Biomedical Engineering and Bioengineering Commons^™

Continuous Production Of Viral Vaccines With A Two-Stage Bioreactor System, Felipe Tapia, Yvonee Genzel, Ingo Jordan, Volker Sandig, Udo Reichl

Integrated Continuous Biomanufacturing II

Continuous processes can be particularly efficient for production of biologicals that are required in large amounts such as viral vaccines. One virus that has received much clinical attention is Modified Vaccinia Ankara virus (MVA), which is a potential platform for expression of recombinant viral antigens and can be used as a vector in gene therapy [1]. Recently, a new MVA virus strain has been successfully propagated at high yields in non-aggregated avian suspension cells [2] allowing the production of MVA virus in continuous bioreactors. MVA is a lytic DNA virus and therefore, continuous production strategies can be implemented using two-stage …

Bioprocess Economics And Optimization Of Continuous And Pre-Packed Disposable Chromatography, Richard Allmendinger, Suzanne Farid

Integrated Continuous Biomanufacturing II

The biotech sector is facing increasing pressures to design more cost-efficient, robust and flexible manufacturing processes. Standard batch chromatography (BATCH) is an established but expensive approach to separate impurities related with both E.coli and mammalian cells expression systems. This study uses a computational framework to investigate if the application of continuous chromatography (CONTI) and disposable technologies can provide a competitive alternative to BATCH and reusable equipment. A set of general assumptions is presented on how some of the key downstream processing characteristics, such as chromatography operating conditions, resin properties and equipment requirements, vary as a function of the chromatography mode …

Mathematical Modeling Of A Bioreactor Producing Epo-Hr Operating In Perfusion Mode, Osman Fernandez, Raydel Alvarez, Ernesto Chico, Adolfo Castillo, Julio Dustet

Integrated Continuous Biomanufacturing II

The interest in use perfusion mode has increased in the last years, due to an increased awareness of perfusion advantages, some general improvement in equipment reliability, and a broadening of operational skills in the biomanufacturing industry. However, mathematical modeling of bioreactors in continuous mode with cell retention (perfusion mode) with concomitant cell bleeding is still emerging, because this scheme has a few applications in global biotechnology industry. The case of study was industrial fermentation process of CHO cells producing EPO-hr. The model involves a total of 7 equations and 19 variables. In order to fix the degrees of freedom were …

Toward Complete Continuity In Antibody Biomanufacture: Multi-Column Continuous Chromatography For Protein A Capture And Mixed Mode Hydroxyapatite Polishing, Anthony Grabski, Tom Van Oosbree, Beth Hammer, Alla Zilberman, Robert Mierendoff

Integrated Continuous Biomanufacturing II

Monoclonal antibodies (mAbs) are a predominant modality for a broad range of clinical indications including oncology and inflammatory diseases. Increasing manufacturing capacity and decreasing cost per purification campaign are critical factors for making antibody therapies more affordable. Cell culture mAb expression levels have steadily increased over the past ten years with titers of five grams per liter frequently achieved. Higher titers reduce production costs and allow processing of kilogram quantities for clinical trials from single use cell culture vessels. Drawbacks of increased titers include higher levels of aggregates, fragments, variants, and process impurities. These combined titer and impurity burdens further …

Strategy For Scaling Semi-Continuous Downstream And Integration Of Process Analytical Tools For Monoclonal Antibody Toxicology, Darshini Shah, Rebecca Chmielowski, Colette Cutler, Hong Li, David Roush, Nihal Tugcu

Integrated Continuous Biomanufacturing II

Currently, continuous processing for biologics is being pursued from a technology perspective in order to increase productivity and decrease cost of manufacture. Disruptive technologies, such as PCC (Periodic Counter-current Chromatography), have been researched extensively in order to reduce the bottleneck for the downstream capture chromatography step. However, there is extremely limited knowledge in how to scale these disruptive technologies for clinical and commercial manufacturing. In this presentation, we demonstrate a strategy for scaling and integration of a PCC capture step into a semi-continuous downstream process (Protein A chromatography-Viral inactivation-Filtration-Anion exchange chromatography). Harvest cell culture fluid (HCCF) feed streams ranging from …

