RESUMO
Liraglutide has been extensively applied in the treatment of type 2 diabetes mellitus (T2DM), but its 11-15 h half-life resulted in daily administration, which led to poor patient compliance. This study aimed to solve this problem by developing liraglutide-loaded microspheres with a 1 month sustained release prepared by the W1/O/W2 method combined with the premix membrane emulsification technique to improve therapeutic efficacy. Remarkably, we found that the amphiphilic properties of liraglutide successfully reduced the oil-water interfacial tension, resulting in a stable primary emulsion and decreasing the level of drug leakage into the external water phase. As a result, exceptional drug loading (>8%) and encapsulation efficiency (>85%) of microspheres were achieved. Furthermore, the uniformity in microsphere size facilitated an in-depth exploration of the structural characteristics of liraglutide-loaded microspheres. The results indicated that the dimensions of the internal cavities of the microspheres were significantly influenced by the size of the inner water droplets in the primary emulsion. A denser and more uniform cavity structure decreased the initial burst release, improving the release process of liraglutide from the microspheres. To evaluate the release behavior of liraglutide from microspheres, a set of in vitro release assays and in vivo pharmacodynamics were performed. The liraglutide-loaded microspheres effectively decreased fasting blood glucose (FBG) levels and hemoglobin A1c (HbA1c) levels while enhancing the pancreatic and hepatic functions in db/db mice. In conclusion, liraglutide sustained-release microspheres showed the potential for future clinical applications in the management of T2DM and provided an effective therapeutic approach to overcoming patient compliance issues.
Assuntos
Preparações de Ação Retardada , Diabetes Mellitus Tipo 2 , Liraglutida , Microesferas , Liraglutida/química , Liraglutida/farmacologia , Diabetes Mellitus Tipo 2/tratamento farmacológico , Animais , Preparações de Ação Retardada/química , Preparações de Ação Retardada/farmacocinética , Preparações de Ação Retardada/farmacologia , Hipoglicemiantes/química , Hipoglicemiantes/administração & dosagem , Hipoglicemiantes/uso terapêutico , Hipoglicemiantes/farmacologia , Camundongos , Glicemia/efeitos dos fármacos , Glicemia/análise , Diabetes Mellitus Experimental/tratamento farmacológico , Masculino , Liberação Controlada de Fármacos , Emulsões/química , Tamanho da PartículaRESUMO
OBJECTIVE: The purpose of this study was to explore the effect of liraglutide on pancreatic islet ß cell apoptosis in rats with type 2 diabetes mellitus (T2DM) and the potential mechanisms. MATERIALS AND METHODS: SD rats were randomly divided into control group, model group, and liraglutide groups (200 and 100 µg/(kg·d)). Rats were fed with high sugar and high-fat diet for 8 weeks, and then streptozotocin (STZ) 40 mg/kg was intraperitoneally injected to establish T2DM model. After successful modeling, rats in the intervention group were given liraglutide through subcutaneous injection for 6 weeks. The indexes of glucose metabolism and lipid metabolism were measured. Apoptosis of islet ß cells was detected by TUNEL. Western blot and RT-PCR were used to detect the protein and mRNA expression levels of IKK ε, NF-κ B, Bcl-2, Bax, IL-6, and Gal-3 in pancreatic tissue. RESULTS: Compared with the control group, the serum FPG, INS, HOMA-IR, TC, TG, LDL-C, IL-6, islet apoptosis rate, glucagon, the positive expression rate of Gal-3, and body weight in the T2DM group were all significantly increased (p<0.05). However, the levels of insulin, SOD, HDL, and HOMA-ß were notably decreased in the T2DM group in comparison with the control group (p<0.05). Moreover, the mRNA and protein expression levels of IKKε, NF-κB, Bax, IL-6, and Bax/Bcl-2 were markedly increased in pancreatic tissue (p<0.05). After liraglutide treatment, these changes were reversed in a dose-dependent manner. CONCLUSIONS: Liraglutide improves pancreatic islet ß cell apoptosis in rats with type 2 diabetes mellitus by inhibiting the IKKε/NF-κB pathway.
