13C tracer analysis reveals the landscape of metabolic checkpoints in human CD8+ T cell differentiation and exhaustion

Abstract

IntroductionNaïve T cells remain in an actively maintained state of quiescence until activation by antigenic signals, upon which they start to proliferate and generate effector cells to initiate a functional immune response. Metabolic reprogramming is essential to meet the biosynthetic demands of the differentiation process, and failure to do so can promote the development of hypofunctional exhausted T cells.MethodsHere we used 13C metabolomics and transcriptomics to study the metabolism of CD8+ T cells in their complete course of differentiation from naïve over stem-like memory to effector cells and in exhaustion-inducing conditions. ResultsThe quiescence of naïve T cells was evident in a profound suppression of glucose oxidation and a decreased expression of ENO1, downstream of which no glycolytic flux was detectable. Moreover, TCA cycle activity was low in naïve T cells and associated with a downregulation of SDH subunits. Upon stimulation and exit from quiescence, the initiation of cell growth and proliferation was accompanied by differential expression of metabolic enzymes and metabolic reprogramming towards aerobic glycolysis with high rates of nutrient uptake, respiration and lactate production. High flux in anabolic pathways imposed a strain on NADH homeostasis, which coincided with engagement of the proline cycle for mitochondrial redox shuttling. With acquisition of effector functions, cells increasingly relied on glycolysis as opposed to oxidative phosphorylation, which was, however, not linked to changes in mitochondrial abundance. In exhaustion, decreased effector function concurred with a reduction in mitochondrial metabolism, glycolysis and amino acid import, and an upregulation of quiescence-associated genes, TXNIP and KLF2, and the T cell suppressive metabolites succinate and itaconate. DiscussionOverall, these results identify multiple metabolic features that regulate quiescence, proliferation and effector function, but also exhaustion of CD8+ T cells during differentiation. Thus, targeting these metabolic checkpoints may be a promising therapeutic strategy for both prevention of exhaustion and promotion of stemness of anti-tumor T cells.

Glycolysis in the tumor microenvironment: a driver of cancer progression and a promising therapeutic target

“…One indicator of the activation of CD8 + T cells is the transition from OXPHOS to aerobic glycolysis, and the mTORC1-HIF-1 pathway is critical for mediating this metabolic transition ( Finlay et al, 2012 ; Kirchmair et al, 2023 ). The expression of glycolytic proteins and the rate of glycolysis were greater in differentiated CD8 + effector T cells than in naïve T cells.…”

“…Glycolysis not only promotes tumor growth but also modifies the TME by impacting immune cell and stromal cell development, activation, and function. Nonmalignant cells, such as immune cells ( Kirchmair et al, 2023 ) and cancer-associated fibroblasts ( Yan et al, 2018 ), undergo metabolic reprogramming during carcinogenesis that is notable for its increased dependence on glycolysis. Additionally, by promoting the production of metabolites and signaling chemicals, glycolysis of tumor cells can positively alter the TME ( Guo et al, 2022 ; Liu et al, 2023 ).…”

Importance of Michaelis Constants for Cancer Cell Redox Balance and Lactate Secretion—Revisiting the Warburg Effect

“…Recent tracer studies have shown that the carbon atoms provided by fed glucose appear in lactate (also produced by the heart muscle) and in other downstream metabolites. In turn, lactate is also routinely utilized by virtually all tissues to feed the TCA cycle and other pathways [28,[64][65][66][67][68][69][70] (see Figure 1). Moreover, lactate can even stimulate its own use by mitochondria by yet unknown mechanisms, independent of its actual use [69].…”