Biopharmaceutical Capacity Planning For Batch And Semi-Continuous Bioprocesses Under Various Strategic Criteria, Cyrus Siganporia, Thomas Daszkowski, Andreas Schluk, Brijesh Rao, Lazaros Papageorgiou, Suzanne Farid

Integrated Continuous Biomanufacturing II

Biopharmaceutical companies with expanding portfolios of commercial therapeutics face increasing pressure to meet market demands whilst minimising costs and capital expenditure. Attaining optimal production plans is made more problematic by portfolios containing products with different production modes: batch and semi-continuous. Semi-continuous perfusion-mode products often exhibit a distinct separation of upstream and downstream manufacturing via an intermediate freezing step. This flexibility adds further complexity which needs to be efficiently overcome during the optimisation process. An added complexity to having different process modes is that changeover times are different, leading to computationally expensive sequence-dependent changeover times. Considering the implications of incorrect capacity …

Development Of A First Generation Perfusion Process And Medium For Continuous Processing Based On Existing Fed-Batch Platform Media, Henry Lin, Samantha Wang, Robert Leighty, Scott Godfrey, Samet Yildirim, Adam Osborne, Flaka Radonigi, Jens Vogel, Jon Coffman

Integrated Continuous Biomanufacturing II

Process intensification leveraging perfusion offers tremendous potential for yield improvement over fed-batch processes for the production of monoclonal antibodies. In the context of continuous processing, the goal is to achieve highly intensified perfusion processes that allow substantial footprint reduction and enable flexible adaptation in new facilities. However developing a perfusion process and medium without prior technology requires leveraging the existing fed-batch platform knowledge. Evolving a medium for perfusion relies on designing suitable mixtures of basal and feed media that serve as adequate starting points for development. Focus on optimization of the medium to decrease byproduct waste, reduce unnecessary cell growth …

Scale Up And Implementation Of A High Density Long-Term Perfusion Suspension Cell Culture In A 250l Single Use Bioreactor, Weichang Zhou, Hang Zhou

Integrated Continuous Biomanufacturing II

As part of efforts to develop a continuous processing platform for biologics manufacturing using a single use bioreactor, we have been focusing on development of several initial unit operations: high density perfusion suspension cell cultures and early product capture steps, in order to realize its potentials of being flexible, improving product quality and lowering costs. These initial unit operations require large volumes and represent the most important part of this processing platform. By integrating these steps into a continuous operation, we can deliver the largest benefits of this processing platform. In this presentation, we will discuss our efforts towards continuous …

From Fed-Batch To Perfusion: Productivity And Quality Considerations For A Late-Stage Program, Sen Xu, Hao Chen

Integrated Continuous Biomanufacturing II

In this poster, we will present the rapid development of a perfusion process with high productivity for late-stage manufacturing. The process uses a Chinese Hamster Ovary (CHO) cell line for monoclonal antibody production. Prior to the development, the clinical supply was manufactured by a fed-batch process with the same cell line.

To achieve desired productivity and comparable quality attributes, we developed a repeated batch shake flask model to rapidly screen media and process conditions. Shake flasks were inoculated at 10 × 10⁶ cells/mL and maintained in a CO₂ incubator with medium exchange for every 48 hours, repeated for …

Continuous Purification Of Hepatitis C Virus-Like Particles By Multi-Column Chromatography, Ricardo Silva, Alex Xenopoulos, Jose Mota, Cristina Peixoto, Manuel Carrondo, Paula Alves

Integrated Continuous Biomanufacturing II

Novel biopharmaceutical products, such as virus-like particles (VLPs) and viral vectors, constitute a challenging task for downstream processing (DSP). Recoveries achieved to reach required purities are significantly inferior compared to more common antibody and other recombinant processes. Therefore, the biopharmaceutical industry is looking for alternative downstream strategies capable of improving purification yields and decreasing cost while maintaining product quality. One of many possible improvements to DSP is to replace single-column batch operation by continuous, or semi-continuous, multi-column chromatography.

