Functionalization of lipid-based formulations for oral delivery of insulin

Insulin is a potent therapeutic molecule used for the treatment of diabetes mellitus type I and late-stage type II. Insulin is usually administered via subcutaneous (s.c.) injection, which bypasses the portal circulation and, in turn, is likely to cause diabetic complications after long-term treatment. For diabetic patients, multiple daily self-injections are associated with discomfort and pain. The limitations of s.c. administration can be overcome by considering oral delivery of insulin as an alternative because of good patient compliance and easier manufacturing than that of sterile injectables. The critical challenge for oral insulin delivery is the low bioavailability owing to poor enzymatic stability and low absorption of insulin in the gastrointestinal tract.

Oral lipid-based formulations are attracting considerable attentions due to their potentials on the protection and facilitation in gastrointestinal absorption of peptides/proteins and their feasibility for functionalization. Initially, a series of Kolliphor® RH40 (RH40)- and Labrasol® (LAB)-based self-emulsifying drug-delivery systems (SEDDS) containing either long-chain (LC) or medium-chain (MC) glycerides were formulated. Insulin, in order to be incorporated in SEDDS, was complexed with soybean phosphatidylcholine (SPC) to form the insulin–SPC complex (ins-SPC). Next, an in vitro model simulating intestinal proteolysis was used to evaluate the ability of the SEDDS to protect insulin against proteolysis. Principal component analysis (PCA) was used to correlate the composition of SEDDS and droplet size after dispersion with the protective effect of SEDDS on the loaded insulin. This study confirmed that some SEDDS could effectively protect the loaded insulin against proteolysis (>60% of insulin remaining after 60 min of proteolysis). Furthermore, SEDDS based on MC glycerides and LAB were superior to those based on LC glycerides and RH40 in protecting insulin against proteolysis. Monoacyl phosphatidylcholine (MAPC), a natural permeation enhancer (PE), and Capmul® MCM C8 were favored for the protective effect of SEDDS on insulin during proteolysis.

Subsequently, three SEDDS containing synthetic PEs and/or natural PEs with a pronounced protective effect on insulin were selected to validate the model predictions of in vitro proteolysis. MCT(RH40) [50% Migyol® 812N, 20% RH40, and 30% Capmul® MCM C8], MCT(M) [20% Captex® 300, 30% LAB, 30% MAPC, and 20% Capmul® MCM C8], and LCT(M) [20% sesame oil, 30% LAB, 30% MAPC, and 20% Maisine® CC] loaded with ins-SPC were subjected to in vitro cell transport and in vivo studies. Insulin permeability was investigated in mucus-secreting Caco-2/HT29-MTX-E12 coculture and Caco-2 cell monolayers. The in vivo efficacy of insulin upon loading into SEDDS was tested in rats via intra-jejunal instillation. MCT(M) significantly increased the in vitro insulin permeability across both cell culture monolayers and decreased rat blood glucose after 0.5 h by 24 ± 11% of the initial value (p < 0.05), whereas the in vivo effect was not significant for MCT(RH40) and LCT(M). Furthermore, the incorporation of the lipase inhibitor Orlistat® into MCT(M) did not significantly change insulin absorption and the following pharmacodynamic effect.

In the last part of the thesis work, MCT(M) was further functionalized by chitosan (Chi) and/or colloidal silica (SiO2) nanoparticles (NPs) to generate various dispersions for functionalized-MCT(M) (f-MCT(M)). The dispersions were evaluated for size/zeta-potential, nano-scale morphology, and in vitro characterization by conducting in vitro proteolysis and cell transport study. The MCT(M) generated fine nanoemulsions, whereas f-MCT(M) dispersions exhibited versatile structures such as multilamellar and oligolamellar vesicles. The obtained lipid vesicles showed the ability to protect insulin against proteolysis, with 30–50% of insulin remaining after 60 min of proteolysis (37 °C). The in vitro transport across the mucus-secreting coculture monolayer suggested that MCT(M) enhanced insulin permeability mainly via the paracellular route. An intra-ileal (i.i.) instillation study in rats showed significant effects of insulin-loaded functionalized-MCT(M) on the glucose-lowering (50–80%), insulin absorption (Frel = 1.3–1.5%), and glucagon modulation, compared to i.i. instillation of insulin solution. Furthermore, the hepatic and peripheric insulin exposures lasted up to 2 h, resulting in the metabolism of a substantial amount of insulin in the liver (45–71%).

Our study revealed the potential of the functionalization of MCT(M) by Chi and SiO2 NPs in improving the intestinal absorption of insulin. The developed CS-SiO2-MCT(M) holds high promise for oral delivery by mimicking the normal physiological insulin secretion. Furthermore, an in vitro proteolysis model with physiological relevance was applied to evaluate the SEDDS efficacy of protection on insulin for oral delivery. The MCT(M) composed of natural and synthetic PEs such as MAPC and LAB, respectively, was validated to increase the in vivo absorption of insulin significantly. The present findings might broaden the biomedical applications of SEDDSs in the oral delivery of insulin, and potentially as well as other biopharmaceuticals.

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Abstract of PhD thesis by Jingying Liu
Department of Pharmacy at the University of Copenhagen
Ask for a copy of the thesis: [email protected]