NAD metabolism-related genes provide prognostic value and potential therapeutic insights for acute myeloid leukemia.

Introduction: Acute myeloid leukemia (AML) is an aggressive blood cancer with high heterogeneity and poor prognosis. Although the metabolic reprogramming of nicotinamide adenine dinucleotide (NAD) has been reported to play a pivotal role in the pathogenesis of acute myeloid leukemia (AML), the prognostic value of NAD metabolism and its correlation with the immune microenvironment in AML remains unclear. Methods: We utilized our large-scale RNA-seq data on 655 patients with AML and the NAD metabolism-related genes to establish a prognostic NAD metabolism score based on the sparse regression analysis. The signature was validated across three independent datasets including a total of 1,215 AML patients. ssGSEA and ESTIMATE algorithms were employed to dissect the tumor immune microenvironment. Ex vivo drug screening and in vitro experimental validation were performed to identify potential therapeutic approaches for the high-risk patients. In vitro knockdown and functional experiments were employed to investigate the role of SLC25A51, a mitochondrial NAD+ transporter gene implicated in the signature. Results: An 8-gene NAD metabolism signature (NADM8) was generated and demonstrated a robust prognostic value in more than 1,800 patients with AML. High NADM8 score could efficiently discriminate AML patients with adverse clinical characteristics and genetic lesions and serve as an independent factor predicting a poor prognosis. Immune microenvironment analysis revealed significant enrichment of distinct tumor-infiltrating immune cells and activation of immune checkpoints in patients with high NADM8 scores, acting as a potential biomarker for immune response evaluation in AML. Furthermore, ex vivo drug screening and in vitro experimental validation in a panel of 9 AML cell lines demonstrated that the patients with high NADM8 scores were more sensitive to the PI3K inhibitor, GDC-0914. Finally, functional experiments also substantiated the critical pathogenic role of the SLC25A51 in AML, which could be a promising therapeutic target. Conclusion: Our study demonstrated that NAD metabolism-related signature can facilitate risk stratification and prognosis prediction in AML and guide therapeutic decisions including both immunotherapy and targeted therapies.

A pH-dependent shift of redox cofactor specificity in a benzyl alcohol dehydrogenase of aromatoleum aromaticum EbN1.

We characterise a reversible bacterial zinc-containing benzyl alcohol dehydrogenase (BaDH) accepting either NAD+ or NADP+ as a redox cofactor. Remarkably, its redox cofactor specificity is pH-dependent with the phosphorylated cofactors favored at lower and the dephospho-forms at higher pH. BaDH also shows different steady-state kinetic behavior with the two cofactor forms. From a structural model, the pH-dependent shift may affect the charge of a histidine in the 2'-phosphate-binding pocket of the redox cofactor binding site. The enzyme is phylogenetically affiliated to a new subbranch of the Zn-containing alcohol dehydrogenases, which share this conserved residue. BaDH appears to have some specificity for its substrate, but also turns over many substituted benzyl alcohol and benzaldehyde variants, as well as compounds containing a conjugated C=C double bond with the aldehyde carbonyl group. However, compounds with an sp3-hybridised C next to the alcohol/aldehyde group are not or only weakly turned over. The enzyme appears to contain a Zn in its catalytic site and a mixture of Zn and Fe in its structural metal-binding site. Moreover, we demonstrate the use of BaDH in an enzyme cascade reaction with an acid-reducing tungsten enzyme to reduce benzoate to benzyl alcohol. KEY POINTS: •Zn-containing BaDH has activity with either NAD + or NADP+ at different pH optima. •BaDH converts a broad range of substrates. •BaDH is used in a cascade reaction for the reduction of benzoate to benzyl alcohol.

The structural basis of the activation and inhibition of DSR2 NADase by phage proteins.

