Oral Delivery Of Peptides Research Articles

Solid lipid particles for oral delivery of peptide and protein drugs III - the effect of fed state conditions on the in vitro release and degradation of desmopressin.

The effect of food intake on the release and degradation of peptide drugs from solid lipid particles is unknown and was therefore investigated in vitro using different fed state media in a lipolysis model. Desmopressin was used as a model peptide and incorporated into solid lipid particles consisting of trimyristin (TG14), tripalmitin (TG16), and tristearin (TG18), respectively. Fasted state and fed state media with varying phospholipid and bile salt concentrations, as well as fed state media with milk and oleic acid glycerides, respectively, were used as the release media. The presence of oleic acid glycerides accelerated the release of desmopressin significantly from all solid lipid particles both in the presence and absence of lipase. The presence of oleic acid glycerides also reduced the degradation rate of desmopressin, probably due to the interactions between the lipids and the protease or desmopressin. Addition of a medium chain triglyceride, trilaurin, in combination with drug-loaded lipid particles diminished the food effect on the TG18 particles, and trilaurin is therefore proposed to be a suitable excipient for reduction of the food effect. Overall, the present study shows that strategies to reduce food effect, such as adding trilaurin, for lipid particle formulations should be considered as drug release from such formulations might be influenced by the presence of food in the gastrointestinal tract.

Chitosan-modified porous silicon microparticles for enhanced permeability of insulin across intestinal cell monolayers

Porous silicon (PSi) based particulate systems are emerging as an important drug delivery system due to its advantageous properties such as biocompatibility, biodegradability and ability to tailor the particles' physicochemical properties. Here, annealed thermally hydrocarbonized PSi (AnnTHCPSi) and undecylenic acid modified AnnTHCPSi (AnnUnTHCPSi) microparticles were developed as a PSi-based platform for oral delivery of insulin. Chitosan (CS) was used to modify the AnnUnTHCPSi microparticles to enhance the intestinal permeation of insulin. Surface modification with CS led to significant increase in the interaction of PSi microparticles with Caco-2/HT-29 cell co-culture monolayers. Compared to pure insulin, the CS-conjugated microparticles significantly improved the permeation of insulin across the Caco-2/HT-29 cell monolayers, with ca. 20-fold increase in the amount of insulin permeated and ca. 7-fold increase in the apparent permeability (Papp) value. Moreover, among all the investigated particles, the CS-conjugated microparticles also showed the highest amount of insulin associated with the mucus layer and the intestinal Caco-2 cells and mucus secreting HT-29 cells. Our results demonstrate that CS-conjugated AnnUnTHCPSi microparticles can efficiently enhance the insulin absorption across intestinal cells, and thus, they are promising microsystems for the oral delivery of proteins and peptides across the intestinal cell membrane.

Formulation Strategies to Improve Oral Peptide Delivery

Delivery of peptides by the oral route greatly appeals due to commercial, patient convenience and scientific arguments. While there are over 60 injectable peptides marketed worldwide, and many more in development, most delivery strategies do not yet adequately overcome the barriers to oral delivery. Peptides are sensitive to chemical and enzymatic degradation in the intestine, and are poorly permeable across the intestinal epithelium due to sub-optimal physicochemical properties. A successful oral peptide delivery technology should protect potent peptides from presystemic degradation and improve epithelial permeation to achieve a target oral bioavailability with acceptable intra-subject variability. This review provides a comprehensive up-to-date overview of the current status of oral peptide delivery with an emphasis on patented formulations that are yielding promising clinical data.

Poly(ethylene glycol)-Mediated Conformational Alteration of α-Chymotrypsin Prevents Inactivation of Insulin by Stabilizing Active Intermediates

Proteolytic enzymes in the gut represent one of the biggest barriers against oral delivery of therapeutic proteins and peptides. In the current study, we explored the effect of poly(ethylene glycol) 400 (PEG 400), a commonly used crowding agent, on insulin degradation mediated by α-chymotrypsin (α-CT). Without PEG 400, insulin was quickly cleaved by α-CT to generate inactive degradation products. In comparison, incorporation of PEG 400 resulted in reaction mixtures with retained biological activity. The analysis on the conformation of α-CT and the local environment of the enzyme's active site unraveled that PEG 400 altered the conformation of α-CT to prevent the inactivation of insulin via stabilization of active intermediates. These findings indicated that PEG 400 may provide a promising addition toward oral delivery of insulin.

