Nicotinamide N-methyltransferase sustains a core epigenetic program that promotes metastatic colonization in breast cancer.

Metastatic colonization of distant organs accounts for over 90% of deaths related to solid cancers, yet the molecular determinants of metastasis remain poorly understood. Here, we unveil a mechanism of colonization in the aggressive basal-like subtype of breast cancer that is driven by the NAD+ metabolic enzyme nicotinamide N-methyltransferase (NNMT). We demonstrate that NNMT imprints a basal genetic program into cancer cells, enhancing their plasticity. In line, NNMT expression is associated with poor clinical outcomes in patients with breast cancer. Accordingly, ablation of NNMT dramatically suppresses metastasis formation in pre-clinical mouse models. Mechanistically, NNMT depletion results in a methyl overflow that increases histone H3K9 trimethylation (H3K9me3) and DNA methylation at the promoters of PR/SET Domain-5 (PRDM5) and extracellular matrix-related genes. PRDM5 emerged in this study as a pro-metastatic gene acting via induction of cancer-cell intrinsic transcription of collagens. Depletion of PRDM5 in tumor cells decreases COL1A1 deposition and impairs metastatic colonization of the lungs. These findings reveal a critical activity of the NNMT-PRDM5-COL1A1 axis for cancer cell plasticity and metastasis in basal-like breast cancer.

Proteomics reveals NNMT as a master metabolic regulator of cancer-associated fibroblasts.

High-grade serous carcinoma has a poor prognosis, owing primarily to its early dissemination throughout the abdominal cavity. Genomic and proteomic approaches have provided snapshots of the proteogenomics of ovarian cancer1,2, but a systematic examination of both the tumour and stromal compartments is critical in understanding ovarian cancer metastasis. Here we develop a label-free proteomic workflow to analyse as few as 5,000 formalin-fixed, paraffin-embedded cells microdissected from each compartment. The tumour proteome was stable during progression from in situ lesions to metastatic disease; however, the metastasis-associated stroma was characterized by a highly conserved proteomic signature, prominently including the methyltransferase nicotinamide N-methyltransferase (NNMT) and several of the proteins that it regulates. Stromal NNMT expression was necessary and sufficient for functional aspects of the cancer-associated fibroblast (CAF) phenotype, including the expression of CAF markers and the secretion of cytokines and oncogenic extracellular matrix. Stromal NNMT expression supported ovarian cancer migration, proliferation and in vivo growth and metastasis. Expression of NNMT in CAFs led to depletion of S-adenosyl methionine and reduction in histone methylation associated with widespread gene expression changes in the tumour stroma. This work supports the use of ultra-low-input proteomics to identify candidate drivers of disease phenotypes. NNMT is a central, metabolic regulator of CAF differentiation and cancer progression in the stroma that may be therapeutically targeted.

Nicotinamide N-methyltransferase mediates lipofibroblast-myofibroblast transition and apoptosis resistance.

Metabolism controls cellular phenotype and fate. In this report, we demonstrate that nicotinamide N-methyltransferase (NNMT), a metabolic enzyme that regulates developmental stem cell transitions and tumor progression, is highly expressed in human idiopathic pulmonary fibrosis (IPF) lungs, and is induced by the pro-fibrotic cytokine, transforming growth factor-ß1 (TGF-ß1) in lung fibroblasts. NNMT silencing reduces the expression of extracellular matrix proteins, both constitutively and in response to TGF-ß1. Furthermore, NNMT controls the phenotypic transition from homeostatic, pro-regenerative lipofibroblasts to pro-fibrotic myofibroblasts. This effect of NNMT is mediated, in part, by the downregulation of lipogenic transcription factors, TCF21 and PPARγ, and the induction of a less proliferative but more differentiated myofibroblast phenotype. NNMT confers an apoptosis-resistant phenotype to myofibroblasts that is associated with the downregulation of pro-apoptotic members of the Bcl-2 family, including Bim and PUMA. Together, these studies indicate a critical role for NNMT in the metabolic reprogramming of fibroblasts to a pro-fibrotic and apoptosis-resistant phenotype and support the concept that targeting this enzyme may promote regenerative responses in chronic fibrotic disorders such as IPF.

