Hepatocellular carcinoma (HCC), with discouraging morbidity and mortality, ranks as one of the most prevalent tumors worldwide. Pyrimidine metabolism is a critical process that regulates DNA and RNA synthesis in cells. It is imperative to investigate the significance of pyrimidine metabolism in liver cancer.
Transcriptome and clinical data were downloaded from the TCGA database and the GEO database. The genes related to pyrimidine metabolism were sourced from the MSigDB. The pyrimidine metabolism-related signature (PMRS) was constructed through Cox regression and Lasso regression and then verified in the external validation set from the ICGC database. Functional enrichment, immune infiltration analysis, drug sensitivity, and Immunophenoscore (IPS) were further implemented to predict the response to immunotherapy. The role of PMRS in the malignant phenotype of hepatocellular carcinoma was explored by conducting a series of in vitro experiments.
Our study developed a four-genes PMRS which demonstrates a substantial correlation with the prognosis of HCC patients, serving as an independent predictor in clinical practice. The result of risk-stratified analysis yielded evidence that low-risk patients experienced more favorable clinical outcomes. The nomogram exhibited remarkable prognostic predictive value. The subsequent results revealed that low-risk patients manifested a more promising response to immunotherapy. Moreover, the results of cell experiments demonstrated that the downregulation of DCK markedly inhibited the malignant phenotype of hepatocellular carcinoma.
Our pyrimidine metabolism-centered prognostic signature accurately predicts overall survival, immune status, and treatment response in hepatocellular carcinoma (HCC) patients, offering innovative insights for precise diagnosis, personalized treatment, and improved prognosis.

UNASSIGNED: Metabolomics has found extensive applications in the field of neurological diseases, significantly contributing to their diagnosis and treatment. However, there has been limited research applying metabolomics to moyamoya disease (MMD). This study aims to investigate and identify differential metabolites associated with MMD.
UNASSIGNED: We employed a liquid chromatography coupled with mass spectrometry (LC-MS) approach, complemented by univariate and multivariate analyses, to discern metabolic biomarkers in cerebrospinal fluid samples. We then compared these biomarkers between MMD patients and healthy controls (Ctl).
UNASSIGNED: Sixteen patients diagnosed with MMD via cerebral angiography and eight healthy controls were enrolled in this study. Comparative analyses, including univariate and multivariate analyses, correlation studies, heatmaps, Volcano Plots, and KEGG pathway enrichment, were performed between MMD patients and controls. As a result, we identified 129 significant differential metabolites in the cerebrospinal fluid between MMD patients and controls. These metabolic biomarkers are associated with various pathways, with notable involvement in purine and pyrimidine metabolism.
UNASSIGNED: Utilizing an LC-MS-based metabolomics approach holds promise for enhancing the clinical diagnosis of MMD. The identified biomarkers offer potential avenues for the development of novel diagnostic methods for MMD and offer fresh insights into the pathogenesis of the disease.

Understanding the molecular underpinnings of disease severity and progression in human studies is necessary to develop metabolism-related preventative strategies for severe COVID-19. Metabolites and metabolic pathways that predispose individuals to severe disease are not well understood. In this study, we generated comprehensive plasma metabolomic profiles in >550 patients from the Longitudinal EMR and Omics COVID-19 Cohort. Samples were collected before (n = 441), during (n = 86), and after (n = 82) COVID-19 diagnosis, representing 555 distinct patients, most of which had single timepoints. Regression models adjusted for demographics, risk factors, and comorbidities, were used to determine metabolites associated with predisposition to and/or persistent effects of COVID-19 severity, and metabolite changes that were transient/lingering over the disease course. Sphingolipids/phospholipids were negatively associated with severity and exhibited lingering elevations after disease, while modified nucleotides were positively associated with severity and had lingering decreases after disease. Cytidine and uridine metabolites, which were positively and negatively associated with COVID-19 severity, respectively, were acutely elevated, reflecting the particular importance of pyrimidine metabolism in active COVID-19. This is the first large metabolomics study using COVID-19 plasma samples before, during, and/or after disease. Our results lay the groundwork for identifying putative biomarkers and preventive strategies for severe COVID-19.

