Selective Antagonist Research Articles

Discovery of novel and selective farnesoid X receptor antagonists through structure-based virtual screening, preliminary structure-activity relationship study, and biological evaluation

Farnesoid X receptor (FXR) is a bile acids receptor and plays a crucial role in regulating bile acids, lipids, and glucose metabolism. Previous research suggests that inhibiting FXR activation can be beneficial in reducing cholesterol and low-density lipoprotein cholesterol (LDL-C) levels, offering potential treatment options for metabolic syndrome with lipid disorders. Herein, we report p-acetylaminobenzene sulfonate derivatives as a novel scaffold of FXR antagonists by multistage screening. Among these derivatives, compound F44-A13 exhibited a half-maximal inhibitory concentration of 1.1 μM. Furthermore, compound F44-A13 demonstrated effective inhibition of FXR activation in cellular assays and exhibited high selectivity over eleven other nuclear receptors. Besides, compound F44-A13 significantly suppressed the regulation of FXR target genes Shp, Besp, and Cyp7a1, while reducing cholesterol levels in human hepatoma HepG2 cells. Pharmacological studies conducted on C57BL/6 mice further confirmed that compound F44-A13 had beneficial effects in reducing cholesterol, triglycerides, and LDL-C levels. These findings highlight that F44-A13 is a highly selective FXR antagonist that might serve as a useful molecule for further FXR studies as well as the development of FXR antagonists for the potential treatment of metabolic diseases with lipid disorders.

Revolutionising CKD Care: Novel Treatment Paradigms and Cutting-edge Strategies

Chronic Kidney Disease (CKD) represents a public health crisis globally. New therapeutic horizons in CKD management encompass a diverse array of options for a more holistic and personalised approach. Sodium-glucose cotransporter-2 (SGLT-2) inhibitors exhibit promising results in preserving kidney function and mitigating cardiovascular risks in patients with or without diabetes. Non-steroidal selective Mineralocorticoid Receptor Antagonists (MRA) present targeted strategies for CKD management. Glucagon-like peptide-1 (GLP-1) receptor agonists offer innovative approaches for glycaemic control and complication reduction. Endothelin Receptor Antagonists contribute to CKD care by modulating endothelial function and reducing kidney damage. Aldosterone synthase inhibitors offer a unique angle, targeting aldosterone production to manage CKD-related complications. Additionally, hypoxia-inducible factor (HIF) inhibitors represent a cutting-edge therapeutic class, addressing the hypoxic conditions associated with kidney damage. This diverse range of advancements underscores a transformative shift towards more personalised and effective CKD treatments, providing newfound optimism for improved patient outcomes. Beyond providing evidence-based insights into recent therapeutic advancements in CKD, this review also delineates contemporary treatment guidelines for CKD. It underscores the imperative for holistic strategies to enhance outcomes for individuals grappling with this complex condition.

Current Understanding of Compulsive Sexual Behavior Disorder and Co-occurring Conditions: What Clinicians Should Know about Pharmacological Options.

Compulsive sexual behavior disorder (CSBD) has recently been recognized as a psychiatric disorder. Pharmacological treatments for CSBD have received little study and thus have limited empirical support. The main objective of the present work is to review existing literature on the efficacy of different drugs on the symptomatology of CSBD, including the subtype of problematic pornography use (PPU). The main pharmacological approaches to treating CSBD have included opioid antagonists (naltrexone and nalmefene), selective serotonin reuptake inhibitors (paroxetine, citalopram, fluoxetine, and sertraline), mood stabilizers (topiramate), tricyclic antidepressants (clomipramine), serotonin antagonist and reuptake inhibitors (nefazodone), and N-acetylcysteine. Since people with CSBD may experience different co-occurring disorders, these should be considered when choosing the best pharmacological treatment. Pharmacological therapy for CSBD/PPU has been suggested as an adjunct to psychological therapies, which, for the moment, have the most empirical evidence. However, to evaluate the efficacy of most of the drugs presented in this narrative review, data to date have only been available from case studies. Thus, empirical support is scant and generalizability of results is limited, highlighting the need for more research in this area.

Subtle Structural Changes across the Boundary between A2AR/A2BR Dual Antagonism and A2BR Antagonism: A Novel Class of 2-Aminopyrimidine-Based Derivatives.

