Discovery of small molecule guanylyl cyclase A receptor positive allosteric modulators

2021 ◽  
Vol 118 (52) ◽  
pp. e2109386118
Author(s):  
S. Jeson Sangaralingham ◽  
Kanupriya Whig ◽  
Satyamaheshwar Peddibhotla ◽  
R. Jason Kirby ◽  
Hampton E. Sessions ◽  
...  
Keyword(s):  
Natriuretic Peptide ◽  
Small Molecule ◽  
Guanylyl Cyclase ◽  
High Throughput Screening ◽  
Metabolic Diseases ◽  
Ex Vivo ◽  
Guanosine Monophosphate ◽  
Cardiomyocyte Hypertrophy ◽  
Orally Bioavailable

The particulate guanylyl cyclase A receptor (GC-A), via activation by its endogenous ligands atrial natriuretic peptide (ANP) and b-type natriuretic peptide (BNP), possesses beneficial biological properties such as blood pressure regulation, natriuresis, suppression of adverse remodeling, inhibition of the renin-angiotensin-aldosterone system, and favorable metabolic actions through the generation of its second messenger cyclic guanosine monophosphate (cGMP). Thus, the GC-A represents an important molecular therapeutic target for cardiovascular disease and its associated risk factors. However, a small molecule that is orally bioavailable and directly targets the GC-A to potentiate cGMP has yet to be discovered. Here, we performed a cell-based high-throughput screening campaign of the NIH Molecular Libraries Small Molecule Repository, and we successfully identified small molecule GC-A positive allosteric modulator (PAM) scaffolds. Further medicinal chemistry structure–activity relationship efforts of the lead scaffold resulted in the development of a GC-A PAM, MCUF-651, which enhanced ANP-mediated cGMP generation in human cardiac, renal, and fat cells and inhibited cardiomyocyte hypertrophy in vitro. Further, binding analysis confirmed MCUF-651 binds to GC-A and selectively enhances the binding of ANP to GC-A. Moreover, MCUF-651 is orally bioavailable in mice and enhances the ability of endogenous ANP and BNP, found in the plasma of normal subjects and patients with hypertension or heart failure, to generate GC-A–mediated cGMP ex vivo. In this work, we report the discovery and development of an oral, small molecule GC-A PAM that holds great potential as a therapeutic for cardiovascular, renal, and metabolic diseases.

scholarly journals Novel enhancers of guanylyl cyclase-A activity via allosteric modulation

2022 ◽  
Author(s):  
Henriette Andresen ◽  
Cristina Pérez-Ternero ◽  
Jerid Robinson ◽  
Deborah M Dickey ◽  
Adrian J Hobbs ◽  
...  
Keyword(s):  
Small Molecules ◽  
Natriuretic Peptide ◽  
Guanylyl Cyclase ◽  
Drug Target ◽  
Ex Vivo ◽  
Cyclic Guanosine Monophosphate ◽  
Guanosine Monophosphate ◽  
Allosteric Binding Sites ◽  
Small Molecule Modulators

Natriuretic peptide receptor (NPR)-A (also known as NPR-A, NPR1 or guanylyl cyclase-A, GC-A) is an attractive but challenging target to activate with small molecules. GC-A is activated by endogenous atrial natriuretic peptide (ANP) and B-type natriuretic peptide (BNP), and this activation leads to the production of cyclic guanosine monophosphate (cGMP). This system plays an important role in the regulation of cardiovascular and renal homeostasis. However, utilization of this receptor as a drug target has so far been limited to peptides, even though small molecule modulators allow oral administration and longer half-life. We have identified small molecular allosteric enhancers of GC-A, which strengthened ANP or BNP activation in various in vitro and ex vivo systems. These compounds do not mediate their actions through previously described allosteric binding sites or via known mechanisms of action. In addition, their selectivity and activity are dependent on only one amino acid in GC-A. Our findings show that there is a novel allosteric binding site on GC-A, which can be targeted by small molecules that increase the signaling effects of ANP and BNP.

