Within the talk novel approaches for the characterization of particulate impurities in biologics will be presented, and compared to state-of-the-art technologies for submicron and subvisible particle analysis. Besides new analytical technologies, the integration of machine learning for data analysis will be presented.

The monitoring and control of process-related impurities is an important element in process development and product manufacturing. USP has developed standards and tools to assist in the quantitation of host cell DNA and host cell proteins.  This presentation will cover USP standards, documentary and reference standards, for host cell DNA quantitation, and new reference materials in development to support quantitation of host cell proteins.

Identification, counting, and minimization of particles continue to be important for the safety and efficacy of biotherapeutics. Protein aggregates, which can arise as pure protein or nucleated on other particle types, present unique measurement challenges. While the standard methods used for lot release are still based on light obscuration methods from the 1990s, significant advances have resulted in improved capabilities over a wide size range. Recent developments will be discussed.

I present strategies for the use of particle reference materials, discussing their strengths and limitations. Both the properties of interest in reference material and its use vary substantially from the size range of nanometers to visible, depending on user expectations, the state of the art in instrumentation, and what is measured. Reference materials developed for size and count are also now finding use for new technologies and methods, such as AI analysis of particle images.

Polysorbates (PS) are used to reduce monoclonal antibody (mAb) aggregation at hydrophobic interfaces. PS can hydrolyze over time to generate free fatty acid (FFA) degradants, primarily lauric acid for PS20 and oleic acid for PS80. This talk discusses key differences in the effect on aggregation of a surface-active mAb by degraded PS20 and PS80, using surface tension studies, static storage behavior, colloidal, and conformational stability of mAbs with degraded PS.

The chemical degradation of polysorbate continues to present a challenge for the development of stable protein formulations. Free radical-mediated mechanisms play a central role in oxidative polysorbate degradation. Polysorbate oxidation can target polysorbate monomers or polysorbate micelles. Here, we show that intra-micellar oxidation reactions dominate polysorbate oxidation even when initiated with hydrophilic oxidants. We further show that specific buffer-derived radicals can play a critical role in polysorbate oxidation.

The degree of protein purity and aggregation are well-established critical quality attributes requiring careful monitoring during the development of monoclonal antibody Drug Products. A strategic approach was utilized to rapidly rescue soluble and insoluble aggregation and enable pulmonary delivery of IgM mAb Drug Product.  Key formulation and delivery considerations to enable Early Development activities will be described.

Mechanistic characterization as to how HCPs persist in drug products is important to refining downstream processing. Weak esterase–mAb interactions potentially enable HCP esterases to evade drug purification processes. Here, we apply state-of-the-art methods such as HRF-MS, native-MS, and ion mobility to establish esterase-mAb binding mechanisms. 

The goal of this presentation is to provide an update on the most recent developments in real-time detection and monitoring of protein aggregation. The focus is on emerging orthogonal methods that may be implemented during product development as well as during large-scale manufacturing.

Monoclonal antibodies at high concentrations interact with each other and with excipients by a variety of mechanisms. We can modulate these interactions by using excipients to change the properties of drug products. Here we look at a few excipients and how they affect mAb formulations.

Development of formulations for high concentration (100+ mg/mL) biotherapeutics poses numerous technical and logistical challenges during formulation and process development. As illustrated in a case study of the development of a 150 mg/mL monoclonal antibody formulation, strategic use of data generated at lower concentrations, utilization of certain predictive assays for stability at high concentrations, and careful molecule selection can enable rapid development of stable high concentration formulations.

Succinate acid/succinate system has an excellent buffering capacity at acidic pH values (4.5-6.0). However, its use in formulating drug products is largely limited due to risk of its components crystallizing and the consequent pH shifts. Physicochemical behavior of succinate system was characterized under pharmaceutically representative conditions. mAbs formulated in de-risked succinate buffer maintained a good stability profile during typical pharmaceutical processing and upon storage, bolstering their wider use in drug products.