7 research outputs found
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Development and validation of an LC-MS/MS method for detection and quantification of in vivo derived metabolites of [Pyr 1 ]apelin-13 in humans
Abstract: [Pyr1]apelin-13 is the predominant apelin peptide isoform in the human cardiovascular system and plasma. To date, few studies have investigated [Pyr1]apelin-13 metabolism in vivo in rats with no studies examining its stability in humans. We therefore aimed to develop an LC-MS/MS method for detection and quantification of intact [Pyr1]apelin-13 and have used this method to identify the metabolites generated in vivo in humans. [Pyr1]apelin-13 (135 nmol/min) was infused into six healthy human volunteers for 120 minutes and blood collected at time 0 and 120 minutes after infusion. Plasma was extracted in the presence of guanidine hydrochloride and analysed by LC-MS/MS. Here we report a highly sensitive, robust and reproducible method for quantification of intact [Pyr1]apelin-13 and its metabolites in human plasma. Using this method, we showed that the circulating concentration of intact peptide was 58.3 ± 10.5 ng/ml after 120 minutes infusion. We demonstrated for the first time that in humans, [Pyr1]apelin-13 was cleaved from both termini but the C-terminal was more susceptible to cleavage. Consequently, of the metabolites identified, [Pyr1]apelin-13(1–12), [Pyr1]apelin-13(1–10) and [Pyr1]apelin-13(1–6) were the most abundant. These data suggest that apelin peptides designed for use as cardiovascular therapeutics, should include modifications that minimise C-terminal cleavage
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Development and validation of an LC-MS/MS method for detection and quantification of in vivo derived metabolites of [Pyr 1 ]apelin-13 in humans
Abstract: [Pyr1]apelin-13 is the predominant apelin peptide isoform in the human cardiovascular system and plasma. To date, few studies have investigated [Pyr1]apelin-13 metabolism in vivo in rats with no studies examining its stability in humans. We therefore aimed to develop an LC-MS/MS method for detection and quantification of intact [Pyr1]apelin-13 and have used this method to identify the metabolites generated in vivo in humans. [Pyr1]apelin-13 (135 nmol/min) was infused into six healthy human volunteers for 120 minutes and blood collected at time 0 and 120 minutes after infusion. Plasma was extracted in the presence of guanidine hydrochloride and analysed by LC-MS/MS. Here we report a highly sensitive, robust and reproducible method for quantification of intact [Pyr1]apelin-13 and its metabolites in human plasma. Using this method, we showed that the circulating concentration of intact peptide was 58.3 ± 10.5 ng/ml after 120 minutes infusion. We demonstrated for the first time that in humans, [Pyr1]apelin-13 was cleaved from both termini but the C-terminal was more susceptible to cleavage. Consequently, of the metabolites identified, [Pyr1]apelin-13(1–12), [Pyr1]apelin-13(1–10) and [Pyr1]apelin-13(1–6) were the most abundant. These data suggest that apelin peptides designed for use as cardiovascular therapeutics, should include modifications that minimise C-terminal cleavage
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Evaluation of metabolic and cardiovascular actions of [Pyr1]apelin-13 in patients with Type 2 diabetes mellitus
Name: Dr Petra Sulentic
Thesis title: Evaluation of metabolic and cardiovascular actions of [Pyr1]apelin-13 in patients with Type 2 diabetes mellitus
Type 2 diabetes mellitus (T2DM) is common, long term metabolic disorder characterised by hyperglycaemia (high blood glucose) resulting from insulin resistance (IR) and insulin insufficiency. Type 2 diabetes mellitus and related complications present a significant cause of morbidity and mortality and there is an urgent need to identify novel pathways that may ultimately lead to the development of new therapies to improve blood glucose control and prevent development of diabetic complications. Apelin is a naturally occurring peptide investigated in animal models and humans showing beneficial cardiovascular and metabolic properties. Apelin reduces peripheral vascular resistance (PVR) and increases cardiac index (CI) in healthy volunteers and heart failure patients, and also increases insulin sensitivity in overweight participants. Besides health risks connected to hyperglycaemia, T2DM patients have increased cardiovascular risk, therefore modulation of the apelin signalling pathway may provide a novel cardiometabolic therapeutic approach. Systemic studies were planned to investigate the effects of prolonged intravenous apelin infusions on CI, blood glucose and IR for the first time in patients with T2DM, following hypothesis that apelin would induce beneficial cardiovascular and metabolic effects in that patient group. Firstly, the tolerability and safety of apelin in increasing doses were tested in a pilot study including healthy volunteers. Subsequently, eighteen participants with increased body mass index (BMI) 25–34.9 kg/m2 who served as a model of IR and nine T2DM patients underwent a series of randomised, double blind, saline controlled, prolonged infusion studies. [Pyr1]apelin-13 was infused systemically for two hours with measurements of cardiovascular and metabolic parameters. Results showed that in participants with increased BMI, compared to saline control, [Pyr1]apelin-13 in dose 30 nmol/min caused a significant rise in CI and stroke volume index (SVI), whilst reducing peripheral vascular resistance (PVR) and mean arterial pressure (MAP). After a mixed meal, [Pyr1]apelin-13 also significantly reduced plasma C peptide without inducing significant changes in glucose or insulin plasma level. In patients with T2DM, [Pyr1]apelin-13 in the equivalent [Pyr1]apelin-13 dose of 30 nmol/min and compared to saline control, also increased SVI and CI and reduced PVR without affecting MAP and levels of insulin, glucose or C-peptide. In T2DM group a possible delay in gastric emptying was observed, representing a novel finding in this condition and requiring further investigations. In summary, following these pilot studies results, there is a reason to believe that patients with T2DM have the potential to benefit from apelin mediated vasodilation, increased CI and better glucose homeostasis combined with possible delay in gastric emptying. Therefore, targeting apelin signalling represents novel pathways yet to be explored and developed therapeutically with further studies required to investigate those effects
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Development and validation of an LC-MS/MS method for detection and quantification of in vivo derived metabolites of [Pyr1]apelin-13 in humans.
