Asparagine signals mitochondrial respiration and can be targeted to impair tumour growth

Abstract

Mitochondrial respiration is critical for cell proliferation. In addition to producing ATP via the electron transport chain (ETC), respiration is required for the generation of TCA cycle-derived biosynthetic precursors, such as aspartate, an essential substrate for nucleotide synthesis.Because mTORC1 coordinates availability of biosynthetic precursors with anabolic metabolism, including nucleotide synthesis, a link between respiration and mTORC1 is fitting. Here we show that in addition to depleting intracellular aspartate, ETC inhibition depletes aspartate-derived asparagine and impairs mTORC1 activity. Providing exogenous asparagine restores mTORC1 activity, nucleotide synthesis, and proliferation in the context of ETC inhibition without restoring intracellular aspartate in a panel of cancer cell lines. As a therapeutic strategy, the combination of ETC inhibitor metformin, which limits tumour asparagine synthesis, and either asparaginase or dietary asparagine restriction, which limit tumour asparagine consumption, effectively impairs tumour growth in several mouse models of cancer. Because environmental asparagine is sufficient to restore proliferation with respiration impairment, both in vitro and in vivo, our findings suggest that asparagine synthesis is a fundamental purpose of mitochondrial respiration. Moreover, the results suggest that asparagine signals active respiration to mTORC1 to communicate biosynthetic precursor sufficiency and promote anabolism. IntroductionRecent literature has demonstrated that asparagine is important for amino acid homeostasis, maintenance of mTORC1 activity, and tumour progression 1-3 . Asparagine regulation of mTORC1 activity and downstream anabolism may explain the clinical efficacy of extracellularacting asparaginase in the treatment of leukaemias that obtain most of their asparagine from the environment. Most solid tumours, however, are capable of synthesizing asparagine via asparagine synthetase (ASNS), making them less responsive to asparaginase 4 . In addition, elevated ASNS expression, and presumably increased de novo asparagine synthesis, accompanies asparaginase resistance in leukaemia 5 . Because cells can obtain asparagine from the environment or synthesize asparagine de novo, targeting both sources may be required to effectively exploit tumour asparagine dependence.Cellular respiration couples nutrient oxidation to ATP production through oxidative phosphorylation. Although most cancer cells convert the majority of consumed glucose to lactate, concurrent respiration is essential: suppressing respiration through ETC inhibition impairs proliferation [6][7][8][9] . Recent literature has shown that ATP synthesis via the ETC is dispensable for cancer cell proliferation. Rather, aspartate synthesis requirements explain the reliance of proliferating cells on respiration 6,7 . Electron acceptors are limiting upon ETC inhibition, resulting in compromised NAD+ recycling, impaired flux through the TCA cycle, and depletion of TCA cycle-derived aspartate. Supplementing cell cu...

Glutamine reliance in cell metabolism

“…Although the clinical effect of L-asparaginase clearly indicates that asparagine is crucial for tumor survival and metastasis 52 , the importance of asparagine beyond protein synthesis and the mechanism by which asparagine supplementation enhances glutamine-associated metabolism are less well understood. Recently, asparagine has been reported to function as an exchange factor needed for the uptake of other amino acids that are required for mTORC1 activation 53 and for enhanced nucleotide biosynthesis under mitochondrial electron chain transport system impairment 54 . Further investigation is needed to explain the considerable mechanistic importance of asparagine in cancer metabolism.…”

Mitochondrial dynamics and oxidative phosphorylation as critical targets in cancer

“…Overall, the importance of ETC-dependent redox balance has been confirmed across a range of cancer contexts. To note, it has been proposed that the main role of the ETC was to generate aspartate for further conversion to asparagine via the action of asparagine synthetase (ASNS) (Krall et al 2021) (Fig. 3B, left panel).…”

“…Higher ASNS expression can correlate with poorer patient survival in breast, bladder and squamous head and neck cancers as well as renal clear cell carcinoma patients. In this way, gene targeting of ASNS, dietary restriction of asparagine and reduction of bioavailable asparagine (by treatment with l-asparaginase) are being explored therapeutically in pre-clinical breast and lung cancer mouse models (Hettmer et al 2015, Knott et al 2018, Krall et al 2021. In this regard, asparagine limitation has been a longestablished strategy to treat certain types of leukaemia that are highly dependent upon asparagine availability (Richards & Kilberg 2006).…”

The biological underpinnings of therapeutic resistance in pancreatic cancer

“…Like pyruvate, asparagine has similarly been shown to promote therapeutic resistance and support tumor growth upon inhibition of complex I (Fig. 2B; Halbrook et al 2020;Krall et al 2020). Asparagine promotes therapeutic resistance and permits tumor growth by supporting biosynthesis and aspartate availability, in both mTOR-dependent and -independent manners.…”