Fungal infections cause serious problems in many aspects of human life; especially infections by fungal species represent problems in immunocompromised patients. Current antifungal antibiotics target various metabolic pathways, predominantly the cell wall or cellular membrane. However, numerous compounds are available to combat fungal infections, their efficacy is far from being satisfactory and some of them display substantial toxicity. The emerging resistance represents a serious issue as well; thus, there is a considerable need for new anti-fungal compounds with lower toxicity and higher effectiveness. One of the unique antifungal antibiotics is sordarin, the only known compound that acts on the fungal translational machinery per se. It has been shown that sordarin inhibits protein synthesis at the elongation step of the translational cycle, acting on eukaryotic elongation-factor-2. In this review, we are aiming to deliver a robust scientific platform promoting the development of antifungal compounds, especially focusing on molecular action of sordarin.
Background and Purpose T-type calcium channels, mainly the Cav3.2 subtype, are important contributors to the nociceptive signaling pathway. We investigated their involvement in inflammation and related pain-like symptoms. Experimental Approach The involvement of Cav3.2 and T-type channels was investigated using genetic and pharmacological inhibition to assess mechanical allodynia/hyperalgesia and edema development in two murine inflammatory pain models. The location of Cav3.2 involved in pain-like symptoms was studied in mice with Cav3.2 knocked out in C-low threshold mechanoreceptors (C-LTMR) and the use of ABT-639, a peripherally restricted T-type channel inhibitor. The anti-edematous effect of Cav3.2 inhibition was investigated in chimeric mice with immune cells deleted for Cav3.2. Lymphocytes and macrophages from either green fluorescent protein-targeted Cav3.2 or KO mice were used to determine the expression of Cav3.2 protein and the functional status of the cells. Key Results We showed the role of Cav3.2 channels in the development of pain-like symptoms and edema in the two murine inflammatory pain models. For the first time, we provide evidence of the involvement of Cav3.2 channels located on C-LTMRs in inflammatory pain at both peripheral and primary afferent terminals at the spinal level. We showed that Cav3.2 channels located in T cells and macrophages contribute to the inflammatory process. Conclusion and Implications This work highlights the crucial role of Cav3.2 channels in inflammation and related pain and suggests that targeting Cav3.2 channels with pharmacological agents could be an attractive and readily evaluable strategy in a clinical trial to relieve chronic inflammatory pain in affected patients.
Lopinavir combined with ritonavir were reported to benefit the patients with SARS by reducing the viral loads. However, in the latest clinical trials, no benefit was observed with lopinavir-ritonavir treatment beyond standard care in patients with COVID-19. We comment here that this disappointed result of clinical trial might result from the low volume of the lung distribution of lopinavir. The major reasons were listed below: 1) The binding affinity of ACE2 with SARS-CoV-2 spike protein is ~10- to 20-fold higher than the binding affinity of ACE2 with SARS-CoV spike protein, indicating that SARS-CoV-2 can enter AT2 cells in lung much easier than SARS-CoV. Therefore, the viral loads of SARS-CoV-2 might be much higher than viral loads of SARS-CoV in the lung tissue. 2) The concentration of lopinavir in the lung tissue was 1.18 μg equiv/ml in rats. The low volume of the lung distribution of lopinavir might not be enough to inhibit the coronavirus replication due to the high viral loads in the lung tissue. 3) In contrast, the concentration of chloroquine in the lung tissue was much higher (30.76 ± 0.85 μg equiv/ml) in rats, which might lead to its clinical and virologic benefits in the treatment of COVID-19 patients. Together, we proposed here that anti-SARS-CoV-2 drug repurposing studies should pay more attentions to the lung tissue distribution of antiviral drugs. The efficacy of antiviral drugs might depend on their lung tissue distributions
Brain mineralocorticoid receptors (MR) mediate effects of aldosterone in relation to salt homeostasis, and of glucocorticoid stress hormones corticosteroids in the context of stress adaptation. Brain stem MRs respond to aldosterone, while forebrain MRs mediate rapid and delayed MR-mediated glucocorticoids effects in conjunction with the glucocorticoid receptor. MR-mediated effects depend on gender, genetic variations and environmental influences. Disturbed MR activity by chronic stress or in certain (endocrine) diseases can cause deleterious effects on affective state, cognitive and behavioural function in susceptible individuals. High MR activation may have protective effects in healthy individuals, whereas dysregulated high MR activity during a stress response would require treatment with mineralocorticoid receptor antagonists (MRAs). Here, we discuss recent pharmacological and genetic developments, from the molecular underpinnings of MR signaling and function, to pharmacological interventions in the clinic. Improved understanding of MR dependent pathways will help to improve glucocorticoid therapy, unwanted side effects and psychiatric symptoms.
