Development of biopharmaceutical proteins (e.g. monoclonal antibodies) is mainly performed in Chinese Hamster Ovary (CHO) cells. Currently, the main purification step remains the capture step which is mostly a Protein A affinity step. The new Fibro nanofibers might be a new approach for the efficient and fast processing of the monoclonal antibody product as capture step.

Nanofitins are non-antibody protein scaffold selected for their (i) specificity to capture only product of interest & ensure high purity, (ii) appropriate affinity to allow elution under target-compatible conditions, (iii) high stability for conserved performance after multiple column regenerations. The affinity columns have been tested by GSK in comparison with their current vaccine’s purification process, resulting in less DSP steps (1 capture step instead of 3) and higher yields (x2).

The main goal in designing affinity ligands is to discover or improve functionality, such as stability or selectivity. In this talk, we will show how rationally designed chemical combinatorial libraries support the development of robust peptidomimetics that can be easily adapted to several targets and to chromatographic and non-chromatographic methods.

Single domain antibodies, or nanobodies, are attractive modalities for therapeutic and diagnostic applications. Their small size allows better tissue penetration and targeting of epitopes inaccessible to standard antibodies. Combining a novel cellulose-based matrix named Fibro PrismA and an adaptable auto-sampler with a liquid chromatography system facilitated the successful implementation of an automated high-throughput downstream processing platform, providing the flexibility and efficiency required for the purification of nanobodies.

Advances in membrane technology facilitated the generation of novel Protein A membrane devices with high capacity, suitable for affinity chromatography operations at short residence times. Using affinity membranes, high productivity operation modes have been attained on standard liquid handling chromatography systems at lab and pilot scale. We'll show the results of a GORE Protein Capture Device pilot scale evaluation targeting maximum productivity within typical operating pressures.

Virus-based biotherapeutics have progressed considerably in the last years. To address some purification process bottlenecks, as the removal of the residual DNA impurities, a universal purification tool by applying an innovative chromatographic media was implemented. Also, innovative analytical technologies to assess product quality and contribute for a more efficient manufacturing of oncolytic enveloped viruses will be discussed.

Over the last two years, the bioprocessing industry has been focused on the rapid development of mRNA production processes. This increased interest in gene therapies has also resulted in new considerations for manufacture of AAV and extracellular vesicles. This talk aims to discuss the challenges the industry faces when developing advanced therapies for future medicines, with case study data covering our own experiences working in this field.

• The effectiveness of continuous production of jetted agarose resins
• Recent developments and commercialization of jetted Protein A resins

• Expanding jetting technology to polymers in ion exchange and other chemistries

Chromatographic separation of biopharmaceuticals is carried out using soft porous and compressible resin particles. This restricts the flow rates, and in turn the productivity of columns packed with such particles. Also, the speed of separation is limited by pore mass transport. These problems could be solved using unconventional cuboid (or box-shaped) chromatography devices. In this presentation, the use of such devices for separation of protein biopharmaceuticals will be discussed.

FDA guidelines on continuous manufacturing suggest measuring and modeling the residence time distribution of an entire process chain as part of the process characterization. The residence time distribution in counter-current chromatography protein A chromatography was determined by pulse injection of a fluorescent-labeled monoclonal antibody. The tracer spreads in the elution peak of the different cycles and a very wide RTD is obtained as suggested by the model.

A proposed technology that gained interest for the downstream processing to replace chromatography systems is precipitation. For a successful development and implementation of a precipitation process, it is important to acquire process data as fast as possible. We successfully designed and developed millidevices for continuous protein precipitation to determine the most appropriate conditions, such as dosage time of the precipitating agent, resulting filterability, and apparent solubility of impurities and antibodies. The simple and flexible design allows a continuous, controlled, and precise addition of the precipitating agent without valves and reducing the number of pumps required.

Understanding the influence of mixing behavior of protein refolding operations will contribute to a better understanding of scalability. Here, we applied computational fluid dynamics (CFD) and performed further trials with a scale-down model (SDM) to increase insight into the main flow pattern of a two-phase system. Based on the obtained results, we proved that reliable SDM representative for the performance of manufacturing equipment together with high-throughput (HTP) analytics are key factors for fast process development for tomorrow´s biotherapeutics.

The omnipresent enzyme Horseradish peroxidase (HRP) is still produced from hairy root cultures in a time-consuming and inefficient way. I will present a novel and scalable recombinant HRP production process in Escherichia coli that yields a highly pure, active, and homogeneous single isoenzyme. We successfully developed a multi-step inclusion body process giving a final yield of 960 mg active HRP/L culture medium with a purity of = 99 %. Our preparation of recombinant, unglycosylated HRP from E. coli is a viable alternative to the enzyme from plants and highly interesting for therapeutic applications.

This presentation will focus on the development of high-affinity, highly specific diagnostic and therapeutic binding molecules based on the creation of two novel phage displayed libraries of a uniquely stable and hydrophilic protein scaffold. This novel scaffold has molecular properties designed to achieve significant impact as a new generation of powerful tools in areas of clinical need, both as diagnostic and therapeutic entities. Processing of specific candidates will be described.

Increasing cell culture and chromatography performance, virus filtration feeds have a higher protein concentration. The high concentration and the resulting potential for aggregation, can cause lower filterability. However, the main benefit is the reduced volume and filtration surface area, providing the nanofilter can handle such challenging solutions. Planova BioEX exhibits high throughput with high concentration mAb solutions and an adsorptive prefiltration can improve the filterability by removing aggregates.

Multiple purification technologies have been developed to expedite process development and expand manufacturing capacity. Affinity resins can be developed with breakthrough success rates and timelines. Pre-packed column reproducibility and configurability focuses suite resources on productivity. System flow path and hardware technologies improve gradient control, shear and dead volume towards overall recovery yield and bioactivity. Technical concepts for each platform will be discussed.