Clonal Heterogeneity Supports Mitochondrial Metabolism in Pancreatic Cancer

Abstract

Pancreatic ductal adenocarcinoma (PDA) is characterized by a heterogenous and densely fibrotic microenvironment. This limits functional vasculature and diffusion of nutrients through the tumor 1,2 . Accordingly, pancreatic cancer cells develop numerous metabolic adaptations to survive and proliferate in nutrient austere conditions 3-7 . Subtypes of PDA have been characterized by transcriptional and functional differences [8][9][10][11][12] , which have been reported to exist within the same tumor 13-15 . However, it remains unclear if this diversity extends to metabolic programming. Here, using a combination of metabolomic profiling and functional interrogation of metabolic dependencies, we identify two distinct metabolic subclasses within neoplastic populations isolated from a single pancreatic tumor. Furthermore, these populations are poised for metabolic crosstalk, and in examining this, we find an unexpected role for asparagine in maintaining cell proliferation following mitochondrial inhibition. Functionally, when challenged by mitochondrial inhibition, asparagine supplementation increases intracellular levels of asparagine and aspartate, a rate limiting biosynthetic precursor [16][17][18] . Conversely, depletion of extracellular asparagine with PEG-asparaginase sensitizes pancreatic tumors to mitochondrial targeting with phenformin. Together, these data extend the concept of metabolic diversity to neoplastic populations within individual tumors, while illustrating a new method of intratumoral communication that supports tumor fitness 19,20 . Finally, the combination of asparaginase with mitochondrial inhibition could provide a powerful new strategy for this difficult to treat disease. Introduction:The tumor microenvironment in pancreatic ductal adenocarcinoma (PDA) is a complex ecosystem 21 , with diverse populations of fibroblasts and immune cells functioning to create a niche supporting cancer cell survival and tumor growth 19,20,[22][23][24][25][26][27][28] . Accordingly, this allows for many and varied cooperative intratumoral crosstalk interactions in the microenvironment 29,30 . These extend to direct metabolic support of cancer cells from adjacent fibroblasts and tumor associated macrophages [31][32][33][34][35] .The majority of PDAs express mutant Kras, so early efforts to understand metabolism in pancreatic cancer focused on the cell intrinsic metabolic rewiring downstream of Kras signaling pathways [36][37][38] . In addition, extensive metabolic profiling of a large set of PDA cell lines revealed *

Asparagine: A Metabolite to Be Targeted in Cancers

“…Recently, the same group found that the ability of asparagine to activate mTORC1 represents a key adaptive strategy for tumor cells to mitigate ETC inhibition-induced stress, which is coupled to ATF4 activation [ 84 ]. This work provides an alternative explanation for the synergy between ETC inhibition and asparagine restriction [ 79 ] ( Figure 3 A), and also brings up the question of whether asparagine or aspartate is the most limiting metabolite to drive ETC inhibition-associated phenomenon in tumor cells. Furthermore, in addition to functioning as an exchange factor for mTORC1 activation, asparagine can also activate mTORC1 directly through an ADP-ribosylation factor 1 (Arf1)-dependent but Rag GTPase-independent mechanism [ 85 ] ( Figure 3 A).…”

“…The ability of asparagine to modulate tumor progression can also be reflected at the stage when tumor cells respond to therapeutics. A recent study (in preprint) shows that clonal heterogeneity within a tumor tissue can contribute to therapeutic resistance through asparagine biosynthesis [ 79 ]. In a mouse pancreatic tumor model, the authors discovered metabolic heterogeneity within a tumor tissue, which leads to differential responses to electron transfer chain inhibitors (ETCi).…”

“…Through comprehensive metabolic analysis, the manuscript identified asparagine as a key metabolite secreted from the resistant clones, which can feed the sensitive clones to drive their resistance to ETCi. Since a major tumor suppressive effect of ETCi is via restricting aspartate production [ 81 , 82 ], the manuscript found that asparagine supplementation can preserve aspartate pool in sensitive clones likely by diverging aspartate from asparagine biosynthesis [ 79 ]. Furthermore, L-asparaginase treatment sensitizes tumor cells to phenformin, a complex I inhibitor, in an allograft mouse pancreatic tumor model in vivo.…”

A Targeted Bioinformatics Assessment of Adrenocortical Carcinoma Reveals Prognostic Implications of GABA System Gene Expression

“…These data support the existence of a GABA shunt-centric inter-tumoral metabolic heterogeneity among the ACC cohort. Interestingly, an intra-tumoral heterogeneity of GABA levels has recently been reported in clonal cell lines of distinct metabolic phenotypes (e.g., glycolytic vs mitochondrial) derived from single tumor ex vivo mouse pancreatic cancer cells [ 50 ]. Evaluation of both intra- and inter-tumoral heterogeneity of the GABA shunt may be important for understanding its role in mitochondrial metabolic heterogeneity, a feature of tumors that is increasingly recognized to influence cancer growth, progression, and drug treatment sensitivity [ 51 ].…”

Impact of cancer metabolism on therapy resistance – Clinical implications

“…Recent studies have shown upregulation of asparagine synthetase (ASNS) (Williams et al, 2020) or aspartate/glutamate transporter SLC1A3 (Sun et al, 2019b) as resistance mechanisms to asparaginase in several solid tumor types. Another study has reported that Asn metabolism mediates resistance to OXPHOS inhibitors in PaC (Halbrook et al, 2020) and identifies Asndepleting modalities (genetic silencing of ASNS and depletion of exogenous Asn upon asparaginase treatment) as potential therapeutic strategies to re-sensitize pancreatic tumor cells to phenformin treatment. In addition, it has been shown that Phenformin is a genuine tumor disruptor not only by producing hypoglycemia due to caloric restriction via energysensing AMP-activated protein kinase, but also as a blocker of the mTOR regulatory complex (Rubiño et al, 2019).…”