Assuntos
Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Tipo 2/tratamento farmacológico , Hipoglicemiantes/farmacologia , Células Secretoras de Insulina/efeitos dos fármacos , Ilhotas Pancreáticas/efeitos dos fármacos , Liraglutida/farmacologia , Animais , Apoptose/efeitos dos fármacos , Diabetes Mellitus Experimental/induzido quimicamente , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 2/induzido quimicamente , Diabetes Mellitus Tipo 2/metabolismo , Hipoglicemiantes/administração & dosagem , Hipoglicemiantes/química , Injeções Intraperitoneais , Injeções Subcutâneas , Células Secretoras de Insulina/fisiologia , Peptídeos e Proteínas de Sinalização Intracelular/antagonistas & inibidores , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Ilhotas Pancreáticas/patologia , Liraglutida/administração & dosagem , Liraglutida/química , Masculino , NF-kappa B/antagonistas & inibidores , NF-kappa B/metabolismo , Ratos , Ratos Sprague-Dawley , Estreptozocina/administração & dosagemRESUMO
Peptide and protein drug molecules fold into higher order structures (HOS) in formulation and these folded structures are often critical for drug efficacy and safety. Generic or biosimilar drug products (DPs) need to show similar HOS to the reference product. The solution NMR spectroscopy is a non-invasive, chemically and structurally specific analytical method that is ideal for characterizing protein therapeutics in formulation. However, only limited NMR studies have been performed directly on marketed DPs and questions remain on how to quantitively define similarity. Here, NMR spectra were collected on marketed peptide and protein DPs, including calcitonin-salmon, liraglutide, teriparatide, exenatide, insulin glargine and rituximab. The 1D 1H spectral pattern readily revealed protein HOS heterogeneity, exchange and oligomerization in the different formulations. Principal component analysis (PCA) applied to two rituximab DPs showed consistent results with the previously demonstrated similarity metrics of Mahalanobis distance (DM) of 3.3. The 2D 1H-13C HSQC spectral comparison of insulin glargine DPs provided similarity metrics for chemical shift difference (Δδ) and methyl peak profile, i.e., 4 ppb for 1H, 15 ppb for 13C and 98% peaks with equivalent peak height. Finally, 2D 1H-15N sofast HMQC was demonstrated as a sensitive method for comparison of small protein HOS. The application of NMR procedures and chemometric analysis on therapeutic proteins offer quantitative similarity assessments of DPs with practically achievable similarity metrics.
Assuntos
Peptídeos/química , Preparações Farmacêuticas/química , Proteínas/química , Calcitonina/química , Exenatida/química , Insulina Glargina/química , Liraglutida/química , Ressonância Magnética Nuclear Biomolecular/métodos , Conformação Proteica , Rituximab/química , Teriparatida/químicaRESUMO
Biologics such as peptides and proteins possess a number of attractive attributes that make them particularly valuable as therapeutics, including their high activity, high specificity, and low toxicity. However, one of the key challenges associated with this class of drugs is their propensity to aggregate. Given the safety and immunogenicity concerns related to polypeptide aggregates, it is particularly important to sensitively detect aggregates in therapeutic drug formulations as part of the quality control process. Here, we report the development of conformation-specific antibodies that recognize polypeptide aggregates composed of a GLP-1 receptor agonist (liraglutide) and their integration into a sensitive immunoassay for detecting liraglutide amyloid fibrils. We sorted single-chain antibody libraries against liraglutide fibrils using yeast surface display and magnetic-activated cell sorting, and identified several antibodies with high conformational specificity. Interestingly, these antibodies cross-react with amyloid fibrils formed by several other polypeptides, revealing that they recognize molecular features common to different types of fibrils. Moreover, we find that our immunoassay using these antibodies is >50-fold more sensitive than the conventional method for detecting liraglutide aggregation (Thioflavin T fluorescence). We expect that our systematic approach for generating a sensitive, aggregate-specific immunoassay can be readily extended to other biologics to improve the quality and safety of formulated drug products.
Assuntos
Amiloide/química , Evolução Molecular Direcionada , Composição de Medicamentos , Peptídeo 1 Semelhante ao Glucagon/química , Liraglutida/química , Agregados Proteicos , Anticorpos de Cadeia Única/química , Humanos , Anticorpos de Cadeia Única/genéticaRESUMO
The delivery of peptides or protein drugs via the oral route has always presented a significant challenge. Here, nanoparticles for the oral delivery of liraglutide are prepared. The nanoparticles are composed of the biodegradable carrier materials chitosan and poly-N-(2-hydroxypropyl) methacrylamide (pHPMA). In addition, CSKSSDYQC (CSK) and hemagglutinin-2 (HA2) are introduced into the particles to improve the in vivo bioavailability of liraglutide. The size of the nanoparticles is less than 200 nm, and the encapsulation efficiency is approximately 80%. Compared with the subcutaneously injected liraglutide solution group (100%), the relative bioavailability of the nanoparticle group modified with CSK and HA2 reached 10.12%, which is 2.53 times that of the oral liraglutide solution group. In vivo imaging results showed that pHPMA/HA2-CSK chitosan nanoparticles (pHPMA/HA-CCNPs) are retained in the gastrointestinal tract for up to 12 h, which is beneficial for oral absorption. CSK and HA2 modified pHPMA/chitosan nanoparticles significantly improved liraglutide oral bioavailability and therefore have the potential to be applied for oral administration of peptides and proteins.