scholarly journals Betulinic Acid Protects DOX-Triggered Cardiomyocyte Hypertrophy Response through the GATA-4/Calcineurin/NFAT Pathway

Molecules ◽  
2020 ◽  
Vol 26 (1) ◽  
pp. 53
Author(s):  
Jung Joo Yoon ◽  
Chan Ok Son ◽  
Hye Yoom Kim ◽  
Byung Hyuk Han ◽  
Yun Jung Lee ◽  
...  
Keyword(s):  
Heart Failure ◽  
Cardiac Hypertrophy ◽  
Natriuretic Peptide ◽  
Betulinic Acid ◽  
Cardiomyocyte Hypertrophy ◽  
Ros Generation ◽  
H9c2 Cells ◽  
Myosin Light Chain 2 ◽  
Cardiovascular Morbidity And Mortality

Cardiac hypertrophy is a major risk factor for heart failure and leads to cardiovascular morbidity and mortality. Doxorubicin (DOX) is regarded as one of the most potent anthracycline antibiotic agents; however, its clinical usage has some limitations because it has serious cardiotoxic side effects such as dilated cardiomyopathy and congestive heart failure. Betulinic acid (BA) is a pentacyclic-cyclic lupane-type triterpene that has been reported to have anti-bacterial, anti-inflammatory, anti-vascular neogenesis, and anti-fibrotic effects. However, there is no study about its direct effect on DOX induced cardiac hypertrophy and apoptosis. The present study aims to investigate the effect of BA on DOX-induced cardiomyocyte hypertrophy and apoptosis in vitro in H9c2 cells. The H9c2 cells were stimulated with DOX (1 µM) in the presence or absence of BA (0.1–1 μM) and incubated for 24 h. The results of the present study indicated that DOX induces the increase cell surface area and the upregulation of hypertrophy markers including atrial natriuretic peptide (ANP), B-type natriuretic peptide (BNP), beta-myosin heavy chain (β-MHC), and Myosin Light Chain-2 (MLC2) in H9c2 cells. However, the pathological hypertrophic responses were downregulated after BA treatment. Moreover, phosphorylation of JNK, ERK, and p38 in DOX treated H9c2 cells was blocked by BA. As a result of measuring the change in ROS generation using DCF-DA, BA significantly inhibited DOX-induced the production of intracellular reactive oxygen species (ROS) when BA was treated at a concentration of over 0.1 µM. DOX-induced activation of GATA-4 and calcineurin/NFAT-3 signaling pathway were remarkably improved by pre-treating of BA to H9c2 cells. In addition, BA treatment significantly reduced DOX-induced cell apoptosis and protein expression levels of Bax and cleaved caspase-3/-9, while the expression of Bcl-2 was increased by BA. Therefore, BA can be a potential treatment for cardiomyocyte hypertrophy and apoptosis that lead to sudden heart failure.

SOMCL-863, a novel, selective and orally bioavailable small-molecule c-Met inhibitor, exhibits antitumor activity both in vitro and in vivo

2014 ◽  
Vol 351 (1) ◽  
pp. 143-150 ◽  
Author(s):  
Lu Wang ◽  
Jing Ai ◽  
Yanyan Shen ◽  
Haotian Zhang ◽  
Xia Peng ◽  
...  
Keyword(s):  
Antitumor Activity ◽  
Small Molecule ◽  
Met Inhibitor ◽  
Orally Bioavailable

Abstract 16339: Mitochondrial Function in Animal and Novel Human Disease Models of Tachycardiomyopathy

