Activation of Relaxin Family Receptor 1 from Different Mammalian Species by Relaxin Peptide and Small-Molecule Agonist ML290

Huang, Zaohua; Myhr, Courtney; Bathgate, Ross A. D.; Ho, Brian A.; Bueno, Amaya; Hu, Xin; Xiao, Jingbo; Southall, Noel; Barnaeva, Elena; Agoulnik, Irina U.; Marugán, Juan; Ferrer, Marc; Agoulnik, Alexander I.

doi:10.3389/fendo.2015.00128

Cited by 19 publications

(22 citation statements)

References 29 publications

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“…ML290 appears to be an RXPF1 agonist in humans, monkey's and pigs, and an antagonist in mice, suggesting species bias in ML290 signaling. [118,119,120] However, studies with mouse RXFP1 replaced with humanized RXFP1 have shown that animal models can be developed to study the effects of ML290 in smaller rodents. [121] In contrast to RLX, ML290 has been shown to have a half-life in the heart and plasma of ~8 hours, opening the possibility of longer lasting effects with less treatment time.…”

Section: Relaxin Mimeticsmentioning

confidence: 99%

Cardioprotective actions of relaxin

Martin

Romero

Salama

2019

Molecular and Cellular Endocrinology

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Relaxin, a hormone of pregnancy, has shown broad cardioprotective effects including anti-fibrotic (reversed excess TGFβ signaling), anti-arrhythmic (Nav1.5 and INa upregulation and Cx43 phosphorylation and trafficking to intercalated disks) and anti-inflammatory properties (reduced IL-1β and IL-6). While relaxin's anti-fibrotic effects are thought to occur through the SMAD2/3/TGFβ pathway, there is a general lack of understanding of relaxin's mode of action to increase sodium current and alter connexin43 localization to suppress arrhythmias. Based on a rat model of aging, we tested the hypothesis that relaxin acts through activation of Wnt/βcatenin signaling to mediate its effects on Nav1.5 and Cx43 and to regulate collagen expression. We show for the first time that relaxin activates canonical Wnt signaling (increased nuclear βcatenin) to increase Nav1.5 in isolated cardiomyocytes. Block of canonical Wnt signaling (via Dikkopf-1) abrogated relaxin's effect on both Nav1.5 in cardiomyocytes and suppression of excess collagen from fibroblasts. Further, we show that relaxin significantly suppressed expression of Dikkopf-1 in isolated cardiomyocytes and whole LV tissue. In addition, show that relaxin suppressed the age-associated genetic upregulation in inflammatory markers in a sexdependent manner. Finally, we tested the hypothesis that relaxin would be an effective therapy in a rat model of pulmonary arterial hypertension and show that it reduced occlusion of small pulmonary arteries, myocardial and lung fibrosis and reversed the cardiac phenotype of ventricular arrhythmia or arrest. These results suggest that relaxin signals through multiple

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Section: Relaxin Mimeticsmentioning

confidence: 99%

Cardioprotective actions of relaxin

Martin

Romero

Salama

2019

Molecular and Cellular Endocrinology

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“…The mouse and rat RXFP1 are not activated by ML290, but monkey and pig RXFP1 are fully activated. 32 Sequence alignment shows that monkey and pig RXFP1 have the same e GT motif within ECL3 as hRXFP1, whereas mouse and rat RXFP1 contain residues DS/T at that position. Again, this finding reiterates the importance of the GT motif in receptor activation.…”

Section: Resultsmentioning

confidence: 99%

Structural Insights into the Activation of Human Relaxin Family Peptide Receptor 1 by Small-Molecule Agonists

Myhr²,

Huang³

et al. 2016

Biochemistry

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The GPCR relaxin family peptide receptor 1 (RXFP1) mediates the action of relaxin peptide hormone including its tissue remodeling and antifibrotic effects. The peptide hormone has a short half-life in plasma, limiting its therapeutic utility. However, small molecule agonists of human RXFP1 can overcome this limitation and may provide a useful therapeutic approach especially for chronic diseases such as heart failure and fibrosis. The first small molecule agonists of RXFP1 were recently identified from a high throughput screen using a homogenous cell-based cAMP assay. Optimization of the hit compounds resulted in a series of highly potent and RXFP1 selective agonists with low cytotoxicity, and excellent in vitro ADME and pharmacokinetic properties. Here we undertook extensive site-directed mutagenesis studies in combination with computational modeling analysis to probe the molecular basis of the small molecule binding to RXFP1. The results showed that the agonists bind to an allosteric site of RXFP1 in a manner that closely interacts with the seven transmembrane domain (TM7) and the third extracellular loop (ECL3). A number of residues were found to play an important role in the agonist binding and receptor activation including a hydrophobic region at TM7 consisting of W664, F668 and L670. The G659/T660 motif within ECL3 is crucial to the observed species selectivity of the agonists for RXFP1. The receptor binding and activation effects by the small molecule agonist ML290 were compared with the cognate ligand relaxin peptide, providing valuable insights on the structural basis and molecular mechanism of activation and selectivity of the agonists for RXFP1.

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“…This is perhaps not surprising as ML290 interacts with human RXFP1 at a site distinct from that of H2-RLX, with the low density lipoprotein class A (LDLa) domain of RXFP1 being dispensable for activity 17 . Furthermore, the actions of ML290 are species-specific, with activity at human, macaque and pig RXFP1 but no agonist action at the mouse, rat, hamster or guinea pig receptor 36 . Amino acid differences in the third extracellular loop of the seven-transmembrane region of RXFP1 are responsible for this specificity.…”

Section: Discussionmentioning

confidence: 99%

In search of a small molecule agonist of the relaxin receptor RXFP1 for the treatment of liver fibrosis

McBride

Hoy

Bamford³

et al. 2017

Sci Rep

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The peptide hormone human relaxin-2 (H2-RLX) has emerged as a potential therapy for cardiovascular and fibrotic diseases, but its short in vivo half-life is an obstacle to long-term administration. The discovery of ML290 demonstrated that it is possible to identify small molecule agonists of the cognate G-protein coupled receptor for H2-RLX (relaxin family peptide receptor-1 (RXFP1)). In our efforts to generate a new medicine for liver fibrosis, we sought to identify improved small molecule functional mimetics of H2-RLX with selective, full agonist or positive allosteric modulator activity against RXFP1. First, we confirmed expression of RXFP1 in human diseased liver. We developed a robust cellular cAMP reporter assay of RXFP1 signaling in HEK293 cells transiently expressing RXFP1. A high-throughput screen did not identify further specific agonists or positive allosteric modulators of RXFP1, affirming the low druggability of this receptor. As an alternative approach, we generated novel ML290 analogues and tested their activity in the HEK293-RXFP1 cAMP assay and the human hepatic cell line LX-2. Differences in activity of compounds on cAMP activation compared with changes in expression of fibrotic markers indicate the need to better understand cell- and tissue-specific signaling mechanisms and their disease-relevant phenotypes in order to enable drug discovery.

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Activation of Relaxin Family Receptor 1 from Different Mammalian Species by Relaxin Peptide and Small-Molecule Agonist ML290

Cited by 19 publications

References 29 publications

Cardioprotective actions of relaxin

Cardioprotective actions of relaxin

Structural Insights into the Activation of Human Relaxin Family Peptide Receptor 1 by Small-Molecule Agonists

In search of a small molecule agonist of the relaxin receptor RXFP1 for the treatment of liver fibrosis

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