Assuntos
Acrilamidas/química , Quitosana/química , Hemaglutininas/química , Liraglutida/química , Administração Oral , Animais , Disponibilidade Biológica , Portadores de Fármacos/química , Trato Gastrointestinal/efeitos dos fármacos , Espectroscopia de Ressonância Magnética , Camundongos , Nanomedicina/métodos , Nanopartículas/química , Tamanho da Partícula , Peptídeos/química , Ratos , Ratos Sprague-DawleyRESUMO
A simple, fast and high-throughput LC-tandem mass spectrometry method was developed and validated to simultaneously measure liraglutide and insulin degludec in rat plasma. After protein precipitation, plasma samples were subjected to gradient elution using an InertSustain Bio C18 column with 1000/20/1 water/acetonitrile/formic acid (v/v/v) and 1000/1 acetonitrile/formic acid (v/v) as the mobile phase. The method was validated from 1.00 to 500 ng/mL of liraglutide and insulin degludec. Further, the extraction recovery from the plasma was 41.8%-49.2% for liraglutide and 56.5%-69.7% for insulin degludec. Intra- and inter-day precision of liraglutide was 3.5%-9.4% and 8.4%-9.8%, respectively, whereas its accuracy was between -12.6% and -1.3%. Intra- and inter-day precision of insulin degludec was 5.2%-13.6% and 11.8%-19.1%, respectively, showing an accuracy between -3.0% and 9.9%. As a result, the method was successfully applied to a pharmacokinetics study of liraglutide and insulin degludec following a single-dose subcutaneous administration to rats.
Assuntos
Cromatografia Líquida/métodos , Insulina de Ação Prolongada/sangue , Liraglutida/sangue , Espectrometria de Massas em Tandem/métodos , Animais , Estabilidade de Medicamentos , Insulina de Ação Prolongada/química , Insulina de Ação Prolongada/farmacocinética , Limite de Detecção , Modelos Lineares , Liraglutida/química , Liraglutida/farmacocinética , Ratos , Reprodutibilidade dos TestesRESUMO
PURPOSE: To design and stabilize Liraglutide loaded poly (lactic-co-glycolic acid) nanoparticles (PLGA NPs) proper for oral administration. METHODS: PLGA NPs were prepared by means of double emulsion solvent evaporation method and optimized by applying 7-factor 2-level Plackett-Burman screening design. RESULTS: Spherical shaped NPs with homogeneous distribution, 188.95 nm particle size and 51.81% encapsulation efficiency were obtained. Liraglutide was successfully entrapped in the NPs while maintaining its native amorphous nature, and its structural integrity as well. CONCLUSION: Lira-PLGA NPs with the required Critical Quality Attributes (CQAs) were successfully designed by implementing a 7-factor 8-run Plackett Burman design into the extended Quality by Design (QbD) model, to elucidate the effect of formulation and process variables on the particle size, size-distribution, encapsulation efficiency and surface charge. As the developed nanoparticles maintained the native structure of the active pharmaceutical ingredient (API), they are promising compositions for the further development for the oral delivery of Lira. Graphical Abstract.
Assuntos
Portadores de Fármacos/química , Hipoglicemiantes/química , Liraglutida/química , Nanopartículas/química , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/química , Administração Oral , Liberação Controlada de Fármacos , Emulsões , Hipoglicemiantes/administração & dosagem , Liraglutida/administração & dosagem , Tamanho da PartículaRESUMO
Chitosan coated calcium-alginate nanocapsules were developed for oral sustained delivery of liraglutide. The effect of coating components including sodium alginate, calcium chloride, and chitosan concentrations on the particle size was studied based on response surface methodology. The beads were characterized by dynamic light scattering, scanning and transmission electron microscopy as well as Fourier transform infrared spectroscopy. It was shown that the diameter of the formed beads was most dependent on the encapsulation technique and alginate concentration. SEM revealed spherical and smooth particles of up to 100â¯nm diameter for the optimum composition of alginate 0.5%, chitosan 0.5% and calcium chloride 0.5% in the volume ratio of 3:1:1. The resulting bead formulation had a loading efficiency of 92.5% and loading capacity of 54.16%. The nanocapsules exhibited stability of 92.4% and 72.3% over freeze drying and subsequent 60â¯days storage at 4⯰C, respectively. In-vitro release studies in simulated gastrointestinal conditions were carried out in a sequential technique and the amount of drug release was found to be 59.1% after 6â¯h. The results of this study demonstrated that chitosan coated calcium-alginate nanocapsules hold promise as a potential natural biodegradable polymer-based oral carrier of liraglutide for better management of diabetes.