The brain is the most cholesterol rich organ in the body containing about 25% of the body’s free cholesterol. Cholesterol cannot pass the blood brain barrier and be imported or exported directly, instead it is synthesised in situ and metabolised to oxysterols, oxidised forms of cholesterol, which can pass the blood brain barrier. 24S-Hydroxycholesterol is the dominant oxysterol in brain after parturition but during development a myriad of other oxysterols are produced which persist as minor oxysterols after birth. During both development and in later life, oxysterols and other sterols interact with a variety of different receptors, including nuclear receptors e.g. liver X receptors; membrane bound G protein-coupled receptors e.g. smoothened; the endoplasmic reticulum resident proteins e.g. INSIG (insulin induced gene), or the cholesterol sensing protein SCAP (SREBP cleavage activating protein); and the ligand-gated ion channel N-methyl-D-aspartate receptors found in nerve cells. In this review we summaries the different oxysterols (neuro-oxysterol) and sterols (neuro-sterols) found in the central nervous system whose biological activity is transmitted via these different classes of protein receptors.
Sodium glucose co-transporter 2 inhibitors (SGLT-2i’s) significantly improve cardiovascular outcome in both diabetic and non-diabetic patients. Preclinical studies suggest that SGLT-2i’s directly affect endothelial function in a glucose-independent manner. The effects of SGLT-2i’s include reduction of oxidative stress and inflammatory reaction in endothelial cells. Furthermore, SGLT2i’s have been shown to restore endothelial-related vasodilation and to regulate angiogenesis. The favorable cardiovascular effects of SGLT-2i’s might be mediated via multiple pathways: 1) by inhibition of the overactive sodium-hydrogen exchanger; 2) by reduction of nicotinamide adenine dinucleotide phosphate oxidases expression; 3) by alleviation of mitochondrial injury; 4) by the suppression of inflammatory-related signaling pathways (e.g. by affecting nuclear factor kappa beta); 5) by modulation of glycolysis, as well as 6) by restoring impaired nitric oxide bioavailability. This review focuses on the most recent progress and existing gaps in preclinical investigations concerning the direct effects of SGLT-2i’s on endothelial dysfunction and their underlying mechanisms.
Muscle protein catabolism in patients with diabetic nephropathy (DN) results in striking losses of muscle proteins, which increases morbidity and mortality risks. Emerging evidence shows that short-chain fatty acids (SCFAs) play an important role in the maintenance of health and disease development. Recently, the connection between butyrate (a SCFA) and DN has been revealed, although the relationship between butyrate and muscle atrophy is still not clear. In our study, we found a significant decrease in butyrate in DN using metabolomics analyses. The addition of butyrate remarkably intestinal barrier function. Concurrently, butyrate could alleviate muscle atrophy and promote PI3K/AKT/mTOR signals, and suppress oxidative stress and autophagy in the skeletal muscle of db/db mice as well as high glucose/lipopolysaccharide (HG/LPS)-induced C2C12 cells. To further explore the mechanism, we found that GPR43, the key SCFAs signaling molecule, was significantly decreased in the skeletal muscle of db/db mice and HG/LPS-induced C2C12 cells. Overexpression of GPR43 could activate PI3K/AKT/mTOR signals and inhibit oxidative stress and autophagy in HG/LPS-induced C2C12 cells. Silencing of GPR43 blocked PI3K/AKT/mTOR signals improved by butyrate, as well as suppression of oxidative stress and reduction of autophagy. Ultimately, butyrate alleviated muscle atrophy in DN via GPR43-mediated PI3K/AKT/mTOR pathway
Acute respiratory distress syndrome (ARDS) is the main cause of morbidity and mortality in Coronavirus disease 19 (Covid-19) for which as of now there is no effective treatment. ARDS is caused and sustained by an uncontrolled inflammatory activation characterized by a massive release of cytokines (cytokine storm), diffuse lung edema, inflammatory cell infiltraton and disseminated coagulation. Macrophage and T lymphocyte dysfunction plays a central role in this syndrome. In several experimental in vitro and in vivo models, many of these pathophysiological changes are triggered by stimulation of the P2X7 receptor. We hypothesize that this receptor might be an ideal candidate to target in Covid-19-associated ARDS.
