2020

Technology to Transform AAV Manufacture

Adenoassociated virus (AAV) vectors are a popular choice for modern gene therapies because of their favorable safety profile, low immunogenicity, and the ease with which they can be transduced into different cell and tissue types. An AAV genome is a single strand of DNA comprising a replication (rep) gene, which encodes regulatory proteins involved in genome replication, and a capsid (cap) gene, which produces three capsid proteins. However, AAVs cannot replicate alone. In nature, AAV shares an exquisite relationship with…

Streamlined Polishing and Viral Clearance Using a New Hybrid, Biomimetic, Single-Use Anion Exchanger

Flow-through anion-exchange (AEX) chromatography is used frequently in biopharmaceutical purification processes for reduction of net–negatively charged host-cell proteins (HCPs) and viruses as part of a validated viral clearance strategy (1, 2). AEX column chromatography is the technology most often used for electrostatic viral clearance, particularly in commercial-scale biopharmaceutical manufacturing, for which columns have a long-established history of reliable and well-understood performance (3). Still, validation of HCP and viral clearance by AEX columns in biopharmaceutical processes involves complexities that contribute significantly…

Simple and Effective Method for Purification of DMT-On Oligonucleotides Using HIC Resins

Within the biopharmaceutical industry, oligonucleotide drug pipelines have increased significantly because of the effectiveness of such drug products in treating devastating diseases. Downstream specialists need improved purification techniques for such highly valuable materials. Dimethoxytrityl (DMT) is used to synthesize oligonucleotides and temporarily mask the characteristic chemistry of the 5Í´-hydroxy functional group. DMT can be left on an oligonucleotide following synthesis to provide stability to a molecule during subsequent processing. Herein, we describe a novel, effective, and high-recovery method for purifying…

eBook: Mixed-Mode Chromatography for Purification of Biopharmaceuticals

Mixed-mode chromatography offers several advantages in downstream processing of biotherapeutics. Mixed-mode chromatography resins use ligands that are capable of at least two modes of interaction with solutes such as hydrophobic, ion exchange, and metal affinity. The interactions between stationary and mobile phases that result from those combinations enhance chromatographic selectivity, facilitating separation efficiencies that are not possible using other chromatography media. As this eBook illustrates, the multimodal approach can save developers time and money by enabling robust purification of biopharmaceuticals…

Endotoxin Questions and Answers

Endotoxin — also known as lipopolysaccharide (LPS) — is a large molecule made up of a lipid and a polysaccharide portion found in the outer membrane of each Gram-negative bacterial cell. Examination of endotoxins is necessary for microbial and animal cell-culture operations that produce active pharmaceutical ingredients (APIs) and vaccines. If a cell culture is contaminated with endotoxins, it could affect cell growth and function (thus potentially affecting product quality). Biopharmaceutical companies need to understand the endotoxin levels within their…

Ask the Expert: Mass Spectrometry for Relative and Absolute Quantitation of Process-Related Impurities

Qualitative and quantitative analyses of process-related impurities are critical to manufacturing a safe, high-quality drug product. During his 17 June 2020 “Ask the Expert” webcast, Steven Broome (BioPharmaSpec) described using mass spectrometry (MS) for such analyses. Focusing on isopropyl-β-D-thiogalactopyranoside (IPTG), kanamycin, and host-cell proteins (HCPs), Broome overviewed best practices for using MS to detect and quantitate low levels of impurities. Broome’s Presentation Chemicals used in biotherapeutic production can be present as impurities in the resulting drug product. These impurities can…

eBook: Raw Material Control Strategy — Leveraging Knowledge of Material Attributes and Data Analytics as Key Elements

Ensuring pharmaceutical quality begins with in-depth understanding of process/platform capabilities, which is informed by knowledge gained through product and process development, subject-matter expertise, and lessons learned from experience. And all outside factors that can affect manufacturing outcomes must be taken into consideration. Extra vigilance is necessary for understanding potential sources of variation and maintaining robust control strategies to ensure process consistency — and ultimately product quality for patients. Biomanufacturing unit operations require multiple raw materials that must be documented as…

eBook: Antibody–Drug Conjugates — Refining Product Designs for Improved Outcomes

Antibody–drug conjugates (ADCs) seek to partner the target specificity of antibodies with the cell-killing punch of chemotherapy drugs. Researchers identify antibodies that bind to proteins found predominantly or exclusively on the surfaces of cancer cells. The cells can absorb the ADC into their interiors, where the chemical environment or enzymes detach the drug from the antibody, freeing it to wreak havoc. Although nine ADCs have received US Food and Drug Administration (FDA) or European Medicines Agency (EMA) approval (and many…

Updating the Economics of Biologics Manufacturing with 5,000-L Single-Use Bioreactors: A Paradigm Shift

Single-use technologies enable a flexibility and modularity effectively unattainable with more traditional stainless-steel technologies, particularly in upstream bioprocesses. Single-use bioreactors up to 2,000 L are employed largely in preclinical- and clinical-stage bioprocesses to leverage this flexibility. As products reach commercial maturity, scales larger than 2,000 L frequently become desirable to take advantage of economies of scale. With the typical upper limit of single-use bioreactors at 2,000 L, this has traditionally meant transfer to stainless-steel systems. The introduction of the Thermo…

eBook: Viral Vector Purification — A Discussion of Current Challenges and Methods

Adenoassociated viral (AAV) vectors have become synonymous with gene therapy delivery. However, because they are produced in such small quantities and because their upstream processes carry comparatively large amounts of host-cell DNA and other impurities, AAV purification can be challenging. Several researchers have applied different chromatographic strategies, but no universal method has been adopted in the biopharmaceutical industry. This eBook features a discussion among several industry experts that explores challenges specific to AAV purification, shedding light on whether current strategies…