Reducing biopharmaceutical manufacturing costs through continuous processing in a flexible J.POD® facility.

In this work, process models were developed to capture the impact of biomanufacturing costs on a commercial scale and emphasize the way in which facility design and operation must balance meeting product demand while minimizing production costs. Using a scenario-based modeling approach, several facility design strategies were evaluated, including a traditional large stainless-steel facility and a small footprint, portable-on-demand (POD)-based facility. Bioprocessing platforms were compared by estimating their total production costs across different facility types and specifically illustrating how continuous bioprocessing has gained in popularity as a novel and cost-effective approach to manufacture high-quality biopharmaceuticals. The analysis showed how fluctuations in market demand have a dramatic effect on manufacturing costs and plant utilization, with far-reaching implications on the total cost to patients.

Recovery modeling of tangential flow systems.

The demand for increased formulation concentrations for protein therapeutics puts a significant strain on already existing tangential flow filtration (TFF) systems that were constructed with lower protein concentration targets as part of their design criteria. TFF is commonly used to buffer exchange and concentrate the product to the appropriate drug substance concentration. Analyzing the ability of an existing TFF system to process under conditions outside its original design specifications can be challenging. In this analysis, we present a systematic approach to assess the operational limits of a TFF process with consideration of system performance parameters for changing process targets. In two new engineering diagrams, the recovery efficiency diagram and the operating space plot, all relevant operational constraints and parameters are related to allow rapid process fit evaluation. The engineering assessment of TFF systems presented in this article allows a rational review of system limitations during process fit evaluations of existing TFF systems. It also provides a rational basis for targeted system upgrades and setting system design specifications for the design of new systems if existing systems are found inadequate.

An alternate diafiltration strategy to mitigate protein precipitation for low solubility proteins.

Application of the minimum diafiltration (DF) time solution for a monoclonal antibody resulted in a 20-h process time rather than the expected 12 h. Further investigation indicated high turbidity associated with a product solubility issue that caused a flux decline. As a result, the gel flux model and the associated minimum DF time were not predictive. Multiwell plate solubility screening confirmed that the protein passed through a region of low solubility during the ultrafiltration step. Multiple approaches to address this issue were considered and a new strategy involving variable volume diafiltration (VVDF) was developed. Process modeling and simulation were used to predict performance and to select a value of the DF ratio control parameter (buffer flow/permeate flow = 0.65). Feasibility testing at the bench and pilot scales confirmed that the new strategy reduced solubility issues, fit within existing manufacturing tank volume and system area constraints, matched model predictions, and did not present significant implementation issues. Recommendations are made regarding the general value of this strategy, when it should be used, and how to implement it.

Use of MMV as a Single Worst-Case Model Virus in Viral Filter Validation Studies.

Typical platform processes for biopharmaceutical products derived from animal cell lines include a parvovirus filtration unit operation to provide viral safety assurance of the drug product. The industry has adopted this platform unit operation and gained a wider understanding of its performance attributes, leading to the possibility of streamlined approaches to virus clearance validation. Here, the concept of virus validation on a parvovirus-grade filter with a single worst-case model virus is presented. Several lines of evidence, including published literature and Amgen's own data, support the use of a parvovirus, such as mouse minute virus (MMV), as a worst-case model virus to assess virus removal by parvovirus filters. The evidence presented includes a discussion of the design and manufacture of virus filters with a size exclusion mechanism for removal. Next, the characteristics of different model viruses are compared and a risk assessment on the selection of the relevant model viruses for clearance studies is presented. Finally, a comprehensive summary of literature and Amgen data is provided, comparing the clearance of larger viruses against MMV. Together, this analysis provides a strong scientific rationale for the use of a single, worst-case model virus for assessing virus removal by parvovirus filters, which will ultimately allow for more efficient and streamlined viral clearance study designs. LAY ABSTRACT: Demonstrating the virus clearance capability of a purification process is an important aspect of biopharmaceutical process development. A key component of the viral safety of the process is the inclusion of a parvovirus-grade filter as an effective and robust virus removal step. Traditional methodologies for viral clearance studies have been based on a conservative, data-intensive approach, but recent trends in the field of virus clearance and process development show evolution towards streamlined and more efficient study designs that are based on understanding the mechanism of viral clearance by downstream unit operations. The publication of scientific datasets and awareness of the underlying mechanisms involved with these unit operations have fueled this trend. Here, the concept of virus validation on a parvovirus-grade filter using a parvovirus as single, worst-case model virus is presented. Multiple lines of evidence are provided to support this proposal, including a review of published literature and Amgen historical data. The adoption of this approach provides benefits in terms of cost savings for executing viral clearance studies, but it also simplifies the necessary dataset and focuses on only supplying value-added information to demonstrate the viral safety of the process.

Multipronged approach to managing beta-glucan contaminants in the downstream process: control of raw materials and filtration with charge-modified nylon 6,6 membrane filters.

(1→3)-ß-D-Glucans (beta-glucans) have been found in raw materials used in the manufacture of recombinant therapeutics. Because of their biological activity, beta-glucans are considered process contaminants and consequently their level in the product needs to be controlled. Although beta-glucans introduced into the cell culture process can readily be removed by bind-and-elute chromatography process steps, beta-glucans can also be introduced into the purification process through raw materials containing beta-glucans as well as leachables from filters made from cellulose. This article reports a multipronged approach to managing the beta-glucan contamination in the downstream process. Raw material screening and selection can be used to effectively limit the level of beta-glucan introduced into the downstream process. Placement of a cellulosic filter upstream of the last bind-and-elute column step or effective preuse flushing can also limit the level of contaminant introduced. More importantly, this article reports the active removal of beta-glucan from the downstream process when necessary. It was discovered that the Posidyne(®) filter, a charge-modified nylon 6,6 membrane filter, was able to effectively remove beta-glucans from buffers at relatively low pH and salt concentrations. An approach of using low beta-glucan buffer components combined with filtration of the buffer with a Posidyne membrane has been successfully demonstrated at preparative scale. Additionally, the feasibility of active removal of beta-glucan from in-process product pools by Posidyne membrane filtration has also been demonstrated. Based on the data presented, a mechanism for binding is proposed, as well as a systematic approach for sizing of the Posidyne filter.

Qualification of a novel inline spiking method for virus filter validation.

Virus filters are widely used in bioprocessing to reduce the risk of virus contamination in therapeutics. The small pores required to retain viruses are sensitive to plugging by trace contaminants and frequently require inline adsorptive prefiltration. Virus spiking studies are required to demonstrate virus removal capabilities of the virus filter using scale down filters. If prefiltration removes viruses and interferes with the measurement of virus filter LRV, the standard approach is to batch prefilter the protein solution, spike with virus, and then virus filter. For a number of proteins, batch prefiltration leads to increased plugging and significantly lower throughputs than inline prefiltration. A novel inline spiking method was developed to overcome this problem. This method allows the use of inline prefiltration with direct measurement of virus filter removal capabilities. The equipment and its operation are described. The method was tested with three different protein feeds, two different parvovirus filters, two virus injection rates; a salt spike, a bacteriophage spike, and two mammalian virus spikes: MMV and xMuLV. The novel inline method can reliably measure LRV at throughputs representative of the manufacturing process. It is recommended for applications where prefiltration is needed to improve throughput, prefiltration significantly reduces virus titer, and virus filter throughput is significantly reduced using batch vs. inline prefiltration. It can even help for the case where the virus preparation causes premature plugging.