Vapor-Phase Hydrogen Peroxide Uptake by Silicone Tubing and Primary Packaging Components during Protein Drug Product Aseptic Filling: Impact of Pretreatment and Sterilization Process.

In the vapor-phase hydrogen peroxide (VPHP)-sanitized environment, VPHP uptake by product-contacting components could eventually lead to undesired oxidation of biological drug products. Silicone tubing and primary packaging materials are prominent examples of such product-contacting surfaces that are typically processed/sterilized prior to use. This study investigated the VPHP-uptake tendency of these components and how their respective processing/sterilization methods affect the uptake behaviors. Silicone tubing that was sterilized via autoclave or gamma irradiation exhibited different VPHP uptake patterns-decreased uptake rates post autoclaving vs. increased uptake rates post gamma irradiation. The reduced uptake tendency of autoclaved tubing is maintained for 14 days after sterilization, whereas the uptake tendency of irradiated tubing was mostly reversed to normal levels 1 month after irradiation. Empty glass vials adsorbed hydrogen peroxide via the diffusion of VPHP into the vial with high vial-to-vial variability. Vial pretreatment (i.e., depyrogenation) and surface hydrophilicity/hydrophobicity impacted the uptake tendency. Stoppers and empty syringes also adsorbed hydrogen peroxide but at a relatively low level. The uptake behavior of these components appeared to correlate with water levels at the surface (i.e., hydrophilicity). This study provides process development scientists and engineers an in-depth understanding of the VPHP uptake by critical product-contacting surfaces so that they can mitigate the impact on drug product quality.LAY ABSTRACT: This study investigated vapor-phase hydrogen peroxide (VPHP) absorption by biopharmaceutical drug products via VPHP uptake by critical product-contacting components during the aseptic manufacturing process with a focus on various pretreatments and processing of these components. Sterilization of silicone tubing by gamma irradiation or autoclaving resulted in different VPHP uptake profiles with different effect durations. Primary packaging components, such as vials, syringes, and stoppers, also showed different levels of VPHP uptake with surface hydrophilicity/hydrophobicity playing a critical role. These outcomes suggested that VPHP uptake is a complex phenomenon and should be carefully considered to minimize its impact on product quality. The approach and outcome of this study can benefit scientists and engineers who develop biological product manufacturing processes by providing an in-depth understanding of drug product process risks.

Stability Evaluation of Hydrogen Peroxide Uptake Samples from Monoclonal Antibody Drug Product Aseptically Filled in Vapor Phase Hydrogen Peroxide-Sanitized Barrier Systems: A Case Study.

During the manufacture of a monoclonal antibody drug product, which was aseptically filled within a vapor phase hydrogen peroxide-sanitized isolator, samples were taken to investigate the hydrogen peroxide uptake behaviors. Surprisingly, the samples had no detectable hydrogen peroxide (most results below the limit of detection). This finding was later attributed to hydrogen peroxide decomposition after the samples were stored frozen at -20°C for two weeks before testing. This case study highlights the criticality of storage conditions for hydrogen peroxide-containing samples and summarizes an investigation on hydrogen peroxide stability in water and in three monoclonal antibody solutions having a wide protein concentration range (30-200 mg/mL). Samples were stored at three temperatures (-70°C, -20°C, or 2-8°C) for up to 28 days to assess the impact of protein concentration and storage temperature on hydrogen peroxide decomposition rates. Hydrogen peroxide degraded slightly more rapidly with increasing protein concentration independent of storage condition. When stored at -20°C, hydrogen peroxide was least stable and degraded faster than when stored at 2-8°C. Hydrogen peroxide was most stable when the samples were stored at -70°C. Overall, this case study brings the hydrogen peroxide stability issue to the attention of process development scientists and engineers and offers a valuable lesson learned during process development.LAY ABSTRACT: The use of vapor phase hydrogen peroxide as a sanitizing agent for isolator and cleanroom decontamination has become common in recent years. Because of the potential impact of residual hydrogen peroxide on biopharmaceutical product quality, hydrogen peroxide uptake behaviors and mechanisms during the manufacturing process within these barriers need to be evaluated and understood. Samples taken from various small-scale and manufacturing-scale hydrogen peroxide uptake studies are often stored frozen before testing. This case study reports an important and interesting finding about hydrogen peroxide stability in samples collected for hydrogen peroxide uptake investigation, and it demonstrates the relationship between hydrogen peroxide stability and storage temperature, storage duration, and monoclonal antibody concentration. The approach and outcome of this study are expected to benefit scientists and engineers who develop biologic product manufacturing processes by providing a better understanding of drug product process challenges and appropriate sample storage.

Processing Impact on Monoclonal Antibody Drug Products: Protein Subvisible Particulate Formation Induced by Grinding Stress.

Subvisible particle formation in monoclonal antibody drug product resulting from mixing and filling operations represents a significant processing risk that can lead to filter fouling and thereby lead to process delays or failures. Several previous studies from our lab and others demonstrated the formation of subvisible particulates in mAb formulations resulting from mixing operations using some bottom-mounted mixers or stirrer bars. It was hypothesized that the stress (e.g., shear/cavitation) derived from tight clearance and/or close contact between the impeller and shaft was responsible for protein subvisible particulate generation. These studies, however, could not distinguish between the two surfaces without contact (tight clearance) or between two contacting surfaces (close contact). In the present study we expand on those findings and utilize small-scale mixing models that are able to, for the first time, distinguish between tight clearances and tight contact. In this study we evaluated different mixer types including a top-mounted mixer, several impeller-based bottom-mounted mixers, and a rotary piston pump. The impact of tight clearance/close contact on subvisible particle formation in at-scale mixing platforms was demonstrated in the gap between the impeller and drive unit as well as between the piston and the housing of the pump. Furthermore, small-scale mixing models based on different designs of magnetic stir bars that mimic the tight clearance/close contact of the manufacturing-scale mixers also induced subvisible particles in mAb formulations. Additional small-scale models that feature tight clearance but no close contact (grinding) suggested that it is the repeated grinding/contacting of the moving parts and not the presence of tight clearance in the processing equipment that is the root cause of protein subvisible particulate formation. When multiple mAbs, Fabs (fragment antigen binding), or non-antibody related proteins were mixed in the small-scale mixing model, for molecules investigated, it was observed that mAbs and Fabs appear to be more susceptible to particle formation than non-antibody-related proteins. In the grinding zone, mAb/Fab molecules aggregated into insoluble particles with neither detectable soluble aggregates nor fragmented species. This investigation represents a step closer to the understanding of the underlying stress mechanism leading to mAb subvisible particulate formation as the result of drug product processing.LAY ABSTRACT: Mixing and fill finish are important unit operations in drug product manufacturing for compounding (dilution, pooling, homogenization, etc.) and filling into primary packaging containers (vials, pre-filled syringes, etc.), respectively. The current trend in adopting bottom-mounted mixers as well as low fill-volume filling systems has raised concerns about their impact on drug product quality and process performance. However, investigations into the effects of their use for biopharmaceutical products, particularly monoclonal antibody formulations, are rarely published. The purpose of this study is three-fold: (1) to revisit the impact of bottom-mounted mixer design on monoclonal antibody subvisible particle formation; (2) to identify the root cause for subvisible particle formation; and (3) to fully utilize available particle analysis tools to demonstrate the correlation between particle count in the solution and filter fouling during sterile filtration. The outcomes of this study will benefit scientists and engineers who develop biologic product manufacturing processes by providing a better understanding of drug product process challenges.