The structure/function of apoprotein A-I mimetic peptides

To review recent advances in our understanding of the mechanism of action of apoprotein A-I (apoA-I) mimetic peptides and improved methods for the oral delivery of peptides. The apoA-I mimetic peptides are based on the structure of the major apoprotein of HDL with the expectation that they may also mimic some of the antiatherogenic functions of HDL. Recent work has provided insight into mechanisms by which they may be antioxidative and anti-inflammatory. In addition, recent work has shifted the focus of the site of action of the mimetic peptides to the small intestine from the plasma and HDL and suggests modulation of bioactive oxidized lipids in the intestine by the peptides may be a major antiatherogenic pathway. The development of transgenic tomatoes expressing an apoA-I mimetic peptide is a significant advance in the oral delivery of peptides as therapies for cardiovascular disease and other chronic inflammatory disorders. In the past year, there have been important advances in the field of apoA-I mimetic peptides, including the oral delivery of bioactive peptides. Further work is required to fully understand the molecular basis for the effect of the peptide on the intestine and bioactive oxidized lipids.

Strategies To Deliver Peptide Drugs to the Brain

Neurological diseases such as neurodegeneration, pain, psychiatric disorders, stroke, and brain cancers would greatly benefit from the use of highly potent and specific peptide pharmaceuticals. Peptides are especially desirable because of their low inherent toxicity. The presence of the blood brain barrier (BBB), their short duration of action, and their need for parenteral administration limits their clinical use. However, over the past decade there have been significant advances in delivering peptides to the central nervous system. Angiopep peptides developed by Angiochem (Montreal, Canada), transferrin antibodies developed by ArmaGen (Santa Monica, USA), and cell penetrating peptides have all shown promise in delivering therapeutic peptides across the BBB after intravenous administration. Noninvasive methods of delivering peptides to the brain include the use of chitosan amphiphile nanoparticles for oral delivery and nose to brain strategies. The uptake of the chitosan amphiphile nanoparticles by the gastrointestinal epithelium is important for oral peptide delivery. Finally protecting peptides from plasma degradation is integral to the success of most of these peptide delivery strategies.

Solid Lipid Particles for Oral Delivery of Peptide and Protein Drugs II – The Digestion of Trilaurin Protects Desmopressin from Proteolytic Degradation

To investigate the in vitro release and degradation of desmopressin from saturated triglyceride microparticles under both lipolytic and proteolytic conditions. The release of desmopressin from different solid lipid microparticles in the absence and presence of a microbial lipase and protease was determined. Trilaurin (TG12), trimyristin (TG14), tripalmitin (TG16), and tristearin (TG18) were used as lipid excipients to produce solid lipid microparticles. In the presence of lipase, the rate of drug release from different lipid particles was in the order of TG14 > TG16 > TG18, which is the same rank order as the lipid degradation rate. A reverse rank order was found for the protection of desmopressin from enzymatic degradation due to spatial separation of desmopressin from the protease. TG12 accelerated the release of desmopressin from all lipid particles when added as either drug-free microparticles to the lipolysis medium or incorporated in TG16 particles. Additionally, TG12 particles protected desmopressin from degradation when present in the lipolysis medium with the other lipid microparticles. TG12 is a very interesting lipid for oral lipid formulations containing peptides and proteins as it alters release and degradation of the incorporated desmopressin. The present study demonstrates the possibility of bio-relevant in vitro evaluation of lipid-based solid particles.

Basics and recent advances in peptide and protein drug delivery.

While the peptide and protein therapeutic market has developed significantly in the past decades, delivery has limited their use. Although oral delivery is preferred, most are currently delivered intravenously or subcutaneously due to degradation and limited absorption in the gastrointestinal tract. Therefore, absorption enhancers, enzyme inhibitors, carrier systems and stability enhancers are being studied to facilitate oral peptide delivery. Additionally, transdermal peptide delivery avoids the issues of the gastrointestinal tract, but also faces absorption limitations. Due to proteases, opsonization and agglutination, free peptides are not systemically stable without modifications. This review discusses oral and transdermal peptide drug delivery, focusing on barriers and solutions to absorption and stability issues. Methods to increase systemic stability and site-specific delivery are also discussed.