Circulation ◽  
2020 ◽  
Vol 142 (Suppl_3) ◽  
Author(s):  
Michael G Paulus ◽  
Kathrin Renner ◽  
Steffen Pabel ◽  
Gabriela Pietrzyk ◽  
Andreas Luchner ◽  
...  
Keyword(s):  
Oxidative Phosphorylation ◽  
Human Disease ◽  
Citrate Synthase ◽  
Ex Vivo ◽  
Pacemaker Implantation ◽  
Induced Pluripotent Stem Cell ◽  
Cardiomyocyte Hypertrophy ◽  
Ex Vivo Model ◽  
Ablation Therapy

Introduction: Clinical significance of tachycardiomyopathy (TCM) increased with trials on catheter ablation therapy. Myocardial biopsies from patients show disturbed mitochondrial architecture. Hypothesis: TCM involves mitochondrial dysfunction. Methods: First, TCM was investigated in an animal model: pacemaker implantation in 7 rabbits was followed by tachypacing for 30 days (TCM), 7 animals served as sham-operated controls (SHAM). Second, results of the animal study were evaluated for their translational perspective for human disease using a novel model of induced pluripotent stem cell-derived cardiomyocytes (iPS-CM), derived from 4 healthy donors. IPS-CM were paced with 120 bpm (TACH) or 60 bpm (CTRL) for 7 days in vitro. Targeted transcriptomics, high-resolution respirometry and flow cytometry (MitoSOX Red) were performed. To account for variations between cell differentiations, experiments on iPS-CM were carried out in a paired design. Results: TCM showed LV dilatation and dysfunction (ΔLVEDD +5.3±0.2mm; ΔFS -19±8%; TCM-SHAM; p<0.001). Histological findings resembled human disease entailing cardiomyocyte hypertrophy (CSA 519±32μm 2 vs. 413±21μm 2 , p<0.01) without fibrosis (hydroxyproline content, p=0.52). Mitochondrial transcriptome of TCM was characterized by downregulation of 10 antioxidative enzymes (e.g. GPX3, fold change (FC) 0.4; TCM/SHAM; p<0.05) as well as mitochondrial carriers, including ADP/ATP- and NADH-shuttling (SLC25A4, FC 0.7; SLC25A12, FC 0.8; p<0.01). As transcriptomics implied impaired substrate import, respirometry was performed in whole tissue. In support of our findings on the transcriptome level, mitochondrial oxidative phosphorylation capacity decreased in TCM (133±13 vs. 170±16 pmol·O 2 ·s -1 ·mg -1 ·tissue, p<0.05). Similarly, oxidative phosphorylation was reduced in iPS-CM (995±738 vs. 1838±901 pmolO 2 ·s -1 ·IU -1 citrate synthase activity, TACH vs. CTRL, p<0.01). Concurrently, tachypacing increased mitochondrial superoxide emission in iPS-CM (MFI 491±206 vs. 301±119, p<0.05). Conclusions: Persistent tachycardia alters two mitochondrial key functions in an animal and a novel human ex vivo model: oxidative phosphorylation capacity is reduced, while superoxide emission increases.

scholarly journals Novel LCAT (Lecithin:Cholesterol Acyltransferase) Activator DS-8190a Prevents the Progression of Plaque Accumulation in Atherosclerosis Models

2020 ◽  
Author(s):  
Masato Sasaki ◽  
Mina Delawary ◽  
Hidetaka Sakurai ◽  
Hideki Kobayashi ◽  
Naoki Nakao ◽  
...  
Keyword(s):  
Small Molecule ◽  
High Throughput Screening ◽  
Affinity Purification ◽  
Atherosclerotic Lesion ◽  
Hdl Cholesterol ◽  
Lecithin:Cholesterol Acyltransferase ◽  
Lesion Area ◽  
Acyltransferase Activity ◽  
Hdl Function

