Trends in upstream and downstream process development for. Aggregates in monoclonal antibody manufacturing processes. Aggregates in mab therapeutics pose a risk to patients. Analysis of monoclonal antibodies and their fragments by size. Monoclonal antibodies are expensive to develop and to manufacture. Reliable, timely and costeffective monoclonal antibody production and purification programs. Removing aggregates in monoclonal antibody purification. Review aggregates in monoclonal antibody manufacturing processes mar. Stress factors in mab drug substance production processes. Evolution of the monoclonal antibody purification platform. As a result, aggregation has to be monitored throughout process development and manufacturing of a mab product. There is a clear need to reduce costs and manufacturing risk in order to.
Capto adhere is a multimodal ion exchanger and is designed to be used as a second or third step in monoclonal antibody mab puri. In the broad sense, it refers to the entire process of creating a usable specific antibody, including steps of immunogen preparation, immunization, hybridoma creation, collection, screening, isotyping, purification, and labeling for direct use in a particular method. Extremes of ph, ionic strength, temperature, concentration, shear forces, and other processing conditions can lead to increased aggregation. Nonsizebased membrane chromatographic separation and. The term antibody production has both general and specific meanings. Production processes for monoclonal antibodies intechopen. Biomanufacturing of monoclonal antibodies mab involves a number of unit operations, including cell culture in a bioreactor followed by chromatography and filtration. Recovery and purification process development for monoclonal. In custom antibody development, once target selection.
Today, antibody concentrations of 35 gl are achieved routinely and some companies have attained up to 10 gl in fedbatch processes 5,6,8. Aggregation kinetics for monoclonal antibody products. The control and avoidance of aggregation in the manufacturing process are needed because aggregates affect drug performance and safety97,98. The agilent advancebio sec column, used with the agilent 1260 biolc system, proved capable of delivering high resolution analysis of mab aggregates with short run times. Analysis of monoclonal antibodies and their fragments by. Aggregates in monoclonal antibody manufacturing processes marava. The study of aggregation propensity of a monoclonal antibody mab and its. Aggregation of mabs continues to be a major problem in their developability.
Size exclusion chromatography analysis of antibody drug. Impact of media and antifoam selection on monoclonal. Aggregates in monoclonal antibody manufacturing processes ifm. Here, we explore the potential of predicting and reducing the aggregation propensity of monoclonal antibodies. Monoclonal antibodies mabs are the fastestgrowing biological therapeutics with important applications ranging from cancers, autoimmunity diseases and metabolic disorders to. To investigate antibody stability and formation of modified species under upstream processing conditions. Mab aggregates may form during protein expression, downstream processing, and storage, and may be caused. Given the high cell densities achievable in both mammalian cell culture and microbial processes, primary recovery can be a significant challenge at lab, pilot and commercial manufacturing scales. Current trends in monoclonal antibody development and. Aggregation mechanism of an igg2 and two igg1 monoclonal antibodies at low ph. Factors influencing biotherapeutic monoclonal antibody. Watch our experts discuss which steps in the biomanufacturing process are involved in the formation or removal of aggregates and industryleading approaches.
Thus, values obtained from sec may not actually represent the true aggregate levels in the protein drug container, so there is a continued effort to. Taken together, these developments are driving the industry to explore alternative separation technologies as a future manufacturing strategy. Adcs travel to target cells, where the antibody binds to its antigen expressed on the cell surface. The stability of 11 purified monoclonal human igg1 and igg4 antibodies, including an igg1based bispecific crossmab, was compared in downscale mixing stress models. Jan 16, 20 to investigate antibody stability and formation of modified species under upstream processing conditions. Since the approval of the first therapeutic monoclonal antibody in 1986. Due to their broad application range, monoclonal antibodies mab are used worldwide in a variety. Quality attributes of monoclonal antibodies and effect of cell culture. This study demonstrated the suitability of cationexchange laterallyfed membrane chromatography lfmc for rapid and highresolution separation of monoclonal antibody mab aggregates. Monoclonal antibodies mabs are the fastestgrowing biological therapeutics with important applications ranging from cancers, autoimmunity diseases and metabolic disorders to emerging infectious diseases.