Targeting nicotinamide N-methyltransferase decreased aggressiveness of osteosarcoma cells.

BACKGROUND: Osteosarcoma (OS) is a primary bone malignancy that mostly affects young people, characterized by high metastatic potential, and a marked chemoresistance that is responsible for disease relapse in most patients. Therefore, it is necessary to identify novel molecules to setup targeted strategies to improve the clinical outcome. The enzyme nicotinamide N-methyltransferase (NNMT) catalyses the N-methylation of nicotinamide and other analogs, playing a crucial role in the biotransformation of drugs and xenobiotics. NNMT overexpression was reported in a wide variety of cancers, and several studies demonstrated that is able to promote cell proliferation, migration and resistance to chemotherapy. The aim of this study was to explore the potential involvement of NNMT in OS. METHODS: Immunohistochemical analyses have been performed to evaluate NNMT expression in selected OS and healthy bone tissue samples. Subsequently, OS cell lines have been transfected with vectors targeting NNMT mRNA (shRNAs) and the impact of this downregulation on migration, cell proliferation, and response to chemotherapeutic treatment was also analysed by wound healing, MTT, SRB and Trypan blue assays, respectively. RESULTS: Results showed that OS samples display a significantly higher NNMT expression compared with healthy tissue. Preliminary results suggest that NNMT silencing in OS cell lines is associated to a decrease of cell proliferation and migration, as well as to enhanced sensitivity to chemotherapy. Data obtained showed that NNMT may represent an interesting marker for OS detection and a promising target for effective anti-cancer therapy.

Enhancing nicotinamide N-methyltransferase bisubstrate inhibitor activity through 7-deazaadenosine and linker modifications.

Nicotinamide N-methyltransferase (NNMT) catalyzes the transfer of a methyl group from S-adenosylmethionine (SAM) to nicotinamide (NAM) and other pyridine-related compounds and is involved in various metabolic processes in the human body. In addition, abnormal expression of NNMT occurs under various pathological conditions such as cancer, diabetes, metabolic disorders, and neurodegenerative diseases, making it a promising drug target worthy of in-depth research. Small-molecule NNMT inhibitors with high potency and selectivity are necessary chemical tools to test biological hypotheses and potential therapies. In this study, we developed a series of highly active NNMT inhibitors by modifying N7 position of adenine. Among them, compound 3-12 (IC50 = 47.9 ± 0.6 nM) exhibited potent inhibitory activity and also had an excellent selectivity profile over a panel of human methyltransferases. We showed that the N7 position of adenine in the NNMT bisubstrate inhibitor was a modifiable site, thus offering insights into the development of NNMT inhibitors.

NNMT enriches for AQP5+ cancer stem cells to drive malignant progression in early gastric cardia adenocarcinoma.

OBJECTIVE: Early gastric cardia adenocarcinoma (EGCA) is a highly heterogeneous cancer, and the understanding of its classification and malignant progression is limited. This study explored the cellular and molecular heterogeneity in EGCA using single-cell RNA sequencing (scRNA-seq). DESIGN: scRNA-seq was conducted on 95 551 cells from endoscopic biopsies of low-grade intraepithelial neoplasia, well/moderately/poorly differentiated EGCA and their paired adjacent nonmalignant biopsy samples. Large-scale clinical samples and functional experiments were employed. RESULTS: Integrative analysis of epithelial cells revealed that chief cells, parietal cells and enteroendocrine cells were rarely detected in the malignant epithelial subpopulation, whereas gland and pit mucous cells and AQP5+ stem cells were predominant during malignant progression. Pseudotime and functional enrichment analyses showed that the WNT and NF-κB signalling pathways were activated during the transition. Cluster analysis of heterogeneous malignant cells revealed that NNMT-mediated nicotinamide metabolism was enriched in gastric mucin phenotype cell population, which was associated with tumour initiation and inflammation-induced angiogenesis. Furthermore, the expression level of NNMT was gradually increased during the malignant progression and associated with poor prognosis in cardia adenocarcinoma. Mechanistically, NNMT catalysed the conversion of nicotinamide to 1-methyl nicotinamide via depleting S-adenosyl methionine, which led to a reduction in H3K27 trimethylation (H3K27me3) and then activated the WNT signalling pathway to maintain the stemness of AQP5+ stem cells during EGCA malignant progression. CONCLUSION: Our study extends the understanding of the heterogeneity of EGCA and identifies a functional NNMT+/AQP5+ population that may drive malignant progression in EGCA and could be used for early diagnosis and therapy.

