Abstract
Transport of biologically active molecules across tight epithelial barriers is a major challenge preventing therapeutic peptides from oral drug delivery. Here, we identify a set of synthetic glycosphingolipids that harness the endogenous process of intracellular lipid-sorting to enable mucosal absorption of the incretin hormone GLP-1. Peptide cargoes covalently fused to glycosphingolipids with ceramide domains containing C6:0 or smaller fatty acids were transported with 20-100-fold greater efficiency across epithelial barriers in vitro and in vivo. This was explained by structure-function of the ceramide domain in intracellular sorting and by the affinity of the glycosphingolipid species for insertion into and retention in cell membranes. In mice, GLP-1 fused to short-chain glycosphingolipids was rapidly and systemically absorbed after gastric gavage to affect glucose tolerance with serum bioavailability comparable to intraperitoneal injection of GLP-1 alone. This is unprecedented for mucosal absorption of therapeutic peptides, and defines a technology with many other clinical applications.
Highlights
One of the major challenges for applying protein and peptide biologics to clinical medicine is the lack of rational and efficient methods to circumvent epithelial and endothelial cell barriers separating large molecules from target tissues
To test if GM1 glycosphingolipids can be harnessed for biologic drug delivery, we first developed a non-degradable all D-isomer reporter peptide for structure-function studies on the ceramide domain
The peptide-coupled GM1 species containing cis-unsaturated or short fatty acid ceramide domains sorted into small cytoplasmic vesicles and basolateral membranes consistent with the recycling and transcytotic pathways, whereas the peptide-coupled GM1 species containing saturated long fatty acid ceramide domains did not; they were sorted into larger cytoplasmic puncta consistent with the late endosome/lysosome instead (Figure 1—figure supplement 1C)
Summary
One of the major challenges for applying protein and peptide biologics to clinical medicine is the lack of rational and efficient methods to circumvent epithelial and endothelial cell barriers separating large molecules from target tissues. The same is true for transport of protein and peptide cargoes across tight endothelial barriers that separate blood from tissue - typified by the blood-brain barrier (Abbott, 2013; Pardridge, 2015; Preston et al, 2014; Lajoie and Shusta, 2015). We address these problems by testing structure-function of the glycosphingolipids for their intracellular
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