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Journal of Medicinal Chemistry May 2024Cofactor mimicry represents an attractive strategy for the development of enzyme inhibitors but can lead to off-target effects due to the evolutionary conservation of...
Cofactor mimicry represents an attractive strategy for the development of enzyme inhibitors but can lead to off-target effects due to the evolutionary conservation of binding sites across the proteome. Here, we uncover the ADP-ribose (ADPr) hydrolase NUDT5 as an unexpected, noncovalent, off-target of clinical BTK inhibitors. Using a combination of biochemical, biophysical, and intact cell NanoBRET assays as well as X-ray crystallography, we confirm catalytic inhibition and cellular target engagement of NUDT5 and reveal an unusual binding mode that is independent of the reactive acrylamide warhead. Further investigation of the prototypical BTK inhibitor ibrutinib also revealed potent inhibition of the largely unstudied NUDIX hydrolase family member NUDT14. By exploring structure-activity relationships (SARs) around the core scaffold, we identify a potent, noncovalent, and cell-active dual NUDT5/14 inhibitor. Cocrystallization experiments yielded new insights into the NUDT14 hydrolase active site architecture and inhibitor binding, thus providing a basis for future chemical probe design.
Topics: Humans; Pyrophosphatases; Agammaglobulinaemia Tyrosine Kinase; Structure-Activity Relationship; Crystallography, X-Ray; Protein Kinase Inhibitors; Pyrazoles; Piperidines; Drug Discovery; Pyrimidines; Adenine; Models, Molecular; Enzyme Inhibitors
PubMed: 38635563
DOI: 10.1021/acs.jmedchem.4c00072 -
Mediators of Inflammation 2023Ulcerative colitis (UC) is an inflammatory bowel disease of unknown cause that typically affects the colon and rectum. Innate intestinal immunity, including macrophages,...
Ulcerative colitis (UC) is an inflammatory bowel disease of unknown cause that typically affects the colon and rectum. Innate intestinal immunity, including macrophages, plays a significant role in the pathological development of UC. Using the CIBERSORT algorithm, we observed elevated levels of 22 types of immune cell infiltrates, as well as increased M1 and decreased M2 macrophages in UC compared to normal colonic mucosa. Weighted gene coexpression network analysis (WGCNA) was used to identify modules associated with macrophages and UC, resulting in the identification of 52 macrophage-related genes (MRGs) that were enriched in macrophages at single-cell resolution. Consensus clustering based on these 52 MRGs divided the integrated UC cohorts into three subtypes. Machine learning algorithms were used to identify ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1), sodium- and chloride-dependent neutral and basic amino acid transporter B(0+) (SLC6A14), and 3-hydroxy-3-methylglutaryl-CoA synthase 2 (HMGCS2) in the training set, and their diagnostic value was validated in independent validation sets. Gene set enrichment analysis (GSEA) and gene set variation analysis (GSVA) revealed the main biological effects, and that interleukin-17 was one of several signaling pathways enriched by the three genes. We also constructed a competitive endogenous RNA (CeRNA) network reflecting a potential posttranscriptional regulatory mechanism. Expression of diagnostic markers was validated in vivo and in biospecimens, and our immunohistochemistry (IHC) results confirmed that HMGCS2 gradually decreased during the transformation of UC to colorectal cancer. In conclusion, ENPP1, SLC6A14, and HMGCS2 are associated with macrophages and the progression of UC pathogenesis and have good diagnostic value for patients with UC.
Topics: Humans; Colitis, Ulcerative; Rectum; Macrophages; Intestinal Mucosa
PubMed: 38633005
DOI: 10.1155/2023/4373840 -
Experimental and Therapeutic Medicine May 2024Despite advances in surgical treatment techniques and chemotherapy-including anti-angiogenic and immune poly (ADP-ribose) polymerase inhibitors, the 5-year survival rate... (Review)
Review
Despite advances in surgical treatment techniques and chemotherapy-including anti-angiogenic and immune poly (ADP-ribose) polymerase inhibitors, the 5-year survival rate in ovarian cancer (OC) remains low. The reasons for this are the diagnosis of cancer in advanced clinical stages, chemoresistance and cancer recurrence. New therapeutic approaches are being developed, including the search for new biomarkers that are also targets for targeted therapy. The present review describes new molecular markers with relevance to targeted therapy, which to date have been studied only in experimental research. These include the angiogenic protein angiopoietin-2, the transmembrane glycoprotein ectonucleotide pyrophosphatase/phosphodiesterase 1, the adhesion protein E-cadherin, the TIMP metallopeptidase inhibitor 1 and Kruppel-like factor 7. Drugs affecting cancer stem cells (CSCs) in OC, such as metformin and salinomycin, as well as inhibitors of CSCs markers aldehyde dehydrogenase 1 (with the drug ATRA) and the transcription factor Nanog homeobox (microRNA) are also discussed. A new approach to prevention and possible therapies under investigation such as development of vaccines containing a subpopulation of CD117(+) and CD44(+) stem cells with a promising option for use in women with OC was described.