A single-column batch chromatographic operation used for capture is limited by the dynamic binding capacity (DBC) of the target product. For high-value …

Innovations In Bioreactor Operational Modes – Hybrid Semi-Continuous Processes To Push Beyond The Limits Of Conventional Fed-Batch Cultures, Gregory Hiller, Matthew Gagnon, Ana Maria Ovalle, Wenge Wang, Elizabeth Eydelman

Integrated Continuous Biomanufacturing II

Due to the limits of feed volume addition and the problem of amino acid counter ion, miscellaneous osmolyte, and cell growth inhibitor accumulation, the fed-batch mode of bioreactor operation for CHO cell production of protein therapeutics is inherently limited with respect to the cell densities and productivities that are achievable. Continuous or perfusion culture with cell retention can overcome some of these limitations, but suffers from the disadvantages of large volume media consumption, long times to reach peak cell densities, and complications with cell retention devices. We will describe hybrid versions of continuous culture that overcome many of these limitations …

Bhk Cells Physiological Response To Spin-Filter Stress Condition, Bruno Labate, Aldo Tonso

Integrated Continuous Biomanufacturing II

Mammalian cells are the preferable platform for secreted pharmaceutical proteins due to the compatibility of posttranslational modifications performed by these cells with therapeutic applications. However, to obtain a desirable quantity of these proteins, high cell concentrations are also needed. Baby hamster kidney BHK-21/C13 cells, adapted to single-cell suspension culture growth were cultivated in batch mode in a Biostat B (Sartorius AG, Germany) bioreactor with a 1,5 L working volume vessel. The temperature was controlled at 37 ºC, pH at 7.2 with CO­₂, agitation at 80 rpm and dissolved oxygen at 50% of air saturation. The culture medium used …

Mab Product Consistency Achieved In Long Duration Microfiltration-Based Cho Perfusion Process, Douglas Rank, Christopher Martin, Lee Madrid, Michael Phillips

Integrated Continuous Biomanufacturing II

Perfusion processes have traditionally been used for the generation of unstable proteins in cell culture systems. The use of perfusion for production of stable proteins has been limited by low product concentration, media costs, and system complexity. However, with the advent of new single-use technology, cell culture media specifically formulated to support high density perfusion, and high-producing cell lines, perfusion processes are gaining widespread industry attention. Additionally, perfusion processes are considered an integral part of the “Factory of the Future” vision through enabling continuous processing while delivering a product effluent with consistent product quality and concentration.

In this study, we …

Evaluation Of A Continuous Chromatography Process Through Process Modeling And Resin Characterization, Ketki Behere, Seongkyu Yoon

Integrated Continuous Biomanufacturing II

Increased product yield at reduced cost and time has been the driving force of any manufacturing process. However, inevitable increase in future demand for biopharmaceutical drugs, along with intensified competition and stringent regulatory laws have exhibited an imperative need for established platform processes. In this regard, the implementation of continuous processing, in whole or in components, has demonstrated to increase the manufacturing productivity with key impact on speed, cost and facility implications. While, upstream operations have confirmed increased productivity by implementation of continuous processing components, such as perfusion culture, similar adaptation for downstream processes has been limited. We have evaluated …

Continuous Production Of Proteins: Integration Of Polishing Using Mcsgp, Fabian Steinebach, Daniel Karst, Massimo Morbidelli

Integrated Continuous Biomanufacturing II

Continuous counter-current processes for polishing steps overcome the purity-yield tradeoff of classical batch chromatography by internal recycling of overlapping regions of the chromatogram. Particularly in the case of challenging separation tasks this leads for the same product quality to significantly higher yields and productivity. Processes using these fundamental principals have been successfully implemented in the chemical industry up to multi-ton production.

