Downstream Development

Special Report: Turning Discoveries into Products — Developability Assessments and Highly Efficient Process Design

High costs and long timelines for biopharmaceutical development are cause for reflecting on how best to allocate resources from the earliest discovery stage through critical go–no-go junctures. With inputs ranging from science, engineering, and economics, the coined term developability becomes the synthesis of answers to such questions as How well does the target represent a disease state? Does manipulating that state bring about improvement? Does the molecule behave as expected in living systems? What can be done about the emergence of independent safety, toxicology, and/or immunogenicity warning signs? Can the molecule…

The Secret Life of Protein A

Affinity chromatography with protein A has become the foundation for purification of nearly every therapeutic IgG in commercial production. One of the features most responsible for its success has been its compelling simplicity. IgG binds. Contaminants do not. Load, wash, and elute pure IgG. In the real world, however, protein A does not elute pure IgG. It typically contains several hundred to a few thousand parts per million (ppm) contamination by host-cell proteins (HCPs) and other contaminants. Numerous studies demonstrate…

Identification and Quantification of Heat-Shock Protein 70: A Major Host-Cell Protein Contaminant from HEK Host Cells

Recombinant therapeutic proteins are commonly produced by cell lines such as Chinese hamster ovary (CHO), human embryonic kidney (HEK) 293, murine myeloma (NS0), and Escherichia coli bacterial cells. Host-cell proteins (HCPs) are indigenous proteins produced by those expression hosts and considered to be process-related impurities generated from the cell culture process (1). HCPs are potentially harmful and immunogenic to patients, and they can compromise the stability of protein drugs (2–4). For those reasons, HCPs must be consistently removed or reduced…

Challenges in Implementing Quality By Design: An Industry Perspective

In the fall of 2004, the US Food and Drug Administration (FDA) published a final report entitled Pharmaceutical CGMPs for the 21st Century: A Risk-Based Approach (1). This publication set the groundwork for a prospective risk‑based approach to pharmaceutical product development. It was published on the heels of a November 2003 agreement between the FDA and the International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use (ICH) to develop an internationally harmonized plan for developing…

Optimization and Scale-Up of HCIC-Based MAb Purification Processes, Part 2

In multistep schemes, hydrophobic charge-induction chromatography (HCIC) has been shown to contribute effectively to clearance of Chinese hamster ovary (CHO) host-cell proteins (CHOPs), DNA, and viruses. When used for capture chromatography, HCIC can provide better aggregate clearance than protein A sorbents can. Chen et al. enhanced clearance of aggregates, CHOPs, and product- related impurities by controlling HCIC based on both pH and the presence of binding-promoting salt in the wash and elution buffers used (1). Taken together with our findings…

An Industrial Platform Solution for Antibody Fragment Purification

Compared with traditional approaches such as chemotherapy and radiotherapy, monoclonal antibodies (MAbs) have become the most successful cancer treatments in the past 20 years (1). With great clinical success in many therapeutic areas, MAbs now account for >40% of the entire biotechnology drug market, and sales are projected to be >US$160 billion over the next few years in the United States alone (2). More than 35 MAbs have been approved for clinical use, and hundreds more are filling industry development…

Optimization and Scale-Up of HCIC-Based MAb Purification Processes, Part 1

Monoclonal antibodies (MAbs) serve important medical needs in cancer treatment as well as that of autoimmune and infectious diseases (1). Antibodies are also widely used in clinical diagnostic assays. They can be coated on solid surfaces to bind specific analytes, conjugated to reporter molecules (either as whole antibodies or fragments) for analyte detection, used in sensitivity panels for lot-release testing, and supplied as positive controls in diagnostic kits (2). Our study evaluates the use of hydrophobic charge-induction chromatography (HCIC) for…

Foundation Elements for Cell Therapy Smart Scaling

Cell therapy is the injection of cellular material into patients. The injected cell-therapy product (CTP) usually consists of intact living cells. In recent years, cell therapies have evolved and matured, moving from academia to industry. That maturation is reflected in the number of open clinical trials that include the term cell therapy in their descriptions: To date, there are more than 8,700 open trials listed on the US National Institutes of Health’s online database (clinicaltrials.gov), most of which are in…

A Multidisciplinary Approach to Manufacturing Biotherapeutics

Optimizing antibody manufacturing processes has gone beyond the first-order goal of achieving elevated protein titers and now also focuses on understanding biologic and manufacturing process variables that define cellular machinery and protein quality. A holistic approach to biotherapeutic manufacturing incorporates several applied disciplines such as biology, engineering, process control, signal processing, and modeling to reduce the “black-box” model of cell- based protein production into an operational design space. This is in line with the US Food and Drug Administration’s quality…

Affinity Capture of F(ab’)2 Fragments: Using Twin-Column Countercurrent Chromatography

Antibody fragments are potent active drug substances (1–4). Because they lack glycosylation, they can be produced using different biological expression systems, including yeast and microbial systems as well as mammalian cells. These molecules are interesting as biopharmaceuticals because they are smaller than full-size antibodies and therefore may penetrate better into different tissues. Antibody fragments are cleared faster in biological systems because they lack the Fc antibody structural region (4). However, fragments may be conjugated to increase their size for improved…