Many Western countries have been affected by the outbreak of COVID-19. Italy has been particularly hit at the beginning of the pandemic, immediately after China. In Italy and elsewhere women seem to be less affected then men by severe/fatal COVID-19 infection, regardless of their age. Despite the evidence that women and men are different fort this infection, very few studies consider different therapeutic approaches for the two sexes. Undoubtedly, understanding the mechanisms at the bases of these differences may help to find appropriate and sex specific therapies. Here we consider that other mechanisms but estrogen protection are involved. Several X-linked genes (such as ACE2) and Y-linked genes (SRY, SOX9) may explain sex differences. Cardiovascular comorbidities are among the major enhancers of virus lethality. In addition, the number of sex-independent non-genetic factors that can change susceptibility and mortality is enormous, and many other factors are likely to be considered, including gender and cultural habits in different countries.
Skin drug delivery is an emerging route in the drug development, due to its great advantages, thus leading to an urgent need to understand the behaviour of active pharmaceutical ingredients into/through the skin. This knowledge is crucial in the early stages of product design and development. Yet, given the skin barrier properties as one of the first body’s natural defence systems, it can act as an obstacle to the successful outcome of a skin drug therapy. To unravel the mechanisms underlying this barrier, reductionist strategies have designed several models with different levels of complexity and integrity, using non-biological and biological components. Besides the detail of information and resemblance to the in vivo Human skin that each in vitro model offers, the technical and economic efforts required should be considered when selecting the most adequate model for the intended research. This review provides an outline of the most commonly applied skin models, including healthy and diseased conditions, lab developed systems and commercialized models, their advantages and limitations and, also an overview of the new trends in skin engineered models.
Background and Purpose: Despite availability of a variety of treatment options, many asthma patients have poorly controlled disease with frequent exacerbations. Proteinase-activated receptor-2 (PAR2) has been identified in pre-clinical animal models as important to asthma initiation and progression following allergen exposure. Proteinase activation of PAR2 induces intracellular Ca2+, mitogen activated protein kinase (MAPK) and -arrestin signaling the airway, leading to both inflammatory and protective effects. We have developed C391, a potent PAR2 antagonist effective in blocking peptidomimetic- and trypsin-induced PAR2 signaling in vitro as well as reducing inflammatory PAR2-associated pain in vivo. We hypothesized that PAR2 reduction with C391 would attenuate allergen-induced asthma indicators in murine models. Experimental Approach: We evaluated the ability for C391 to alter Alternaria alternata-induced PAR2 signaling pathways in vitro using a human airway epithelial cell line that naturally expresses PAR2 (16HBE14o-) and a transfected embryonic cell line (HEK 293). We next evaluated the ability for C391 to reduce A. alternata-induced asthma indicators in vivo in two murine strains. Key Results: C391 blocked A. alternata-induced, PAR2-dependent Ca2+ and MAPK signaling in 16HBE14o- cells, as well as -arrestin recruitment in HEK 293 cells. C391 effectively attenuated A. alternata-induced inflammation, mucus production, mucus cell hyperplasia and airway hyperresponsiveness in acute asthma murine models. Conclusions and Implications: To our knowledge, this is the first demonstration of pharmacological intervention of PAR2 to reduce allergen-induced asthma indicators in vivo. These data support further development of PAR2 antagonists as potential first-in-class allergic asthma drugs.
