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Pancreatic ductal adenocarcinoma (PDA) is a lethal disease notoriously resistant to therapy 1 , 2 . This is mediated in part by a complex tumour microenvironment 3 , low vascularity 4 , and metabolic aberrations 5 , 6 . Although altered metabolism drives tumour progression, the spectrum of metabolites used as nutrients by PDA remains largely unknown. Here we identified uridine as a fuel for PDA in glucose-deprived conditions by assessing how more than 175 metabolites impacted metabolic activity in 21 pancreatic cell lines under nutrient restriction. Uridine utilization strongly correlated with the expression of uridine phosphorylase 1 (UPP1), which we demonstrate liberates uridine-derived ribose to fuel central carbon metabolism and thereby support redox balance, survival and proliferation in glucose-restricted PDA cells. In PDA, UPP1 is regulated by KRAS–MAPK signalling and is augmented by nutrient restriction. Consistently, tumours expressed high UPP1 compared with non-tumoural tissues, and UPP1 expression correlated with poor survival in cohorts of patients with PDA. Uridine is available in the tumour microenvironment, and we demonstrated that uridine-derived ribose is actively catabolized in tumours. Finally, UPP1 deletion restricted the ability of PDA cells to use uridine and blunted tumour growth in immunocompetent mouse models. Our data identify uridine utilization as an important compensatory metabolic process in nutrient-deprived PDA cells, suggesting a novel metabolic axis for PDA therapy. A metabolite screen of pancreatic cells shows that pancreatic cancer cells metabolize uridine-derived ribose via UPP1, supporting redox balance, survival and proliferation.

BackgroundCholangiocarcinoma (CCA) is a very difficult-to-treat cancer. Chemotherapies are little effective and response to immune checkpoint inhibitors is limited. Therefore, new therapeutic strategies need to be identified.ObjectiveWe characterised the enzyme protein arginine-methyltransferase 5 (PRMT5) as a novel therapeutic target in CCA.DesignWe evaluated the expression of PRMT5, its functional partner MEP50 and methylthioadenosine phosphorylase (MTAP)—an enzyme that modulates the sensitivity of PRMT5 to pharmacological inhibitors—in human CCA tissues. PRMT5-targeting drugs, currently tested in clinical trials for other malignancies, were assessed in human CCA cell lines and organoids, as well as in two immunocompetent CCA mouse models. Transcriptomic, proteomic and functional analyses were performed to explore the underlying antitumoural mechanisms.ResultsPRMT5 and MEP50 proteins were correlatively overexpressed in most CCA tissues. MTAP was absent in 25% of intrahepatic CCA. PRMT5-targeting drugs markedly inhibited CCA cell proliferation, synergising with cisplatin and gemcitabine and hindered the growth of cholangiocarcinoma organoids. PRMT5 inhibition blunted the expression of oncogenic genes involved in chromatin remodelling and DNA repair, consistently inducing the formation of RNA loops and promoting DNA damage. Treatment with PRMT5-targeting drugs significantly restrained the growth of experimental CCA without adverse effects and concomitantly induced the recruitment of CD4 and CD8 T cells to shrinking tumourous lesions.ConclusionPRMT5 and MEP50 are frequently upregulated in human CCA, and PRMT5-targeting drugs have significant antitumoural efficacy in clinically relevant CCA models. Our findings support the evaluation of PRMT5 inhibitors in clinical trials, including their combination with cytotoxic and immune therapies.

Thymidine phosphorylase (TYMP), a protein found in both prokaryotic and eukaryotic cells, is encoded by a gene located in the q13 region of chromosome 22. With a relative molecular mass of 55,000, TYMP exists as a homodimer. Recent research has increasingly illuminated the diverse functions of TYMP. It is known to facilitate platelet activation, osteoclast differentiation, and angiogenesis. Mutations in the TYMP gene are linked to mitochondrial neurogastrointestinal encephalomyopathy. Beyond its physiological roles, TYMP contributes significantly to tumor growth and cancer progression, where it promotes angiogenesis, modulates epigenetic genes, inhibits apoptosis, and acts as a critical enzyme in the nucleoside metabolic rescue pathway. Moreover, TYMP holds substantial implications in cancer treatment and prognosis. Given its involvement in cancer progression, TYMP inhibitors may prove valuable in inhibiting tumor growth and metastasis. Interestingly, while TYMP can drive tumor growth, certain concentrations of TYMP also enhance the cytotoxic effects of chemotherapy drugs such as 5-fluorouracil (5-FU). Although challenges exist—such as the potential disruption of normal physiological functions when inhibiting TYMP—the protein remains a promising target for cancer treatment. Ongoing research on TYMP could deepen our understanding of human physiology and the pathogenesis of cancer and open new avenues for therapeutic interventions. This article provides a comprehensive review of TYMP’s structure, physiological functions, and its role in tumorigenesis and anti-tumor therapy.

