Targeting cancer’s sweet spot: UGP2 as a therapeutic vulnerability

ABSTRACT

Understanding the mechanisms governing metabolic reprogramming that underlie potential vulnerabilities in cancer cells is key to developing novel therapeutic strategies. The catalytic enzyme UDP-glucose pyrophosphorylase 2 (UGP2) drives the production of UDP-glucose. Our recent work demonstrated the crucial role of UGP2 in cancer growth and its regulation of cellular metabolic processes.

UGP2 in disease

Intractable pancreatic ductal adenocarcinomas (PDACs) reprogram their metabolism to survive in harsh tumor microenvironments. The metabolism of glucose can support many anabolic processes that depend on the generation of UDP-glucose by the enzyme UDP-glucose pyrophosphorylase 2 (UGP2). Model organisms such as Dictyostelium and Arabidopsis possess two genes with UGP activity, but these have relatively low homology with the single human UGP2 that can catalyze UDP-glucose from glucose-1-phosphate (Glc-1-P).^1,2 Zebrafish with ugp2 depletion modeled developmental phenotypes induced by reduced enzymatic activity and demonstrated that organisms with impaired UGP2 function during development can remain viable.³ While complete deletion of Ugp2 is embryonic lethal in mice, homozygous genomic alterations eliminating the start codon of the short isoform of UGP2 in humans have been causally linked to developmental epileptic encephalopathy.³

Data from the Cancer Genome Atlas revealed that high UGP2 expression correlated with worse prognosis in PDAC, non-small cell lung cancer, and stomach adenocarcinoma, but not hepatocellular carcinoma. UGP2 expression was also positively correlated with poor prognosis in gallbladder cancer and glioma. In our study, we demonstrated that UGP2 expression correlates strongly with poor prognosis in PDAC tumors, particularly in early phase and low-grade tumors.⁴ We further found that partial depletion of UGP2 in a panel of PDAC lines halted their growth in two-dimensional culture, had a stronger effect in three-dimensional culture, and largely halted tumor growth and proliferation in xenograft models. Interestingly, while mutant Kirsten rat sarcoma (KRAS)-expressing MCF10A cells relied on UGP2 to form colonies, non-transformed MCF10A cells lacking an oncogene did not. This result suggests that UGP2 is not universally essential in all systems. The non-essentiality of UGP2 is further supported by CRISPR knockout data in the DepMap database, opening the possibility of avoiding global stromal toxicity when inhibiting UGP2 in tumors. UGP2 inhibitors under development could be excellent tools to dissect the variability in dependency on UGP2 and the variable correlations between UGP2 expression and prognostic outcomes across cancers with different tissues of origin.

Regulation of UGP2 expression

Despite the central role of UGP2 in carbohydrate synthesis and metabolism, little is known about the regulatory signals that control UGP2 expression and activity (Figure 1a). In yeast, UGP has been shown to be phosphorylated and inhibited by the Per-Arnt-Sim kinase (PASK).⁵ In Arabidopsis, sucrose feeding and cold exposure, as well as phosphate deficiency stress, leads to UGP upregulation,⁶ whereas okadaic acid, an inhibitor of protein phosphatases 1 (PP1) and 2A (PP2A) inhibits the expression of UGP. In mouse embryonic fibroblasts, Ugp2 is transcriptionally regulated via its promoter by mixed lineage leukemia 1 (MLL1) regulation of H3K4 trimethylation and enforced expression of UGP2 partially rescued the effects of MLL1 deletion on increasing the efficacy of the glycosylation inhibitor tunicamycin.⁷

Figure 1.

(a), Mechanisms of UDP-glucose pyrophosphorylase 2 (UGP2) regulation and action in cancer. GALE, UDP-galactose-4-epimerase; GALT, galactose-1-phosphate uridyltransferase; GFPT1, glutamine-fructose-6-phosphate transaminase 1; GMPPB, GDP-mannose pyrophosphorylase (b); MLL1, mixed lineage leukemia 1; MPI, mannose phosphate isomerase; PGM3, phosphoglucomutase 3; PMM2, phosphomannomutase 2. (b); TEAD, TEA domain transcription factor; YAP, yes-associated protein. (b), Gene ontology of proteins with UGP2-regulated N-glycosylation.⁴