Nicotinamide N-methyltransferase upregulation contributes to palmitate-elicited peroxisome proliferator-activated receptor transactivation in hepatocytes.

Peroxisome proliferator-activated receptor γ (PPARγ) plays a pivotal role in regulating lipid metabolism and hepatic PPARγ transactivation contributes to fatty liver development. Fatty acids (FAs) are well-known endogenous ligands for PPARγ. Palmitate, a 16-C saturated FA (SFA) and the most abundant SFA in human circulation, is a strong inducer of hepatic lipotoxicity, a central pathogenic factor for various fatty liver diseases. In this study, using both alpha mouse liver 12 (AML12) and primary mouse hepatocytes, we investigated the effects of palmitate on hepatic PPARγ transactivation and underlying mechanisms, as well as the role of PPARγ transactivation in palmitate-induced hepatic lipotoxicity, all of which remain ambiguous currently. Our data revealed that palmitate exposure was concomitant with both PPARγ transactivation and upregulation of nicotinamide N-methyltransferase (NNMT), a methyltransferase catalyzing the degradation of nicotinamide, the predominant precursor for cellular NAD+ biosynthesis. Importantly, we discovered that PPARγ transactivation by palmitate was blunted by NNMT inhibition, suggesting that NNMT upregulation plays a mechanistic role in PPARγ transactivation. Further investigations uncovered that palmitate exposure is associated with intracellular NAD+ decline and NAD+ replenishment with NAD+-enhancing agents, nicotinamide and nicotinamide riboside, obstructed palmitate-induced PPARγ transactivation, implying that cellular NAD+ decline resulted from NNMT upregulation represents a potential mechanism behind palmitate-elicited PPARγ transactivation. At last, our data showed that the PPARγ transactivation marginally ameliorated palmitate-induced intracellular triacylglycerol accumulation and cell death. Collectively, our data provided the first-line evidence supporting that NNMT upregulation plays a mechanistic role in palmitate-elicited PPARγ transactivation, potentially through reducing cellular NAD+ contents.NEW & NOTEWORTHY Hepatic PPARγ transactivation contributes to fatty liver development. Saturated fatty acids (SFAs) induce hepatic lipotoxicity. Here, we investigated whether and how palmitate, the most abundant SFA in the human blood, affects PPARγ transactivation in hepatocytes. We reported for the first time that upregulation of nicotinamide N-methyltransferase (NNMT), a methyltransferase catalyzing the degradation of nicotinamide, the predominant precursor for cellular NAD+ biosynthesis, plays a mechanistic role in regulating palmitate-elicited PPARγ transactivation through reducing intracellular NAD+ contents.

Nicotinamide N-methyltransferase ameliorates renal fibrosis by its metabolite 1-methylnicotinamide inhibiting the TGF-ß1/Smad3 pathway.

Chronic kidney disease (CKD), a disease involving damage to the kidney structure and function, is a global public health problem. Tubulointerstitial fibrosis (TIF) is both an inevitable pathological change in individuals with CKD and a driving force in the progression of renal fibrosis. Nicotinamide N-methyltransferase (NNMT) and its metabolite 1-methylnicotinamide (MNAM) have been shown to protect against lipotoxicity-induced kidney tubular injury. However, the biological roles of NNMT and MNAM in regulating TIF remain elusive. This study aimed to investigate the protective effect of NNMT and MNAM on TIF and the mechanisms involved. We explored the functions and mechanisms of NNMT and MNAM in TIF, as well as the interaction between NNMT and MNAM, using unilateral ureteral obstruction (UUO) mice and cultured mouse tubular epithelial cells (mTECs) stimulated with transforming growth factor-ß1 (TGF-ß1). Several important findings were obtained as follows: (1) NNMT expression was upregulated in the kidneys of UUO mice and TGF-ß1-induced mTECs, and this upregulation was proposed to be a protective compensatory response to TIF. (2) MNAM was a potentially effective antifibrotic and anti-inflammatory medication in UUO mice. (3) The antifibrotic effect of NNMT overexpression was exerted by increasing the concentration of MNAM. (4) The renoprotective role of MNAM depended on the selective blockade of the interaction of Smad3 with TGFß receptor I. Overall, our study shows that NNMT is involved in the development and progression of CKD and that its metabolite MNAM may be a novel inhibitor of the TGF-ß1/Smad3 pathway with great therapeutic potential for CKD.