Background and Purpose: Many pain-triggering nociceptor neurons express TRPV1 or TRPA1, cation-selective channels with large pores that enable permeation of QX-314, a cationic analogue of lidocaine. Co-application of QX-314 with TRPV1 or TRPA1 activators can silence nociceptors. We now describe BW-031, a novel more potent cationic sodium channel inhibitor, test whether its application alone can inhibit the pain associated with tissue inflammation, and whether this strategy can also inhibit cough. Experimental Approach: We characterized BW-031 inhibition of sodium channels and tested BW-031 in three models of inflammatory pain: rat paw inflammation produced by Complete Freund’s Adjuvant injection or surgical incision and a mouse paw UV burn model. We also tested the ability of BW-031 to inhibit cough induced by inhalation of dilute citric acid in guinea pigs. Key Results: BW-031 inhibited Nav1.7 and Nav1.1 channels with ~6-fold greater potency than QX-314 when introduced inside cells and entered capsaicin-activated TRPV1 expressing sensory neurons. BW-031 inhibited inflammatory pain in all three models, producing more effective and longer-lasting inhibition of pain than QX-314 in the mouse UV burn model. BW-031 was also effective in reducing cough counts by 78-90% when applied intratracheally under isoflurane anesthesia or by aerosol inhalation in awake guinea pigs with airway inflammation produced by ovalbumin sensitization. Conclusion and Implications: BW-031 a novel cationic sodium channel inhibitor can be applied locally as a single agent to inhibit inflammatory pain and also effectively inhibits cough in a guinea pig model of nociceptor-activated cough, suggesting a new clinical approach to treating cough.
As human spaceflight continues with extended mission durations, the demand of effective and safe drugs is going to increase. To date, the medications used during missions (for space motion sickness, sleep disturbances, allergies, pain and sinus congestion) are administered under the assumption that they act similarly as on the Earth. During spaceflights however fluid shifts, muscle and bone loss, immune system dysregulation and changes in the gastrointestinal tract and metabolism are documented. These alterations may change the pharmacokinetics (PK) and pharmacodynamics. The information gained from bed-rest studies and from inflight observations is partial and demonstrates variability in drug PK. The objectives of this review are to report: i) the impact of the space environmental stressors on human physiology in relation to PK; ii) the state-of-the-art on experimental data in space and/or in ground-based models; iii) the validation of ground-based models for PK studies; and iv) the identification of possible research gaps.
The insulin receptor is a membrane protein responsible for regulation of nutrient balance and therefore an attractive target in the treatment of diabetes and metabolic syndrome. Pharmacology of the insulin receptor involves two distinct mechanisms, (1) activation of the receptor by insulin mimetics that bind in the extracellular domain and (2) inhibition of the receptor tyrosine kinase enzymatic activity in the cytoplasmic domain. While a complete structural picture of the full-length receptor comprising the entire sequence covering extracellular, transmembrane, juxtamembrane and cytoplasmic domains is still elusive, recent progress through cryoelectron microscopy has made it possible to describe the initial insulin ligand binding events at atomistic detail. We utilize this opportunity to obtain structural insights into the pharmacology of the insulin receptor. To this end, we conducted a comprehensive docking study of known ligands to the new structures of the receptor. Through this approach, we provide an in-depth, structure-based review of human insulin receptor pharmacology in light of the new structures.
Schizophrenia remains a sizable socioeconomic burden that continues to be treated with therapeutics based on 70-year old science. All currently approved therapeutics primarily target the dopamine D2 receptor to achieve their efficacy. Whilst dopaminergic dysregulation is a key feature in this disorder, the targeting of dopaminergic machinery has yielded limited efficacy and an appreciable side effect burden. Over the recent decades, numerous drugs that engage non-dopaminergic GPCRs have yielded a promise of efficacy without the deleterious side effect profile, yet none have successfully completed clinical studies and progressed to the market. More recently, there has been increased attention around non-dopaminergic GPCR-targeting drugs, notably KarXT, which demonstrated efficacy in some schizophrenia symptom domains. This provides renewed hope that effective schizophrenia may lay outside of the dopaminergic space. Despite the potential for muscarinic receptor- (and other well-characterised GPCR families) targeting drugs to treat schizophrenia, they are often plagued with complications such as lack of receptor subtype selectivity and peripheral on-target side-effects. Orphan GPCR studies have opened a new avenue of exploration with many demonstrating schizophrenia-relevant mechanisms and a favourable expression profile, thus offering potential for novel drug development. This review discusses centrally-expressed orphan G protein-coupled receptors: GPR3, GPR6, GPR12, GPR52, GPR85, GPR88 and GPR139 and their relationship to schizophrenia. We review their expression, signalling mechanisms and cellular function, in conjunction with small molecule development and structural insights. We seek to provide a snapshot of the growing evidence and development potential of new classes of schizophrenia therapeutics.