Nucleoside phosphorylases (NPs) are pivotal enzymes in the salvage pathway, catalyzing the reversible phosphorolysis of nucleosides to produce nucleobases and α-D-ribose 1-phosphate. Due to their efficiency in catalyzing nucleoside synthesis from purine or pyrimidine bases, these enzymes hold significant industrial importance in the production of nucleoside-based drugs. Given that the thermodynamic equilibrium for purine NPs (PNPs) is favorable for nucleoside synthesis—unlike pyrimidine NPs (PyNPs, UP, and TP)—multi-enzymatic systems combining PNPs with PyNPs, UPs, or TPs are commonly employed in the synthesis of nucleoside analogs. In this study, we report the first development of two engineered bifunctional fusion enzymes, created through the genetic fusion of purine nucleoside phosphorylase I (PNP I) and thymidine phosphorylase (TP) from Thermus thermophilus. These fusion constructs, PNP I/TP-His and TP/PNP I-His, provide an innovative one-pot, single-step alternative to traditional multi-enzymatic synthesis approaches. Interestingly, both fusion enzymes retain phosphorolytic activity for both purine and pyrimidine nucleosides, demonstrating significant activity at elevated temperatures (60–90 °C) and within a pH range of 6–8. Additionally, both enzymes exhibit high thermal stability, maintaining approximately 80–100% of their activity when incubated at 60–80 °C over extended periods. Furthermore, the transglycosylation capabilities of the fusion enzymes were explored, demonstrating successful catalysis between purine (2′-deoxy)ribonucleosides and pyrimidine bases, and vice versa. To optimize reaction conditions, the effects of pH and temperature on transglycosylation activity were systematically examined. Finally, as a proof of concept, these fusion enzymes were successfully employed in the synthesis of various purine and pyrimidine ribonucleoside and 2′-deoxyribonucleoside analogs, underscoring their potential as versatile biocatalysts in nucleoside-based drug synthesis.

Malignant pleural mesothelioma (MPM), associated with unfavorable outcomes, is closely associated with asbestos exposure. Early detection and treatment are critical to prolong survival of patients with MPM because of the rapid progression and resistance to treatment. The recently defined malignant mesothelioma in situ (MIS) has been gaining increasing attention with advances in genome-based methods including fluorescence in situ hybridization (FISH) as well as immunohistochemistry. We herein report the case of a MIS in a 73-year-old male with a history of asbestos exposure presenting with massive pleural effusion in the right thoracic cavity. Video-assisted thoracoscopic surgery with pleural biopsy of the right side revealed a single layer of atypical mesothelial cells without invasive lesions by hematoxylin and eosin staining. However, these mesothelial cells exhibited a loss of methylthioadenosine phosphorylase (MTAP) by immunohistochemistry and homozygous deletion of CDKN2A (p16) by FISH, leading to the diagnosis of MIS.

Glycoside Hydrolase family 65 (GH65) is a unique family of carbohydrate-active enzymes. It is the first protein family to bring together glycoside hydrolases, glycoside phosphorylases and glycosyltransferases, thereby spanning a broad range of reaction types. These enzymes catalyze the hydrolysis, reversible phosphorolysis or synthesis of various α-glucosides, typically α-glucobioses or their derivatives. In this review, we present a comprehensive overview of the diverse reaction types and substrate specificities found in family GH65. We describe the determinants that control this remarkable diversity, as well as the applications of GH65 enzymes for carbohydrate synthesis. Key points • GH65 is the first CAZy family to contain hydrolases, phosphorylases and transferases • Distinct residues and loops are determinants of substrate specificity in family GH65 • GH65 enzymes hold strong potential for carbohydrate synthesis via coupled reactions