Assuntos
Quitosana/química , Diabetes Mellitus/tratamento farmacológico , Composição de Medicamentos , Liraglutida/uso terapêutico , Administração Oral , Alginatos/química , Alginatos/uso terapêutico , Plásticos Biodegradáveis/química , Plásticos Biodegradáveis/uso terapêutico , Quitosana/uso terapêutico , Portadores de Fármacos , Humanos , Liraglutida/química , Nanocápsulas/química , Nanocápsulas/uso terapêutico , Polímeros/química , Polímeros/uso terapêutico , Espectroscopia de Infravermelho com Transformada de FourierRESUMO
Risk Assessment (RA) is the key element of the Quality by Design (QbD) approach recommended by the pharmaceutical regulatory bodies. This research paper aimed to implement the regulatory requirements, the QbD thinking and the RA from the first steps of the oral peptide formulation development. The authors intended to give a general recommendation about the application possibilities of this methodology, to demonstrate the risk factors and the required decision points. Later, this paper presents a concrete development in practice. This case study shows the QbD and RA based early phase development of the GLP 1 analog, Liraglutide, an antidiabetic peptide drug mainly used in the treatment of type 2 Diabetes Mellitus. The objective here was to design Liraglutide encapsulated polymeric nanoparticles for oral delivery and the progress of their RA based development is presented. In this case, the particle size, the encapsulation efficiency, and the drug loading were found as the most critical quality attributes, the polymer concentration, the drug concentration, the w2/o ratio, the stabilizer concentration and polymer type were identified by the criticality rating as having the greatest impact on the product quality among the critical material attributes, finally the sonication time and sonication power were selected as the most critical elements of the production process. The results showed the importance of the risk factor-focused development in the oral peptide pharmaceutical formulations, and underlined the importance of the profound planning phase even in such cases. The formulation of an oral peptide delivery system is associated with several risks, but their priority ranking helps to focus on the resources (human, financial, time) related to the final product quality aimed at.
Assuntos
Sistemas de Liberação de Medicamentos , Desenho de Fármacos , Hipoglicemiantes/química , Liraglutida/química , Nanopartículas/química , Química Farmacêutica , Peptídeos , Polímeros/química , Medição de RiscoRESUMO
A simple, sensitive and selective LC-MS/MS method was developed for the quantitative analysis of liraglutide and validated in rat plasma. Human insulin was used as the internal standard. After a simple protein precipitation step, liraglutide was chromatographically separated using an InertSustain Bio C18 column with mobile phases comprising acetonitrile with 0.1% formic acid (A) and water with 0.1% formic acid (B). Detection was achieved using positive ion electrospray ionization in multiple-reaction monitoring (MRM) mode. Good linearity was observed in the concentration range 0.5-250â¯ng/mL (r2â¯>â¯0.99). The intra- and inter-day precision values (expressed as relative standard deviation, RSD) of liraglutide ranged from 1.97-7.63% and 5.25-11.9, respectively. The accuracy (expressed as relative error, RE) ranged from -8.79-11.4%. Both the recovery and matrix effect were within acceptable limits. This method was successfully applied for the pharmacokinetics study of liraglutide in rats after subcutaneous administration.