In our study, we identified the Yes-associated protein (YAP)-TEA domain transcription factor (TEAD) complex as a direct transcriptional regulator of UGP2 in PDAC cells and found that the expression levels and patterns of YAP and UGP2 were positively correlated in PDAC patient samples.⁴ When YAP was depleted to minimal levels, UGP2 expression was decreased but not absent, suggesting the existence of other UGP2 regulatory factors. We observed that MCF10A cells engineered to express activated KRAS mutants displayed elevated UGP2 transcripts and proteins.⁴ The intermediary regulatory steps between KRAS activation and UGP2 expression remain unclear, but the phenotype was not reversible by mitogen-activated protein kinase kinase (MAP2K1, best known as MEK) inhibition suggesting that it does not act through mitogen-activated protein kinase (MAPK1, best known as ERK). Although YAP is not a direct downstream effector of KRAS, YAP and KRAS share related regulatory patterns, and in murine models constitutive YAP activity is able to rescue growth of mutant KRAS-driven tumors that have been depleted of their driver oncogene.

Functional roles of UGP2

UDP-glucose can contribute to multiple metabolic pathways in cells through the activity of several enzymes.² UDP-glucose ceramide glycosyltransferase, located in the Golgi apparatus, can utilize UDP-glucose for the de novo production of glucosylceramide, which is the precursor for all glycosphingolipids. UDP-glucose-6-dehydrogenase converts UDP-glucose to UDP-glucuronate, contributing to hyaluronan or heparan sulfate synthesis. In addition, glycogen synthase can use UDP-glucose for the synthesis of glycogen, a multi-branched polymer of glucose, that serves as a form of energy storage in case of glucose shortage. We have shown that in PDAC cells, UGP2 acts as a positive regulator of glycogen, particularly under nutrient-depleted conditions.⁴ When UGP2 activity is lacking, the Leloir pathway, consisting of galactokinase, galactose-1-phosphate uridyltransferase, and UDP-galactose-4-epimerase (GALE), can generate UDP-glucose through the metabolism of galactose. The liver exhibits high expression of GALE and is a site of active galactose metabolism as well as glycogen synthesis, suggesting that UGP2 may have a distinct effect on metabolism in this tissue and may explain why UGP2 shows a distinct prognosis status in hepatocellular carcinoma.

UDP-glucose can also be used as a substrate for N-glycosylation, representing a major mechanism through which UGP2 functions in cancer cells. Fibroblasts lacking UGP2 are deficient in the biosynthesis of glycoconjugates including N-glycans, O-glycans, and glycosphingolipids.⁸ Furthermore, according to DepMap, the genes that show the highest codependencies with UGP2 include phosphoglucomutase 3 and glutamine-fructose-6-phosphate transaminase 1, which generate UDP-N-acetylglucosamine (UDP-GlcNAc), and mannose phosphate isomerase, phosphomannomutase 2, and GDP-mannose pyrophosphorylase B, which generate GDP-mannose. UDP-GlcNAc and GDP-mannose are substrates for O- and N-glycosylation, respectively, suggesting that the regulation of the glycosylation pathway by UGP2 in cancer cells is functionally relevant for cancer cell survival. In PDAC cells, UGP2 regulates glycosylation of several proteins and pathways relevant to cancer biology, as elucidated by gene ontology analsysis (Figure 1b).⁴ Of particular note is the epidermal growth factor receptor (EGFR), whose function is deeply ingrained in the growth and proliferation of cancer cells, and multiple integrin proteins, which have roles in cell migration and metastasis. UGP2 has been reported to regulate invasive phenotypes and correlate with metastatic potential.⁹ Individual functions of each of the multiple UGP2-regulated modifications identified on EGFR N361 remain to be elucidated. The site is proximal to the EGF-binding region, suggesting that it could be involved in regulating ligand binding.¹⁰ Long-term UGP2 loss in culture or xenograft tumors leads to decreases in total EGFR protein, suggesting a potential role in its stability and degradation.⁴

Thus, UGP2 is a key player in regulating multiple interrelated aspects of cancer metabolism and post-translational modifications. Employing UGP2 inhibitors in mechanistic studies can help to further elucidate the role of UGP2 as a growing player in the biology of human cancer.

Funding Statement

A.L.W. was a Damon Runyon Fellow, supported by the Damon Runyon Cancer Research Foundation through postdoctoral fellowship number DRG-2214-15. The research reported in this publication was supported by the National Cancer Institute of the NIH under Award K99CA226363 and R00CA226363 (to A.L.W.) and the National Research Foundation of Korea Grant NRF-2020R1C1C1013220 (to S.E.K.).

Disclosure statement

A.L.W. received research funding from Oncogenuity, Inc. for an unrelated project.