Characterization of tumor metabolism with spatial information contributes to our understanding of complex cancer metabolic reprogramming, facilitating the discovery of potential metabolic vulnerabilities that might be targeted for tumor therapy. However, given the metabolic variability and flexibility of tumors, it is still challenging to characterize global metabolic alterations in heterogeneous cancer. Here, we propose a spatially resolved metabolomics approach to discover tumor-associated metabolites and metabolic enzymes directly in their native state. A variety of metabolites localized in different metabolic pathways were mapped by airflow-assisted desorption electrospray ionization mass spectrometry imaging (AFADESI-MSI) in tissues from 256 esophageal cancer patients. In combination with in situ metabolomics analysis, this method provided clues into tumor-associated metabolic pathways, including proline biosynthesis, glutamine metabolism, uridine metabolism, histidine metabolism, fatty acid biosynthesis, and polyamine biosynthesis. Six abnormally expressed metabolic enzymes that are closely associated with the altered metabolic pathways were further discovered in esophageal squamous cell carcinoma (ESCC). Notably, pyrroline-5-carboxylate reductase 2 (PYCR2) and uridine phosphorylase 1 (UPase1) were found to be altered in ESCC. The spatially resolved metabolomics reveal what occurs in cancer at the molecular level, from metabolites to enzymes, and thus provide insights into the understanding of cancer metabolic reprogramming.

The cellulose-rich corncob residue (CCR) is an abundant and renewable agricultural biomass that has been under-exploited. In this study, two strategies were compared for their ability to transform CCR into cello-oligosaccharides (COS). The first strategy employed the use of endo-glucanases. Although selected endo-glucanases from GH9, GH12, GH45, and GH131 could release COS with degrees of polymerization from 2 to 4, the degrading efficiency was low. For the second strategy, first, CCR was efficiently depolymerized to glucose and cellobiose using the cellulase from Trichoderma reesei . Then, using these simple sugars and sucrose as the starting materials, phosphorylases from different microorganisms were combined to generate COS to a level up to 100.3 g/L with different patterns and degrees of polymerization. Using tomato as a model plant, the representative COS obtained from Ba SP (a sucrose phosphorylase from Bifidobacterium adolescens ), Cu CbP (a cellobiose phosphorylase from Cellulomonas uda ), and Cc CdP (a cellodextrin phosphorylase from Clostridium cellulosi ) were shown to be able to promote plant growth. The current study pointed to an approach to make use of CCR for production of the value-added COS. Key points • Sequential use of cellulase and phosphorylases effectively generated cello-oligosaccharides from corncob residue . • Cello-oligosaccharides patterns varied in accordance to cellobiose/cellodextrin phosphorylases . • Spraying cello-oligosaccharides promoted tomato growth . Graphical Abstract

Prostatic luminal epithelial cells secrete high levels of acetylated polyamines into the prostatic lumen, sensitizing them to perturbations of connected metabolic pathways. Enhanced flux is driven by spermidine/spermine N1-acetyltransferase (SSAT) activity, which acetylates polyamines leading to their secretion and drives biosynthetic demand. The methionine salvage pathway recycles one-carbon units lost to polyamine biosynthesis to the methionine cycle to overcome stress. Prostate cancer (CaP) relies on methylthioadenosine phosphorylase (MTAP), the rate-limiting enzyme, to relieve strain. Here, we show that inhibition of MTAP alongside SSAT upregulation is synergistic in androgen sensitive and castration recurrent CaP models in vitro and in vivo. The combination treatment increases apoptosis in radical prostatectomy ex vivo explant samples. This unique high metabolic flux through polyamine biosynthesis and connected one carbon metabolism in CaP creates a metabolic dependency. Enhancing this flux while simultaneously targeting this dependency in prostate cancer results in an effective therapeutic approach potentially translatable to the clinic. Prostate cancer cells depend on MTAP, the rate-limiting enzyme involved in the methionine salvage pathway, to cope with increased polyamine biosynthesis. Here, the authors show that inducing upregulation of polyamine biosynthesis and targeting MTAP synergize to increase apoptosis in prostate cancer cells.

Inhibition of glycogen phosphorylase (Pyg) – a regulatory enzyme of glycogen phosphorolysis – influences memory formation in rodents. We have previously shown that 2-week intraperitoneal administration of a Pyg inhibitor BAY U6751 stimulated the “rejuvenation” of the hippocampal proteome and dendritic spines morphology and improved cognitive skills of old mice. Given the tedious nature of daily intraperitoneal drug administration, in this study we investigated whether a single dose of BAY U6751 could induce enduring behavioral effects. Obtained results support the efficacy of such treatment in significantly improving the cognitive performance of 20-22-month-old mice. Metabolomic analysis of alterations observed in the hippocampus, cerebellum, and cortex reveal that the inhibition of glycogen phosphorolysis impacts not only glucose metabolism but also various other metabolic processes.