Correction: Iannuzzi et al. Fyn Tyrosine Kinase Elicits Amyloid Precursor Protein Tyr682 Phosphorylation in Neurons from Alzheimer's Disease Patients. Cells 2020, 9, 1807.

Error in Figure [...].

Semaphorin 3A Increases in the Plasma of Women with Diminished Ovarian Reserve Who Respond Better to Controlled Ovarian Stimulation.

Semaphorin 3A (SEMA3A) plays a crucial role in the development, differentiation, and plasticity of specific types of neurons that secrete Gonadotropin-Releasing Hormone (GnRH) and regulates the acquisition and maintenance of reproductive competence in humans and mice. Its insufficient expression has been linked to reproductive disorders in humans, which are characterized by reduced or failed sexual competence. Various mutations, polymorphisms, and alternatively spliced variants of SEMA3A have been associated with infertility. One of the common causes of infertility in women of reproductive age is diminished ovarian reserve (DOR), characterized by a reduced ovarian follicular pool. Despite its clinical significance, there are no universally accepted diagnostic criteria or therapeutic interventions for DOR. In this study, we analyzed the SEMA3A plasma levels in 77 women and investigated their potential role in influencing fertility in patients with DOR. The results revealed that the SEMA3A levels were significantly higher in patients with DOR than in healthy volunteers. Furthermore, the SEMA3A levels were increased in patients who underwent fertility treatment and had positive Beta-Human Chorionic Gonadotropin (ßHCG) values (ß+) after controlled ovarian stimulation (COS) compared to those who had negative ßHCG values (ß-). These findings may serve as the basis for future investigations into the diagnosis of infertility and emphasize new possibilities for the SEMA3A-related treatment of sexual hormonal dysfunction that leads to infertility.

TNF-α Levels Are Increased in Patients with Subjective Cognitive Impairment and Are Negatively Correlated with ß Amyloid-42.

The role of tumor necrosis factor-α (TNF-α) in Alzheimer's disease (AD) has recently become a topic of debate. TNF-α levels increase in the blood of patients with AD, and amyloid beta (Aß) plaques contain TNF-α deposits. The therapeutic efficacy of blocking TNF-α in patients with AD remains controversial as it is mostly based on preclinical studies. Thus, whether and how TNF-α contributes to amyloidogenic processes in AD is still an open question to be addressed. We analyzed plasma TNF-α and Aß42 levels in patients with subjective cognitive impairment (SCI), mild cognitive impairment (MCI), and AD, and in healthy volunteers (HLT). In addition, we performed correlation analysis to evaluate whether changes in plasma TNF-α levels correlate with cognitive decline, Aß42 levels, age, and BMI, which are all factors considered to contribute to or predispose individuals to AD. We found that TNF-α and Aß42 plasma levels were higher in patients with AD than in HLT individuals. High TNF-α levels were also observed in patients with SCI, in whom TNF-α and Aß42 levels were negatively correlated. Notably, TNF-α did not affect the amyloidogenic pathway in human microglial cultures exposed to 48 h of incubation, although it did trigger neuroinflammatory processes. These results imply that high TNF-α levels are more likely to be a clinical condition linked to AD than are direct contributors. Nonetheless, elevated levels of TNF-α in early-stage patients, like those with SCI and MCI, may provide a distinguishing feature for identifying clinical profiles that are at risk of having a poorer outcome in AD and could benefit from tailored therapies.

Correction: Iannuzzi et al. Might Fibroblasts from Patients with Alzheimer's Disease Reflect the Brain Pathology? A Focus on the Increased Phosphorylation of Amyloid Precursor Protein Tyr682 Residue. Brain Sci. 2021, 11, 103.

In the original article [...].

Advances in peripheral blood biomarkers of patients with Alzheimer's disease: Moving closer to personalized therapies.

