Oral delivery of proteins such as peptide therapeutics remains a major challenge for the pharmaceutical industry. Recently we reported the enhanced transport of oral insulin across rat intestine in vivo using peptides that that were rationally designed to induce myosin light chain (MLC) phosphorylation. These permeable inhibitors of MLC phosphatase (PIP peptides) transiently open intestinal tight junctions that exist between adjunct epithelial cells. In doing so, PIP peptides can be used to dynamically enhance solute uptake through the paracellular route, proving a promising new, mechanism-based approach for enhancing peptide therapeutic absorption following oral administration.
Presently, the medical field and pharmaceutical industry are facing a diabetes epidemic on a global scale with most patients poorly controlling their blood sugar levels, leading to devastating morbidity issues and ultimately advanced mortality. Better management of blood sugar is of paramount concern to limit this ever-increasing burden on health care systems. Oral delivery of insulin, and more recently incretin molecules such as GLP-1 secreted by intestinal epithelial cells in response to food intake, could provide a simpler needle-free method of treating diabetes, with the possibility of an enhanced clinical benefit. Orally administered insulin absorbed across the intestinal mucosa and would be delivered to the liver by the portal circulation, just as insulin is normally delivered to the liver by the pancreas of a non-diabetic human. Thus, successful oral administration of insulin and GLP-1 are now considered more than just a strategy to improve patient compliance and comfort, but essential to effectively combat the diabetes epidemic. PIP peptides will likely lead to exciting opportunities for oral delivery of therapeutic agents such as insulin and GLP-1, and to transformative treatments for diabetic patients.
These peptides act by a defined mechanism of action, without causing any intestinal damage, by direct actions on the epithelium after topical application. Clinical utilization of agents enhancing paracellular permeability without first understanding their mechanisms of permeabilization is challenging both because of potential safety issues and translation to humans from pre-clinical models. PIP peptides overcome previous challenges to clinical development strategies for oral insulin and GLP-1 dosage forms because they act transiently and locally following topical application, and do not appear to affect other tissues prior to their metabolism and elimination from the body. Most importantly, PIP peptides work through a defined and endogenous pathway, do not appear to incite cell damage or an extensive co-uptake of bacterial components (like lipopolysaccharides) present in the intestinal lumen.
Our initial studies using a simple administration into the intestine of rats have shown 3-4% bioavailability for insulin. We believe that formulation efforts to enhance the stability of the peptides therapeutic to be delivered could increase this level of bioavailability significantly. While our initial studies have used insulin to demonstrate the feasibility of PIP peptides, we are interested in exploring a spectrum of therapeutic peptides for oral delivery. Although only insulin has been tested to date we would expect the PIP peptides to be capable of delivering other peptides and small proteins orally. We are seeking industrial partners for this technology and are keen to validate the PIP peptides in a large animal study.
Taverner A1, Dondi R1, Almansour K1, Laurent F1, Owens SE2, Eggleston IM1, Fotaki N1, Mrsny RJ3.[expand title=”Show Affiliations”]
- Department of Pharmacy and Pharmacology, University of Bath, Bath BA2 7AY, UK.
- Welsh School of Pharmacy, Cardiff University, Cardiff, CF10 3XF, UK.
- Department of Pharmacy and Pharmacology, University of Bath, Bath BA2 7AY, UK. Electronic address: [email protected] [/expand]
The intestinal epithelium functions to effectively restrict the causal uptake of luminal contents but has been demonstrated to transiently increase paracellular permeability properties to provide an additional entry route for dietary macromolecules. We have examined a method to emulate this endogenous mechanism as a means of enhancing the oral uptake of insulin. Two sets of stable Permeant Inhibitor of Phosphatase (PIP) peptides were rationally designed to stimulate phosphorylation of intracellular epithelial myosin light chain (MLC) and screened using Caco-2 monolayers in vitro. Apical application of PIP peptide 640, designed to disrupt protein– protein interactions between protein phosphatase 1 (PP1) and its regulator CPI-17, resulted in a reversible and non-toxic transient reduction in Caco-2 monolayer trans-epithelial electric resistance (TEER) and opening of the paracellular route to 4 kDa fluorescent dextran but not 70 kDa dextran in vitro. Apical application of PIP peptide 250, designed to impede MYPT1-mediated regulation of PP1, also decreased TEER in a reversible and non-toxic manner but transiently opened the paracellular route to both 4 and 70 kDa fluorescent dextrans. Direct injection of PIP peptides 640 or 250 with human insulin into the lumen of rat jejunum caused a decrease in blood glucose levels that was PIP peptide and insulin dose-dependent and correlated with increased pMLC levels. Systemic levels of insulin suggested approximately 3–4% of the dose injected into the intestinal lumen was absorbed, relative to a subcutaneous injection. Measurement of insulin levels in the portal vein showed a time window of absorption that was consistent with systemic concentration-time profiles and approximately 50% first-pass clearance by the liver. Monitoring the uptake of a fluorescent form of insulin suggested its uptake occurred via the paracellular route. Together, these studies add validation to the presence of an endogenous mechanism used by the intestinal epithelium to dynamically regulate its paracellular permeability properties and better define the potential to enhance the oral delivery of biopharmaceuticals via a transient regulation of an endogenous mechanism controlling the intestinal paracellular barrier.
Copyright © 2015. Published by Elsevier B.V.Go To J Control Release.