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Metabolic pathway analysis using stable isotopes in patients with cancer

Nature Reviews Cancer volume 23, pages 863–878 (2023)Cite this article

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Abstract

Metabolic reprogramming is central to malignant transformation and cancer cell growth. How tumours use nutrients and the relative rates of reprogrammed pathways are areas of intense investigation. Tumour metabolism is determined by a complex and incompletely defined combination of factors intrinsic and extrinsic to cancer cells. This complexity increases the value of assessing cancer metabolism in disease-relevant microenvironments, including in patients with cancer. Stable-isotope tracing is an informative, versatile method for probing tumour metabolism in vivo. It has been used extensively in preclinical models of cancer and, with increasing frequency, in patients with cancer. In this Review, we describe approaches for using in vivo isotope tracing to define fuel preferences and pathway engagement in tumours, along with some of the principles that have emerged from this work. Stable-isotope infusions reported so far have revealed that in humans, tumours use a diverse set of nutrients to supply central metabolic pathways, including the tricarboxylic acid cycle and amino acid synthesis. Emerging data suggest that some activities detected by stable-isotope tracing correlate with poor clinical outcomes and may drive cancer progression. We also discuss current challenges in isotope tracing, including comparisons of in vivo and in vitro models, and opportunities for future discovery in tumour metabolism.

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Fig. 1: Basic concepts of isotope labelling.
Fig. 2: Metabolic rewiring in different kinds of human cancer.
Fig. 3: Shared and divergent metabolic properties in cultured cells, mice and patients.
Fig. 4: Limitations of stable-isotope tracing studies in patients with cancer.
Fig. 5: Future directions of research in cancer metabolism in vivo.

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Data availability

The data supporting the findings displayed in Fig. 3 are available in Supplementary Table 1.

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Acknowledgements

R.J.D. is supported by the Howard Hughes Medical Institute and by US National Cancer Institutes grants R35CA22044901, P50CA196516-06A1 and 2P50CA070907-21A1. B.F. is supported by US National Cancer Institutes grant R00CA237724.

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Author notes

  1. These authors contributed equally: Caroline R. Bartman, Brandon Faubert.

  2. These authors jointly supervised this work: Joshua D. Rabinowitz, Ralph J. DeBerardinis.

Authors and Affiliations

  1. Lewis-Sigler Institute for Integrative Genomics, Princeton University, Princeton, NJ, USA

    Caroline R. Bartman & Joshua D. Rabinowitz

  2. Department of Medicine, Section of Hematology/Oncology, University of Chicago, Chicago, IL, USA

    Brandon Faubert

  3. Howard Hughes Medical Institute and Children’s Research Institute, University of Texas Southwestern Medical Center, Dallas, TX, USA

    Ralph J. DeBerardinis

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Contributions

All authors researched the literature and contributed to the conceptualization of the text and figures for this Review. C.R.B. and B.F. wrote the first draft, which was edited by J.D.R. and R.J.D. All authors participated in revising the final version of the paper.

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Correspondence to Joshua D. Rabinowitz or Ralph J. DeBerardinis.

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R.J.D. is a founder and adviser for Atavistik Bioscience and an adviser for Agios Pharmaceuticals, Vida Ventures and Droia Ventures. The other authors declare no competing interests.

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Supplementary information

Glossary

18-Fluorodeoxyglucose-PET

(18FDG-PET). 18Fluorodeoxyglucose (FDG) is an analogue of glucose conjugated with a radioisotope of fluorine (18F). FDG is imported into tissues and tumours using the same transporters that import glucose, for example, GLUT1, and the tracer is retained after phosphorylation by hexokinase. The localized accumulation of 18F can be imaged by positron emission tomography. FDG uptake is increased in many tumours, so this technique can be used to diagnose and stage multiple cancers.

Acetyl-CoA carboxylase

The first enzyme of fatty acid synthesis, which catalyses the carboxylation of acetyl-CoA to produce malonyl-CoA for fatty acid synthesis.