PubMed: 38628658
DOI: 10.3892/etm.2024.12523 -
Journal of Gastroenterology Jun 2024This study evaluated the effectiveness of NUDT15 codon 139 genotyping in optimizing thiopurine treatment for inflammatory bowel disease (IBD) in Japan, using real-world...
BACKGROUND
This study evaluated the effectiveness of NUDT15 codon 139 genotyping in optimizing thiopurine treatment for inflammatory bowel disease (IBD) in Japan, using real-world data, and aimed to establish genotype-based treatment strategies.
METHODS
A retrospective analysis of 4628 IBD patients who underwent NUDT15 codon 139 genotyping was conducted. This study assessed the purpose of the genotyping test and subsequent prescriptions following the obtained results. Outcomes were compared between the Genotyping group (thiopurine with genotyping test) and Non-genotyping group (thiopurine without genotyping test). Risk factors for adverse events (AEs) were analyzed by genotype and prior genotyping status.
RESULTS
Genotyping test for medical purposes showed no significant difference in thiopurine induction rates between Arg/Arg and Arg/Cys genotypes, but nine Arg/Cys patients opted out of thiopurine treatment. In the Genotyping group, Arg/Arg patients received higher initial doses than the Non-genotyping group, while Arg/Cys patients received lower ones (median 25 mg/day). Fewer AEs occurred in the Genotyping group because of their lower incidence in Arg/Cys cases. Starting with < 25 mg/day of AZA reduced AEs in Arg/Cys patients, while Arg/Arg patients had better retention rates when maintaining ≥ 75 mg AZA. Nausea and liver injury correlated with thiopurine formulation but not dosage. pH-dependent mesalamine reduced leukopenia risk in mesalamine users.
CONCLUSIONS
NUDT15 codon 139 genotyping effectively reduces thiopurine-induced AEs and improves treatment retention rates in IBD patients after genotype-based dose adjustments. This study provides data-driven treatment strategies based on genotype and identifies risk factors for specific AEs, contributing to a refined thiopurine treatment approach.
Topics: Humans; Pyrophosphatases; Female; Male; Retrospective Studies; Adult; Middle Aged; Mercaptopurine; Genotype; Inflammatory Bowel Diseases; Japan; Azathioprine; Young Adult; Aged; Immunosuppressive Agents; Adolescent; Risk Factors; Codon; Nudix Hydrolases
PubMed: 38589597
DOI: 10.1007/s00535-024-02099-7 -
Gene Jul 2024Biallelic variants in PPA2 gene cause a rare but lethal mitochondrial disorder. We describe the first four cases reported in Spain of PPA2 disease in two unrelated...
Biallelic variants in PPA2 gene cause a rare but lethal mitochondrial disorder. We describe the first four cases reported in Spain of PPA2 disease in two unrelated families. We have conducted a revision of the clinical history, necropsies, and postmortem genetic testing from probands, and clinical evaluation, genetic testing and blood transcript analysis in family members. All the cases harbored biallelic PPA2 variants in compound heterozygous status. Two brothers from family 1 suffered sudden death after a small first intake of alcohol in 2013 and 2022. The sister remains alive but affected with cardiomyopathy, extensive scar on cardiac imaging, and high sensitivity to alcohol intake. The three siblings carried PPA2 c.290A > G (p.Glu97Gly) novel missense variant and PPA2 c.513C > T (p.Cys171 = ) altering splicing site variant, both probably leading to mRNA degradation based on in-silico and transcript analyses. A teenager from family 2 suffered sudden death after a small intake of alcohol in 2018 and carried PPA2 c.683C > T (p.Pro228Leu) missense and PPA2 c.980_983del (p.Gln327fs) novel frameshift variant, both probably leading to abnormal protein structure. All cases were asymptomatic until adolescence. Furthermore, the sister in family 1 has survived as an asymptomatic adult. PPA2 disease can manifest as cardiac arrest in the young, especially after alcohol exposure. Our results show that PPA2 deficiency can be related to different pathogenicity mechanisms such as abnormal protein structure but also mRNA decay caused by synonymous or missense variants. Strict avoidance of alcohol consumption and early defibrillator implantation might prevent lethal arrhythmias in patients at risk.