Ligand-based in silico identification and biological evaluation of potential inhibitors of nicotinamide N-methyltransferase.

Nicotinamide N-methyltransferase (NNMT) is a protein coding gene, which methylates the nicotinamide (NA) (vitamin B3) to produce 1-methylnicotinamide (MNA). Several studies have suggested that the overexpression of NNMT is associated with different metabolic disorders like obesity and type-2 diabetes thereby making it an important therapeutic target for development of anti-diabetic agents. Here we describe a workflow for identification of new inhibitors of NNMT from a library of small molecules. In this study, we have hypothesized a four-point pharmacophore model based on the pharmacophoric features of reported NNMT inhibitors in the literature. The statistically significant pharmacophore hypothesis was used to explore the Maybridge compound library that resulted in mapping of 1330 hit compounds on the proposed hypothesis. Subsequently, a total of eight high scoring compounds, showing good protein-ligand interactions in the molecular docking study, were selected for biological evaluation of NNMT activity. Eventually, four compounds were found to show significant inhibitory activity for NNMT and can be further explored to design new derivatives around the identified scaffolds with improved activities as NNMT inhibitors.

Nicotinamide N-Methyltransferase inhibits HBV replication by suppressing NR5A1 expression invitro.

Chronic hepatitis B virus (HBV) infection can lead to fibrosis, liver cirrhosis, and primary hepatocellular carcinoma. Investigating host factors that regulate HBV replication helps to identify antiviral targets. In the current study, we identified Nicotinamide N-Methyltransferase gene (NNMT) as a novel factor that regulates HBV transcription. NNMT is up-regulated at both the mRNA and protein levels in HepG2.2.15 cells compared to HepG2 cells. Overexpression of NNMT reduces HBV replication in several cell models, while knockdown of NNMT enhances HBV DNA levels. Mechanistically, NNMT suppresses HBV DNA replication by inhibiting HBV RNA transcription. The region required for the inhibitory effect of NNMT was narrowed to nt 1672-1708 in enhancer II by luciferase assays. On the other hand, ChIP assays and EMSA results showed that NNMT does not bind to this region substantially, either directly or indirectly. Next, a collection of hepatic nuclear receptor transcription factors was screened to determine whether they were affected by NNMT overexpression. NR5A1, a positive regulator of HBV replication, decreased significantly after NNMT overexpression. Collectively, the findings of this study shed light on the regulation of HBV transcription.

Nicotinamide N-methyltransferase expression in squamous cell carcinoma of the vulva.