Aberrantly elevated adenosine in the tumor microenvironment exerts its immunosuppressive functions through adenosine receptors A2AR and A2BR. Antagonism of A2AR and A2BR has the potential to suppress tumor growth. Herein, we report a systemic assessment of the effects of an indole modification at position 4, 5, 6, or 7 on both A2AR/A2BR activity and selectivity of novel 2-aminopyrimidine compounds. Substituting indole at the 4-/5-position produced potent A2AR/A2BR dual antagonism, whereas the 6-position of indole substitution gave highly selective A2BR antagonism. Molecular dynamics simulation showed that the 5-cyano compound 7ai had a lower binding free energy than the 6-cyano compound 7aj due to water-bridged hydrogen bond interactions with E169 or F168 in A2AR. Of note, dual A2AR/A2BR antagonism by compound 7ai can profoundly promote the activation and cytotoxic function of T cells. This work provided a strategy for obtaining novel dual A2AR/A2BR or A2BR antagonists by fine-tuning structural modification.

Loxapine inhibits replication of hepatitis A virus in vitro and in vivo by targeting viral protein 2C.

No antiviral drugs currently are available for treatment of infection by hepatitis A virus (HAV), a causative agent of acute hepatitis, a potentially life-threatening disease. Chemical screening of a small-compound library using nanoluciferase-expressing HAV identified loxapine succinate, a selective dopamine receptor D2 antagonist, as a potent inhibitor of HAV propagation in vitro. Loxapine succinate did not inhibit viral entry nor internal ribosome entry site (IRES)-dependent translation, but exhibited strong inhibition of viral RNA replication. Blind passage of HAV in the presence of loxapine succinate resulted in the accumulation of viruses containing mutations in the 2C-encoding region, which contributed to resistance to loxapine succinate. Analysis of molecular dynamics simulations of the interaction between 2C and loxapine suggested that loxapine binds to the N-terminal region of 2C, and that resistant mutations impede these interactions. We further demonstrated that administration of loxapine succinate to HAV-infected Ifnar1-/- mice (which lack the type I interferon receptor) results in decreases in the levels of fecal HAV RNA and of intrahepatic HAV RNA at an early stage of infection. These findings suggest that HAV protein 2C is a potential target for antivirals, and provide novel insights into the development of drugs for the treatment of hepatitis A.

Uric acid–driven NLRP3 inflammasome activation triggers lens epithelial cell senescence and cataract formation

Excessive uric acid (UA) is associated with age-related cataract. A previous study showed that a high UA level in the aqueous humor stimulated the senescence of lens epithelial cells (LECs), leading to cataract progression. To better understand the underlying mechanisms, we investigated UA-driven senescence in human lens tissue samples obtained during surgery, rat lens organ cultures, and in vivo experiments, using senescence-associated β-galactosidase (SA-β-gal) staining, electronic microscopy, Western blotting, and histological analyses. Initially, we identified markedly higher expressions of NLRP3 and caspase-1 in the lens capsules of hyper-uricemic patients compared to normo-uricemic patients. This increase was accompanied by a significant rise in the SA-β-gal positive rate. We next built a cataract model in which rat lenses in an organ culture system were treated with an increasing dosage of UA. Notably, opacification was apparent in the lenses treated with 800 μM of UA starting on the fifth day. Mechanistically, UA treatment not only significantly induced the expression of NLRP3, caspase-1, and IL-1β, but also upregulated the levels of SA-β-gal and the senescence regulators p53 and p21. These effects were fully reversed, and lens opacification was ameliorated by the addition of MCC950, a selective NLRP3 antagonist. Moreover, an in vivo model showed that intravitreal UA injection rapidly induced cataract phenotypes within 21 days, an effect significantly mitigated by co-injection with MCC950. Together, our findings suggest that targeting the UA-induced NLRP3 inflammasome with MCC950 could be a promising strategy for preventing cataract formation associated with inflammageing.

