1.1 Summary
Antibodies, a main component of the immune response, have been recognized, more than a century ago, for their proven therapeutic value. The hybridoma fusion technology, proposed in the early 1970s, for the first time gave easy access to the production and engineering of murine monoclonal antibodies. The potential of these new molecules, as laboratory tools, was largely exploited during the two following decades. At present antibodies, still omnipresent in both diagnostic and research domains, have progressively come to dominate the field of immunotherapy. New technologies, such as phage display, humanized transgenic mice, and repertoire mining, have been proposed, allowing for the isolation of fully human antibodies. The natural complexity of the antibody molecules and the rapid implementation of engineering methodologies helped in making them ideal candidates for new applications and for the solution of complex immunotherapeutic challenges. The first chapter is a current update on the different antibodyâderived molecules as well as a survey of the latest antibody engineering technologies. In addition the chapter reviews the critical issue of the development of expression systems suitable for largeâscale and costâeffective production of recombinant antibodies.
1.2 Introduction
The historical roots of immunotherapy trace their origins to the end of the nineteenth century. In collaboration with ShibasaburĆ Kitasato, the first bacteriologist to succeed in cultivation of Clostridium tetani, Emil Adolf von Behring demonstrated the efficacy of âsoâcalledâ antitoxins to protect animals against tetanus and diphtheria [1, 2]. Although not immediately recognized, the discovery was adapted to the treatment of diphtheriaâstricken children and was found to significantly reduce mortality [3]. For some years serum therapy was widely adopted before being gradually replaced by active immunization. Several decades passed before the composition, physiological behavior, and chemical nature of the antitoxin components were thoroughly characterized.
These components, referred to as gamma globulins according to their electrophoretic mobility and as immunoglobulins and antibodies according to their immunological function, were at the center of multiple investigations involving a steadily increasing number of renowned scientific teams. From the early 1950s until now, the definition of antibody (Ab) became more precise, as well as the genetic aspects behind the creation of their molecular diversity. In parallel, advances in cellular biology were under way around the world, allowing a better understanding of the precise role of B cells in the immune response. Paving the way toward the advent of modern immunotherapy, hybridoma technology described by Köhler and Milstein in 1975 gave, for the first time, easy access to murine monoclonal antibodies (mAbs) [4]. mAbs were recognized as revolutionary laboratory tools from their inception, although the in vivo applications and therapeutic potential of these molecules were still controversial, raising some skepticism in the early 1980s. Nevertheless, significant progress in molecular biology techniques allowed mAbs to move from research to diagnostics and applications in therapy.
More recently, monoclonal and recombinant antibodies have become the focus of new technologies, such as bacteriophage or other in vitro display techniques, mice or larger animal transgenesis, and other technologies permitting direct access to fully human antibodies. Importantly, the pressing need for large quantities of mAbs was a major driver for the development and optimization of recombinant protein production systems. The contribution and complementarity of these different approaches will be considered in the context of largeâscale industrial production of therapeutic mAbs.
1.3 New Markets for Old Antibodies, Old Markets for New Antibodies
To date, more than 40 mAbs have been approved by the US Food and Drug Administration (USFDA) or European Medicines Agency (EMA) for therapeutic applications: 5 are of murine origin, including 2 bispecific constructs, 8 chimeric, 18 humanized, and 13 fully human. Thanks to the novel technologies available [5], fully human antibodies are rapidly taking over the market, and it seems probable that even humanized molecules will be marginalized in the coming years. The composition of the earlyâstage antibody pipeline is indicative of this trend. In terms of revenue, substantial returns have been realized both on existing and new markets. A good illustration is provided by recent data from Lawrence and Lahteenmaki: [6] among the top 10 selling biological drugs of 2014, 5 are mAbs. HumiraÂź (adalimumab, AbbVie Inc., North Chicago, IL, the United States), with applications to several conditions (such as rheumatoid arthritis, juvenile rheumatoid arthritis, Crohnâs disease, psoriatic arthritis, psoriasis, ankylosing spondylitis, and ulcerative colitis), is number one, with a total revenue exceeding $10.5 billion. RemicadeÂź (infliximab, Janssen Biotech, Inc., Titusville, NJ, the United States) for multiple indications (rheumatoid arthritis, psoriatic arthritis, ulcerative colitis, Crohnâs disease, ankylosing spondylitis, and severe or disabling plaque psoriasis) is in second place with a total revenue over $9.2 billion. RituxanÂź (rituximab, Biogen Idec Inc., Cambridge, MA, the United States, and Genentech USA, Inc., San Francisco, CA, the United States) for multiple indications (rheumatoid arthritis, chronic lymphocytic leukemia/smallâcell lymphocytic lymphoma, nonâHodgkinâs lymphoma, antineutrophil cytoplasmic antibodyâassociated vasculitis, indolent nonâHodgkinâs lymphoma, and diffuse large Bâcell lymphoma) is found in the sixth position with a total revenue over $7.5 billion. AvastinÂź (bevacizumab, Genentech USA, Inc., San Francisco, CA, the United States) and HerceptinÂź (trastuzumab, Genentech USA, Inc., San Francisco, CA, the United States) are, respectively, ranking seventh and ninth positions with individual revenues fluctuating between $6.8 and $7.0 billion.
Beside the success stories of some blockbusters, mAbs are set to play a role in the rapid control of emergent diseases. A striking example is the 2014 Ebola outbreak, for which more than 20 laboratories and research groups around the world, including those from Canada, Japan, Israel, Uganda, and the United States, are working simultaneously to develop therapeutic mAbs against the virus. The dire state of emergency will no doubt facilitate and shorten the approval process. A mixture of three mAbs known as ZMapp (LeafBio, Inc., San Diego, CA, the United States), never tested in humans, was exceptionally accepted despite the fact that very little is known regarding the safety and effectiveness of this treatment.
1.3.1 Intellectual Property
One possible explanation for the great commercial success of therapeutic antibodies is the proper management of intellectual property and accurate designation of strategies for protecting antibodies or antibodyâderived products [7]. This is particularly meaningful if we consider the emblematic and wellâknown Cabilly patent filed by Genentech, one of the most ubiquitous patents in biotechnology, which covers a fundamental method for the production of therapeutic recombinant mAbs that cannot be ignored by anyone planning to commercialize an antibody [8]. Besides the uniqueness of this example, one should not underestimate the inextricably complex patent situation governing the recombinant antibody world. As a matter of illustration of this complexity, according to Sandercock and Storz [9], it might be possible to patent the precise definition of an epitope with the possibility to claim laterâgeneration antibodies targeting the same epitope.
1.3.2 Biosimilars
A consequence of the astronomical commercial value of mAbs associated with the patent expiry date (or estimated expiry date) is their attractiveness as candidates for biosimilar drugs [10]. The European Union (EU) has been the first to establish a regulatory framework for marketing authorization application (MAA) and has named âcopycatâ biotherapeutic products with the term biosimilars, a term also recently adopted by the USFDA [11].
Biosimilars offer a highly attractive mechanism for reducing the cost of medical care and thus should be considered positively [12]. Nevertheless, the abbreviated approval pathway associated with their adoption requires rein...