Liposomes coated with thiolated chitosan enhance oral peptide delivery to rats

The aim of the present study was the in vivo evaluation of thiomer-coated liposomes for an oral application of peptides. For this purpose, salmon calcitonin was chosen as a model drug and encapsulated within liposomes. Subsequently, the drug loaded liposomes were coated with either chitosan–thioglycolic acid (CS–TGA) or an S-protected version of the same polymer (CS–TGA–MNA), leading to an increase in the particle size of about 500nm and an increase in the zeta potential from approximately −40mV to a maximum value of about +44mV, depending on the polymer. Coated liposomes were demonstrated to effectively penetrate the intestinal mucus layer where they came in close contact with the underlying epithelium. To investigate the permeation enhancing properties of the coated liposomes ex vivo, we monitored the transport of fluoresceinisothiocyanate-labeled salmon calcitonin (FITC-sCT) through rat small intestine. Liposomes coated with CS–TGA–MNA showed the highest effect, leading to a 3.8-fold increase in the uptake of FITC-sCT versus the buffer control. In vivo evaluation of the different formulations was carried out by the oral application of 40μg of sCT per rat, either encapsulated within uncoated liposomes, CS–TGA-coated liposomes or CS–TGA–MNA-coated liposomes, or given as a solution serving as negative control. The blood calcium level was monitored over a time period of 24h. The highest reduction in the blood calcium level, to a minimum of 65% of the initial value after 6h, was achieved for CS–TGA–MNA-coated liposomes. Comparing the areas above curves (AAC) of the blood calcium levels, CS–TGA–MNA-coated liposomes led to an 8.2-fold increase compared to the free sCT solution if applied orally in the same concentration. According to these results, liposomes coated with S-protected thiomers have demonstrated to be highly valuable carriers for enhancing the oral bioavailability of salmon calcitonin.

Solid lipid particles for oral delivery of peptide and protein drugs I – Elucidating the release mechanism of lysozyme during lipolysis

The mechanism of protein release from solid lipid particles was investigated by a new lipolysis model in a biorelevant medium containing both bile salts and phospholipids. Lysozyme, a model protein, was formulated into solid lipid particles using four different types of lipids, two triglycerides with different chain-length of fatty acyl groups i.e. trimyristin (TG14) and tristearin (TG18), and two lipid blends dominated by diglycerides and monoglycerides, respectively. The release of lysozyme from the solid lipid particles and the lipid hydrolysis process were assessed in the lipolysis model, while the change in particle surface during the lipolysis process was evaluated using scanning electron microscopy. The lysozyme release profiles from TG14 and TG18 as well as diglyceride particles correlated well with the release of free fatty acids from the lipid particles during the lipolysis and therefore exhibited a lipase-mediated degradation-based release mechanism. The release of lysozyme from monoglyceride particles was independent on lipase degradation due to the instability of the lipid matrix in the lipolysis medium. In conclusion, the established lipolysis model is successfully used to elucidate the drug release mechanism from solid lipid particles and can potentially be used in rational selection of lipid excipients for oral delivery of peptide/protein drugs.

<I>N</I>-Octyl-<I>N</I>-Arginine Chitosan Micelles as an Oral Delivery System of Insulin

N-octyl-N-Arginine chitosan (OACS) was synthesized in an attempt to combine the permeation enhancing effects of arginine-rich peptides and the drug loading capacity of the amphipathic polymers for insulin oral delivery. OACS self-assembled micelles of insulin were prepared by the conventional stirring technique, which were characterized by Dynamic light scattering, transmission electron microscopy and differential scanning calorimetry. Molecular docking by Discovery studio software confirmed that the interactions between OACS and insulin were mostly electrostatic in nature. In vitro, the result of the degradation experiment by enzyme showed that the OACS has a relative protective effect for insulin from proteolyses. Compared to the insulin solution, OACS micelles increased the Caco-2 cell's internalization by up to 22.3 folds. In vivo, the pharmacological activity PA% of series OACS-insulin micelles ranged from 7.7%-16.8%. Meanwhile by increasing arginine degree of the substitution both the uptake in Caco-2 cells and the hypoglycemic effect in diabetic rats were enhanced. Therefore, it is concluded that using arginine polymeric micelles for the enhancement of oral insulin delivery is a promising approach for the oral peptide delivery.

Amphiphilic polyallylamine based polymeric micelles for siRNA delivery to the gastrointestinal tract: In vitro investigations

Targeting the gastrointestinal (GI) tract represents a promising strategy for local or systemic delivery of nucleic acid-based therapeutics. The development of a nano-carrier for siRNA delivery via the GI tract would enable localised therapy for a range of gastrointestinal diseases. Previously, nano-sized polymeric micelles (PM) formed by a variety of amphiphilic poly(allylamine) (PAA) derivatives have shown potential for oral delivery of hydrophobic drugs and bioactive peptides. The aim of this study was to evaluate the ability of these amphiphilic PAA-based PM for siRNA delivery via the GI tract. The physicochemical characteristics of PAA·siRNA transfection complexes and their biological efficacy in vitro were investigated. Physicochemical profiles demonstrated that PAAs and siRNA self-assembled into complexes with nano-sized diameters (150-300 nm) and cationic surface charge (+ 20 to 30 mV). The PAA·siRNA complexes were stable in the presence of salt solutions and simulated gastric/intestinal fluids. In undifferentiated Caco-2 cells, PAA·siRNA complexes achieved up to 35% cellular uptake, with successful siRNA release from endosomes/lysosomes and significant luciferase gene knockdown. These results highlight the potential of these nano-sized PM for siRNA oral delivery via the GI tract for treatment of gastrointestinal diseases.

