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1.
Blood Adv ; 7(9): 1754-1761, 2023 05 09.
Article in English | MEDLINE | ID: mdl-36521029

ABSTRACT

Cancer cells take up amino acids from the extracellular space to drive cell proliferation and viability. Similar mechanisms are applied by immune cells, resulting in the competition between conventional T cells, or indeed chimeric antigen receptor (CAR) T cells and tumor cells, for the limited availability of amino acids within the environment. We demonstrate that T cells can be re-engineered to express SLC7A5 or SLC7A11 transmembrane amino acid transporters alongside CARs. Transporter modifications increase CAR T-cell proliferation under low tryptophan or cystine conditions with no loss of CAR cytotoxicity or increased exhaustion. Transcriptomic and phenotypic analysis reveals that downstream, SLC7A5/SLC7A11-modified CAR T cells upregulate intracellular arginase expression and activity. In turn, we engineer and phenotype a further generation of CAR T cells that express functional arginase 1/arginase 2 enzymes and have enhanced CAR T-cell proliferation and antitumor activity. Thus, CAR T cells can be adapted to the amino acid metabolic microenvironment of cancer, a hitherto recognized but unaddressed barrier for successful CAR T-cell therapy.


Subject(s)
Neoplasms , Receptors, Chimeric Antigen , Humans , T-Lymphocytes , Receptors, Chimeric Antigen/metabolism , Receptors, Antigen, T-Cell/genetics , Arginase/genetics , Arginase/metabolism , Large Neutral Amino Acid-Transporter 1/metabolism , Neoplasms/metabolism , Amino Acids/metabolism , Tumor Microenvironment
2.
Cell Rep ; 38(5): 110320, 2022 02 01.
Article in English | MEDLINE | ID: mdl-35108535

ABSTRACT

The demands of cancer cell proliferation alongside an inadequate angiogenic response lead to insufficient oxygen availability in the tumor microenvironment. Within the mitochondria, oxygen is the major electron acceptor for NADH, with the result that the reducing potential produced through tricarboxylic acid (TCA) cycle activity and mitochondrial respiration are functionally linked. As the oxidizing activity of the TCA cycle is required for efficient synthesis of anabolic precursors, tumoral hypoxia could lead to a cessation of proliferation without another means of correcting the redox imbalance. We show that in hypoxic conditions, mitochondrial pyrroline 5-carboxylate reductase 1 (PYCR1) activity is increased, oxidizing NADH with the synthesis of proline as a by-product. We further show that PYCR1 activity is required for the successful maintenance of hypoxic regions by permitting continued TCA cycle activity, and that its loss leads to significantly increased hypoxia in vivo and in 3D culture, resulting in widespread cell death.


Subject(s)
Cell Proliferation/physiology , Neoplasms/metabolism , Oxygen/metabolism , Pyrroline Carboxylate Reductases/metabolism , Citric Acid Cycle/physiology , Humans , Mitochondria/metabolism , Proline/metabolism , Tumor Microenvironment , delta-1-Pyrroline-5-Carboxylate Reductase
3.
Amino Acids ; 53(12): 1779-1788, 2021 Dec.
Article in English | MEDLINE | ID: mdl-34291343

ABSTRACT

Proline is a non-essential amino acid with key roles in protein structure/function and maintenance of cellular redox homeostasis. It is available from dietary sources, generated de novo within cells, and released from protein structures; a noteworthy source being collagen. Its catabolism within cells can generate ATP and reactive oxygen species (ROS). Recent findings suggest that proline biosynthesis and catabolism are essential processes in disease; not only due to the role in new protein synthesis as part of pathogenic processes but also due to the impact of proline metabolism on the wider metabolic network through its significant role in redox homeostasis. This is particularly clear in cancer proliferation and metastatic outgrowth. Nevertheless, the precise identity of the drivers of cellular proline catabolism and biosynthesis, and the overall cost of maintaining appropriate balance is not currently known. In this review, we explore the major drivers of proline availability and consumption at a local and systemic level with a focus on cancer. Unraveling the main factors influencing proline metabolism in normal physiology and disease will shed light on new effective treatment strategies.