Recently, measurable peripheral biomarkers in the plasma of patients with Alzheimer's disease (AD) have gained considerable clinical interest. Several studies have identified one or more blood signatures that may facilitate the development of novel diagnostic and therapeutic strategies. For instance, changes in peripheral amyloid ß42 (Aß42) levels have been largely investigated in patients with AD and correlated with the progression of the pathology, although with controversial results. In addition, tumor necrosis factor α (TNFα) has been identified as an inflammatory biomarker strongly associated with AD, and several studies have consistently suggested the pharmacological targeting of TNFα to reduce systemic inflammation and prevent neurotoxicity in AD. Moreover, alterations in plasma metabolite levels appear to predict the progression of systemic processes relevant to brain functions. In this study, we analyzed the changes in the levels of Aß42, TNFα, and plasma metabolites in subjects with AD and compared the results with those in healthy elderly (HE) subjects. Differences in plasma metabolites of patients with AD were analyzed with respect to Aß42, TNFα, and the Mini-Mental State Examination (MMSE) score, searching for plasma signatures that changed simultaneously. In addition, the phosphorylation levels of the Tyr682 residue of the amyloid precursor protein (APP), which we previously proposed as a biomarker of AD, were measured in five HE and five AD patients, in whom the levels of Aß42, TNFα, and two plasma lipid metabolites increased simultaneously. Overall, this study highlights the potential of combining different plasma signatures to define specific clinical phenotypes of patient subgroups, thus paving the way for the stratification of patients with AD and development of personalized approaches.

Semaphorin 3A influences neuronal processes that are altered in patients with autism spectrum disorder: Potential diagnostic and therapeutic implications.

Autism spectrum disorder (ASD) is a pervasive disorder that most frequently manifests in early childhood and lasts for their entire lifespan. Several behavioural traits characterise the phenotype of patients with ASD, including difficulties in reciprocal social communication as well as compulsive/repetitive stereotyped verbal and non-verbal behaviours. Although multiple hypotheses have been proposed to explain the aetiology of ASD and many resources have been used to improve our understanding of ASD, several aspects remain largely unexplored. Class 3 semaphorins (SEMA3) are secreted proteins involved in the organisation of structural and functional connectivity in the brain that regulate synaptic and dendritic development. Alterations in brain connectivity and aberrant neuronal development have been described in some patients with ASD. Mutations and polymorphisms in SEMA3A and alterations in its receptors and signalling have been associated with some neurological disorders such as schizophrenia and epilepsy, which are comorbidities in ASD, but also with ASD itself. In addition, SEMA3A is a key regulator of the immune response and neuroinflammatory processes, which have been found to be dysregulated in mothers of children who develop ASD and in affected patients. In this review, we highlight neurodevelopmental-related processes in which SEMA3A is involved, which are altered in ASD, and provide a viewpoint emphasising the development of strategies targeting changes in the SEMA3A signal to identify patterns of anomalies distinctive of ASD or to predict the prognosis of affected patients.

A Targeted Mass Spectrometry Approach to Identify Peripheral Changes in Metabolic Pathways of Patients with Alzheimer's Disease.

Alzheimer's disease (AD), a neurodegenerative disorder, is the most common cause of dementia in the elderly population. Since its original description, there has been intense debate regarding the factors that trigger its pathology. It is becoming apparent that AD is more than a brain disease and harms the whole-body metabolism. We analyzed 630 polar and apolar metabolites in the blood of 20 patients with AD and 20 healthy individuals, to determine whether the composition of plasma metabolites could offer additional indicators to evaluate any alterations in the metabolic pathways related to the illness. Multivariate statistical analysis showed that there were at least 25 significantly dysregulated metabolites in patients with AD compared with the controls. Two membrane lipid components, glycerophospholipids and ceramide, were upregulated, whereas glutamic acid, other phospholipids, and sphingolipids were downregulated. The data were analyzed using metabolite set enrichment analysis and pathway analysis using the KEGG library. The results showed that at least five pathways involved in the metabolism of polar compounds were dysregulated in patients with AD. Conversely, the lipid pathways did not show significant alterations. These results support the possibility of using metabolome analysis to understand alterations in the metabolic pathways related to AD pathophysiology.