DSR2, a Sir2 domain-containing protein, protects bacteria from phage infection by hydrolyzing NAD+. The enzymatic activity of DSR2 is triggered by the SPR phage tail tube protein (TTP), while suppressed by the SPbeta phage-encoded DSAD1 protein, enabling phages to evade the host defense. However, the molecular mechanisms of activation and inhibition of DSR2 remain elusive. Here, we report the cryo-EM structures of apo DSR2, DSR2-TTP-NAD+ and DSR2-DSAD1 complexes. DSR2 assembles into a head-to-head tetramer mediated by its Sir2 domain. The C-terminal helical regions of DSR2 constitute four partner-binding cavities with opened and closed conformation. Two TTP molecules bind to two of the four C-terminal cavities, inducing conformational change of Sir2 domain to activate DSR2. Furthermore, DSAD1 competes with the activator for binding to the C-terminal cavity of DSR2, effectively suppressing its enzymatic activity. Our results provide the mechanistic insights into the DSR2-mediated anti-phage defense system and DSAD1-dependent phage immune evasion.

The T120P or M172V mutation on rv2172c confers high level para-aminosalicylic acid resistance in Mycobacterium tuberculosis.

Although para-aminosalicylic acid (PAS) has been used to treat tuberculosis for decades, mechanisms of resistance to this drug in Mycobacterium tuberculosis (M. tuberculosis) clinical isolates have not been thoroughly investigated. Previously, we found that decreased methylenetetrahydrofolate reductase (MTHFR) activity of Rv2172c led to increased sensitivity to antifolates in M. tuberculosis. In this study, we collected the genome-sequencing data of 173 PAS-resistant and 803 PAS-sensitive clinical isolates and analyzed rv2172c mutations in those 976 isolates. The results showed that two mutations (T120P and M172V) on rv2172c could be identified in a certain proportion (6.36%) of PAS-resistant isolates. The results of AlphaFold2 prediction indicated that the T120P or M172V mutation might affect the enzymatic activity of Rv2172c by influencing nicotinamide adenine dinucleotide (NADH) binding, and this was verified by subsequent biochemical analysis, demonstrating the role of residues Thr120 and Met172 on NADH binding and enzymatic activity of Rv2172c. In addition, the effect of rv2172c T120P or M172V mutation on methionine production and PAS resistance was determined in M. tuberculosis. The results showed that both T120P and M172V mutations caused increased intracellular methionine concentrations and high level PAS resistance. In summary, we discovered new molecular markers and also a novel mechanism of PAS resistance in M. tuberculosis clinical isolates and broadened the understanding of the NADH-dependent MTHFR catalytic mechanism of Rv2172c in M. tuberculosis, which will facilitate the molecular diagnosis of PAS resistance and also the development of new drugs targeting Rv2172c.

STAT5 Is Necessary for the Metabolic Switch Induced by IL-2 in Cervical Cancer Cell Line SiHa.

The tumor cells reprogram their metabolism to cover their high bioenergetic demands for maintaining uncontrolled growth. This response can be mediated by cytokines such as IL-2, which binds to its receptor and activates the JAK/STAT pathway. Some reports show a correlation between the JAK/STAT pathway and cellular metabolism, since the constitutive activation of STAT proteins promotes glycolysis through the transcriptional activation of genes related to energetic metabolism. However, the role of STAT proteins in the metabolic switch induced by cytokines in cervical cancer remains poorly understood. In this study, we analyzed the effect of IL-2 on the metabolic switch and the role of STAT5 in this response. Our results show that IL-2 induces cervical cancer cell proliferation and the tyrosine phosphorylation of STAT5. Also, it induces an increase in lactate secretion and the ratio of NAD+/NADH, which suggest a metabolic reprogramming of their metabolism. When STAT5 was silenced, the lactate secretion and the NAD+/NADH ratio decreased. Also, the expression of HIF1α and GLUT1 decreased. These results indicate that STAT5 regulates IL-2-induced cell proliferation and the metabolic shift to aerobic glycolysis by regulating genes related to energy metabolism. Our results suggest that STAT proteins modulate the metabolic switch in cervical cancer cells to attend to their high demand of energy required for cell growth and proliferation.

Ultrathin silica-tiling on living cells for chemobiotic catalysis.