Maintenance of dNTPs pools in Trypanosoma brucei is dependent on both biosynthetic and degradation pathways that together ensure correct cellular homeostasis throughout the cell cycle which is essential for the preservation of genomic stability. Both the salvage and de novo pathways participate in the provision of pyrimidine dNTPs while purine dNTPs are made available solely through salvage. In order to identify enzymes involved in degradation here we have characterized the role of a trypanosomal SAMHD1 orthologue denominated TbHD82. Our results show that TbHD82 is a nuclear enzyme in both procyclic and bloodstream forms of T. brucei. Knockout forms exhibit a hypermutator phenotype, cell cycle perturbations and an activation of the DNA repair response. Furthermore, dNTP quantification of TbHD82 null mutant cells revealed perturbations in nucleotide metabolism with a substantial accumulation of dATP, dCTP and dTTP. We propose that this HD domain-containing protein present in kinetoplastids plays an essential role acting as a sentinel of genomic fidelity by modulating the unnecessary and detrimental accumulation of dNTPs.

This present study was conducted to provide evidence and an explanation for the apoptosis that occurs in the marine rotifer Brachionus plicatilis when facing 2,2\',4,4\'-tetrabromodiphenyl ether (BDE-47) stress. Metabolomics analysis showed that aminoacyl-tRNA biosynthesis, valine, leucine and isoleucine biosynthesis, and arginine biosynthesis were the top three sensitive pathways to BDE-47 exposure, which resulted in the reduction in the amino acid pool level. Pyrimidine metabolism and purine metabolism pathways were also significantly influenced, and the purine and pyrimidine content were obviously reduced in the low (0.02 mg/L) and middle (0.1 mg/L) concentration groups while increased in the high (0.5 mg/L) concentration group, evidencing the disorder of nucleotide synthesis and decomposition in B. plicatilis. The biochemical detection of the key enzymes in purine metabolism and pyrimidine metabolism showed the downregulation of Glutamine Synthetase (GS) protein expression and the elevation of Xanthine Oxidase (XOD) activity, which suggested the impaired DNA repair and ROS overproduction. The content of DNA damage biomarker (8-OHdG) increased in treatment groups, and the p53 signaling pathway was found to be activated, as indicated by the elevation of the p53 protein expression and Bax/Bcl-2 ratio. The ROS scavenger (N-acetyl-L-cysteine, NAC) addition effectively alleviated not only ROS overproduction but also DNA damage as well as the activation of apoptosis. The combined results backed up the speculation that purine metabolism and pyrimidine metabolism alteration play a pivotal role in BDE-47-induced ROS overproduction and DNA damage, and the consequent activation of the p53 signaling pathway led to the observed apoptosis in B. plicatilis.

Small cell lung cancers (SCLCs) rapidly resist cytotoxic chemotherapy and immune checkpoint inhibitor (ICI) treatments. New, non-cross-resistant therapies are thus needed. SCLC cells are committed into neuroendocrine lineage then maturation arrested. Implicating DNA methyltransferase 1 (DNMT1) in the maturation arrests, we find (1) the repression mark methylated CpG, written by DNMT1, is retained at suppressed neuroendocrine-lineage genes, even as other repression marks are erased; (2) DNMT1 is recurrently amplified, whereas Ten-Eleven-Translocation 2 (TET2), which functionally opposes DNMT1, is deleted; (3) DNMT1 is recruited into neuroendocrine-lineage master transcription factor (ASCL1, NEUROD1) hubs in SCLC cells; and (4) DNMT1 knockdown activated ASCL1-target genes and released SCLC cell-cycling exits by terminal lineage maturation, which are cycling exits that do not require the p53/apoptosis pathway used by cytotoxic chemotherapy. Inhibiting DNMT1/corepressors with clinical compounds accordingly extended survival of mice with chemorefractory and ICI-refractory, p53-null, disseminated SCLC. Lineage commitment of SCLC cells can hence be leveraged into non-cytotoxic therapy able to treat chemo/ICI-refractory SCLC.

CAD is a large, 2225 amino acid multienzymatic protein required for de novo pyrimidine biosynthesis. Pathological CAD variants cause a developmental and epileptic encephalopathy which is highly responsive to uridine supplements. CAD deficiency is difficult to diagnose because symptoms are nonspecific, there is no biomarker, and the protein has over 1000 known variants. To improve diagnosis, we assessed the pathogenicity of 20 unreported missense CAD variants using a growth complementation assay that identified 11 pathogenic variants in seven affected individuals; they would benefit from uridine treatment. We also tested nine variants previously reported as pathogenic and confirmed the damaging effect of seven. However, we reclassified two variants as likely benign based on our assay, which is consistent with their long-term follow-up with uridine. We found that several computational methods are unreliable predictors of pathogenic CAD variants, so we extended the functional assay results by studying the impact of pathogenic variants at the protein level. We focused on CAD\'s dihydroorotase (DHO) domain because it accumulates the largest density of damaging missense changes. The atomic-resolution structures of eight DHO pathogenic variants, combined with functional and molecular dynamics analyses, provided a comprehensive structural and functional understanding of the activity, stability, and oligomerization of CAD\'s DHO domain. Combining our functional and protein structural analysis can help refine clinical diagnostic workflow for CAD variants in the genomics era.