Adenosine A2B receptor agonist improves epidermal barrier integrity in a murine model of epidermal hyperplasia

Adenosine regulates multiple physiological processes through the activation of four receptor subtypes, of which the A2B adenosine receptor (A2BAR) has the lowest affinity for adenosine. Being the adenosine receptor subtype most prominently expressed in epidermis, we recently described the antiproliferative and anti-inflammatory effect of the selective A2BAR agonist BAY60–6583 (BAY) in human keratinocytes stimulated with 12-O-tetradecanoylphorbol-13-acetate (TPA), so we sought to establish the effect of topical application of BAY in a model of murine epidermal hyperplasia.Topical application of BAY (1 or 10 μg/site) prevented the inflammatory reaction and skin lesions induced by TPA, minimizing hyperproliferation and acanthosis, as well as the expression of specific markers of proliferative keratinocytes. On the other hand, pre-treatment with the selective A2BAR antagonist, PSB-1115 (PSB, 5 or 50 μg/site) reversed these beneficial effects. Additionally, BAY application normalized the expression of epidermal barrier proteins, whose integrity is altered in inflammatory skin diseases, while treatment with the antagonist alone worsened it.Our results, besides confirming the anti-inflammatory and antiproliferative effects of the A2BAR agonist, further demonstrate a role of A2BAR activation to preserve the epidermal barrier. Therefore, the activation of A2BAR may constitute a possible new pharmacological target for the treatment of skin inflammatory diseases such as psoriasis.

A REVIEW ON RECENT ADVANCES IN TRANSDERMAL DRUG DELIVERY SYSTEMS OF TAMSULOSIN

Tamsulosin is a highly selective α1-adrenoreceptor antagonist. It has been developed to treat signs and symptoms of benign prostatic hyperplasia. Tamsulosin is absorbed quickly and completely in intestinal mucosa and is eliminated gradually after oral administration, which might generate some side effects as postural hypotension in number of patients. Transdermal drug delivery systems were developed for prolonged tamsulosin delivery in order to control its bioavailability and minimize its side effects. Hence, the present review aims to discuss thoroughly the various transdermal drug delivery systems of tamsulosin investigated in recent years. This review also discusses the skin as a route of drug administration, technologies in transdermal drug delivery along with different techniques used in the preparation of transdermal delivery systems of tamsulosin and their effects on its release and permeation.

Evolutionarily conserved mechanisms regulating stress-induced neutrophil redistribution in fish.

Stress may pose a serious challenge to immune homeostasis. Stress however also may prepare the immune system for challenges such as wounding or infection, which are likely to happen during a fight or flight stress response. In common carp (Cyprinus carpio L.) we studied the stress-induced redistribution of neutrophils into circulation, and the expression of genes encoding CXC chemokines known to be involved in the regulation of neutrophil retention (CXCL12) and redistribution (CXCL8), and their receptors (CXCR4 and CXCR1-2, respectively) in blood leukocytes and in the fish hematopoietic organ - the head kidney. The potential involvement of CXC receptors and stress hormone receptors in stress-induced neutrophil redistribution was determined by an in vivo study with selective CXCR inhibitors and antagonists of the receptors involved in stress regulation: glucocorticoid/mineralocorticoid receptors (GRs/MRs), adrenergic receptors (ADRs) and the melanocortin 2 receptor (MC2R). The stress-induced increase of blood neutrophils was accompanied by a neutrophil decrease in the hematopoietic organs. This increase was cortisol-induced and GR-dependent. Moreover, stress upregulated the expression of genes encoding CXCL12 and CXCL8 chemokines, their receptors, and the receptor for granulocytes colony-stimulation factor (GCSFR) and matrix metalloproteinase 9 (MMP9). Blocking of the CXCR4 and CXCR1 and 2 receptors with selective inhibitors inhibited the stress-induced neutrophil redistribution and affected the expression of genes encoding CXC chemokines and CXCRs as well as GCSFR and MMP9. Our data demonstrate that acute stress leads to the mobilization of the immune system, characterized by neutrophilia. CXC chemokines and CXC receptors are involved in this stress-induced redistribution of neutrophils from the hematopoietic tissue into the peripheral blood. This phenomenon is directly regulated by interactions between cortisol and the GR/MR. Considering the pivotal importance of neutrophilic granulocytes in the first line of defense, this knowledge is important for aquaculture, but will also contribute to the mechanisms involved in the stress-induced perturbation in neutrophil redistribution as often observed in clinical practice.