Harnessing the power of cell biocatalysis for sustainable chemical synthesis requires rational integration of living cells with the modern synthetic catalysts. Here, we develop silica-tiling strategy that constructs a hierarchical, inorganic, protocellular confined nanospace around the individual living cell to accommodate molecularly accessible abiotic catalytic sites. This empowers the living microorganisms for new-to-nature chemical synthesis without compromising the cellular regenerative process. Yeast cell, a widely used biocatalyst, is upgraded via highly controlled self-assembly of 2D-bilayer silica-based catalytic modules on cell surfaces, opening the avenues for diverse chemobiotic reactions. For example, combining [AuPt]-catalyzed NADH regeneration, light-induced [Pd]-catalyzed C-C cross-coupling or lipase-catalyzed esterification reactions-with the natural ketoreductase activity inside yeast cell. The conformal silica bilayer provides protection while allowing proximity to catalytic sites and preserving natural cell viability and proliferation. These living nanobiohybrids offer to bridge cell's natural biocatalytic capabilities with customizable heterogeneous metal catalysis, enabling programmable reaction sequences for sustainable chemical synthesis.

Discovery and characterization of a new class of NAD+-independent SIRT1 activators.

The activity of sirtuin 1 (SIRT1, a member of the NAD+-dependent deacetylases family) decreases during aging as NAD+ levels naturally decline, thus increasing the risk of several age-associated diseases. Several sirtuin-activating compounds (STACs) have been developed to counteract the age-associated reduction in SIRT1 activity, and some of them are currently under development in clinical trials. STACs induce SIRT1 activation, either through allosteric activation of the enzyme in the presence of NAD+, or by increasing NAD+ levels by inhibiting its degradation or by supplying a key precursor in biosynthesis. In this study, we have identified (E)-2'-des-methyl sulindac analogues as a novel class of STACs that act also in the absence of NAD+, a peculiar behavior demonstrated through enzymatic and mass spectrometry experiments, both in vitro and in cell lines. The activation of the SIRT1 pathway was confirmed in vivo through gene expression and metabolomics analysis. Our data suggest that these compounds could serve as candidate leads for a novel therapeutic strategy aimed at addressing a key metabolic deficiency that may contribute to metabolic and age-associated diseases.

ER calcium depletion as a key driver for impaired ER-to-mitochondria calcium transfer and mitochondrial dysfunction in Wolfram syndrome.

Wolfram syndrome is a rare genetic disease caused by mutations in the WFS1 or CISD2 gene. A primary defect in Wolfram syndrome involves poor ER Ca2+ handling, but how this disturbance leads to the disease is not known. The current study, performed in primary neurons, the most affected and disease-relevant cells, involving both Wolfram syndrome genes, explains how the disturbed ER Ca2+ handling compromises mitochondrial function and affects neuronal health. Loss of ER Ca2+ content and impaired ER-mitochondrial contact sites in the WFS1- or CISD2-deficient neurons is associated with lower IP3R-mediated Ca2+ transfer from ER to mitochondria and decreased mitochondrial Ca2+ uptake. In turn, reduced mitochondrial Ca2+ content inhibits mitochondrial ATP production leading to an increased NADH/NAD+ ratio. The resulting bioenergetic deficit and reductive stress compromise the health of the neurons. Our work also identifies pharmacological targets and compounds that restore Ca2+ homeostasis, enhance mitochondrial function and improve neuronal health.

Mechanoresponsive Protein Crystals for NADH Recycling in Multicycle Enzyme Reactions.

NAD(H)-dependent enzymes play a crucial role in the biosynthesis of pharmaceuticals and fine chemicals, but the limited recyclability of the NAD(H) cofactor hinders its more general application. Here, we report the generation of mechano-responsive PEI-modified Cry3Aa protein crystals and their use for NADH recycling over multiple reaction cycles. For demonstration of its practical utility, a complementary Cry3Aa protein particle containing genetically encoded and co-immobilized formate dehydrogenase for NADH regeneration and leucine dehydrogenase for catalyzing the NADH-dependent l-tert-leucine (l-tert-Leu) biosynthesis has been produced. When combined with the PEI-modified Cry3Aa crystal, the resultant reaction system could be used for the efficient biosynthesis of l-tert-Leu for up to 21 days with a 10.5-fold improvement in the NADH turnover number.