Topics: Adolescent; Adult; Female; Humans; Male; Alcohol Drinking; Death, Sudden, Cardiac; Mutation, Missense; Pedigree; Spain; Mitochondrial Proteins; Inorganic Pyrophosphatase
PubMed: 38582264
DOI: 10.1016/j.gene.2024.148437 -
Free Radical Biology & Medicine Jun 2024Dysregulated ecto-nucleotide pyrophosphatase/phosphodiesterase (ENPP) family occurs in metabolic reprogramming pathological processes. Nonetheless, the epigenetic...
BACKGROUND
Dysregulated ecto-nucleotide pyrophosphatase/phosphodiesterase (ENPP) family occurs in metabolic reprogramming pathological processes. Nonetheless, the epigenetic mechanisms by which ENPP family impacts NAFLD, also known as metabolic dysfunction-associated steatotic liver disease (MASLD), is poorly appreciated.
METHODS
We investigated the causes and consequences of ENPP1 promoter hypomethylation may boost NAFLD using NAFLD clinical samples, as well as revealed the underlying mechanisms using high-fat diet (HFD) + carbon tetrachloride (CCl) induced mouse model of NAFLD and FFA treatment of cultured hepatocyte.
RESULTS
Herein, we report that the expression level of ENPP1 are increased in patients with NAFLD liver tissue and in mouse model of NAFLD. Hypomethylation of ENPP1, is associated with the perpetuation of hepatocyte autophagy and liver fibrosis in the NAFLD. ENPP1 hypomethylation is mediated by the DNA demethylase TET3 in NAFLD liver fibrosis and hepatocyte autophagy. Additionally, knockdown of TET3 methylated ENPP1 promoter, reduced the ENPP1 expression, ameliorated the experimental NAFLD. Mechanistically, TET3 epigenetically promoted ENPP1 expression via hypomethylation of the promoter. Knocking down TET3 can inhibit the hepatocyte autophagy but an overexpression of ENPP1 showing rescue effect.
CONCLUSIONS
We describe a novel epigenetic mechanism wherein TET3 promoted ENPP1 expression through promoter hypomethylation is a critical mediator of NAFLD. Our findings provide new insight into the development of preventative measures for NAFLD.
Topics: Animals; Humans; Male; Mice; Autophagy; Carbon Tetrachloride; Diet, High-Fat; Dioxygenases; Disease Models, Animal; DNA Methylation; Epigenesis, Genetic; Hepatocytes; Liver Cirrhosis; Mice, Inbred C57BL; Non-alcoholic Fatty Liver Disease; Phosphoric Diester Hydrolases; Promoter Regions, Genetic; Proto-Oncogene Proteins; Pyrophosphatases
PubMed: 38582229
DOI: 10.1016/j.freeradbiomed.2024.04.207 -
International Journal of Biological... May 2024The emergence of multidrug resistance has provided a great challenge to treat nosocomial infections, which have become a major health threat around the globe. Lipid A... (Review)
Review
The emergence of multidrug resistance has provided a great challenge to treat nosocomial infections, which have become a major health threat around the globe. Lipid A (an active endotoxin component), the final product of the Raetz lipid A metabolism pathway, is a membrane anchor of lipopolysaccharide (LPS) of the gram-negative bacterial outer membrane. It shields bacterial cells and serves as a protective barrier from antibiotics, thereby eliciting host response and making it difficult to destroy. UDP-2,3-diacylglucosamine pyrophosphate hydrolase (LpxH), a crucial peripheral membrane enzyme of the Raetz pathway, turned out to be the potential target to inhibit the production of Lipid A. This review provides a comprehensive compilation of information regarding the structural and functional aspects of LpxH, as well as its analogous LpxI and LpxG. In addition, apart from by providing a broader understanding of the enzyme-inhibitor mechanism, this review facilitates the development of novel drug candidates that can inhibit the pathogenicity of the lethal bacterium.
Topics: Gram-Negative Bacteria; Pyrophosphatases; Lipid A; Enzyme Inhibitors; Humans
PubMed: 38574903
DOI: 10.1016/j.ijbiomac.2024.131327 -
Cancer Medicine Apr 2024Studies have shown that some single nucleotide polymorphisms (SNPs) could serve as excellent markers in foretelling the treatment outcome of interferon (IFN) in...
BACKGROUND
Studies have shown that some single nucleotide polymorphisms (SNPs) could serve as excellent markers in foretelling the treatment outcome of interferon (IFN) in myeloproliferative neoplasms (MPN). However, most work originated from western countries, and data from different ethnic populations have been lacking.
METHODS
To gain insights, targeted sequencing was performed to detect myeloid-associated mutations and SNPs in eight loci across three genes (IFNL4, IFN-γ, and inosine triphosphate pyrophosphatase [ITPA]) to explore their predictive roles in our cohort of 21 ropeginterferon alpha-2b (ROPEG)-treated MPN patients, among whom real-time quantitative PCR was also performed periodically to monitor the JAK2V617F allele burden in 19 JAK2V617F-mutated cases.