5-HT2A receptors are involved in the pharmaco-toxicological effects of the synthetic cannabinoids JWH-018 and 5F-PB22: In vivo studies in mice

Over the last years, Synthetic Cannabinoids (SCs) have been among the largest and most frequently seized groups of Novel Psychoactive Substances (NPS). These substances have been frequently detected in biological samples from patients involved in several intoxication and death cases. Their serious adverse effects have been related to their action as potent agonist of cannabinoid CB1 receptors. However, evidence concerning the potential interaction between SCs and serotoninergic mechanisms has emerged. Therefore, this study aims to evaluate the involvement of 5-HT2A receptors in the effects induced by acute systemic administration of 1-pentyl-3-(1-naphthoyl)indole (JWH-018; 1 mg/kg) and quinolin-8-yl 1-pentyfluoro-1H-indole-3-8-carboxylate (5F-PB22; 1 mg/kg). Sensorimotor (visual, acoustic, and tactile) responses, pain threshold (acute mechanical and thermal nociception), core temperature, breath rate and motor performance (stepping activity) have been assessed in CD-1 male mice. The present results pointed out that both substances deeply alter sensorimotor responses, nociceptive threshold, core temperature, breath rate and motor activity in mice. Noteworthy, pretreatment with the selective 5-HT2A receptors antagonist MDL100907 (0.1 mg/kg) at least partially prevented sensorimotor disruption, antinociception and hypothermic effects. Conversely, the respiratory and motor impairment was not prevented. Thus, it states the relevance of serotoninergic 5-HT2A mechanisms on pharmaco-toxicological effects induced by SCs.

Emerging therapeutic strategies in cancer therapy by HDAC inhibition as the chemotherapeutic potent and epigenetic regulator.

In developing new cancer medications, attention has been focused on novel epigenetic medicines called histone deacetylase (HDAC) inhibitors. Our understanding of cancer behavior is being advanced by research on epigenetics, which also supplies new targets for improving the effectiveness of cancer therapy. Most recently published patents emphasize HDAC selective drugs and multitarget HDAC inhibitors. Though significant progress has been made in emerging HDAC selective antagonists, it is urgently necessary to find new HDAC blockers with novel zinc-binding analogues to avoid the undesirable pharmacological characteristics of hydroxamic acid. HDAC antagonists have lately been explored as a novel approach to treating various diseases, including cancer. The complicated terrain of HDAC inhibitor development is summarized in this article, starting with a discussion of the many HDAC isotypes and their involvement in cancer biology, followed by a discussion of the mechanisms of action of HDAC inhibitors, their current level of development, effect of miRNA, and their combination with immunotherapeutic.

Exploring Biginelli-based scaffolds as A2B adenosine receptor antagonists: Unveiling novel structure-activity relationship trends, lead compounds, and potent colorectal anticancer agents

Antagonists of the A2B adenosine receptor have recently emerged as targeted anticancer agents and immune checkpoint inhibitors within the realm of cancer immunotherapy. This study presents a comprehensive evaluation of novel Biginelli-assembled pyrimidine chemotypes, including mono-, bi-, and tricyclic derivatives, as A2BAR antagonists. We conducted a comprehensive examination of the adenosinergic profile (both binding and functional) of a large compound library consisting of 168 compounds. This approach unveiled original lead compounds and enabled the identification of novel structure-activity relationship (SAR) trends, which were supported by extensive computational studies, including quantum mechanical calculations and free energy perturbation (FEP) analysis. In total, 25 molecules showed attractive affinity (Ki < 100 nM) and outstanding selectivity for A2BAR. From these, five molecules corresponding to the new benzothiazole scaffold were below the Ki < 10 nM threshold, in addition to a novel dual A2A/A2B antagonist. The most potent compounds, and the dual antagonist, showed enantiospecific recognition in the A2BAR. Two A2BAR selective antagonists and the dual A2AAR/A2BAR antagonist reported in this study were assessed for their impact on colorectal cancer cell lines. The results revealed a significant and dose-dependent reduction in cell proliferation. Notably, the A2BAR antagonists exhibited remarkable specificity, as they did not impede the proliferation of non-tumoral cell lines. These findings support the efficacy and potential that A2BAR antagonists as valuable candidates for cancer therapy, but also that they can effectively complement strategies involving A2AAR antagonism in the context of immune checkpoint inhibition.

