Case Study in the Design of a Surrogate Solution for Use in Biopharmaceutical Drug Product Process Development.

The use of an aqueous-based surrogate solution in at-scale process development for biopharmaceutical drug products enables significant reduction in the usage of costly drug substance and improves confidence in initial drug product production runs performed using active biotherapeutic. Strategies for the formulation design of a surrogate solution that is representative of the unit operations in a typical drug product manufacturing process for a biopharmaceutical are presented herein, and a case study for the development of a surrogate solution for an example protein drug product is discussed. The surrogate was shown to have similar physical attributes to the drug product, including viscosity, surface tension, and density.

Plasmonic measurements of monoclonal antibody self-association using self-interaction nanoparticle spectroscopy.

One of the most significant challenges in developing therapeutic monoclonal antibodies (mAbs) is their unpredictable solubilities and viscosities at the high concentrations required for subcutaneous delivery. This challenge has motivated the development of screening assays that rapidly identify mAb variants with minimal self-association propensities and/or formulation conditions that suppress mAb self-association. Here we report an improved version of self-interaction nanoparticle spectroscopy (SINS)capable of characterizing both repulsive and attractive self-interactions between diverse mAbs.

A systematic multitechnique approach for detection and characterization of reversible self-association during formulation development of therapeutic antibodies.

In addition to controlling typical instabilities such as physical and chemical degradations, understanding monoclonal antibodies' (mAbs) solution behavior is a key step in designing and developing process and formulation controls during their development. Reversible self-association (RSA), a unique solution property in which native, reversible oligomeric species are formed as a result of the noncovalent intermolecular interactions has been recognized as a developability risk with the potential to negatively impact manufacturing, storage stability, and delivery of mAbs. Therefore, its identification, characterization, and mitigation are key requirements during formulation development.

A systematic multitechnique approach for detection and characterization of reversible self-association during formulation development of therapeutic antibodies.

In addition to controlling typical instabilities such as physical and chemical degradations, understanding monoclonal antibodies' (mAbs) solution behavior is a key step in designing and developing process and formulation controls during their development. Reversible self-association (RSA), a unique solution property in which native, reversible oligomeric species are formed as a result of the noncovalent intermolecular interactions has been recognized as a developability risk with the potential to negatively impact manufacturing, storage stability, and delivery of mAbs. Therefore, its identification, characterization, and mitigation are key requirements during formulation development.

Formulation development of therapeutic monoclonal antibodies using high-throughput fluorescence and static light scattering techniques: role of conformational and colloidal stability.

In this work, we describe the application of two different high-throughput screening (HTS) techniques that can be used to determine protein stability during early formulation development. Differential scanning fluorescence (DSF) and differential static light scattering (DSLS) are used to determine the conformational and colloidal stability of therapeutic monoclonal antibodies (mAbs) during thermal denaturation in a high-throughput fashion. DSF utilizes SYPRO Orange, a polarity-sensitive extrinsic fluorescent probe, to monitor protein unfolding.

Buffer-dependent fragmentation of a humanized full-length monoclonal antibody.

During storage stability studies of a monoclonal antibody (mAb) it was determined that the primary route of degradation involved fragmentation into lower molecular weight species. The fragmentation was characterized with size-exclusion high performance liquid chromatography (SE-HPLC), SDS-PAGE, and matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry. Fragmentation proceeded via hydrolysis, likely catalyzed by trace metal ions, of a peptide bond in the hinge region of the mAb's heavy chain, which produced two prominent low molecular weight species during storage: a single, free Fab fragment and a Fab + Fc fragment.

Effect of polyanions on the structure and stability of repifermin (keratinocyte growth factor-2).

The interaction of several of the fibroblast growth factors (FGFs) with polyanions is thought to be of physiological significance and has been exploited to create more stable pharmaceutical formulations of FGF-1 and -2. The extent of such phenomena throughout the 23-member FGF family is, however, unknown. In these studies, we examine the effect of several polyanions on the structure and stability of keratinocyte growth factor 2 (KGF-2, FGF-10), a candidate for use as a wound-healing agent.