The paradigm of amyloid precursor protein in amyotrophic lateral sclerosis: The potential role of the 682YENPTY687 motif.

Neurodegenerative diseases are characterized by the progressive decline of neuronal function in several brain areas, and are always associated with cognitive, psychiatric, or motor deficits due to the atrophy of certain neuronal populations. Most neurodegenerative diseases share common pathological mechanisms, such as neurotoxic protein misfolding, oxidative stress, and impairment of autophagy machinery. Amyotrophic lateral sclerosis (ALS) is one of the most common adult-onset motor neuron disorders worldwide. It is clinically characterized by the selective and progressive loss of motor neurons in the motor cortex, brain stem, and spinal cord, ultimately leading to muscle atrophy and rapidly progressive paralysis. Multiple recent studies have indicated that the amyloid precursor protein (APP) and its proteolytic fragments are not only drivers of Alzheimer's disease (AD) but also one of the earliest signatures in ALS, preceding or anticipating neuromuscular junction instability and denervation. Indeed, altered levels of APP peptides have been found in the brain, muscles, skin, and cerebrospinal fluid of ALS patients. In this short review, we discuss the nature and extent of research evidence on the role of APP peptides in ALS, focusing on the intracellular C-terminal peptide and its regulatory motif 682YENPTY687, with the overall aim of providing new frameworks and perspectives for intervention and identifying key questions for future investigations.

An increase in Semaphorin 3A biases the axonal direction and induces an aberrant dendritic arborization in an in vitro model of human neural progenitor differentiation.

BACKGROUND: Semaphorins (Sema) belong to a large family of repellent guidance cues instrumental in guiding axons during development. In particular, Class 3 Sema (Sema 3) is among the best characterized Sema family members and the only produced as secreted proteins in mammals, thereby exerting both autocrine and paracrine functions. Intriguingly, an increasing number of studies supports the crucial role of the Sema 3A in hippocampal and cortical neurodevelopment. This means that alterations in Sema 3A signaling might compromise hippocampal and cortical circuits and predispose to disorders such as autism and schizophrenia. Consistently, increased Sema 3A levels have been detected in brain of patients with schizophrenia and many polymorphisms in Sema 3A or in the Sema 3A receptors, Neuropilins (Npn 1 and 2) and Plexin As (Plxn As), have been associated to autism. RESULTS: Here we present data indicating that when overexpressed, Sema 3A causes human neural progenitors (NP) axonal retraction and an aberrant dendritic arborization. Similarly, Sema 3A, when overexpressed in human microglia, triggers proinflammatory processes that are highly detrimental to themselves as well as NP. Indeed, NP incubated in microglia overexpressing Sema 3A media retract axons within an hour and then start suffering and finally die. Sema 3A mediated retraction appears to be related to its binding to Npn 1 and Plxn A2 receptors, thus activating the downstream Fyn tyrosine kinase pathway that promotes the threonine-serine kinase cyclin-dependent kinase 5, CDK5, phosphorylation at the Tyr15 residue and the CDK5 processing to generate the active fragment p35. CONCLUSIONS: All together this study identifies Sema 3A as a critical regulator of human NP differentiation. This may imply that an insult due to Sema 3A overexpression during the early phases of neuronal development might compromise neuronal organization and connectivity and make neurons perhaps more vulnerable to other insults across their lifespan.

Tandem Mass Spectrometry as Strategy for the Selective Identification and Quantification of the Amyloid Precursor Protein Tyr682 Residue Phosphorylation Status in Human Blood Mononuclear Cells.