Pyrimidine metabolism is a hallmark of cancer and will soon become an essential part of cancer therapy. In the tumor microenvironment, cells reprogram pyrimidine metabolism intrinsically and extracellularly, thereby promoting tumorigenesis. Metabolites in pyrimidine metabolism have a significant impact on promoting cancer advancement and modulating immune system responses. In preclinical studies and practical clinical applications, critical targets in pyrimidine metabolism are acted upon by drugs to exert promising therapeutic effects on tumors. However, the pyrimidine metabolism in breast cancer (BC) is still largely underexplored. In this study, 163 credible pyrimidine metabolism-related genes (PMGs) were retrieved, and their somatic mutations and expression levels were determined. In addition, by using The Cancer Genome Atlas (TCGA) and the Molecular Taxonomy of Breast Cancer International Consortium (METABRIC) databases, 12 PMGs related to the overall survival (OS) were determined using the univariate Cox regression analysis. Subsequently, by performing the LASSO Cox hazards regression analysis in the 12 PMGs in TCGA-BRCA dataset, we developed a prognosis nomogram using eight OS-related PMGs and then verified the same in the METABRIC, GSE96058, GSE20685, GSE42568 and GSE86166 data. Moreover, we validated relationships between the pyrimidine metabolism index (PMI) and the survival probability of patients, essential clinical parameters, including the TNM stage and the PAM50 subtypes. Next, we verified the predictive capability of the optimum model, including the signature, the PAM50 subtype, and age, using ROC analysis and calibration curve, and compared it with other single clinical factors for the predictive power of benefit using decision curve analysis. Finally, we investigated the potential effects of pyrimidine metabolism on immune checkpoints, tumor-infiltrating immune cells, and cytokine levels and determined the potential implications of pyrimidine metabolism in BC immunotherapy. In conclusion, our findings suggest that pyrimidine metabolism has underlying prognostic significance in BC and can facilitate a new management approach for patients with different prognoses and more precise immunotherapy.

New antimicrobials are needed for the treatment of extensively drug-resistant Acinetobacter baumannii. The de novo pyrimidine biosynthetic enzyme dihydroorotate dehydrogenase (DHODH) is a validated drug target for malaria and human autoimmune diseases. We provide genetic evidence that A. baumannii DHODH (AbDHODH) is essential for bacterial survival in rodent infection models. We chemically validate the target by repurposing a unique library of ~450 triazolopyrimidine/imidazopyrimidine analogs developed for our malaria DHODH program to identify 21 compounds with submicromolar activity on AbDHODH. The most potent (DSM186, DHODH IC50 28 nM) had a minimal inhibitory concentration of ≤1 µg/ml against geographically diverse A. baumannii strains, including meropenem-resistant isolates. A structurally related analog (DSM161) with a long in vivo half-life conferred significant protection in the neutropenic mouse thigh infection model. Encouragingly, the development of resistance to these compounds was not identified in vitro or in vivo. Lastly, the X-ray structure of AbDHODH bound to DSM186 was solved to 1.4 Å resolution. These data support the potential of AbDHODH as a drug target for the development of antimicrobials for the treatment of A. baumannii and potentially other high-risk bacterial infections.

Pyrimidine 5\'-nucleotidase deficiency is a rare erythrocyte enzymopathy. Here we report two cases of hemolytic anemia in brothers of Polish origin that are associated with a very rare mutation. Heterozygous deletion in the NT5C3A gene (c.444_446delGTT), inherited most likely from their asymptomatic mother, resulted in a single amino acid residue deletion (p.F149del) in cytosolic pyrimidine 5\'-nucleotidase. However, only the mutated transcript was present in the reticulocyte transcriptome of both patients. Only residual activity of pyrimidine 5\'-nucleotidase in the brothers\' erythrocytes could be observed when compared with the controls, including their asymptomatic father and sister. Western blot showed no sign of the presence of 5\'-nucleotidase protein in the erythrocytes of both studied patients. The 2.5-fold reduction of the purine/pyrimidine ratio observed only in the brothers\' erythrocytes confirms the correlation of the results of molecular analysis, including whole-exome sequencing, with the phenotype of the pyrimidine 5\'-nucleotidase deficiency. Altogether, our results may substantiate the hypothesis of the heterogeneity of the molecular basis of the defect involving both the mutation presented here and negative regulation of expression of the \"normal\" allele.