Approaches for enhancing oral bioavailability of peptides and proteins

Oral delivery of peptide and protein drugs faces immense challenge partially due to the gastrointestinal (GI) environment. In spite of considerable efforts by industrial and academic laboratories, no major breakthrough in the effective oral delivery of polypeptides and proteins has been accomplished. Upon oral administration, gastrointestinal epithelium acts as a physical and biochemical barrier for absorption of proteins resulting in low bioavailability (typically less than 1-2%). An ideal oral drug delivery system should be capable of (a) maintaining the integrity of protein molecules until it reaches the site of absorption, (b) releasing the drug at the target absorption site, where the delivery system appends to that site by virtue of specific interaction, and (c) retaining inside the gastrointestinal tract irrespective of its transitory constraints. Various technologies have been explored to overcome the problems associated with the oral delivery of macromolecules such as insulin, gonadotropin-releasing hormones, calcitonin, human growth factor, vaccines, enkephalins, and interferons, all of which met with limited success. This review article intends to summarize the physiological barriers to oral delivery of peptides and proteins and novel pharmaceutical approaches to circumvent these barriers and enhance oral bioavailability of these macromolecules.

Biodegradable pH-responsive polyacrylic acid derivative hydrogels with tunable swelling behavior for oral delivery of insulin

Novel biodegradable and pH-sensitive hydrogels composed of four types of pH-sensitive polyacrylic acid derivatives (PAAD) and a biodegradable poly(l-glutamic acid) (PGA) crosslinker were synthesized and characterized for oral delivery of proteins or peptides. The swelling ratios of hydrogels in buffer solutions showed a pH-dependent profile at different pH values. Insulin was loaded into the hydrogels as a model protein. The in vitro drug release experiment was carried out at different pH values and the release data suggested that both the pH and the type of the AAD unit played important roles in the drug release behaviors of the hydrogels. In vitro MTT assay indicated that the hydrogels displayed good cytocompatibility. After oral administration of insulin-loaded hydrogel particles to streptozotocin-induced diabetic rats at 60 IU/kg, the fasting plasma glucose level was reduced continuously to 67.4% within 7 h. These results indicated that the hydrogels are potential new vehicles for oral delivery of drugs.

Drug Carriers for Oral Delivery of Peptides and Proteins: Accomplishments and Future Perspectives

Effective formulation for peptide and protein delivery through the oral route has always been the critical effort with the advent of biotechnology. Stability, enzymatic degradation and ineffective absorption are common difficulties found for conventional dosage forms. As a result, new drug-delivery approaches are used to circumvent these limitations and enhance effective oral drug delivery. Some of these technologies have reached late stages of clinical trials and promising results will be available in the near future. This review covers, in general, the recent carriers reported in literature.

Oral insulin delivery in rats by nanoparticles prepared with non-toxic solvents

Nanoparticles (NPs) have shown a certain potential to overcome the drawbacks of oral peptide delivery in the gastrointestinal tract such as low peptide stability and permeability. The preparation of insulin loaded NPs was carried out with Eudragit RL or RS dissolved in different non-toxic polyethylene glycol (PEG) derivatives. The use of these non-toxic solvents allowed the design of an one step NP preparation method where insulin retained its full biological activity as it was proven in vitro and in vivo. The insulin trapping NPs were in a size range of around 150-250 nm and exhibited a pH-dependent release. The type of solvent did not distinctly influence the particle properties or insulin stability but modified significantly the performance in vivo in rats, NPs prepared with glycofurol led to a bioavailability of F=1.4 ± 1.0% after oral administration while NPs prepared with PEG 300 were hardly efficient (F=0.3 ± 0.5%). In all cases t(max) was shifted to 2h compared to 1h after subcutaneous insulin solution. In general, we believe that the method presented here is a promising way to encapsulate sensitive drugs, especially for the production of peptide loaded NPs.