Assuntos
Cromatografia Líquida/métodos , Liraglutida/sangue , Liraglutida/farmacocinética , Espectrometria de Massas em Tandem/métodos , Animais , Estabilidade de Medicamentos , Feminino , Modelos Lineares , Liraglutida/química , Masculino , Ratos , Ratos Sprague-Dawley , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
The long-acting glucagon-like peptide-1 analogue liraglutide has proven efficiency in the management of type 2 diabetes and also has beneficial effects on cardiovascular diseases. Liraglutide's protracted action highly depends on its capacity to bind to albumin via its palmitic acid part. However, in diabetes, albumin can undergo glycation, resulting in impaired drug binding. Our objective in this study was to assess the impact of human serum albumin (HSA) glycation on liraglutide affinity. Using fluorine labeling of the drug and 19F NMR, we determined HSA affinity for liraglutide in two glycated albumin models. We either glycated HSA in vitro by incubation with glucose (G25- or G100-HSA) or methylglyoxal (MGO-HSA) or purified in vivo glycated HSA from the plasma of diabetic patients with poor glycemic control. Nonglycated commercial HSA (G0-HSA) and HSA purified from plasma of healthy individuals served as controls. We found that glycation decreases affinity for liraglutide by 7-fold for G100-HSA and by 5-fold for MGO-HSA compared with G0-HSA. A similarly reduced affinity was observed for HSA purified from diabetic individuals compared with HSA from healthy individuals. Our results reveal that glycation significantly impairs HSA affinity to liraglutide and confirm that glycation contributes to liraglutide's variable therapeutic efficiency, depending on diabetes stage. Because diabetes is a progressive disease, the effect of glycated albumin on liraglutide affinity found here is important to consider when diabetes is managed with this drug.
Assuntos
Peptídeo 1 Semelhante ao Glucagon , Liraglutida/química , Albumina Sérica Humana/química , Diabetes Mellitus/sangue , Glicosilação , Humanos , Liraglutida/farmacocinética , Ressonância Magnética Nuclear Biomolecular , Ligação Proteica , Albumina Sérica Humana/metabolismoRESUMO
Glucagon-like peptide-1 (GLP-1) is secreted by intestinal L-cells following food intake, and plays an important role in glucose homeostasis due to its stimulation of glucose-dependent insulin secretion. Further, GLP-1 is also associated with protective effects on pancreatic ß-cells and the cardiovascular system, decreased appetite, and weight loss, making GLP-1 derivatives an exciting treatment for typeâ 2 diabetes and obesity. Despite these benefits, wild-type GLP-1 exhibits a short circulation time due to its poor metabolic stability and rapid renal clearance, and must be administered by injection, making it a poor therapeutic agent. Many strategies have been used to improve the circulation time of GLP-1 (e.g., mutations, unnatural amino acids, depot formulations, use of exendin-4 sequences, and fusions with high-molecular-weight proteins or polymers), with its therapeutic utility further improved by adding agonist activity for gastric inhibitory peptide and glucagon receptors. This minireview focuses on strategies that have been used to improve the pharmacokinetics of GLP-1 and provides an overview of GLP-1-based therapeutics in the pipeline.
Assuntos
Diabetes Mellitus Tipo 2/tratamento farmacológico , Peptídeo 1 Semelhante ao Glucagon/uso terapêutico , Hipoglicemiantes/uso terapêutico , Animais , Preparações de Ação Retardada , Exenatida/administração & dosagem , Exenatida/uso terapêutico , Peptídeo 1 Semelhante ao Glucagon/agonistas , Peptídeo 1 Semelhante ao Glucagon/análogos & derivados , Peptídeo 1 Semelhante ao Glucagon/farmacocinética , Meia-Vida , Humanos , Hipoglicemiantes/química , Hipoglicemiantes/farmacocinética , Liraglutida/química , Liraglutida/uso terapêutico , Hormônios Peptídicos/química , Hormônios Peptídicos/farmacocinética , Hormônios Peptídicos/uso terapêutico , Relação Estrutura-AtividadeRESUMO
INTRODUCTION: Type 2 diabetes (T2DM) is associated with significant morbidity and mortality. Obesity is one of the main risk factors for T2DM and its management requires a multidisciplinary approach, which may include pharmacotherapy. AREAS COVERED: In this paper, data on efficacy, tolerability and safety of FDA-approved pharmacotherapies for obesity (orlistat, phentermine/topiramate extended-release, lorcaserin, bupropion sustained release/naltrexone sustained release and liraglutide) are reviewed, focusing on individuals with type 2 diabetes. EXPERT OPINION: Obesity is the major pathophysiologic driver of T2DM; conversely 5-10% weight loss leads to significant improvement in glycemic control, lipids and blood pressure. Weight loss maintenance is difficult with lifestyle interventions alone and may require adjunctive therapies. There is good evidence for the efficacy and tolerability of approved anti-obesity pharmacotherapies in individuals with T2DM, with current cardiovascular safety data being most favorable for liraglutide, orlistat and lorcaserin. Given the link between obesity and T2DM, a weight-centric therapeutic approach including use of weight reducing anti-diabetic therapies, and anti-obesity pharmacotherapies is both intuitive and rational to improve glycemic and other metabolic outcomes in patients with T2DM.