Metabolic engineering of Komagataella phaffii for the efficient utilization of methanol.

BACKGROUND: Komagataella phaffii, a type of methanotrophic yeast, can use methanol, a favorable non-sugar substrate in eco-friendly bio-manufacturing. The dissimilation pathway in K. phaffii leads to the loss of carbon atoms in the form of CO2. However, the ΔFLD strain, engineered to lack formaldehyde dehydrogenase-an essential enzyme in the dissimilation pathway-displayed growth defects when exposed to a methanol-containing medium. RESULTS: Inhibiting the dissimilation pathway triggers an excessive accumulation of formaldehyde and a decline in the intracellular NAD+/NADH ratio. Here, we designed dual-enzyme complex with the alcohol oxidase1/dihydroxyacetone synthase1 (Aox1/Das1), enhancing the regeneration of the formaldehyde receptor xylulose-5-phosphate (Xu5P). This strategy mitigated the harmful effects of formaldehyde accumulation and associated toxicity to cells. Concurrently, we elevated the NAD+/NADH ratio by overexpressing isocitrate dehydrogenase in the TCA cycle, promoting intracellular redox homeostasis. The OD600 of the optimized combination of the above strategies, strain DF02-1, was 4.28 times higher than that of the control strain DF00 (ΔFLD, HIS4+) under 1% methanol. Subsequently, the heterologous expression of methanol oxidase Mox from Hansenula polymorpha in strain DF02-1 resulted in the recombinant strain DF02-4, which displayed a growth at an OD600 4.08 times higher than that the control strain DF00 in medium containing 3% methanol. CONCLUSIONS: The reduction of formaldehyde accumulation, the increase of NAD+/NADH ratio, and the enhancement of methanol oxidation effectively improved the efficient utilization of a high methanol concentration by strain ΔFLD strain lacking formaldehyde dehydrogenase. The modification strategies implemented in this study collectively serve as a foundational framework for advancing the efficient utilization of methanol in K. phaffii.

Novel insight into nicotinamide adenine dinucleotide and related metabolites in cancer patients undergoing surgery.

Nicotinamide adenine dinucleotide (NAD +) plays a pivotal role in numerous cellular functions. Reduced NAD + levels are postulated to be associated with cancer. As interest in understanding NAD + dynamics in cancer patients with therapeutic applications in mind grows, there remains a shortage of comprehensive data. This study delves into NAD + dynamics in patients undergoing surgery for different digestive system cancers. This prospective study enrolled 99 patients with eight different cancers. Fasting blood samples were obtained during the perioperative period. The concentrations of NAD + , nicotinamide mononucleotide (NMN), and nicotinamide riboside were analyzed using tandem mass spectrometry. After erythrocyte volume adjustment, NAD + remained relatively stable after surgery. Meanwhile, NMN decreased the day after surgery and displayed a recovery trend. Interestingly, liver and pancreatic cancer patients exhibited poor postoperative NMN recovery, suggesting a potential cancer type-specific influence on NAD + metabolism. This study illuminated the behavior of NAD + in surgically treated cancer patients. We identified which cancer types have particularly low levels and at what point depletion occurs during the perioperative period. These insights suggest the need for personalized NAD + supplementation strategies, calibrated to individual patient needs and treatment timelines. Clinical trial registration jRCT1020210066.

Exploring nicotinamide adenine dinucleotide precursors across biosynthesis pathways: Unraveling their role in the ovary.