The molecular pathways involved in the development of vulvar squamous cell carcinoma (SCC) cancer are not completely known. Nicotinamide N-methyltransferase (NNMT) is a cytosolic enzyme associated with tumorigenesis and metastasis in a variety of cancers. Its role in vulvar cancer has not been studied, previously. Vulvar SCC, high and low grade squamous intraepithelial lesions (SILs) and benign squamous hyperplasia were analysed immunohistochemically. The mean staining score for vulvar SCC was significantly higher than the score for vulvar squamous hyperplasia (p<.001). The mean relapse-free survival for patients with low and high NNMT expression was 41.4 months (95% CI: 25.6-57.2) and 19.8 months (95% CI: 3.0-36.6), respectively (p=.035). The mean disease-specific survival for patients with low and high NNMT expression was 75.8 months (95% CI: 57.5-94.2) and 27.8 months (95% CI 12.2-43.4), respectively (p=.015). Although quite preliminary, this study showed that NNMT expression was elevated in vulvar SCC compared to benign and premalignant lesions. Additionally, elevated NNMT expression was associated with poor survival. Impact StatementWhat is already known on this subject? Nicotinamide N-methyltransferase (NNMT) is a methyltransferase, associated with tumour progression, spread and poor prognosis in a variety of cancers. Its upregulation can lead to DNA hypomethylation, which can in turn result in the activation of proto-oncogenes and deactivation of tumour suppressor genes.What do the results of this study add? Although quite preliminary, this study showed that NNMT expression was elevated in vulvar SCC compared to benign and premalignant lesions. Additionally, elevated NNMT expression was associated with poor survival.What are the implications of these findings for clinical practice and/or further research? NNMT has been regarded as a potential target of cancer therapy and its role in vulvar cancer has not been studied, previously. This is the first study to investigate the expression of NNMT in vulvar cancer and associate NNMT elevation with poor survival. NNMT can further be investigated as a possible target of vulvar cancer therapy.

Exploring Unconventional SAM Analogues To Build Cell-Potent Bisubstrate Inhibitors for Nicotinamide N-Methyltransferase.

Nicotinamide N-methyltransferase (NNMT) methylates nicotinamide and has been associated with various diseases. Herein, we report the first cell-potent NNMT bisubstrate inhibitor II399, demonstrating a Ki of 5.9 nM in a biochemical assay and a cellular IC50 value of 1.9 µM. The inhibition mechanism and cocrystal structure confirmed II399 engages both the substrate and cofactor binding pockets. Computational modeling and binding data reveal a balancing act between enthalpic and entropic components that lead to II399's low nM binding affinity. Notably, II399 is 1 000-fold more selective for NNMT than closely related methyltransferases. We expect that II399 would serve as a valuable probe to elucidate NNMT biology. Furthermore, this strategy provides the first case of introducing unconventional SAM mimics, which can be adopted to develop cell-potent inhibitors for other SAM-dependent methyltransferases.

Nicotinamide N-methyltransferase upregulation via the mTORC1-ATF4 pathway activation contributes to palmitate-induced lipotoxicity in hepatocytes.

Defined as the dysfunction and/or cell death caused by toxic lipids accumulation in hepatocytes, hepatic lipotoxicity plays a pathological role in nonalcoholic fatty liver disease. The cellular and molecular mechanisms underlying lipotoxicity remain to be elucidated. In this study, using AML12 cells, a nontransformed murine hepatocyte cell line, exposed to palmitate (a 16-C saturated fatty acid) as an experimental model, we investigated the role and mechanisms of nicotinamide N-methyltransferase (NNMT), a methyltransferase catalyzing nicotinamide methylation and degradation, in hepatic lipotoxicity. We initially identified activating transcription factor 4 (ATF4) as a major transcription factor for hepatic NNMT expression. Here, we demonstrated that palmitate upregulates NNMT expression via activating ATF4 in a mechanistic target of rapamycin complex 1 (mTORC1)-dependent mechanism in that mTORC1 inhibition by both Torin1 and rapamycin attenuated ATF4 activation and NNMT upregulation. We further demonstrated that the mTORC1-dependent ATF4 activation is an integral signaling event of unfolded protein response (UPR) as both ATF4 activation and NNMT upregulation by tunicamycin, a well-documented endoplasmic reticulum (ER) stress inducer, are blunted when hepatocytes were pretreated with Torin1. Importantly, our data uncovered that NNMT upregulation contributes to palmitate-induced hepatotoxicity as NNMT inhibition, via either pharmacological (NNMT inhibitors) or genetic approach (siRNA transfection), provided protection against palmitate lipotoxicity. Our further mechanistic exploration identified protein kinase A (PKA) activation to contribute, at least, partially to the protective effect of NNMT inhibition against lipotoxicity. Collectively, our data demonstrated that NNMT upregulation by the mTORC1-ATF4 pathway activation contributes to the development of lipotoxicity in hepatocytes.

Patients with low nicotinamide N-methyltransferase expression benefit significantly from bevacizumab treatment in ovarian cancer.