Peptide-enhanced oral delivery of therapeutic peptides and proteins

Systemic therapy upon oral delivery of biologics, such as peptide and protein drugs is limited due to their large molecular size, their low enzymatic stability and their inability to cross the intestinal epithelium. Ways to overcome the epithelial barrier include the use of peptide-based excipients such as the cell penetrating peptides (CPPs) and the tight junction modulating peptides (TJMPs), which are able to translocate across the cellular membranes in a non-disruptive way or reversibly modulate the integrity of intercellular tight junctions (TJs), respectively. However, because of the harsh environment throughout the gastrointestinal (GI) tract, chemical stability is an inherent challenge when employing amino acid-based excipients for oral delivery, and multiple approaches have been investigated to improve this. The exact mechanisms of transepithelial translocation are discussed, and it is believed that CPP-mediated translocation involves transcytosis and/or direct translocation through the epithelial cells; whereas TJMP-mediated translocation is dependent on interaction with transmembrane or peripheral TJ proteins. This review focuses on the CPPs and the TJMPs currently employed as excipients for oral delivery of peptide and protein drugs highlighting recent studies and the most promising compounds from these classes of peptide excipients.

The Oral Delivery of Peptides and Proteins: Established Versus Recently Patented Approaches

Over the past 30 years there has been significant research into technologies that promote the delivery of high molecular weight, poor membrane-permeable compounds across the gut. Most work has concentrated on the delivery of peptides and proteins. However, technologies have also been applied to compounds such as poorly membrane-permeable small molecules, heparin and oligonucleotides. Much of this research has been characterized by early promise with many systems showing positive results in animal studies. Success in man has proven more elusive. In 2011, however, the oral delivery of peptides took one step closer to commercial reality when Tarsa Therapeutics announced that it had achieved positive Phase III data for oral recombinant salmon calcitonin. This article reviews the current development status of oral delivery systems for peptides and proteins and examines recent patent activity in this field based mainly on US patents issued in the last 2-3 years.

In Vitro and In Vivo Evaluation of a Water-in-Oil Microemulsion System for Enhanced Peptide Intestinal Delivery

Peptide and protein drugs have become the new generation of therapeutics, yet most of them are only available as injections, and reports on oral local intestinal delivery of peptides and proteins are quite limited. The aim of this work was to develop and evaluate a water-in-oil (w/o) microemulsion system in vitro and in vivo for local intestinal delivery of water-soluble peptides after oral administration. A fluorescent labeled peptide, 5-(and-6)-carboxytetramethylrhodamine labeled HIV transactivator protein TAT (TAMRA-TAT), was used as a model peptide. Water-in-oil microemulsions consisting of Miglyol 812, Capmul MCM, Tween 80, and water were developed and characterized in terms of appearance, viscosity, conductivity, morphology, and particle size analysis. TAMRA-TAT was loaded and its enzymatic stability was assessed in modified simulated intestinal fluid (MSIF) in vitro. In in vivo studies, TAMRA-TAT intestinal distribution was evaluated using fluorescence microscopy after TAMRA-TAT microemulsion, TAMRA-TAT solution, and placebo microemulsion were orally gavaged to mice. The half-life of TAMRA-TAT in microemulsion was enhanced nearly three-fold compared to that in the water solution when challenged by MSIF. The treatment with TAMRA-TAT microemulsion after oral administration resulted in greater fluorescence intensity in all intestine sections (duodenum, jejunum, ileum, and colon) compared to TAMRA-TAT solution or placebo microemulsion. The in vitro and in vivo studies together suggested TAMRA-TAT was better protected in the w/o microemulsion in an enzyme-containing environment, suggesting that the w/o microemulsions developed in this study may serve as a potential delivery vehicle for local intestinal delivery of peptides or proteins after oral administration.

Phospholipid vesicle–bound lysozyme to enhance permeability in human intestinal cells

Background: Oral peptide and protein drug delivery still remain the area of challenges for pharmaceutical scientists due to their low stability and permeability in gastrointestinal (GI) tract. In this study phospholipid vesicle–bound lysozyme were prepared and assessed for their physicochemical properties, secondary structure, and permeation across Caco-2 cells.Results: Lysozyme was found to be substantially bound onto negatively charged vesicles via electrostatic interaction as evidenced by zeta potential measurements regardless of cholesterol content. In contrast, the size of phospholipid vesicle–bound lysozyme became larger with the increasing cholesterol content. The secondary structure of vesicle-bound lysozyme examined by FTIR was unchanged compared to that in buffer solution. The apparent permeability of vesicle-bound lysozyme across Caco-2 cells monolayer was significantly enhanced with a size dependent manner compared to that of solution.Conclusion: The permeation across Caco-2 cell monolayers of phospholipid vesicle–bound lysozyme was demonstrated to be significantly enhanced with a size-dependent manner.