In an integrated continuous process, with a perfusion bioreactor and a continuous capture process, the product quality of the capture product is different compared to classical batch processing. The lower residence time in the bioreactor and the higher loading …

Exometabolome Characterization Of High Cell Density Culture Perfusion And Optimization Of The Cell Specific Perfusion Rate, Veronique Chotteau, Leila Zamani, Ye Zhang, Magnus Aberg, Anna Lindahl, Axel Mie

Integrated Continuous Biomanufacturing II

High cell density perfusion has the potential to generate intensified processes. A target for such perfusion process can be to maintain a steady state culture around 100 x 10⁶ cells/mL. A challenge for the development of these processes is to minimize the medium consumption, to reduce the costs associated with this raw material and with the harvest processing. This is particularly true for processes at densities such as 100 x 10⁶ cells/mL for which several reactor volumes per day are typically necessary.

We have shown that cell specific perfusion rate (CSPR) is an approach, which has proven to …

Continuous Downstream Processing Of A Monoclonal Antibody Using Periodic Counter Current Chromatography (Pcc) And Straight Through Processing (Stp), Hans Blom, Christer Eriksson, Annika Forss, Helena Skoglar

Integrated Continuous Biomanufacturing II

There is an increased interest to perform process intensification in order to reduce costs and improve throughput in the development and production of monoclonal antibodies (MAbs). One solution to these demands can be to implement continuous or semi-continuous downstream processing. New emerging technologies such as periodic counter-current (PCC) chromatography and straight through processing (STP) are entering the market. Here, these two technologies were evaluated in a continuous three step chromatography MAb process. The capture step was performed with protein A media (resin) on a 3 column PCC chromatography system followed by two polishing steps which were connected in series with …

Integrated Solutions For Continuous Processing In Mobius® Bioreactor Systems, Andrew Clutterbuck

Integrated Continuous Biomanufacturing II

Throughout the bioprocess industry, the use of disposable systems such as bioreactors is playing an increasingly significant role. Manufactures are recognising the strategic benefits of single use systems in both reducing production costs and increasing overall manufacturing throughput.

Over the past few years there has been increasing demand for larger scale disposable bioreactor systems, to satisfy the manufacturing demands and an emphasis on suppliers such as Merck Millipore to demonstrate robust and reproducible scale-up within these systems.

This presentation will explore the design philosophies behind the Mobius^® bioreactors and will also give an overview of the novel SensorReady external …

Optical Enzymatic Sensors For Continuous Monitoring Of Bioreactors, Kenneth Reardon, Brian Heinze, Trenton Danna

Integrated Continuous Biomanufacturing II

Continuous monitoring of biotechnological processes is important for control and optimization of quality and productivity. Typically, samples must be removed from the cultivation and analyzed in a laboratory to determine the concentrations of substrates and products. These time-delayed data cannot be used for real-time process control. The Process Analytical Technology initiative of the FDA supports the use of on-line measurement techniques for process development, production, and quality. An overview of sensors will be presented, focusing on in-situ sensors that contact the biological medium, and external sensors that contact the medium either optically (ex situ) or via a sterile (and disposable) …

Salt-Tolerant Cation Exchange Hd-Sb Hydrogel Membrane: Mab Purification Performance In Flowthrough Mode, Dharmesh Kanani, Daniel Luo, Nirali Paghdal, Navneet Sidhu, Annabel Shang, Jim Stout

Integrated Continuous Biomanufacturing II

Development of Protein A based purification platforms have simplified the downstream processing of monoclonal antibodies (mAb), the largest component of biopharmaceuticals. The ever increasing titer of the cell culture process is putting more pressure on the downstream process to further increase its productivity. The classical Protein A based mAb purification platform consists of two polishing steps, bind and elute cation-exchanger and flowthrough anion-exchanger. Cation-exchange chromatography is very efficient at the separation of HCP, leached Protein A and product-related impurities such as aggregates and fragments in bind and elute mode. However, anion-exchange chromatography is a proven technology to remove DNA, viruses, …