Natural Nicotinamide Adenine Dinucleotide (NAD+) precursors have attracted much attention due to their positive effects in promoting ovarian health. However, their target tissue, synthesis efficiency, advantages, and disadvantages are still unclear. This review summarizes the distribution of NAD+ at the tissue, cellular and subcellular levels, discusses its biosynthetic pathways and the latest findings in ovary, include: (1) NAD+ plays distinct roles both intracellularly and extracellularly, adapting its distribution in response to requirements. (2) Different precursors differs in target tissues, synthetic efficiency, biological utilization, and adverse effects. Importantly: tryptophan is primarily utilized in the liver and kidneys, posing metabolic risks in excess; nicotinamide (NAM) is indispensable for maintaining NAD+ levels; nicotinic acid (NA) constructs a crucial bridge between intestinal microbiota and the host with diverse functions; nicotinamide riboside (NR) and nicotinamide mononucleotide (NMN) increase NAD+ systemically and can be influenced by delivery route, tissue specificity, and transport efficiency. (3) The biosynthetic pathways of NAD+ are intricately intertwined. They provide multiple sources and techniques for NAD+ synthesis, thereby reducing the dependence on a single molecule to maintain cellular NAD+ levels. However, an excess of a specific precursor potentially influencing other pathways. In addition, Protein expression analysis suggest that ovarian tissues may preferentially utilize NAM and NMN. These findings summarize the specific roles and potential of NAD+ precursors in enhancing ovarian health. Future research should delve into the molecular mechanisms and intervention strategies of different precursors, aiming to achieve personalized prevention or treatment of ovarian diseases, and reveal their clinical application value.

in vivo and in situ robust quantitative optical biopsy of hepatocellular carcinoma and metastasis based on two-photon autofluorescence imaging.

Two-photon autofluorescence (TPAF) imaging is able to offer precise cellular metabolic information with high spatiotemporal resolution, making it a promising biopsy tool. The technique is greatly hampered by the complexity of either the optical system or data processing. Here, the excitation wavelength was optimized to simultaneously excite both flavin adenine dinucleotide and nicotinamide adenine dinucleotide and eliminate the unexpected TPAF. The optical redox ratio (ORR) images were robustly achieved without additional calibration under the optimized single-wavelength excitation. The in vitro, ex vivo, and in vivo biopsy by the TPAF method were systematically studied and compared using hepato-cellular carcinoma and metastasis as examples. It was demonstrated that the proposed TPAF method simplified the optical system, improved the robustness of ORR, and enabled early-stage cancer diagnosis, showing distinguished advantages as compared with previous methods.

Image Sensing of Gaseous Acetone Using Secondary Alcohol Dehydrogenase-Immobilized Mesh for Exhaled Air.

Human-borne acetone is a potent marker of lipid metabolism. Here, an enzyme immobilization method for secondary alcohol dehydrogenase (S-ADH), which is suitable for highly sensitive and selective biosensing of acetone, was developed, and then its applicability was demonstrated for spatiotemporal imaging of concentration distribution. After various investigations, S-ADH-immobilized meshes could be prepared with less than 5% variation by cross-linking S-ADH with glutaraldehyde on a cotton mesh at 40 °C for 15 min. Furthermore, high activity was obtained by adjusting the concentration of the coenzyme nicotinamide adenine dinucleotide (NADH) solution added to the S-ADH-immobilized mesh to 500 µM and the solvent to a potassium phosphate buffer solution at pH 6.5. The gas imaging system using the S-ADH-immobilized mesh was able to image the decrease in NADH fluorescence (ex 340 nm, fl 490 nm) caused by the catalytic reaction of S-ADH and the acetone distribution in the concentration range of 0.1-10 ppm-v, including the breath concentration of healthy people at rest. The exhaled breath of two healthy subjects at 6 h of fasting was quantified as 377 and 673 ppb-v, which were consistent with the values quantified by gas chromatography-mass spectrometry.

Dissemination of pathogenic bacteria is reinforced by a MARTX toxin effector duet.