Assuntos
Fármacos Antiobesidade/uso terapêutico , Diabetes Mellitus Tipo 2/complicações , Obesidade/tratamento farmacológico , Fármacos Antiobesidade/química , Fármacos Antiobesidade/farmacologia , Benzazepinas/química , Benzazepinas/farmacologia , Benzazepinas/uso terapêutico , Ensaios Clínicos como Assunto , Composição de Medicamentos , Humanos , Lactonas/química , Lactonas/farmacologia , Lactonas/uso terapêutico , Liraglutida/química , Liraglutida/farmacologia , Liraglutida/uso terapêutico , Orlistate , Fentermina/química , Fentermina/farmacologia , Fentermina/uso terapêutico , Redução de Peso/efeitos dos fármacosRESUMO
To develop novel long-acting antidiabetics with improved therapeutic efficacy, two glucagon-like peptide-1 (GLP-1) analogs were constructed through the hybridization of key sequences of GLP-1, xenGLP-1B, exendin-4, and lixisenatide. Hybrids 1 and 2 demonstrated enhanced in vitro and in vivo biological activities and were further site-specifically lipidized at lysine residues to achieve prolonged duration of action and less frequent administration. Compared with their native peptides, compounds 3-6 showed similar in vitro activities but impaired in vivo acute hypoglycemic potencies due to decreased aqueous solubility and retarded absorption in vivo. To circumvent these issues, compound 3 (xenoglutide) was selected to be self-associated with sterically stabilized micelles (SSM). The α-helix and solubility of xenoglutide were significantly improved after self-associated with SSM. Notably, the improved physicochemical characteristics of xenoglutide-SSM led to revival of acute hypoglycemic ability without affecting its long-term glucose-lowering activity. Most importantly, preclinical studies demonstrated improved therapeutic effects and safety of xenoglutide-SSM in diabetic db/db mice. Our work suggests the SSM incorporation as an effective approach to improve the pharmacokinetic and biological properties of hydrophobicity peptide drugs. Furthermore, our data clearly indicate xenoglutide-SSM as a novel nanomedicine for the treatment of type 2 diabetics.
Assuntos
Diabetes Mellitus Tipo 2/tratamento farmacológico , Ácidos Graxos/química , Peptídeo 1 Semelhante ao Glucagon/química , Hipoglicemiantes/uso terapêutico , Nanomedicina/métodos , Animais , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Ingestão de Alimentos/efeitos dos fármacos , Exenatida , Peptídeo 1 Semelhante ao Glucagon/uso terapêutico , Teste de Tolerância a Glucose , Humanos , Hipoglicemiantes/química , Liraglutida/química , Liraglutida/uso terapêutico , Masculino , Camundongos , Micelas , Peptídeos/química , Peptídeos/uso terapêutico , Peçonhas/química , Peçonhas/uso terapêuticoRESUMO
Liraglutide is a glucagon-like peptide-1 analog for the treatment of type 2 diabetes. Major interference in plasma of human and animals and low fragment signal in tandem mass spectrometry are the main difficulties encountered in the bioanalysis of liraglutide. In this study, by combining differential mobility spectrometry (DMS) with multiple ion monitoring detection (MIM), a liquid chromatography differential mobility spectrometry tandem mass spectrometry with multiple ion monitoring detection (LC-DMS-MIM) method was developed for the quantitation of liraglutide in dog plasma. Mixed anion-exchange solid-phase extraction was used for sample preparation. The parameters of DMS were meticulously optimized to increase the signal-to-noise ratio of the analyte. The assay was linear in the range 1-100 ng/mL with good accuracy and precision. The lower limit of quantitation (LLOQ, the lowest standard on the calibration curve) of this method was 1 ng/mL. The research reveals that DMS is an effective tool for the elimination of interference in bioanalysis and that LC-DMS-MIM has better specificity and higher signal-to-noise ratio than classical liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the bioanalysis of liraglutide. Graphical abstract Process for the bioanalysis of liraglutide by liquid chromatography differential mobility spectrometry tandem mass spectrometry with multiple ion monitoring detection.