Background: Alzheimer's disease (AD) is a devastating neurodegenerative disease without guidelines for early diagnosis or personalized treatment. Previous studies have highlighted a crucial role of increasing phosphorylation levels of the amyloid precursor protein (APP) Tyr682 residue in predicting neuronal deficits in AD patients. However, the lack of a method for the identification and quantification of Tyr682 phosphorylation levels prevents its potential clinical applications. Methods: Here we report a method to identify and quantify APP Tyr682 phosphorylation levels in blood mononuclear cells of AD patients by tandem mass spectrometry (tMS). Results: This method showed excellent sensitivity with detection and quantification limits set respectively at 0.035 and 0.082 ng injected for the phosphorylated peptide and at 0.02 and 0.215 ng injected for the non-phosphorylated peptide. The average levels of both peptides were quantified in transfected HELA cells (2.48 and 3.53 ng/µg of protein, respectively). Preliminary data on 3 AD patients showed quantifiable levels of phosphorylated peptide (0.10-0.15 ng/µg of protein) and below the LOQ level of non-phosphorylated peptide (0.13 ng/µg of protein). Conclusion: This method could allow the identification of patients with increased APP Tyr682 phosphorylation and allow early characterization of molecular changes prior to the appearance of clinical signs.

Challenges in LC-MS-based metabolomics for Alzheimer's disease early detection: targeted approaches versus untargeted approaches.

BACKGROUND: Alzheimer's disease (AD) is one of the most common causes of dementia in old people. Neuronal deficits such as loss of memory, language and problem-solving are severely compromised in affected patients. The molecular features of AD are Aß deposits in plaques or in oligomeric structures and neurofibrillary tau tangles in brain. However, the challenge is that Aß is only one piece of the puzzle, and recent findings continue to support the hypothesis that their presence is not sufficient to predict decline along the AD outcome. In this regard, metabolomic-based techniques are acquiring a growing interest for either the early diagnosis of diseases or the therapy monitoring. Mass spectrometry is one the most common analytical platforms used for detection, quantification, and characterization of metabolic biomarkers. In the past years, both targeted and untargeted strategies have been applied to identify possible interesting compounds. AIM OF REVIEW: The overall goal of this review is to guide the reader through the most recent studies in which LC-MS-based metabolomics has been proposed as a powerful tool for the identification of new diagnostic biomarkers in AD. To this aim, herein studies spanning the period 2009-2020 have been reported. Advantages and disadvantages of targeted vs untargeted metabolomic approaches have been outlined and critically discussed.

Metabolic Syndrome and Autophagy: Focus on HMGB1 Protein.

Metabolic syndrome (MetS) affects the population worldwide and results from several factors such as genetic background, environment and lifestyle. In recent years, an interplay among autophagy, metabolism, and metabolic disorders has become apparent. Defects in the autophagy machinery are associated with the dysfunction of many tissues/organs regulating metabolism. Metabolic hormones and nutrients regulate, in turn, the autophagy mechanism. Autophagy is a housekeeping stress-induced degradation process that ensures cellular homeostasis. High mobility group box 1 (HMGB1) is a highly conserved nuclear protein with a nuclear and extracellular role that functions as an extracellular signaling molecule under specific conditions. Several studies have shown that HMGB1 is a critical regulator of autophagy. This mini-review focuses on the involvement of HMGB1 protein in the interplay between autophagy and MetS, emphasizing its potential role as a promising biomarker candidate for the early stage of MetS or disease's therapeutic target.

Might Fibroblasts from Patients with Alzheimer's Disease Reflect the Brain Pathology? A Focus on the Increased Phosphorylation of Amyloid Precursor Protein Tyr682 Residue.

Alzheimer's disease (AD) is a devastating neurodegenerative disorder with no cure and no effective diagnostic criteria. The greatest challenge in effectively treating AD is identifying biomarkers specific for each patient when neurodegenerative processes have not yet begun, an outcome that would allow the design of a personalised therapeutic approach for each patient and the monitoring of the therapeutic response during the treatment. We found that the excessive phosphorylation of the amyloid precursor protein (APP) Tyr682 residue on the APP 682YENPTY687 motif precedes amyloid ß accumulation and leads to neuronal degeneration in AD neurons. We proved that Fyn tyrosine kinase elicits APP phosphorylation on Tyr682 residue, and we reported increased levels of APP Tyr682 and Fyn overactivation in AD neurons. Here, we want to contemplate the possibility of using fibroblasts as tools to assess APP Tyr682 phosphorylation in AD patients, thus making the changes in APP Tyr682 phosphorylation levels a potential diagnostic strategy to detect early pathological alterations present in the peripheral cells of AD patients' AD brains.