[Pyr1]apelin-13 is the predominant apelin peptide isoform in the human cardiovascular system and plasma. To date, few studies have investigated [Pyr1]apelin-13 metabolism in vivo in rats with no studies examining its stability in humans. We therefore aimed to develop an LC-MS/MS method for detection and quantification of intact [Pyr1]apelin-13 and have used this method to identify the metabolites generated in vivo in humans. [Pyr1]apelin-13 (135 nmol/min) was infused into six healthy human volunteers for 120 minutes and blood collected at time 0 and 120 minutes after infusion. Plasma was extracted in the presence of guanidine hydrochloride and analysed by LC-MS/MS. Here we report a highly sensitive, robust and reproducible method for quantification of intact [Pyr1]apelin-13 and its metabolites in human plasma. Using this method, we showed that the circulating concentration of intact peptide was 58.3 ± 10.5 ng/ml after 120 minutes infusion. We demonstrated for the first time that in humans, [Pyr1]apelin-13 was cleaved from both termini but the C-terminal was more susceptible to cleavage. Consequently, of the metabolites identified, [Pyr1]apelin-13(1-12), [Pyr1]apelin-13(1-10) and [Pyr1]apelin-13(1-6) were the most abundant. These data suggest that apelin peptides designed for use as cardiovascular therapeutics, should include modifications that minimise C-terminal cleavage
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International Union of Basic and Clinical Pharmacology XXX. Structure and pharmacology of the apelin receptor with a recommendation that Elabela/Toddler is a second endogenous peptide ligand.
The predicted protein encoded by the APJ gene discovered in 1993 was originally classified as a class A G protein coupled orphan receptor but was subseqently paired with a novel peptide ligand, apelin-36 in 1998. Substantial research identified a family of shorter peptides activating the apelin receptor including apelin -17, apelin-13 and [Pyr1]apelin-13, with the latter peptide predominating in human plasma and cardiovascular system. A range of pharmacological tools have been developed including radiolabelled ligands, analogues with improved plasma stability, peptides and small molecules including biased agonists and antagonists, leading to the recommendation that the APJ gene be renamed APLNR and encodes the apelin receptor protein. Recently, a second endogenous ligand has been identified and called Elabela/Toddler, a 54 amino acid peptide originally identified in the genomes of fish and humans but misclassified as non-coding. This precursor is also able to be cleaved to shorter sequences (32, 21 and 11 amino acids), and all are able to activate the apelin receptor and are blocked by apelin receptor antagonists. This review summarises the pharmacology of these ligands and the apelin receptor, highlights the emerging physiological and pathophysiological roles in a number of diseases and recommends that Elabela/Toddler is a second endogenous peptide ligand of the apelin receptor protein.British Heart Foundation [TAF 03, ; FS/14/59/31282, CR, FS/17/61/33473]; Medical Research Council [MC_PC_14116]; Wellcome Trust [WT107715/Z/15/Z ], Wellcome Trust Programme in Metabolic and Cardiovascular Disease [096822/Z/11/Z , 203814/Z/16/A]; CRUK [C33616/A27229 ], Cambridge Biomedical Research Centre Biomedical Resources Grant [RG85315], University of Cambridge
International Union of Basic and Clinical Pharmacology XXX. Structure and pharmacology of the apelin receptor with a recommendation that Elabela/Toddler is a second endogenous peptide ligand.
The predicted protein encoded by the APJ gene discovered in 1993 was originally classified as a class A G protein coupled orphan receptor but was subseqently paired with a novel peptide ligand, apelin-36 in 1998. Substantial research identified a family of shorter peptides activating the apelin receptor including apelin -17, apelin-13 and [Pyr1]apelin-13, with the latter peptide predominating in human plasma and cardiovascular system. A range of pharmacological tools have been developed including radiolabelled ligands, analogues with improved plasma stability, peptides and small molecules including biased agonists and antagonists, leading to the recommendation that the APJ gene be renamed APLNR and encodes the apelin receptor protein. Recently, a second endogenous ligand has been identified and called Elabela/Toddler, a 54 amino acid peptide originally identified in the genomes of fish and humans but misclassified as non-coding. This precursor is also able to be cleaved to shorter sequences (32, 21 and 11 amino acids), and all are able to activate the apelin receptor and are blocked by apelin receptor antagonists. This review summarises the pharmacology of these ligands and the apelin receptor, highlights the emerging physiological and pathophysiological roles in a number of diseases and recommends that Elabela/Toddler is a second endogenous peptide ligand of the apelin receptor protein.British Heart Foundation [TAF 03, ; FS/14/59/31282, CR, FS/17/61/33473]; Medical Research Council [MC_PC_14116]; Wellcome Trust [WT107715/Z/15/Z ], Wellcome Trust Programme in Metabolic and Cardiovascular Disease [096822/Z/11/Z , 203814/Z/16/A]; CRUK [C33616/A27229 ], Cambridge Biomedical Research Centre Biomedical Resources Grant [RG85315], University of Cambridge