BACKGROUND: The role of nicotinamide N-methyltransferase (NNMT) in ovarian cancer is still elusive. Our aim is to explore the expression of NNMT in ovarian cancer and to assess its association with patient prognosis and treatment response. METHODS: We first analyzed the differential expression of NNMT among fallopian tube epithelium, primary ovarian cancers, metastatic ovarian cancers, and recurrent ovarian cancers using Gene Expression Ominus (GEO) database (GSE10971, GSE30587, GSE44104 and TCGA datasets). Then, we assessed the association of NNMT expression with clinical and molecular parameters using CSIOVDB database and GSE28739 dataset. Next, we evaluate the association of NNMT expression with the prognosis of ovarian cancer patients in both GSE9891 dataset and TCGA dataset. Finally, GSE140082 dataset was used to explore the association of NNMT expression with bevacizumab response. RESULTS: NNMT expression was significantly elevated in lymphovascular space invasion (LVSI)-positive ovarian cancers compared with that in LVSI-negative ovarian cancers (TCGA dataset, P < 0.05), Moreover, increased expression of NNMT was associated with increased tumor stage, grade, and mesenchymal molecular subtype (CSIOVDB database). Survival analysis indicated that increased expression of NNMT was associated with a reduced OS in both GSE9891 dataset (HR: 2.28, 95%CI: 1.51-3.43, Log-rank P < 0.001) and TCGA dataset (HR: 1.55, 95%CI: 1.02-2.36, Log-rank P = 0.039). Multivariate analysis further confirmed the negative impact of NNMT expression on OS in ovarian cancer patients in those two datasets. Furthermore, the NNMT-related nomogram showed that NNMT shared a larger contribution to OS, compared with debulking status. More interestingly, bevacizumab conferred significant improvements in OS for patients with low NNMT expression (HR: 0.56, 95%CI: 0.31-0.99, Log-rank P = 0.049). In contrast, patients with high NNMT expression didn't benefit from bevacizumab treatment significantly (HR: 0.85, 95%CI: 0.48-1.49, Log-rank P = 0.561). NNMT expression was positively correlated with the expression of genes, LDHA and PGAM1, involved in Warburg effect. CONCLUSIONS: In conclusion, NNMT expression is associated with the aggressive behavior of ovarian cancer, correlates with a poor prognosis, and is predictive of sensitivity to bevacizumab treatment.

Atorvastatin and pravastatin stimulate nitric oxide and reactive oxygen species generation, affect mitochondrial network architecture and elevate nicotinamide N-methyltransferase level in endothelial cells.

Statins belong to the most often prescribed medications, which efficiently normalise hyperlipidaemia and prevent cardiovascular complications in obese and diabetic patients. However, beside expected therapeutic results based on the inhibition of 3-hydroxyl-3-methylglutaryl-CoA reductase, these drugs exert multiple side effects of poorly understood characteristic. In this study, side effects of pravastatin and atorvastatin on EA.hy926 endothelial cell line were investigated. It was found that both statins activate proinflammatory response, elevate nitric oxide and reactive oxygen species (ROS) generation and stimulate antioxidative response in these cells. Moreover, only slight stimulation of the mitochondrial biogenesis and significant changes in the mitochondrial network organisation have been noted. Although biochemical bases behind these effects are not clear, they may partially be explained as an elevation of AMP-activated protein kinase (AMPK) activity and an increased activating phosphorylation of sirtuin 1 (Sirt1), which were observed in statins-treated cells. In addition, both statins increased nicotinamide N-methyltransferase (NNMT) protein level that may explain a reduced fraction of methylated histone H3. Interestingly, a substantial reduction of the total level of histone H3 in cells treated with pravastatin but not atorvastatin was also observed. These results indicate a potential additional biochemical target for statins related to reduced histone H3 methylation due to increased NNMT protein level. Thus, NNMT may directly modify gene activity.

Nicotinamide N-Methyltransferase in Acquisition of Stem Cell Properties and Therapy Resistance in Cancer.