The Estrogen Receptor Alpha Regulates the Sex-dependent Expression and Pronociceptive Role of Bestrophin-1 in Neuropathic Rats

Bestrophin-1, a calcium-activated chloride channel (CaCC), is involved in neuropathic pain; however, it is unclear whether it has a dimorphic role in female and male neuropathic rats. This study investigated if 17β-estradiol and estrogen receptor alpha (ERα) activation regulate bestrophin-1 activity and expression in neuropathic rats. Neuropathic pain was induced by L5-spinal nerve transection (SNT). Intrathecal administration of CaCCinh-A01 (.1–1 µg), a CaCC blocker, reversed tactile allodynia induced by SNT in female but not male rats. In contrast, T16Ainh-A01, a selective anoctamin-1 blocker, had an equal antiallodynic effect in both sexes. SNT increased bestrophin-1 protein expression in injured L5 dorsal root ganglia (DRG) in female rats but decreased bestrophin-1 protein in L5 DRG in male rats. Ovariectomy prevented the antiallodynic effect of CaCCinh-A01, but 17β-estradiol replacement restored it. The effect of CaCCinh-A01 was prevented by intrathecal administration of MPP, a selective ERα antagonist, in rats with and without prior hormonal manipulation. In female rats with neuropathy, ovariectomy prevented the increase in bestrophin-1 and ERα protein expression, while 17β-estradiol replacement allowed for an increase in both proteins in L5 DRG. Furthermore, ERα antagonism (with MPP) prevented the increase in bestrophin-1 and ERα protein expression. Finally, ERα activation with PPT, an ERα selective activator, induced the antiallodynic effect of CaCCinh-A01 in neuropathic male rats and prevented the reduction in bestrophin-1 protein expression in L5 DRG. In summary, data suggest ERα activation is necessary for bestrophin-1’s pronociceptive action to maintain neuropathic pain in female rats. PerspectiveThe mechanisms involved in neuropathic pain differ between male and female animals. Our data suggest that ERα is necessary for expression and function of bestrophin-1 in neuropathic female but not male rats. Data support the idea that a therapeutic approach to relieving neuropathic pain must be based on patient's gender.

Unveiling the antinociceptive mechanisms of Methyl-2-(4-chloro-phenyl)-5-benzoxazoleacetate: insights from nociceptive assays in mice.

Methyl-2-(4-chloro- phenyl)-5-benzoxazoleacetate (MCBA), a synthetic benzoxazole derivative with established antipsoriatic efficacy, was investigated for potential antinociceptive effects. This study employs various nociceptive assays in mice to elucidate MCBA's antinociceptive mechanisms. MCBA's antinociceptive potential was tested against various nociception models induced by formalin, glutamate, capsaicin, a transient receptor potential vanilloid 1 (TRPV1) receptor agonist, and phorbol 12-myristate 13-acetate, a protein kinase C (PKC) activator. It was then assessed using the hot plate test and examined within the acetic acid-induced writhing test. During the acetic acid-induced writhing test, MCBA was pre-challenged against selective receptor antagonists such as naloxone, caffeine, atropine, yohimbine, ondansetron, and haloperidol. It was also pre-challenged with ATP-sensitive potassium channel inhibitor (glibenclamide) to further elucidate its antinociceptive mechanism. The results showed that oral administration of MCBA led to a dose-dependent and significant inhibition (p < 0.05) of nociceptive effects across all evaluated models at doses of 60, 120, and 240 mg/kg. Moreover, the efficacy of MCBA's antinociceptive potential was significantly counteracted (p < 0.0001) by specific antagonists: (i) directed at adenosinergic, alpha-2 adrenergic, and cholinergic receptors using caffeine, yohimbine, and atropine, respectively; and (ii) targeting ATP-sensitive potassium channels, employing glibenclamide. Antagonists aimed at opioidergic and serotoninergic receptors (naloxone and ondansetron, respectively) had poor utility in inhibiting antinociceptive activity. Conversely, the dopaminergic receptor antagonist haloperidol potentiated locomotor abnormalities associated with MCBA treatment. MCBA-induced antinociception involves modulation of glutamatergic-, TRVP1 receptors- and PKC-signaling pathways. It impacts adenosinergic, alpha-2 adrenergic, and cholinergic receptors and opens ATP-sensitive potassium channels.

Cortico-striatal gamma oscillations are modulated by dopamine D3 receptors in dyskinetic rats.