Aggregation mechanism of an igg2 and two igg1 monoclonal. Monoclonal antibodies mab or moab are antibodies that are made by identical immune cells that are all clones of a unique parent cell. Schematic manufacturing process of monoclonal antibodies from cell. The small molecules attached to the mab are often relatively hydrophobic. Table 1 summarizes potential sources of aggregate formation during. Aggregate content of cht xt eluate mab s significant reduction of the mab s aggregate was achieved unosphere supra eluate. Antibodydrug conjugates adcs are monoclonal antibodies coupled to cytotoxic agents with stable linkers. Introduction to antibody production and purification. For example, we have seen issues in manufacturing, such as quality, process. Aggregation and chemical modification of monoclonal. Review aggregates in monoclonal antibody manufacturing processes marava.
For the final biopharmaceutical product approval and subsequent manufacturing processes, a comprehensive characterization of mab purity, aggregate forms, and charge variants is required by the regulatory agency. One of these molecules was further evaluated in realistic bioreactor stress models and in cell culture fermentations. Protein a chromatography increases monoclonal antibody aggregation rate during subsequent low ph virus inactivation hold. A read is counted each time someone views a publication summary such as the title, abstract, and list of authors, clicks on a figure, or views or downloads the fulltext. In the broad sense, it refers to the entire process of creating a usable specific antibody, including steps of. The stability of 11 purified monoclonal human igg1 and igg4. Prediction and reduction of the aggregation of monoclonal antibodies. It has been more than a decade since the industry started establishing the process platform.
Adcs travel to target cells, where the antibody binds to its antigen expressed on. Largescale manufacturing of pharmaceutical antibodies. Antibody aggregation could be triggered by partial unfolding of its domains, leading to monomermonomer. Monoclonal antibodies mabs are a successful class of therapeutic products used increasingly in the past 1520 years, but the manufacture of safe and effective mab drug products provides many challenges to downstream molecule purification. Monoclonal antibodies can have monovalent affinity. Guideline on development, production, characterisation and. To demonstrate the analysis of monoclonal antibody mab aggregates.
These include production cell culture, harvest, antibody capture by protein a, virus. Aggregates have been characterized in mab purification and formulation. Mar 25, 2018 aggregates in monoclonal antibody manufacturing processes. Current trends in monoclonal antibody development and manufacturing will provide readers with an understanding of what is currently being done in the industry to develop, manufacture, and.
Integrated flowthrough purification for therapeutic. Predicting aggregation propensity and monitoring aggregates. Pdf aggregates in monoclonal antibody manufacturing. Clarification technologies for monoclonal antibody. But this method can provide misleading results because aggregates can dissociate or form during sec andor adsorb to the column resin. Jan 09, 2018 the formation of aggregates is a common but highly undesirable occurrence during monoclonal antibody purification. Processing impact on monoclonal antibody drug products. Largescale manufacturing of pharmaceutical antibodies is a complex activity that requires considerable effort in both process and analytical development. Protein a chromatography pac is commonly used as an efficient capture step in monoclonal antibody mab separation processes.
Preparative separation of monoclonal antibody aggregates. Imbaratto, cell culture manufacturing, covance biotechnology services, inc. Overall purification performance of cht xt chromatography is summarized in table 1. A systematic framework to optimize and control monoclonal. Low ph elution can induce mab aggregation in protein a chromatography 7, 8. Understanding and managing aggregates are critical to your monoclonal antibody mab process. Pdf monoclonal antibodies have proved to be a highly successful class of therapeutic products. This 31 st article in the elements of biopharmaceutical production series focuses on evolution of the purification platform for manufacturing of mab therapeutics. Antibody aggregation can occur at various stages including the up and downstream processes in manufacturing, formulation, and longterm storage 5, 6.