Putting Cells in Motion: Advantages of Endogenous Boosting of BDNF Production.

Motor exercise, such as sport or musical activities, helps with a plethora of diseases by modulating brain functions in neocortical and subcortical regions, resulting in behavioural changes related to mood regulation, well-being, memory, and even cognitive preservation in aging and neurodegenerative diseases. Although evidence is accumulating on the systemic neural mechanisms mediating these brain effects, the specific mechanisms by which exercise acts upon the cellular level are still under investigation. This is particularly the case for music training, a much less studied instance of motor exercise than sport. With regards to sport, consistent neurobiological research has focused on the brain-derived neurotrophic factor (BDNF), an essential player in the central nervous system. BDNF stimulates the growth and differentiation of neurons and synapses. It thrives in the hippocampus, the cortex, and the basal forebrain, which are the areas vital for memory, learning, and higher cognitive functions. Animal models and neurocognitive experiments on human athletes converge in demonstrating that physical exercise reliably boosts BDNF levels. In this review, we highlight comparable early findings obtained with animal models and elderly humans exposed to musical stimulation, showing how perceptual exposure to music might affect BDNF release, similar to what has been observed for sport. We subsequently propose a novel hypothesis that relates the neuroplastic changes in the human brains after musical training to genetically- and exercise-driven BDNF levels.

The Emerging Therapeutic Potential of Nitro Fatty Acids and Other Michael Acceptor-Containing Drugs for the Treatment of Inflammation and Cancer.

Nitro fatty acids (NFAs) are endogenously generated lipid mediators deriving from reactions of unsaturated electrophilic fatty acids with reactive nitrogen species. Furthermore, Mediterranean diets can be a source of NFA. These highly electrophilic fatty acids can undergo Michael addition reaction with cysteine residues, leading to post-translational modifications (PTM) of selected regulatory proteins. Such modifications are capable of changing target protein function during cell signaling or in biosynthetic pathways. NFA target proteins include the peroxisome proliferator-activated receptor γ (PPAR-γ), the pro-inflammatory and tumorigenic nuclear factor-κB (NF-κB) signaling pathway, the pro-inflammatory 5-lipoxygenases (5-LO) biosynthesis pathway as well as soluble epoxide hydrolase (sEH), which is essentially involved in the regulation of vascular tone. In several animal models of inflammation and cancer, the therapeutic efficacy of well-tolerated NFA has been demonstrated. This has already led to clinical phase II studies investigating possible therapeutic effects of NFA in subjects with pulmonary arterial hypertension. Albeit Michael acceptors feature a broad spectrum of bioactivity, they have for a rather long time been avoided as drug candidates owing to their presumed unselective reactivity and toxicity. However, targeted covalent modification of regulatory proteins by Michael acceptors became recognized as a promising approach to drug discovery with the recent FDA approvals of the cancer therapeutics, afatanib (2013), ibrutinib (2013), and osimertinib (2015). Furthermore, the Michael acceptor, neratinib, a dual inhibitor of the human epidermal growth factor receptor 2 and epidermal growth factor receptor, was recently approved by the FDA (2017) and by the EMA (2018) for the treatment of breast cancer. Finally, a number of further Michael acceptor drug candidates are currently under clinical investigation for pharmacotherapy of inflammation and cancer. In this review, we focus on the pharmacology of NFA and other Michael acceptor drugs, summarizing their potential as an emerging class of future antiphlogistics and adjuvant in tumor therapeutics.

Fyn Tyrosine Kinase Elicits Amyloid Precursor Protein Tyr682 Phosphorylation in Neurons from Alzheimer's Disease Patients.