Multiple bacterial genera take advantage of the multifunctional autoprocessing repeats-in-toxin (MARTX) toxin to invade host cells. Secretion of the MARTX toxin by Vibrio vulnificus, a deadly opportunistic pathogen that causes primary septicemia, the precursor of sepsis, is a major driver of infection; however, the molecular mechanism via which the toxin contributes to septicemia remains unclear. Here, we report the crystal and cryo-electron microscopy (EM) structures of a toxin effector duet comprising the domain of unknown function in the first position (DUF1)/Rho inactivation domain (RID) complexed with human targets. These structures reveal how the duet is used by bacteria as a potent weapon. The data show that DUF1 acts as a RID-dependent transforming NADase domain (RDTND) that disrupts NAD+ homeostasis by hijacking calmodulin. The cryo-EM structure of the RDTND-RID duet complexed with calmodulin and Rac1, together with immunological analyses in vitro and in mice, provide mechanistic insight into how V. vulnificus uses the duet to suppress ROS generation by depleting NAD(P)+ and modifying Rac1 in a mutually-reinforcing manner that ultimately paralyzes first line immune responses, promotes dissemination of invaders, and induces sepsis. These data may allow development of tools or strategies to combat MARTX toxin-related human diseases.

The blood serum metabolome profile after different phases of a 4-km cycling time trial: Secondary analysis of a randomized controlled trial.

It has been assumed that exercise intensity variation throughout a cycling time trial (TT) occurs in alignment of various metabolic changes to prevent premature task failure. However, this assumption is based on target metabolite responses, which limits our understanding of the complex interconnection of metabolic responses during exercise. The current study characterized the metabolomic profile, an untargeted metabolic analysis, after specific phases of a cycling 4-km TT. Eleven male cyclists performed three separated TTs in a crossover counterbalanced design, which were interrupted at the end of the fast-start (FS, 600 ± 205 m), even-pace (EP, 3600 ± 190 m), or end-spurt (ES, 4000 m) phases. Blood samples were taken before any exercise and 5 min after exercise cessation, and the metabolomic profile characterization was performed using Nuclear Magnetic Resonance metabolomics. Power output (PO) was also continually recorded. There were higher PO values during the FS and ES compared to the EP (all p < 0.05), which were accompanied by distinct metabolomic profiles. FS showed high metabolite expression in TCA cycle and its related pathways (e.g., glutamate, citric acid, and valine metabolism); whereas, the EP elicited changes associated with antioxidant effects and oxygen delivery adjustment. Finally, ES was related to pathways involved in NAD turnover and serotonin metabolism. These findings suggest that the specific phases of a cycling TT are accompanied by distinct metabolomic profiles, providing novel insights regarding the relevance of specific metabolic pathways on the process of exercise intensity regulation.

Exogenous NADH promotes the bactericidal effect of aminoglycoside antibiotics against Edwardsiella tarda.

The global surge in multidrug-resistant bacteria owing to antibiotic misuse and overuse poses considerable risks to human and animal health. With existing antibiotics losing their effectiveness and the protracted process of developing new antibiotics, urgent alternatives are imperative to curb disease spread. Notably, improving the bactericidal effect of antibiotics by using non-antibiotic substances has emerged as a viable strategy. Although reduced nicotinamide adenine dinucleotide (NADH) may play a crucial role in regulating bacterial resistance, studies examining how the change of metabolic profile and bacterial resistance following by exogenous administration are scarce. Therefore, this study aimed to elucidate the metabolic changes that occur in Edwardsiella tarda (E. tarda), which exhibits resistance to various antibiotics, following the exogenous addition of NADH using metabolomics. The effects of these alterations on the bactericidal activity of neomycin were investigated. NADH enhanced the effectiveness of aminoglycoside antibiotics against E. tarda ATCC15947, achieving bacterial eradication at low doses. Metabolomic analysis revealed that NADH reprogrammed the ATCC15947 metabolic profile by promoting purine metabolism and energy metabolism, yielding increased adenosine triphosphate (ATP) levels. Increased ATP levels played a crucial role in enhancing the bactericidal effects of neomycin. Moreover, exogenous NADH promoted the bactericidal efficacy of tetracyclines and chloramphenicols. NADH in combination with neomycin was effective against other clinically resistant bacteria, including Aeromonas hydrophila, Vibrio parahaemolyticus, methicillin-resistant Staphylococcus aureus, and Listeria monocytogenes. These results may facilitate the development of effective approaches for preventing and managing E. tarda-induced infections and multidrug resistance in aquaculture and clinical settings.

Pathological Changes and Metabolic Adaptation in the Myocardium of Rats in Response to Chronic Variable Mild Stress.