Objective: Enhancement of LCAT (lecithin:cholesterol acyltransferase) activity has possibility to be beneficial for atherosclerosis. To evaluate this concept, we characterized our novel, orally administered, small-molecule LCAT activator DS-8190a, which was created from high-throughput screening and subsequent derivatization. We also focused on its mechanism of LCAT activation and the therapeutic activity with improvement of HDL (high-density lipoprotein) functionality. Approach and Results: DS-8190a activated human and cynomolgus monkey but not mouse LCAT enzymes in vitro. DS-8190a was orally administered to cynomolgus monkeys and dose dependently increased LCAT activity (ca. 2.1-fold in 3 mg/kg group on day 7), resulting in HDL cholesterol elevation without drastic changes of non-HDL cholesterol. Atheroprotective effects were then evaluated using Ldl-r KO × hLcat Tg mice fed a Western diet for 8 weeks. DS-8190a treatment achieved significant reduction of atherosclerotic lesion area (48.3% reduction in 10 mg/kg treatment group). Furthermore, we conducted reverse cholesterol transport study using Ldl-r KO × hLcat Tg mice intraperitoneally injected with J774A.1 cells loaded with [ 3 H]-cholesterol and confirmed significant increases of [ 3 H] count in plasma (1.4-fold) and feces (1.4-fold on day 2 and 1.5-fold on day3) in the DS-8190a–treated group. With regard to the molecular mechanism involved, direct binding of DS-8190a to human LCAT protein was confirmed by 2 different approaches: affinity purification by DS-8190a–immobilized beads and thermal shift assay. In addition, the candidate binding site of DS-8190a in human LCAT protein was identified by photoaffinity labeling. Conclusions: This study demonstrates the potential of DS-8190a as a novel therapeutic for atherosclerosis. This is also the first report describing that a small-molecule direct LCAT activator achieved HDL cholesterol elevation in monkey and reduction of atherosclerotic lesion area with enhanced HDL function in rodent.

scholarly journals Studies to Elucidate the Mechanism of Cardio Protective and Hypotensive Activities of Anogeissus acuminata (Roxb. ex DC.) in Rodents

Molecules ◽  
2020 ◽  
Vol 25 (15) ◽  
pp. 3471
Author(s):  
Fatima Saqib ◽  
Muhammad Arif Aslam ◽  
Khizra Mujahid ◽  
Luigi Marceanu ◽  
Marius Moga ◽  
...  
Keyword(s):  
Creatine Kinase ◽  
Ex Vivo ◽  
Inflammatory Cells ◽  
Left Ventricular ◽  
Guanosine Monophosphate ◽  
Sprague Dawley ◽  
Positive Effects ◽  
Creatine Kinase Mb

Anogeissus acuminata (Roxb. ex DC.) is a folkloric medicinal plant in Asia; including Pakistan; used as a traditional remedy for cardiovascular disorders. This study was planned to establish a pharmacological basis for the trivial uses of Anogeissus acuminata in certain medical conditions related to cardiovascular systems and to explore the underlying mechanisms. Mechanistic studies suggested that crude extract of Anogeissus acuminata (Aa.Cr) produced in vitro cardio-relaxant and vasorelaxant effects in isolated paired atria and aorta coupled with in vivo decrease in blood pressure by invasive method; using pressure and force transducers connected to Power Lab Data Acquisition System. Moreover; Aa.Cr showed positive effects in left ventricular hypertrophy in Sprague Dawley rats observed hemodynamically by a decrease in cardiac cell size and fibrosis; along with absence of inflammatory cells; coupled with reduced levels of angiotensin converting enzyme (ACE) and renin concentration along with increased concentrations of nitric oxide (NO) and cyclic guanosine monophosphate (cGMP). In Acute Myocardial Infarction (AMI) model; creatine kinase (CK), creatine kinase-MB (CK-MB) and lactic acid dehydrogenase (LDH levels) were found to be decreased; along with decreased necrosis; edema and recruitment of inflammatory cells histologically. In vivo and ex vivo studies of Anogeissus acuminata provided evidence of vasorelaxant; hypotensive and cardioprotective properties facilitated through blockage of voltage-gated Ca++ ion channel; validating its use in cardiovascular diseases