RESULTS
ELN response criteria were adopted to designate patients as good responders if they achieved complete hematological responses (CHR) within 1 year (CHR1) or attained major molecular responses (MMR), which occurred in 70% and 45% of the patients, respectively. IFNL4 and IFN-γ gene SNPs were infrequent in our population and were thus excluded from further analysis. Two ITPA SNPs rs6051702 A>C and rs1127354 C>A were associated with an inferior CHR1 rate and MMR rate, respectively. The former seemed to be linked to grade 2 or worse hepatotoxicity as well, although the comparison was of borderline significance only (50%, vs. 6.7% in those with common haplotype, p = 0.053). Twelve patients harbored 19 additional somatic mutations in 12 genes, but the trajectory of these mutations varied considerably and was not predictive of any response.
CONCLUSIONS
Overall, this study provided valuable information on the ethnics- and genetics-based algorithm in the treatment of MPN.
Topics: Humans; Neoplasms; Myeloproliferative Disorders; Treatment Outcome; Haplotypes; Germ Cells; Interferon Lambda; Interleukins
PubMed: 38572926
DOI: 10.1002/cam4.7166 -
Nature Communications Apr 2024Phosphorus (P) limitation of ecosystem processes is widespread in terrestrial habitats. While a few auxiliary metabolic genes (AMGs) in bacteriophages from aquatic...
Phosphorus (P) limitation of ecosystem processes is widespread in terrestrial habitats. While a few auxiliary metabolic genes (AMGs) in bacteriophages from aquatic habitats are reported to have the potential to enhance P-acquisition ability of their hosts, little is known about the diversity and potential ecological function of P-acquisition genes encoded by terrestrial bacteriophages. Here, we analyze 333 soil metagenomes from five terrestrial habitat types across China and identify 75 viral operational taxonomic units (vOTUs) that encode 105 P-acquisition AMGs. These AMGs span 17 distinct functional genes involved in four primary processes of microbial P-acquisition. Among them, over 60% (11/17) have not been reported previously. We experimentally verify in-vitro enzymatic activities of two pyrophosphatases and one alkaline phosphatase encoded by P-acquisition vOTUs. Thirty-six percent of the 75 P-acquisition vOTUs are detectable in a published global topsoil metagenome dataset. Further analyses reveal that, under certain circumstances, the identified P-acquisition AMGs have a greater influence on soil P availability and are more dominant in soil metatranscriptomes than their corresponding bacterial genes. Overall, our results reinforce the necessity of incorporating viral contributions into biogeochemical P cycling.
Topics: Bacteriophages; Ecosystem; Phosphorus; Metagenome; Soil
PubMed: 38565528
DOI: 10.1038/s41467-024-47214-7 -
Talanta Jul 2024Fenton chemistry has aroused widespread concern due to its application in the green oxidation and mineralization of organic wastes. Inorganic pyrophosphatase (PPase)...
Fenton chemistry has aroused widespread concern due to its application in the green oxidation and mineralization of organic wastes. Inorganic pyrophosphatase (PPase) catalyzes the hydrolysis of pyrophosphate ions (PPi) and provides a thermodynamic driving force for many biosynthetic reactions. Fluoride (F) is widely applied to fight against tooth decay and reduce cavities. The electrochemical determination of PPase activity and F was realized based on Fenton chemistry in this work. Glassy carbon electrode modified with poly (azure A) and acetylene black (GCE/PAA-AB) was fabricated. Hydroxyl radicals (∙OH) that were generated from a Cu-catalyzed Fenton-type reaction could oxidize PAA in the near-neutral medium, leading to a great increase of the cathodic peak current (I). A coordination reaction between PPi and Cu exerted a negative effect on Fenton reaction and hindered the I enhancement. Cu-PPi complex was decomposed due to the hydrolysis of PPi induced by PPase, which caused the reappearance of the notably increased current response. F could effectively inhibit PPase activity. As a result, the stable Cu-PPi complex remained and the high I suffered from the decline again. The I difference was used for the highly sensitive determination of PPase activity in the content range of 0.001-20 mU mL with a detection of limit (LOD) at 0.6 μU mL and that of F in the concentration range of 0.01-100 μM with a LOD at 7 nM. The proposed PPase and F sensor displayed a good selectivity, stability and reproducibility, and a high accuracy.
Topics: Fluorides; Iron; Electrochemical Techniques; Hydrogen Peroxide; Copper; Electrodes; Pyrophosphatases; Inorganic Pyrophosphatase; Limit of Detection; Enzyme Assays
PubMed: 38564823
DOI: 10.1016/j.talanta.2024.125943