The activity of nicotinamide N-methyltransferase (NNMT) is tightly linked to the maintenance of the nicotinamide adenine dinucleotide (NAD+) level. This enzyme catalyzes methylation of nicotinamide (NAM) into methyl nicotinamide (MNAM), which is either excreted or further metabolized to N1-methyl-2-pyridone-5-carboxamide (2-PY) and H2O2. Enzymatic activity of NNMT is important for the prevention of NAM-mediated inhibition of NAD+-consuming enzymes poly-adenosine -diphosphate (ADP), ribose polymerases (PARPs), and sirtuins (SIRTs). Inappropriately high expression and activity of NNMT, commonly present in various types of cancer, has the potential to disrupt NAD+ homeostasis and cellular methylation potential. Largely overlooked, in the context of cancer, is the inhibitory effect of 2-PY on PARP-1 activity, which abrogates NNMT's positive effect on cellular NAD+ flux by stalling liberation of NAM and reducing NAD+ synthesis in the salvage pathway. This review describes, and discusses, the mechanisms by which NNMT promotes NAD+ depletion and epigenetic reprogramming, leading to the development of metabolic plasticity, evasion of a major tumor suppressive process of cellular senescence, and acquisition of stem cell properties. All these phenomena are related to therapy resistance and worse clinical outcomes.

Alterations in Kynurenine and NAD+ Salvage Pathways during the Successful Treatment of Inflammatory Bowel Disease Suggest HCAR3 and NNMT as Potential Drug Targets.

A meta-analysis of publicly available transcriptomic datasets was performed to identify metabolic pathways profoundly implicated in the progression and treatment of inflammatory bowel disease (IBD). The analysis revealed that genes involved in tryptophan (Trp) metabolism are upregulated in Crohn's disease (CD) and ulcerative colitis (UC) and return to baseline after successful treatment with infliximab. Microarray and mRNAseq profiles from multiple experiments confirmed that enzymes responsible for Trp degradation via the kynurenine pathway (IDO1, KYNU, IL4I1, KMO, and TDO2), receptor of Trp metabolites (HCAR3), and enzymes catalyzing NAD+ turnover (NAMPT, NNMT, PARP9, CD38) were synchronously coregulated in IBD, but not in intestinal malignancies. The modeling of Trp metabolite fluxes in IBD indicated that changes in gene expression shifted intestinal Trp metabolism from the synthesis of 5-hydroxytryptamine (5HT, serotonin) towards the kynurenine pathway. Based on pathway modeling, this manifested in a decline in mucosal Trp and elevated kynurenine (Kyn) levels, and fueled the production of downstream metabolites, including quinolinate, a substrate for de novo NAD+ synthesis. Interestingly, IBD-dependent alterations in Trp metabolites were normalized in infliximab responders, but not in non-responders. Transcriptomic reconstruction of the NAD+ pathway revealed an increased salvage biosynthesis and utilization of NAD+ in IBD, which normalized in patients successfully treated with infliximab. Treatment-related changes in NAD+ levels correlated with shifts in nicotinamide N-methyltransferase (NNMT) expression. This enzyme helps to maintain a high level of NAD+-dependent proinflammatory signaling by removing excess inhibitory nicotinamide (Nam) from the system. Our analysis highlights the prevalent deregulation of kynurenine and NAD+ biosynthetic pathways in IBD and gives new impetus for conducting an in-depth examination of uncovered phenomena in clinical studies.

Novel Inhibitors of Nicotinamide-N-Methyltransferase for the Treatment of Metabolic Disorders.

Nicotinamide-N-methyltransferase (NNMT) is a cytosolic enzyme catalyzing the transfer of a methyl group from S-adenosyl-methionine (SAM) to nicotinamide (Nam). It is expressed in many tissues including the liver, adipose tissue, and skeletal muscle. Its expression in several cancer cell lines has been widely discussed in the literature, and recent work established a link between NNMT expression and metabolic diseases. Here we describe our approach to identify potent small molecule inhibitors of NNMT featuring different binding modes as elucidated by X-ray crystallographic studies.

Nicotinamide N-methyltransferase overexpression may be associated with poor prognosis in ovarian cancer.