Assuntos
Hipoglicemiantes/análise , Liraglutida/química , Espectrometria de Massas em Tandem/métodos , Animais , Cães , Limite de Detecção , Reprodutibilidade dos TestesRESUMO
In this work, we sought to generate sustained-release injectable microspheres loaded with the GLP-1 analogue liraglutide. Using water-in-oil-in-water double emulsion methods, poly(lactic-co-glycolic acid) (PLGA) microspheres loaded with liraglutide were prepared. The microspheres gave sustained drug release over 30days, with cumulative release of up to 90% reached in vitro. The microspheres were further studied in a rat model of diabetes, and their performance compared with a group given daily liraglutide injections. Reduced blood sugar levels were seen in the microsphere treatment groups, with the results being similar to those obtained with conventional injections between 10 and 25days after the commencement of treatment. After 5 and 30days of treatment, the microspheres seem a little slower to act than the injections. The pathology of the rats' spleen, heart, kidney and lungs was probed after the 30-day treatment period, and the results indicated that the microspheres were safe and had beneficial effects on the liver, reducing the occurrence of fatty deposits seen in untreated diabetic rats. Moreover, in terms of liver, renal and cardiac functions, and blood lipid and antioxidant levels, the microspheres were as effective as the injections. The expression of several proteases linked to the metabolism of aliphatic acids and homocysteine was promoted by the microsphere formulations. Inflammatory markers in the microsphere treatment groups were somewhat higher than the injection group, however. The liraglutide/PLGA microspheres prepared in this work are overall shown to be efficacious in a rat model of diabetes, and we thus believe they have strong potential for clinical use.
Assuntos
Diabetes Mellitus Experimental/tratamento farmacológico , Sistemas de Liberação de Medicamentos , Hipoglicemiantes/administração & dosagem , Ácido Láctico/química , Liraglutida/administração & dosagem , Microesferas , Ácido Poliglicólico/química , Animais , Glicemia/análise , Preparações de Ação Retardada/administração & dosagem , Preparações de Ação Retardada/química , Preparações de Ação Retardada/uso terapêutico , Diabetes Mellitus Experimental/sangue , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/tratamento farmacológico , Composição de Medicamentos , Liberação Controlada de Fármacos , Hemoglobinas Glicadas/análise , Hipoglicemiantes/química , Hipoglicemiantes/uso terapêutico , Insulina/sangue , Liraglutida/química , Liraglutida/uso terapêutico , Masculino , Copolímero de Ácido Poliláctico e Ácido Poliglicólico , Ratos , Ratos Sprague-DawleyRESUMO
Subcutaneous liraglutide-loaded multivesicular liposomes (Lrg-MVLs) were developed as a sustained drug-delivery system for treating diabetes and their properties were characterized. The Lrg-MVLs prepared using a two-step water-in-oil-in-water double emulsification process had a spherical appearance with a mean diameter of 6.69 µm and an encapsulation efficiency of 82.23 ± 4.78% without any initial burst release. Their pharmacodynamics (PD) and pharmacokinetics (PK) were also studied after a single subcutaneous administration to Sprague-Dawley (SD) rats with diabetes. The PD results demonstrated that Lrg-MVLs presented sustained glucose-lowering effects for nearly a week, while the pharmacokinetic parameters showed that the plasma liraglutide concentration of the designed preparation produced Cmax of 81.979 ± 12.140 pg/ml and an MRT0-t of 88.224 ± 3.893 h. Furthermore, retention of Lrg-MVLs at the injection site was studied semiquantitatively by an in vivo imaging system, which can be used to evaluate the drug release from MVLs in vivo. In conclusion, MVLs are a promising carrier for liraglutide and Lrg-MVLs deserve further study for the treatment of diabetes.
Assuntos
Glicemia/efeitos dos fármacos , Preparações de Ação Retardada/administração & dosagem , Preparações de Ação Retardada/química , Diabetes Mellitus Experimental/tratamento farmacológico , Lipossomos/química , Liraglutida/administração & dosagem , Liraglutida/química , Administração Cutânea , Animais , Diabetes Mellitus Experimental/sangue , Portadores de Fármacos/química , Sistemas de Liberação de Medicamentos/métodos , Liberação Controlada de Fármacos , Lipossomos/administração & dosagem , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
Incretin mimetics are set to become a mainstay of typeâ 2 diabetes treatment. By acting on the pancreas and brain, they potentiate insulin secretion and induce weight loss to preserve normoglycemia. Despite this, incretin therapy has been associated with off-target effects, including nausea and gastrointestinal disturbance. A novel photoswitchable incretin mimetic based upon the specific glucagon-like peptide-1 receptor (GLP-1R) agonist liraglutide was designed, synthesized, and tested. This peptidic compound, termed LirAzo, possesses an azobenzene photoresponsive element, affording isomer-biased GLP-1R signaling as a result of differential activation of second messenger pathways in response to light. While the trans isomer primarily engages calcium influx, the cis isomer favors cAMP generation. LirAzo thus allows optical control of insulin secretion and cell survival.