Anaplerotic contribution

Metabolic pathways that contribute to recover or replenish catalytic intermediates, particularly in the TCA cycle. Examples include the use of glutamine or pyruvate to provide 4-carbon and 5-carbon intermediates to the TCA cycle.

Antifolates

Drugs that inhibit folate-using enzymes, including three of the enzymes required for nucleotide biosynthesis. Several chemotherapeutic agents, such as pemetrexed and methotrexate, are antifolates.

Fatty acid synthase

(FASN). A multifunctional enzyme that catalyses several reactions required to produce fatty acids from precursors, including malonyl-CoA and acetyl-CoA.

Folate cycle

A metabolic pathway producing one-carbon metabolites for use in numerous biochemical reactions, including nucleotide synthesis.

Fractional contribution

Refers to the fraction of a metabolite pool arising from a particular precursor. In stable-isotope tracing experiments, transfer of the isotope from the labelled precursor such as [13C]glucose to a metabolite, for example, pyruvate, provides information about the fractional contribution of that tracer to that metabolite.

Hyperpolarization

A molecular state in which the nuclear spin is polarized well beyond thermal equilibrium, increasing the ability to detect the nucleus by NMR. Hyperpolarization of 13C nuclei can be achieved by transferring to 13C the high polarization state of free electrons contained in a radical.

Imaging mass spectrometry

An analytical technique that enables visualization of the spatial distribution of metabolites within a tissue sample.

Lactate dehydrogenase

(LDH). A tetrameric, reversible NAD(H)-dependent enzyme that interconverts pyruvate and lactate.

Metabolic flux analysis

An algorithm to calculate metabolic fluxes in cultured cells, which takes as inputs the consumption and production rates of media nutrients, 13C labelling of cellular metabolites from 13C isotope tracers such as glucose and glutamine and the known reaction structure of metabolic pathways.

Metabolic fluxes

The rates at which metabolites flow through biochemical reactions.

Monocarboxylate transporters

A family of membrane proteins that transport monocarboxylates such as lactate, pyruvate and ketone bodies across the plasma membrane.

Monoclonal gammopathy of undetermined significance

A condition characterized by the accumulation of abnormal monoclonal immunoglobulin in the blood. Monoclonal gammopathy of undetermined significance can be a precursor to multiple myeloma or other disorders.

Nuclear spins

The intrinsic angular momentum of an atomic nucleus.

Nucleoside analogues

Synthetic compounds that structurally mimic endogenous nucleosides and are used as chemotherapeutic agents. These analogues can be incorporated into the DNA of rapidly dividing cells, disrupting DNA replication.

Nutrostat

Cell culture system designed to maintain consistent levels of extracellular nutrients and other metabolites by providing a consistent source of fresh medium to the culture.

Pyruvate carboxylation

A mitochondrial anaplerotic activity in which the enzyme pyruvate carboxylase converts pyruvate to oxaloacetate.

Von Hippel–Lindau (VHL) tumour suppressor

A tumour suppressor whose physiological role is to target the α subunits of hypoxia-inducible factors (HIF1α and HIF2α) for degradation when oxygen is present. In VHL-deficient tumours such as ccRCC, HIF1α and HIF2α are stabilized regardless of oxygen availability. The resulting ‘pseudohypoxic’ state causes chronic expression of genes involved in the hypoxic response.

Warburg metabolism

Named after Otto Warburg and also known as aerobic glycolysis, this classic metabolic phenotype of tumours and proliferating cells involves the brisk conversion of glucose to lactate in the presence of sufficient oxygen to oxidize glucose to CO2.

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Bartman, C.R., Faubert, B., Rabinowitz, J.D. et al. Metabolic pathway analysis using stable isotopes in patients with cancer. Nat Rev Cancer 23, 863–878 (2023). https://doi.org/10.1038/s41568-023-00632-z

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