A novel small-molecule screening strategy identifies mitoxantrone as a RhoGTPase inhibitor

2013 ◽  
Vol 450 (1) ◽  
pp. 55-62 ◽  
Author(s):  
Aurélien Bidaud-Meynard ◽  
Daniela Arma ◽  
Said Taouji ◽  
Michel Laguerre ◽  
Jean Dessolin ◽  
...  
Keyword(s):  
Small Molecule ◽  
High Throughput Screening ◽  
Large Scale ◽  
Topoisomerase Ii ◽  
Screening Strategy ◽  
Filamentous Actin ◽  
Library Screen ◽  
Rac1 Gtpase ◽  
Gtpase Activation

RhoGTPases are GDP/GTP molecular switches that control a wide variety of cellular processes, thereby contributing to many diseases, including cancer. As a consequence, there is great interest in the identification of small-molecule inhibitors of RhoGTPases. In the present paper, using the property of GTP-loaded RhoGTPases to bind to their effectors, we describe a miniaturized and robust assay to monitor Rac1 GTPase activation that is suitable for large-scale high-throughput screening. A pilot compound library screen revealed that the topoisomerase II poison MTX (mitoxantrone) is an inhibitor of Rac1, and also inhibits RhoA and Cdc42 in vitro. We show that MTX prevents GTP binding to RhoA/Rac1/Cdc42 in vitro. Furthermore, MTX strongly inhibits RhoGTPase-mediated F-actin (filamentous actin) reorganization and cell migration. Hence, we report a novel biochemical assay yielding the identification of RhoGTPase inhibitors and we present a proof-of-concept validation with the identification of MTX as a novel pan-RhoGTPase inhibitor.

scholarly journals Dipeptidyl-Peptidase IV Converts Intact B-Type Natriuretic Peptide into Its des-SerPro Form

2006 ◽  
Vol 52 (1) ◽  
pp. 82-87 ◽  
Author(s):  
Inger Brandt ◽  
Anne-Marie Lambeir ◽  
Jean-Marie Ketelslegers ◽  
Marc Vanderheyden ◽  
Simon Scharpé ◽  
...  
Keyword(s):  
Natriuretic Peptide ◽  
Ex Vivo ◽  
Dipeptidyl Peptidase ◽  
Dipeptidyl Peptidase Iv ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Amino Terminal ◽  
Dpp Iv

Abstract Background: Analysis of plasma B-type natriuretic peptide (BNP) has suggested the in vivo formation of a truncated form, BNP (3–32), also called des-SerPro-BNP. The objectives of this study were to investigate (a) whether BNP and other natriuretic peptides are truncated by dipeptidyl-peptidase IV (DPP IV/CD26; EC 3.4.14.5) and (b) whether this truncation affects the susceptibility to cleavage by neutral endopeptidase (NEP; EC 3.4.24.11). Methods: Human BNP (1–32), A-type natriuretic peptide 1–28 (ANP 1–28), and related peptides were incubated with purified DPP IV and with human plasma. In addition, BNP (1–32), BNP (3–32), and ANP (1–28) were subjected to hydrolysis by NEP. Cleavage products were analyzed by mass spectrometry. Results: BNP (1–32) was cleaved by purified DPP IV with a specificity constant of 0.37 × 106 L · mol−1 · s−1. The DPP IV activity in EDTA-plasma was able to truncate BNP (1–32) ex vivo. Addition of Vildagliptin, a specific DPP IV inhibitor, prevented this truncation in a concentration-dependent manner. Under in vitro circumstances in which ANP was hydrolyzed extensively, BNP (1–32) and BNP (3–32) were very resistant to NEP-mediated cleavage. Conclusions: DPP IV cleaves BNP (1–32) with an efficiency higher than or comparable to several known in vivo substrates of the enzyme. Even after loss of the amino-terminal dipeptide, BNP remains highly resistant to cleavage by NEP.

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