Chemometrics in Protein Formulation: Stability Governed by Repulsion and Protein Unfolding.

Therapeutic proteins can be challenging to develop due to their complexity and the requirement of an acceptable formulation to ensure patient safety and efficacy. To date, there is no universal formulation development strategy that can identify optimal formulation conditions for all types of proteins in a fast and reliable manner. In this work, high-throughput characterization, employing a toolbox of five techniques, was performed on 14 structurally different proteins formulated in 6 different buffer conditions and in the presence of 4 different excipients.

study of the interactions between acylated glucagon like-peptide-1 analogues and the native receptor.

We have performed a series of multiple molecular dynamics (MD) simulations of glucagon-like peptide-1 (GLP-1) and acylated GLP-1 analogues in complex with the endogenous receptor (GLP-1R) to obtain a molecular understanding of how fatty acid (FA) chain structure, acylation position on the peptide, and presence of a linker affect the binding. MD simulations were analysed to extract heatmaps of receptor-peptide interaction patterns and to determine the free energy of binding using the molecular mechanics Poisson-Boltzmann surface area (MM-PBSA) approach. The extracted free energies from MM-PBSA calculations are in qualitative agreement with experimentally determined potencies.

An albumin-mediated cholesterol design-based strategy for tuning siRNA pharmacokinetics and gene silencing.

Major challenges for the clinical translation of small interfering RNA (siRNA) include overcoming the poor plasma half-life, site-specific delivery and modulation of gene silencing. In this work, we exploit the intrinsic transport properties of human serum albumin to tune the blood circulatory half-life, hepatic accumulation and gene silencing; based on the number of siRNA cholesteryl modifications. We demonstrate by a gel shift assay a strong and specific affinity of recombinant human serum albumin (rHSA) towards cholesteryl-modified siRNA (Kd>1×10(-7)M) dependent on number of modifications.

Bio-reduction of redox-sensitive albumin conjugates in FcRn-expressing cells.

Disulfide-containing IgG-, Fc-, or albumin-based prodrugs that rely on FcRn-trafficking by endothelial cells for prolonged circulation in the body might be hampered by premature bio-reduction processes during FcRn-mediated recycling events. A detailed bio-reduction analysis of redox-sensitive albumin conjugates in two FcRn-expressing cell lines has been performed. The obtained results indicate that the FcRn-mediated recycling pathway is not (or is only poorly) bio-reducing.