Biomedical Engineering and Bioengineering Commons^™

Comparison Of Bioreactor Systems Operated At High Bacterial Cell Density For The Production Of Lactic Acid: Batch – Cstr – Cstr Cascade – Tubular Reactor, Ulrich Kulozik

Integrated Continuous Biomanufacturing II

Subject of the work was the microbial conversion of lactose, an abundant processing by-product in the dairy industry, to lactic acid. Lactic acid serves as a preservative in many areas in various product sectors, but its potential applications go far beyond this. In industrial applications lactic acid as a renewable material can be applied as a building block for novel technical materials such as biodegradable films for packaging purposes in replacement of materials based on fossile raw materials.

Purpose of the work was to compare different bioreactor systems with regard to achievable lactic acid concentrations and volumetric productivities. The goal …

Continuous Downstream Process Or Connected Batch Process: Which One Makes Most Sense For Biogen?, John Pieracci, Sanchayita Ghose, Venkatesh Natarajan

Integrated Continuous Biomanufacturing II

As biologics-based products move into therapeutic areas with large patient populations and high doses, batch processing may not be able to keep pace with product demands. At Biogen, we have been exploring a number of options that can enable higher productivity of our downstream processes. In addition to a fully “end to end” continuous process, a batch process comprised of several steps connected in series has been evaluated. In this presentation, technologies Biogen has evaluated to enable either continuous or connected processing will be shown. Multi-Column-Chromatography (MCC) for the Protein A capture chromatography step was evaluated in order to maximize …

Continuous Culture In The Age Of Single-Use, Christel Fenge, Melisa Carpio

Integrated Continuous Biomanufacturing II

Continuous culture offers a number of advantages, e.g. higher titres and constant culture environment and is the method of choice for challenging therapeutic proteins such as coagulation factors. Additional opportunities arise for the commercial production of monoclonal antibodies using so called concentrated fed-batch mode where the product is retained in the bioreactor and fresh medium is constantly perfused through the culture and combining it with single use production technologies. This approach is an effective way to reduce time to market as additional scale-up steps after phase III are not required to produce large amounts of product whilst benefiting from the …

Pat Concepts For The Process Monitoring And Control Of Continuous Biomanufacturing, Eike Zimmerman, Jeff Goby, Anoushka Durve

Integrated Continuous Biomanufacturing II

Process Analytical technologies (PAT) are a key component to fully leverage the success of continuous manufacturing. The timely measurement of critical quality attributes and critical process parameters assures that the desired product quality is being consistently manufactured and non-conforming material being identified. PAT tools can be utilized to meet the expectations for in-process monitoring, and allows for identification and isolation of rejected in-process materials. It further supports process development, as continuous and fast response of process to factor changes allows efficient experimentation and increases process understanding within the range of conditions studied during development. Ultimately, PAT will allow Real Time …

Upstream Perfusion Process: Back To The Future, Jean-Marc Bielser, Henri Kornmann, Herve Broly

Integrated Continuous Biomanufacturing II

At an early stage of bioprocess science, continuous operations were the workhorse in the industry. Therefore, EMD-Serono has legacy with continuous upstream processes as many of its molecules such as Rebif are being produced in perfusion mode. These processes from the 90’s consume large volumes of commercial media and demonstrate low productivities. Cells are mainly attached because no performant cell retention device existed for cells in suspension at that time.

Around 10-15 years ago, the industry decided to move on fed-batch operations. EMD-Serono also re-oriented towards fed-batch operations. Molecules such as Erbitux are now being produced in 15,000 L stainless …

Development Of Functionally Closed Downstream Operations For Continuous Biomanufacturing Of Recombinant Therapeutic Proteins, Rohan Patil, Rahul Godawat, Tarl Vetter, Yogesh Waghmare, Konstantin Konstantinov, Veena Warikoo

Integrated Continuous Biomanufacturing II

Traditionally, upstream processing in biologics manufacturing is performed in an aseptic and functionally closed manner while downstream processes are open operations designed to maintain a low-bioburden state. Consequently, the downstream process components such as skids, column hardware, and chromatography resins currently available have been designed for low bioburden rather than aseptic operations. One of the key design elements of Integrated Continuous Biomanufacturing (ICB) is the linkage of upstream and downstream operations and the capability of the system to run for prolonged durations at ambient temperature. Thus, a critical design criterion for ICB is the ability of the system to maintain …

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 …