Alzheimer's disease (AD) is an incurable neurodegenerative disorder with a few early detection strategies. We previously proposed the amyloid precursor protein (APP) tyrosine 682 (Tyr682) residue as a valuable target for the development of new innovative pharmacologic or diagnostic interventions in AD. Indeed, when APP is phosphorylated at Tyr682, it is forced into acidic neuronal compartments where it is processed to generate neurotoxic amyloid ß peptides. Of interest, Fyn tyrosine kinase (TK) interaction with APP Tyr682 residue increases in AD neurons. Here we proved that when Fyn TK was overexpressed it elicited APP Tyr682 phosphorylation in neurons from healthy donors and promoted the amyloidogenic APP processing with Aß peptides accumulation and neuronal death. Phosphorylation of APP at Tyr (pAPP-Tyr) increased in neurons of AD patients and AD neurons that exhibited high pAPP-Tyr also had higher Fyn TK activity. Fyn TK inhibition abolished the pAPP-Tyr and reduced Aß42 secretion in AD neurons. In addition, the multidomain adaptor protein Fe65 controlled the Fyn-mediated pAPP-Tyr, warranting the possibility of targeting the Fe65-APP-Fyn pathway to develop innovative strategies in AD. Altogether, these results strongly emphasize the relevance of focusing on pAPP Tyr682 either for diagnostic purposes, as an early biomarker of the disease, or for pharmacological targeting, using Fyn TKI.

Fyn Tyrosine Kinase as Harmonizing Factor in Neuronal Functions and Dysfunctions.

Fyn is a non-receptor or cytoplasmatic tyrosine kinase (TK) belonging to the Src family kinases (SFKs) involved in multiple transduction pathways in the central nervous system (CNS) including synaptic transmission, myelination, axon guidance, and oligodendrocyte formation. Almost one hundred years after the original description of Fyn, this protein continues to attract extreme interest because of its multiplicity of actions in the molecular signaling pathways underlying neurodevelopmental as well as neuropathologic events. This review highlights and summarizes the most relevant recent findings pertinent to the role that Fyn exerts in the brain, emphasizing aspects related to neurodevelopment and synaptic plasticity. Fyn is a common factor in healthy and diseased brains that targets different proteins and shapes different transduction signals according to the neurological conditions. We will primarily focus on Fyn-mediated signaling pathways involved in neuronal differentiation and plasticity that have been subjected to considerable attention lately, opening the fascinating scenario to target Fyn TK for the development of potential therapeutic interventions for the treatment of CNS injuries and certain neurodegenerative disorders like Alzheimer's disease.

The Y682ENPTY687 motif of APP: Progress and insights toward a targeted therapy for Alzheimer's disease patients.

Alzheimer's disease (AD) is a devastating neurodegenerative disorder for which no curative treatments, disease modifying strategies or effective symptomatic therapies exist. Current pharmacologic treatments for AD can only decelerate the progression of the disease for a short time, often at the cost of severe side effects. Therefore, there is an urgent need for biomarkers able to diagnose AD at its earliest stages, to conclusively track disease progression, and to accelerate the clinical development of innovative therapies. Scientific research and economic efforts for the development of pharmacotherapies have recently homed in on the hypothesis that neurotoxic ß-amyloid (Aß) peptides in their oligomeric or fibrillary forms are primarily responsible for the cognitive impairment and neuronal death seen in AD. As such, modern pharmacologic approaches are largely based on reducing production by inhibiting ß and γ secretase cleavage of the amyloid precursor protein (APP) or on dissolving existing cerebral Aß plaques or to favor Aß clearance from the brain. The following short review aims to persuade the reader of the idea that APP plays a much larger role in AD pathogenesis. APP plays a greater role in AD pathogenesis than its role as the precursor for Aß peptides: both the abnormal cleavage of APP leading to Aß peptide accumulation and the disruption of APP physiological functions contribute to AD pathogenesis. We summarize our recent results on the role played by the C-terminal APP motif -the Y682ENPTY68 motif- in APP function and dysfunction, and we provide insights into targeting the Tyr682 residue of APP as putative novel strategy in AD.