Assuntos
Incretinas/química , Insulina/metabolismo , Liraglutida/farmacologia , Sequência de Aminoácidos , Animais , Células CHO , Linhagem Celular Transformada , Cricetinae , Cricetulus , Diabetes Mellitus Tipo 2/tratamento farmacológico , Receptor do Peptídeo Semelhante ao Glucagon 1/agonistas , Receptor do Peptídeo Semelhante ao Glucagon 1/metabolismo , Secreção de Insulina , Liraglutida/química , Liraglutida/uso terapêutico , Camundongos , Mimetismo Molecular , Dados de Sequência Molecular , Transdução de SinaisRESUMO
The glucagon-like peptide 1 (GLP-1) analog, liraglutide, is a GLP-1 agonist and is used in the treatment of type-2 diabetes mellitus and obesity. From a pharmaceutical perspective, it is important to know the oligomerization state of liraglutide with respect to stability. Compared to GLP-1, liraglutide has an added fatty acid (FA) moiety that causes oligomerization of liraglutide as suggested by small-angle x-ray scattering (SAXS) and multiangle static light scattering (MALS) results. SAXS data suggested a global shape of a hollow elliptical cylinder of size hexa-, hepta-, or octamer, whereas MALS data indicate a hexamer. To elaborate further on the stability of these oligomers and the role of the FA chains, a series of molecular-dynamics simulations were carried out on 11 different hexa-, hepta-, and octameric systems. Our results indicate that interactions of the fatty acid chains contribute noticeably to the stabilization. The simulation results indicate that the heptamer with paired FA chains is the most stable oligomer when compared to the 10 other investigated structures. Theoretical SAXS curves extracted from the simulations qualitatively agree with the experimentally determined SAXS curves supporting the view that liraglutide forms heptamers in solution. In agreement with the SAXS data, the heptamer forms a water-filled oligomer of elliptical cylindrical shape.
Assuntos
Liraglutida/química , Sequência de Aminoácidos , Interações Hidrofóbicas e Hidrofílicas , Liraglutida/metabolismo , Simulação de Dinâmica Molecular , Multimerização Proteica , Espalhamento a Baixo Ângulo , Solventes/química , Água/química , Difração de Raios XRESUMO
AIMS/HYPOTHESIS: We assessed the contribution of glucagon-like peptide-1 (GLP-1) receptor (GLP-1R) signalling to thermogenesis induced by high-fat diet (HFD) consumption. Furthermore, we determined whether brown adipose tissue (BAT) activity contributes to weight loss induced by chronic subcutaneous treatment with the GLP-1R agonist, liraglutide, in a model of diet-induced obesity. METHODS: Metabolic phenotyping was performed using indirect calorimetry in wild-type (WT) and Glp1r-knockout (KO) mice during chow and HFD feeding at room temperature and at thermoneutrality. In a separate study, we investigated the contribution of BAT thermogenic capacity to the weight lowering effect induced by GLP-1 mimetics by administering liraglutide (10 or 30 nmol kg(-1) day(-1) s.c.) to diet-induced obese (DIO) mice for 6 or 4 weeks, respectively. In both studies, animals were subjected to a noradrenaline (norepinephrine)-stimulated oxygen consumption [Formula: see text] test. RESULTS: At thermoneutrality, HFD-fed Glp1r-KO mice had similar energy expenditure (EE) compared with HFD-fed WT controls. However, HFD-fed Glp1r-KO mice exhibited relatively less EE when housed at a cooler standard room temperature, and had relatively lower [Formula: see text] in response to a noradrenaline challenge, which is consistent with impaired BAT thermogenic capacity. In contrast to the loss of function model, chronic peripheral liraglutide treatment did not increase BAT activity as determined by noradrenaline-stimulated [Formula: see text] and BAT gene expression. CONCLUSIONS/INTERPRETATION: These data suggest that although endogenous GLP-1R signalling contributes to increased BAT thermogenesis, this mechanism does not play a significant role in the food intake-independent body weight lowering effect of the GLP-1 mimetic liraglutide in DIO mice.