete p53 loss renders tumour cell lines susceptible to serine and glycine deprivation, the paper finds that cells carrying one mutant, R273H, behave similarly to p53 null cells, but that cells expressing the R248W p53 mutant have retained wild-type p53’s ability to confer resistance to a -SG diet. Tumours containing this particular p53 mutant have selectively retained the arm of the p53 response that allows cells to adapt to serine starvation, due to their ability to switch on the antioxidant response to ROS. Combined with the loss of the cell elimination functions of wild-type p53, this gives rise to more aggressive tumours that are resistant to dietary serine modulation.
These results were recapitulated in xenografts, and their relevance to human disease is underscored by a comparison of survival rates between patients with R248W versus R175H mutations: ~4 and ~7 years respectively.
From these papers, Vousden and colleagues, together with others in the field, had created a convincing case for exploring the therapeutic possibilities of combining dietary restriction of serine with increasing redox stress. With the recent development of PH755, a highly potent and selective inhibitor of PHGDH (the first enzyme in the serine synthesis pathway), it was now possible to look at the effect of completely blocking serine availability in tumours.
In Tajan et al (2021), Vousden and coworkers first demonstrated the efficacy of PH755 in vitro, showing that the growth of a variety of colorectal cancer cell lines carrying different oncogenic mutations was completely inhibited when they were cultured in -SG medium in the presence of the PHGDH inhibitor. The effect was specific to PHGDH inhibition, as it could be phenocopied by CRISPR-mediated knockout of the gene. In intestinal tumour organoids, either from mice or derived from patient colon carcinoma samples, the result held true irrespective of the oncogenic driver mutation: inhibition of PHGDH synergised with -SG medium to become a highly effective inhibitor of organoid growth.
How toxic is the combination of the PHGDH inhibitor and a -SG diet? Wild-type mice fed the combined treatment for 20 days had some weight loss but stayed active and apparently healthy. Crucially, their brains, kidneys and livers appeared histologically normal, suggesting that short-term treatment with the combination does not cause long-term damage.
To mimic a potential therapeutic intervention, mice xenografted with two different tumour cell lines were moved to a -SG diet when tumours became evident, and then treated with the PHGDH inhibitor two to four days later. For both lines, the combination treatment was highly effective, strongly inhibiting tumour growth.
These xenograft models are proof of the concept that limiting the availability of serine and glycine to tumours by targeting the exogenous and endogenous supply of serine can have therapeutic benefit. It remains possible that long term treatment would be toxic, so a short term application for therapy is likely to be required — perhaps as an augmentation of cycles of chemo- or radiotherapy. Further, as discussed above, the nature of the intervention will need to be tailored to the tumour, as there is clearly a spectrum of vulnerability; the molecular mechanisms underlying these differences in vulnerability are now under investigation in multiple laboratories. To develop this and other precision nutrition approaches with the hope of marrying them to established tumour therapies and bringing them to the clinic, Vousden has recently co-founded a company, Faeth Therapeutics.