(W1030-02-08) Optimization of Artificial Colonic Mucus for Drug Diffusion Study

Purpose: To select appropriate polymer for porcine artificial colonic mucus (PACM) formulation which captures diffusion properties of native porcine colonic mucus for drug screening purpose.

Methods: Biosimilar PACM prototypes with focus on polymer selection were generated to mimic porcine native colonic mucus (PNCM) and to improve previous model of PACM with polyacrylic acid (PAA). PACM viscosity & storage modulus were measured with rheometer ARES-G2. Apparent viscosity was measured within the range of 10 to 1000 Pa.s and within 10^-4 to 10^-2 MPa for storage modulus¹. Surface charge and zeta potential were determined using dynamic light scattering (DLS). Binding coefficients of model compounds were calculated using microscale themophoresis (MST). Diffusion coefficients were determined by using particle tracking resulting in mean squared displacement (MSD) based on fluorescence recovery after photobleaching (FRAP).

Results: Previously developed PACM based on PAA was used as the benchmark for the new prototypes of PACM. The selected polymers (hydroxyethylcellulose (HEC), hyaluronic acid (HA), sodium alginate (SA), and pectin (PEC)) were visually studied for their gelling ability and apparent viscosity was determined. From measurements of zeta potential, the PNCM zeta potential value was determined to be -21.9 mV and the closest polymer formulation for this value is PACM based on HEC (-19.4 mV). Binding coefficient of FITC-Dextran 4K+ with PNCM was also closer to the PACM HEC compared to PACM PAA.

Conclusion: HEC was identified as a better polymer than PAA in PACM with closer properties to PNCM. Evaluation of PACM HEC with various methods and approaches based on inherent characteristics and interaction characteristics allows the generation of a platform that mimics native colonic mucus for studying drug diffusion in colonic mucus environment.

References: 1. Barmpatsalou V, Dubbelboer IR, Rodler A, Jacobson M, Karlsson E, Pedersen BL, Bergström CAS. Physiological properties, composition and structural profiling of porcine gastrointestinal mucus. Eur J Pharm Biopharm. 2021 Dec;169:156-167. doi: 10.1016/j.ejpb.2021.10.008. Epub 2021 Oct 20. PMID: 34687897.

Acknowledgements: The work was supported by the European Union’s Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie grant agreement No 956851 and the Swedish Innovation Agency grant 2019-00048.

Figure 1. Rheological determination of the series of PACM explored. All PACMs show viscoelastic profile both in terms of viscosity sweep (A) and oscillatory frequency resulting in storage modulus in linear viscoelastic region (B) around PNCM range. Data for PNCM range were obtained from previous publication by Barmpatsalou et al. 2021.

Figure 2. Measured zeta potentials of the different prototypes of PACM in comparison to PNCM.

Figure 3. Interaction of mucus with compounds and particles. Fitted Kd (dissociation constant) plot of normalized fraction bound against normalized concentration of PNCM, PACM PAA, and PACM HEC (A). Diffusion of fluorescently labeled polystyrene nanobeads with 0.1, 0.2 and 1 µm in size with carboxylate and amine modification as model of negative and positive charge respectively. Size and surface charge are shown to be influencing the particle MSD in artificial colonic mucus environment (B).