Ovarian cancer is the fifth leading cause of cancer-related mortality in women. Nicotinamide N-methyltransferase (NNMT) is a metabolic enzyme and there is growing evidence to suggest that it plays an important role in cancer progression. This is the first study to examine the expression of NNMT in serous ovarian cystadenomas, serous borderline tumours, low grade serous carcinomas (LGSC) and high grade serous carcinomas (HGSC) and investigate the potential independent association of NNMT expression with survival. Tissue samples were analysed immunohistochemically for NNMT expression. The stromal NNMT score was significantly higher in HGSC compared to serous cystadenomas and serous borderline tumours (p < .001, p < .043, respectively). The mean stromal NNMT score of patients with HGSC was significantly higher than patients with LGSC (p = .043). Patients with low expression of NNMT had a significantly higher mean recurrence-free survival than patients with high expression (p = .036). NNMT may support tumour progression in ovarian cancer by promoting desmoplastic stromal tumour reaction. NNMT overexpression may be associated with poor prognosis and can be a therapeutic target in ovarian cancer.IMPACT STATEMENTWhat is already known on this subject? Nicotinamide N-methyltransferase (NNMT) is a cytosolic enzyme that is overexpressed in many malignancies. Its overexpression was shown to lead to histone hypomethylation, which in turn can decrease and increase the expression of tumour suppressor proteins and onco-proteins, respectively. NNMT was also shown to play a role in epithelial-to-mesenchymal transition, which is critical in tumour progression and the stromal tumour reaction. The stromal tumour reaction was recently targeted with promising therapeutic results in ovarian cancer.What do the results of this study add? The expression of NNMT in various ovarian neoplasms including serous cystadenomas, borderline tumours and serous carcinomas has not been studied and independently associated with poor survival, previously. This study suggests that NNMT is progressively overexpressed in the stroma of ovarian neoplasms from benign cysts to HGSCs. NNMT overexpression appears to be independently associated with poor survival in ovarian cancer.What are the implications of these findings for clinical practice and/or further research? The implications of these findings are that NNMT may play an important role in the stromal tumour reaction, and therefore its overexpression may contribute to poor survival. NNMT overexpression may be an important target of ovarian cancer therapy.

Nicotinamide N-methyltransferase decreases 5-fluorouracil sensitivity in human esophageal squamous cell carcinoma through metabolic reprogramming and promoting the Warburg effect.

Esophageal squamous cell carcinoma (ESCC) is a common malignant tumor with poor prognosis. And different individuals respond to the same drug differently. Increasing evidence has confirmed that metabolism reprogramming was involved in the drug sensitivity of tumor cells. However, the potential molecular mechanism of 5-fluorouracil (5-FU) sensitivity remains to be elucidated in ESCC cells. In this study, we found that the 5-FU sensitivity of TE1 cells was lower than that of EC1 and Eca109 cells. Gas chromatography-mass spectrometry analysis results showed that nicotinate and nicotinamide metabolism and tricarboxylic acid cycle were significantly different in these three cell lines. Nicotinamide N-methyltransferase (NNMT), a key enzyme of nicotinate and nicotinamide metabolism, was significantly higher expressed in TE1 cells than that in EC1 and Eca109 cells. Therefore, the function of NNMT on 5-FU sensitivity was analyzed in vitro and in vivo. NNMT downregulation significantly increased 5-FU sensitivity in TE1 cells. Meanwhile, the glucose consumption and lactate production were decreased, and the expression of glycolysis-related enzymes hexokinase 2, lactate dehydrogenase A, and phosphoglycerate mutase 1 were downregulated in NNMT knockdown TE1 cells. Besides, overexpression of NNMT in EC1 and Eca109 cells caused the opposite effects. Moreover, when glycolysis was inhibited by 2-deoxyglucose, the roles of NNMT on 5-FU sensitivity was weakened. In vivo experiments showed that NNMT knockdown significantly increased the sensitivity of xenografts to 5-FU and suppressed the Warburg effect. Overall, these results demonstrated that NNMT decreases 5-FU sensitivity in human ESCC cells through promoting the Warburg effect, suggesting that NNMT may contribute to predict the treatment effects of the clinical chemotherapy in ESCC.