Anti-inflammatory nitro-fatty acids suppress tumor growth by triggering mitochondrial dysfunction and activation of the intrinsic apoptotic pathway in colorectal cancer cells.

Nitro-fatty acids (NFAs) are endogenously occurring lipid mediators exerting strong anti-inflammatory effects and acting as anti-oxidants in a number of animal models of inflammation. These NFA effects are mediated by targeting important regulatory proteins involved in inflammatory processes, such as 5-lipoxygenase, soluble epoxide hydrolase, or NF-κB. In the present study, we investigated the anti-tumorigenic effects of NFAs on colorectal cancer (CRC) cells in cell culture-based experiments and in a murine xenograft model of human CRC. We could show that 9-NOA suppresses the viability of CRC cells (HCT-116 and HT-29) by inducing a caspase-dependent apoptosis via the intrinsic apoptotic pathway. Co-treatment with the pan-caspase inhibitor Q-VD-OPH counteracted the NFA-mediated apoptosis in both cell lines. Furthermore, NFAs affected the cell cycle transition and reduced the oxygen consumption rate (OCR) immediately. On the contrary to their well-known anti-oxidative properties, NFAs mediated the generation of mitochondrial oxidative stress in human CRC cells. Additionally, similar to the cytostatic drug mitomycin, 9-NOA significantly reduced tumor growth in a murine xenograft model of human colorectal cancer. In contrast to the established cytostatic drug, 9-NOA treatment was well tolerated by mice. This study delivers a novel mechanistic approach for nitro-fatty acid-induced inhibition of CRC cell growth by targeting mitochondrial functions such as the mitochondrial membrane potential and mitochondrial respiration. We suggest these naturally occurring lipid mediators as a new class of well tolerated chemotherapeutic drug candidates for treatment of CRC or potentially other inflammation-driven cancer types.

Drug-Mediated Intracellular Donation of Nitric Oxide Potently Inhibits 5-Lipoxygenase: A Possible Key to Future Antileukotriene Therapy.

AIMS: 5-Lipoxygenase (5-LO) is the key enzyme of leukotriene (LT) biosynthesis and is critically involved in a number of inflammatory diseases such as arthritis, gout, bronchial asthma, atherosclerosis, and cancer. Because 5-LO contains critical nucleophilic amino acids, which are sensitive to electrophilic modifications, we determined the consequences of a drug-mediated intracellular release of nitric oxide (NO) on 5-LO product formation by human granulocytes and on 5-LO-dependent pulmonary inflammation in vivo. RESULTS: Clinically relevant concentrations of NO-releasing nonsteroidal anti-inflammatory drugs and other agents releasing NO intracellularly suppress 5-LO product synthesis in isolated human granulocytes via direct S-nitrosylation of 5-LO at the catalytically important cysteines 416 and 418. Furthermore, suppression of 5-LO product formation was observed in ionophore-stimulated human whole blood and in an animal model of pulmonary inflammation. INNOVATION: Here, we report for the first time that drugs releasing NO intracellularly are efficient 5-LO inhibitors in vitro and in vivo at least equivalent to approved 5-LO inhibitors. CONCLUSION: Our findings provide a novel mechanistic strategy for the development of a new class of drugs suppressing LT biosynthesis by site-directed nitrosylation. The results may also help to better understand the well-recognized anti-inflammatory clinically relevant actions of NO-releasing drugs. Furthermore, our study describes in detail a novel molecular mode of action of NO. Rebound Track: This work was rejected during standard peer review and rescued by Rebound Peer Review (Antioxid Redox Signal 16: 293-296, 2012) with the following serving as open reviewers: Angel Lanas, Hartmut Kühn, Joan Clària, Orina Belton. Antioxid. Redox Signal. 28, 1265-1285.