Chronic variable mild stress (CVS) in rats is a well-established paradigm for inducing depressive-like behaviors and has been utilized extensively to explore potential therapeutic interventions for depression. While the behavioral and neurobiological effects of CVS have been extensively studied, its impact on myocardial function remains largely unexplored. To induce the CVS model, rats were exposed to various stressors over 40 days. Behavioral assessments confirmed depressive-like behavior. Biochemical analyses revealed alterations in myocardial metabolism, including changes in NAD+ and NADP+, and NADPH concentrations. Free amino acid analysis indicated disturbances in myocardial amino acid metabolism. Evaluation of oxidative DNA damage demonstrated an increased number of abasic sites in the DNA of rats exposed to CVS. Molecular analysis showed significant changes in gene expression associated with glucose metabolism, oxidative stress, and cardiac remodeling pathways. Histological staining revealed minor morphological changes in the myocardium of CVS-exposed rats, including increased acidophilicity of cells, collagen deposition surrounding blood vessels, and glycogen accumulation. This study provides novel insights into the impact of chronic stress on myocardial function and metabolism, highlighting potential mechanisms linking depression and cardiovascular diseases. Understanding these mechanisms may aid in the development of targeted therapeutic strategies to mitigate the adverse cardiovascular effects of depression.

Inhibition of nicotinamide phosphoribosyltransferase (NAMPT) in cancer: a patent review.

INTRODUCTION: Nicotinamide phosphoribosyltransferase (NAMPT) is the rate-limiting enzyme in the biosynthesis of nicotinamide adenine dinucleotide (NAD) from nicotinamide. In addition to its role as essential redox cofactor, NAD also functions as a substrate for NAD-consuming enzymes, regulating multiple cellular processes such as DNA repair and gene expression, fundamental to sustain energetic needs for tumor growth. In this sense, NAMPT over-expression represents a common strategy that several tumor types adopt to sustain NAD production. In addition to its enzymatic role, NAMPT behaves as cytokine-like protein with pro-inflammatory function. Increasing evidence demonstrated that NAMPT inhibition represents a promising anti-cancer strategy to deplete NAD and impair cellular metabolism in cancer conditions. AREAS COVERED: By using Espacenet, we collected the patents which identified new molecules, compounds, formulations and methods able to inhibit NAMPT from 2007 to date. EXPERT OPINION: Most of the collected patents focused the attention on the ability of different compounds to inhibit the enzymatic activity of NAMPT, lacking other important aspects related to the extracellular role of NAMPT and the ability of alternative enzymes to counteract NAMPT-mediated NAD depletion. It is necessary to consider also these aspects to promote novel strategies and create novel inhibitors and molecules useful as anti-cancer compounds.

Structural characterization of macro domain-containing Thoeris antiphage defense systems.

Thoeris defense systems protect bacteria from infection by phages via abortive infection. In these systems, ThsB proteins serve as sensors of infection and generate signaling nucleotides that activate ThsA effectors. Silent information regulator and SMF/DprA-LOG (SIR2-SLOG) containing ThsA effectors are activated by cyclic ADP-ribose (ADPR) isomers 2'cADPR and 3'cADPR, triggering abortive infection via nicotinamide adenine dinucleotide (NAD+) depletion. Here, we characterize Thoeris systems with transmembrane and macro domain (TM-macro)-containing ThsA effectors. We demonstrate that ThsA macro domains bind ADPR and imidazole adenine dinucleotide (IAD), but not 2'cADPR or 3'cADPR. Combining crystallography, in silico predictions, and site-directed mutagenesis, we show that ThsA macro domains form nucleotide-induced higher-order oligomers, enabling TM domain clustering. We demonstrate that ThsB can produce both ADPR and IAD, and we identify a ThsA TM-macro-specific ThsB subfamily with an active site resembling deoxy-nucleotide and deoxy-nucleoside processing enzymes. Collectively, our study demonstrates that Thoeris systems with SIR2-SLOG and TM-macro ThsA effectors trigger abortive infection via distinct mechanisms.