Mechanistic studies of a cell-permeant peptide designed to enhance myosin light chain phosphorylation in polarized intestinal epithelia

Almansour, Khaled; Taverner, Alistair; Eggleston, Ian M.; Mrsny, Randall J.

doi:10.1016/j.jconrel.2018.03.033

Cited by 20 publications

(24 citation statements)

References 33 publications

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“…They claimed that these peptides can dynamically adjust endogenous mechanisms, inducing myosin light chains (MLCs), opening the tight junctions and facilitating paracellular transition, especially for peptide therapeutics. Almansour et al (in the same group) studied PIP 640 further and confirmed its stability in the intestinal lumen environment and explained its functioning mechanism: PIP 640 selectively enhances the MLC-pS 19 levels of the cytoplasm of enterocytes in the epithelial layer [120]. Apart from the recent studies and the progress made, tight junctions are still one of the main challenges against the absorption of biopharmaceuticals.…”

Section: Challenges Associated With Oral Deliverymentioning

confidence: 99%

Challenges and Recent Progress in Oral Drug Delivery Systems for Biopharmaceuticals

2019

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Routes of drug administration and the corresponding physicochemical characteristics of a given route play significant roles in therapeutic efficacy and short term/long term biological effects. Each delivery method has favorable aspects and limitations, each requiring a specific delivery vehicles design. Among various routes, oral delivery has been recognized as the most attractive method, mainly due to its potential for solid formulations with long shelf life, sustained delivery, ease of administration and intensified immune response. At the same time, a few challenges exist in oral delivery, which have been the main research focus in the field in the past few years. The present work concisely reviews different administration routes as well as the advantages and disadvantages of each method, highlighting why oral delivery is currently the most promising approach. Subsequently, the present work discusses the main obstacles for oral systems and explains the most recent solutions proposed to deal with each issue.

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Section: Challenges Associated With Oral Deliverymentioning

confidence: 99%

Challenges and Recent Progress in Oral Drug Delivery Systems for Biopharmaceuticals

2019

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“…PIP-640 increased fluorescein isothiocyanate dextran 4 kDa (FD4) permeation across Caco-2 monolayers and improved BA of insulin to 4% in rat intestinal instillations [47]. More recently, a structure–activity assessment of PIP 640 showed that residues of glutamic acid and tyrosine are requisite for binding to MYPT1 subunit of the MLCP complex, while substitution of aspartic acid for arginine led to more specific targeting of PP1 subunit, which was associated with greater cytotoxicity [48]. In a follow-on study, PIP 640 was shown to selectively increase total levels of claudin 2 (both the cytoplasm and at the membrane) [49].…”

Section: Recent Highlightsmentioning

confidence: 99%

Application of Permeation Enhancers in Oral Delivery of Macromolecules: An Update

et al. 2019

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The application of permeation enhancers (PEs) to improve transport of poorly absorbed active pharmaceutical ingredients across the intestinal epithelium is a widely tested approach. Several hundred compounds have been shown to alter the epithelial barrier, and although the research emphasis has broadened to encompass a role for nanoparticle approaches, PEs represent a key constituent of conventional oral formulations that have progressed to clinical testing. In this review, we highlight promising PEs in early development, summarize the current state of the art, and highlight challenges to the translation of PE-based delivery systems into safe and effective oral dosage forms for patients.

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“…PIP peptides were synthesized as described previously using an automated Symphony Quartet peptide synthesizer (Gyros Protein Technologies, USA) with Fmoc-protected D-amino acids on Rink amide MBHA resin (0.6 mmol/g loading) (Novabiochem, UK) [20]. First amino acid loading onto the resin was performed manually with N,N’-diisopropylcarbodiimide and 1hydroxybenzotriazole as coupling agents in the presence of diisopropylethylamine (DIEA) (Sigma-Aldrich, UK), with assembly of the remaining amino acids being performed automatically by the peptide synthesizer using (benzotriazol-1-yloxy)-tripyrrolidinophosphonium hexafluorophosphate (Novabiochem, UK) as the coupling reagent with DIEA.…”

Section: Methodsmentioning

confidence: 99%

“…We have described a peptide, termed PIP 640, that can transiently increase myosin light chain phosphorylation at serine 19 (MLC-pS 19 ) to stimulate a TJassociated actomyosin filament contraction to recreate the actions of supra-physiological concentrations of nutrients that acts to dynamically diminish barrier function [19]. Most recently, we have performed positional assessment of amino acid requirements within the PIP 640 sequence to better define its capability to enter intestinal epithelial cells, localize to TJ structures, and interact with MLC phosphatase elements in order to further validate its anticipated MoA [20].…”

Section: Introductionmentioning

confidence: 99%

An intestinal paracellular pathway biased toward positively-charged macromolecules

Almansour

Taverner

Turner

et al. 2018

Journal of Controlled Release

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Lacking an effective mechanism to safely and consistently enhance macromolecular uptake across the intestinal epithelium, prospects for successful development of oral therapeutic peptide drugs remain unlikely. We previously addressed this challenge by identifying an endogenous mechanism that controls intestinal paracellular permeability that can be activated by a peptide, termed PIP 640, which can increase cellular levels of phosphorylated myosin light chain at position S (MLC-pS). Apical application in vitro or luminal application in vivo was shown to increase macromolecular solute transport within minutes that recovered completely within a few hours after removal. We now examine the nature of PIP 640-mediated permeability changes. Confluent Caco-2 cell monolayers treated with PIP 640 enhanced apical-to-basolateral (AB) transport of 4-kDa, but not 10-kDa, dextran. Expression and/or cellular distribution changes of tight junction (TJ) proteins were restricted to increased claudin-2 over a time course that correlated with an apparent shift in its distribution from the nucleus to the membrane fraction of the cell. PIP 640-mediated epithelial changes were distinct from the combined actions of the pro-inflammatory cytokines tumor necrosis factor alpha (TNF-α) and interferon gamma (IFN-γ). While TNF-α/IFN-γ treatment also increased MLC-pS levels, these cytokines enhanced AB transport for 70-kDa dextran and decreased occludin expression at TJs. Claudin-2-dependent changes induced by PIP 640 resulted in an AB transport bias for positively-charged macromolecules demonstrated in vitro using charge variants of 4-kDa dextrans and by comparing transport of salmon calcitonin to exenatide. Comparable outcomes of increased TJ localization of claudin-2 and enhanced transport of these therapeutic peptides that biased toward cationic characteristics was demonstrated in vivo following after intra-luminal injection into rat jejunum. Together, these data have shown a potential mechanism for PIP 640 to enhance paracellular permeability of solutes in the size range of small therapeutic peptides that is biased toward positively-charged solutes.

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Mechanistic studies of a cell-permeant peptide designed to enhance myosin light chain phosphorylation in polarized intestinal epithelia

Cited by 20 publications

References 33 publications

Challenges and Recent Progress in Oral Drug Delivery Systems for Biopharmaceuticals

Challenges and Recent Progress in Oral Drug Delivery Systems for Biopharmaceuticals

Application of Permeation Enhancers in Oral Delivery of Macromolecules: An Update

An intestinal paracellular pathway biased toward positively-charged macromolecules

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