JOURNAL/nrgr/04.03/01300535-202504000-00031/figure1/v/2024-07-06T104127Z/r/image-tiff Long-term levodopa administration can lead to the development of levodopa-induced dyskinesia. Gamma oscillations are a widely recognized hallmark of abnormal neural electrical activity in levodopa-induced dyskinesia. Currently, studies have reported increased oscillation power in cases of levodopa-induced dyskinesia. However, little is known about how the other electrophysiological parameters of gamma oscillations are altered in levodopa-induced dyskinesia. Furthermore, the role of the dopamine D3 receptor, which is implicated in levodopa-induced dyskinesia, in movement disorder-related changes in neural oscillations is unclear. We found that the cortico-striatal functional connectivity of beta oscillations was enhanced in a model of Parkinson's disease. Furthermore, levodopa application enhanced cortical gamma oscillations in cortico-striatal projections and cortical gamma aperiodic components, as well as bidirectional primary motor cortex (M1) ↔ dorsolateral striatum gamma flow. Administration of PD128907 (a selective dopamine D3 receptor agonist) induced dyskinesia and excessive gamma oscillations with a bidirectional M1 ↔ dorsolateral striatum flow. However, administration of PG01037 (a selective dopamine D3 receptor antagonist) attenuated dyskinesia, suppressed gamma oscillations and cortical gamma aperiodic components, and decreased gamma causality in the M1 → dorsolateral striatum direction. These findings suggest that the dopamine D3 receptor plays a role in dyskinesia-related oscillatory activity, and that it has potential as a therapeutic target for levodopa-induced dyskinesia.

Altered purinergic P2X7 and A2B receptors signaling limits macrophage-mediated host defense in schistosomiasis

BackgroundThe occurrence of co-infections during schistosomiasis, a neglected tropical disease, with other parasites have been reported suggesting an impaired host immune defense. Macrophage purinergic P2X7 receptor (P2X7R) play an important role against intracellular pathogens. Therefore, we investigated the P2X7R-mediated phagocytosis and killing capacity of Leishmania amazonensis by macrophages during schistosomiasis in vitro and in vivo. MethodsSwiss and C57BL/6 (Wild type) and P2X7R−/− were randomized in two groups: control (uninfected) and Schistosoma mansoni-infected. Alternatively, control Swiss and S. mansoni-infected mice were also infected with L. amazonensis. ResultsThe pre-treatment of macrophages with the P2X7R antagonist (A74003) or TGF-β reduced the phagocytosis index, mimicking the phenotype of cells from S. mansoni-infected mice and P2X7R−/− mice. Apyrase also reduced the phagocytosis index corroborating the role of ATP to macrophage activation. Moreover, l-arginine-nitric oxide pathway was compromised, which could explain the reduced killing capacity in response to ATP in vitro and in vivo. We found an increased extracellular nucleotide (ATP, ADP and AMP) hydrolysis along with an increased frequency of F4/80+ CD39+ macrophages from the S. mansoni-infected group. Moreover, the content of adenosine in the cell supernatant was higher in the S. mansoni-infected group in relation to controls. Schistosomiasis also increased the expression of macrophage adenosine A2BR. In good accordance, both ADA and the selective A2BR antagonist restored the phagocytosis index of macrophages from S. mansoni-infected group. ConclusionsAltogether, the altered P2X7R and A2BR signaling limits the role of macrophages to host defense against L. amazonensis during schistosomiasis, potentially contributing to the pathophysiology and clinically relevant co-infections.

An Orally Efficacious Thyrotropin Receptor Ligand Inhibits Growth and Metastatic Activity of Thyroid Cancers.

Thyroid-stimulating hormone (or thyrotropin) receptor (TSHR) could be a selective target for small molecule ligands to treat thyroid cancer (TC). We report a novel, orally efficacious ligand for TSHR that exhibits proliferation inhibitory activity against human TC in vitro and in vivo, and inhibition of metastasis in vivo. A35 (NCATS-SM4420; NCGC00241808) was selected from a sublibrary of >200 TSHR ligands. Cell proliferation assays including BrdU incorporation and WST-1, along with molecular docking studies were done. In vivo activity of A35 was assessed in TC cell-derived xenograft (CDX) models with immunocompromised (NSG) mice. Formalin-fixed, paraffin-embedded sections of tumor and lung tissues were observed for the extent of cell death and metastasis. A35 was shown to stimulate cAMP production in some cell types by activating TSHR but not in TC cells, MDA-T32, and MDA-T85. A35 inhibited proliferation of MDA-T32 and MDA-T85 in vitro and in vivo, and pulmonary metastasis of MDA-T85F1 in mice. In vitro, A35 inhibition of proliferation was reduced by a selective TSHR antagonist. Inhibition of CDX tumor growth without decreases in mouse weights and liver function showed A35 to be efficacious without apparent toxicity. Lastly, A35 reduced levels of Ki67 in the tumors and metastatic markers in lung tissues. We conclude that A35 is a TSHR-selective inhibitor of TC cell proliferation and metastasis, and suggest that A35 may be a promising lead drug candidate for the treatment of differentiated TC in humans.

