(523d) Self-Interaction Chromatography As a Tool for Determining Oral Peptide Stability Under GI Conditions

Peptide therapeutics typically display higher specificity to their target binding sites than small molecule drugs and so display fewer side effects and lower toxicity. However, they are conventionally administered via parenteral injections rather than via oral dosage, resulting in lower patient compliance and an increased cost to the consumer. Peptide drugs, in general, exhibit low oral bioavailabilities, in part due to their propensity to aggregate under gastrointestinal (GI) conditions, thereby reducing their already low permeability through the epithelial layer. Peptide aggregation is the self-association of monomeric peptides to form highly-ordered fibrils or amorphous structures and is dependent on several factors including net charge, concentration and temperature. Current techniques for monitoring peptide aggregation, including ultracentrifugation, dynamic light scattering (DLS) and size exclusion chromatography, are time-consuming, to allow for sample aggregation, and necessitate high sample consumption.
Self-interaction chromatography (SIC) involves the immobilisation of a peptide of interest onto the surface of a solid material to form a chromatographic stationary phase, through which the same species is eluted isocratically to measure its retention. The species retention time is indicative of the average peptide interaction strength under the conditions of the mobile phase and may be quantitatively related to the osmotic second virial coefficient (B22), a term known to describe the direction and magnitude of weak protein interactions and thus correlate to their phase behaviour. A positive B22 value indicates net repulsive forces between species and so a preference for the monomeric form, whilst a negative value implies more attractive forces and thus a propensity to aggregate.
Our approach involves the application of SIC to study the stability of Semaglutide under varied intestinal conditions to establish its optimum oral dosage time in the absence of formulation and the effects of net charges and cosolutes on its aggregation propensity. A tresyl-functionalised formyl resin acted as the stationary phase for immobilisation. The results were then measured against DLS measurements of the corresponding conditions to establish the effectiveness of the technique for such an application.
This SIC methodology shows potential for future application in oral peptide formulation studies, requiring minimal expertise and a significant reduction in sample peptide relative to other techniques. In addition, one column, requiring a relatively short time investment for immobilisation, allows for the study of multiple formulations under multiple conditions in quick succession, thereby reducing overall experiment time.