Humanized monoclonal antibody produced by mammalian cell culture may contain significant amounts of antibody dimers and smaller amounts of higher order aggregates. Lang2 1manufacturing science and technology, lonza biologics porrin. Managing aggregates in your monoclonal antibody mab process. Secretory leakage of igg1 aggregates from recombinant chinese. See how oncolumn aggregation is dependent on the type of resin used and how one particular highresolution cation exchange resin allows for the lowest proportion of aggregate formation. The study focuses on very large scale processes in the region of five 5 tonnes per annum of bulk antibody. Monoclonal antibody production a report of the committee on methods of producing monoclonal antibodies institute for laboratory animal research national research council national academy press washington, dc 1999. Largescale production has revolutionized the market for monoclonal antibodies by boosting its production and becoming a more practical method of production. The agilent advancebio sec column, used with the agilent 1260 biolc system, proved capable of. Secretory leakage of igg1 aggregates from recombinant. Hic as a complementary, confirmatory tool to sec for the. Strategies for the assessment of protein aggregates in pharmaceutical biotech product development. Introduction to antibody production and purification thermo. Trends in upstream and downstream process development.
In this talk, herb lutz, global consultant, will discuss the formation of protein. Current trends in monoclonal antibody development and manufacturing will provide readers with an understanding of what is currently being done in the industry to develop, manufacture, and release. This hydrophobicity can be problematic during manufacturing and storage in terms of aggregate formation as well as in analytical characterization of the latter1. Monoclonal antibody production in commercial scale cell culture bioprocessing requires a thorough understanding of the engineering process and components used. Removal of leached protein a, antibody dimers and aggregates da, host cell proteins hcp, viruses and. Given the high cell densities achievable in both mammalian cell culture and microbial processes, primary recovery can be a. Aggregates in the final drug product are undesirable for two major reasons. Pdf aggregates in monoclonal antibody manufacturing processes. Dec 16, 2016 aggregates in monoclonal antibody manufacturing processes biotechnol bioeng 2011, 108 7, 1494508. After completing this chapter, students will be able to. In this talk, herb lutz, global consultant, will discuss the formation of protein aggregates and their. The structure of the monoclonal antibody should be justified with respect to its mechanism of action. The outcomes of this study will benefit scientists and engineers who develop biologic product manufacturing processes by providing a better understanding of drug product.
Review aggregates in monoclonal antibody manufacturing processes mara va. The current trend for cell culture processes is to increase. Upon binding, the full adc can be internalized by a process called receptormediated endocytosis. During the manufacturing process, the protein is exposed to multiple stress. Monoclonal antibodies have proved to be a highly successful class of therapeutic products. Simultaneous removal of leached proteina and aggregates from monoclonal antibody using hydrophobic interaction membrane chromatography. This study demonstrated the suitability of cationexchange laterallyfed membrane chromatography lfmc for rapid and highresolution separation of monoclonal antibody. Size exclusion chromatography analysis of antibody drug conjugates using the agilent 1260 infinity ii bioinert lc system. For the final biopharmaceutical product approval and subsequent manufacturing processes, a comprehensive characterization of mab. Purification is intended to remove impurities, such as protein aggregates, but some such operations may actually generate protein aggregation 1.
Are you considering how best to manage and remove aggregates across your template. Usp manufacturing processes which generate higher titers still take. The formation of aggregates is a common but highly undesirable occurrence during monoclonal antibody purification. Monoclonal antibody mab products have emerged as an extremely important and valuable class of therapeutic products for the treatment of cancer and other diseases such. Preparative separation of monoclonal antibody aggregates by. The primary objective of this project was to use the experimental analysis together with the knowledge of chemical kinetics to build a kinetic model for protein. This 31 st article in the elements of biopharmaceutical production series focuses. In the creation of this study, we used the industry standard modelling software, biosolve process, to address these questions. Removing the aggregates of an acidic monoclonal antibody with. Various economic analyses estimate that process development and clinical manufacturing costs can. Aggregates in monoclonal antibody manufacturing processes biotechnol bioeng 2011, 108 7, 1494508. Selecting the optimal resins for aggregate removal references he x et al.
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