The metabolism of the orexin-1 selective receptor antagonist nivasorexant

Nivasorexant was the first orexin-1 selective receptor antagonist entering clinical development. Despite encouraging preclinical evidence in animal models, a proof-of-concept trial in binge-eating patients recently failed to demonstrate its clinical utility in this population. Across species, nivasorexant clearance was driven by metabolism along seven distinct pathways, five of which were hydroxylation reactions in various locations of the molecule. The exact sites of metabolism were identified by means of mass spectrometry, the use of deuterated analogues, and finally confirmed by chemical references. CYP3A4 was the main cytochrome P450 enzyme involved in nivasorexant metabolism in vitro and accounting for about 90% of turnover in liver microsomes. Minor roles were taken by CYP2C9 and CYP2C19 but individually did not exceed 3–7%. In the rat, nivasorexant was mostly excreted via the bile after extensive metabolism, while urinary excretion was negligible. Only traces of the parent drug were detected in urine, bile, or faeces.

Exploring the therapeutic potential of cannabidiol for sleep deprivation-induced hyperalgesia

Hyperalgesia resulting from sleep deprivation (SD) poses a significant a global public health challenge with limited treatment options. The nucleus accumbens (NAc) plays a crucial role in the modulation of pain and sleep, with its activity regulated by two distinct types of medium spiny neurons (MSNs) expressing dopamine 1 or dopamine 2 (D1-or D2) receptors (referred to as D1-MSNs and D2-MSNs, respectively). However, the specific involvement of the NAc in SD-induced hyperalgesia remains uncertain. Cannabidiol (CBD), a nonpsychoactive phytocannabinoid, has demonstrated analgesic effects in clinical and preclinical studies. Nevertheless, its potency in addressing this particular issue remains to be determined. Here, we report that SD induced a pronounced pronociceptive effect attributed to the heightened intrinsic excitability of D2-MSNs within the NAc in Male C57BL/6N mice. CBD (30 mg/kg, i.p.) exhibited an anti-hyperalgesic effect. CBD significantly improved the thresholds for thermal and mechanical pain and increased wakefulness by reducing delta power. Additionally, CBD inhibited the intrinsic excitability of D2-MSNs both in vitro and in vivo. Bilateral microinjection of the selective D2 receptor antagonist raclopride into the NAc partially reversed the antinociceptive effect of CBD. Thus, these findings strongly suggested that SD activates NAc D2-MSNs, contributing heightened to pain sensitivity. CBD exhibits antinociceptive effects by activating D2R, thereby inhibiting the excitability of D2-MSNs and promoting wakefulness under SD conditions.

Exponential Combination of a and e/g Intracellular Peptide Libraries Identifies a Selective ATF3 Inhibitor.

Activating transcription factor 3 (ATF3) is an activation transcription factor/cyclic adenosine monophosphate (cAMP) responsive element-binding (CREB) protein family member. It is recognized as an important regulator of cancer progression by repressing expression of key inflammatory factors such as interferon-γ and chemokine (C-C motif) ligand 4 (CCL4). Here, we describe a novel library screening approach that probes individual leucine zipper components before combining them to search exponentially larger sequence spaces not normally accessible to intracellular screening. To do so, we employ two individual semirational library design approaches and screen using a protein-fragment complementation assay (PCA). First, a 248,832-member library explored 12 amino acid positions at all five a positions to identify those that provided improved binding, with all e/g positions fixed as Q, placing selection pressure onto the library options provided. Next, a 59,049-member library probed all ten e/g positions with 3 options. Similarly, during e/g library screening, a positions were locked into a generically bindable sequence pattern (AIAIA), weakly favoring leucine zipper formation, while placing selection pressure onto e/g options provided. The combined a/e/g library represents ∼14.7 billion members, with the resulting peptide, ATF3W_aeg, binding ATF3 with high affinity (Tm = 60 °C; Kd = 151 nM) while strongly disfavoring homodimerization. Moreover, ATF3W_aeg is notably improved over component PCA hits, with target specificity found to be driven predominantly by electrostatic interactions. The combined a/e/g exponential library screening approach provides a robust, accelerated platform for exploring larger peptide libraries, toward derivation of potent yet selective antagonists that avoid homoassociation to provide new insight into rational peptide design.