Subject(s)
Neoplasms/metabolism , Proline/metabolism , Animals , Homeostasis/physiology , Humans , Oxidation-Reduction , Protein Biosynthesis/physiology , Reactive Oxygen Species/metabolism
4.
Cancer Res ; 81(13): 3480-3494, 2021 07 01.
Article in English | MEDLINE | ID: mdl-34127497

ABSTRACT

Succinate dehydrogenase is a key enzyme in the tricarboxylic acid cycle and the electron transport chain. All four subunits of succinate dehydrogenase are tumor suppressor genes predisposing to paraganglioma, but only mutations in the SDHB subunit are associated with increased risk of metastasis. Here we generated an Sdhd knockout chromaffin cell line and compared it with Sdhb-deficient cells. Both cell types exhibited similar SDH loss of function, metabolic adaptation, and succinate accumulation. In contrast, Sdhb-/- cells showed hallmarks of mesenchymal transition associated with increased DNA hypermethylation and a stronger pseudo-hypoxic phenotype compared with Sdhd-/- cells. Loss of SDHB specifically led to increased oxidative stress associated with dysregulated iron and copper homeostasis in the absence of NRF2 activation. High-dose ascorbate exacerbated the increase in mitochondrial reactive oxygen species, leading to cell death in Sdhb-/- cells. These data establish a mechanism linking oxidative stress to iron homeostasis that specifically occurs in Sdhb-deficient cells and may promote metastasis. They also highlight high-dose ascorbate as a promising therapeutic strategy for SDHB-related cancers. SIGNIFICANCE: Loss of different succinate dehydrogenase subunits can lead to different cell and tumor phenotypes, linking stronger 2-OG-dependent dioxygenases inhibition, iron overload, and ROS accumulation following SDHB mutation.


Subject(s)
Ascorbic Acid/pharmacology , Homeostasis , Iron/metabolism , Mutation , Oxidative Stress , Succinate Dehydrogenase/physiology , Animals , Antioxidants/pharmacology , Dioxygenases/antagonists & inhibitors , Female , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Mitochondria/metabolism , Mitochondria/pathology , Phenotype , Reactive Oxygen Species
5.
FASEB J ; 34(1): 303-315, 2020 01.
Article in English | MEDLINE | ID: mdl-31914648

ABSTRACT

Mutations in succinate dehydrogenase (SDH) lead to the development of tumors in a restricted subset of cell types, including chromaffin cells and paraganglia. The molecular basis for this specificity is currently unknown. We show that loss of SDH activity in a chromaffin cell model does not perturb complex I function, retaining the ability to oxidize NADH within the electron transport chain. This activity supports continued oxidation of substrates within the tricarboxylic acid (TCA) cycle. However, due to the block in the TCA cycle at SDH, the high glutamine oxidation activity is only maintained through an efflux of succinate. We also show that although the mitochondria of SDH-deficient cells are less active per se, their higher mass per cell results in an overall respiratory rate that is comparable with wild-type cells. Finally, we observed that when their mitochondria are uncoupled, SDH-deficient cells are unable to preserve their viability, suggesting that the mitochondrial metabolic network is unable to compensate when exposed to additional stress. We therefore show that in contrast to models of SDH deficiency based on epithelial cells, a chromaffin cell model retains aspects of metabolic "health," which could form the basis of cell specificity of this rare tumor type.


Subject(s)
Chromaffin Cells/metabolism , Electron Transport Complex I/metabolism , Mitochondria/metabolism , Mitochondrial Diseases/metabolism , Neoplasms/metabolism , Succinate Dehydrogenase/physiology , Animals , Chromaffin Cells/pathology , Humans , Male , Mice , Mice, Knockout , Mitochondria/pathology , Mitochondrial Diseases/pathology , Mutation , NAD/metabolism , Neoplasms/pathology , Succinate Dehydrogenase/genetics , Succinate Dehydrogenase/metabolism , Transcriptome
6.
Br J Cancer ; 122(2): 150-156, 2020 01.
Article in English | MEDLINE | ID: mdl-31819187

ABSTRACT

An abundant supply of amino acids is important for cancers to sustain their proliferative drive. Alongside their direct role as substrates for protein synthesis, they can have roles in energy generation, driving the synthesis of nucleosides and maintenance of cellular redox homoeostasis. As cancer cells exist within a complex and often nutrient-poor microenvironment, they sometimes exist as part of a metabolic community, forming relationships that can be both symbiotic and parasitic. Indeed, this is particularly evident in cancers that are auxotrophic for particular amino acids. This review discusses the stromal/cancer cell relationship, by using examples to illustrate a number of different ways in which cancer cells can rely on and contribute to their microenvironment - both as a stable network and in response to therapy. In addition, it examines situations when amino acid synthesis is driven through metabolic coupling to other reactions, and synthesis is in excess of the cancer cell's proliferative demand. Finally, it highlights the understudied area of non-proteinogenic amino acids in cancer metabolism and their potential role.


Subject(s)
Amino Acids/metabolism , Cell Proliferation/genetics , Energy Metabolism/genetics , Neoplasms/metabolism , Amino Acids/genetics , Humans , Neoplasms/genetics , Protein Biosynthesis/genetics , Tumor Microenvironment/genetics
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