Background and Purpose The cyclic nucleotides cAMP and cGMP are ubiquitous second messengers participating in the regulation of several biological processes. Interference of cNMP signalling is linked to multiple diseases and thus is an important component of pharmaceutical research. The existing optogenetic toolbox in C. elegans is restricted to soluble adenylyl cyclases, the membrane-bound Blastocladiella CyclOp and hyperpolarizing rhodopsins, yet missing are membrane-bound photoactivatable adenylyl cyclases and hyperpolarizers on the basis of K+-currents. Experimental Approach For the characterization of the photoactivatable nucleotidyl cyclases, we expressed the proteins alone or in combination with cyclic-nucleotide gated channels in C. elegans muscle cells and cholinergic motor neurons. To investigate the extent of optogenetic cNMP production and the ability of the systems to de- or hyperpolarize the cells, we performed behavioural analyses (locomotion, muscle contraction) and measured the cNMP content in vitro. Key Results We implemented Catenaria CyclOp as a new tool for cGMP production, allowing fine-control of cGMP levels. As photoactivatable membrane-bound adenylyl cyclases, we established YFP::BeCyclOp(A-2x) and YFP::CaCyclOp(A-2x), enabling more specific optogenetic cAMP signalling compared to soluble ACs. For the hyperpolarization of excitable cells by K+-currents, we introduced the cAMP-gated K+-channel SthK from Spirochaeta thermophila with either bPAC or BeCyclOp(A-2x), and the Blastocladiella emersonii cGMP-gated K+-channel BeCNG1 with BeCyclOp. Conclusion and Implications We established a comprehensive suite of optogenetic tools for cNMP manipulation for the nematode, which will be useful for applications in many cell types, including sensory neurons which use mainly cGMP as second messenger, and for potent hyperpolarization using K+-ions.
Peptides play a key role in controlling many physiological and neurobiological pathways. Many bioactive peptides require a C-terminal α-amide for full activity. The bifunctional enzyme catalyzing α-amidation, peptidylglycine α-amidating monooxygenase (PAM), is the sole enzyme responsible for amidated peptide biosynthesis, from Chlamydomonas reinhardtii to Homo sapiens. Many neuronal and endocrine functions are dependent upon amidated peptides; additional amidated peptides are growth promoters in tumors. The amidation reaction occurs in two steps, glycine α-hydroxylation followed by dealkylation to generate the α-amide product. Currently, most potentially useful inhibitors target the first reaction, which is rate-limiting. PAM is a membrane-bound enzyme that visits the cell surface during peptide secretion. PAM is then used again in the biosynthetic pathway, meaning that cell-impermeable inhibitors or inactivators could have therapeutic value for the treatment of cancer or psychiatric abnormalities. To date, inhibitor design has not fully exploited the structures and mechanistic details of PAM.
Substance use disorder (SUD) is a chronic condition with maintained abuse of a substance leading to physiological and psychological alterations and often changes in cognitive and social behaviours. Current therapies mainly consist of psychotherapy coupled with medication; however, alarmingly high relapse rates reveal the shortcomings of the current standard of care. The signalling and expression profile, and neurological function of the serotonin 2C receptor (5-HT2C receptor) make it an ideal candidate of interest for the treatment of SUD. This is further corroborated by pre-clinical and clinical evidence of therapeutically relevant compounds acting at the 5-HT2C receptor. Notwithstanding, drug binding at closely related serotonin receptor subtypes has impeded drug development. More recently, psychedelics, which broadly act at 5-HT2 receptors, have indicated promising potential for the treatment of SUD, implicating in part, the 5-HT2C receptor. The modern psychedelic movement has rekindled therapeutic interest in the 5-HT2C receptor, resulting in an influx of new studies, especially structural analyses. This review delves into the structural, molecular and cellular mechanisms governing the 5-HT2C receptor function, in the context of SUD. This provides the basis of the preclinical and clinical evidence for their role in SUD and highlights the potential for future exploration.