Nanobodies (12–15 kDa) are antibody fragments derived from naturally occurring heavy-chain–only antibodies from camels and llamas ( 9, 13). ScFvs are single-chain antibody fragments (28 kDa) that contain variable regions of MAb heavy (VH) and light (VL) Ig chains fused together (using antibody engineering) by a flexible peptide linker that usually contains 10–25 amino acids ( 10). Of all such molecules, most clinical progress has been made with scFvs and nanobodies (Table 2). They are less costly to produce than conventional MAbs because they can be chemically synthesized or manufactured using nonmammalian expression systems. All those are thermally stable, soluble, and maintain their binding and antigenic specificity. These include single-domain (dAbs), nanobody (from camelids), single-chain variable fragment (scFv), antigen-binding fragment (Fab) ( 10), Avibody, minibody, CH2D domain, Fcab ( 11, 12), and bispecific T-cell engager (BiTE) molecules. Small antibody-based formats derived from preexisting MAbs were the first type of protein scaffolds to be developed. Some advantages that protein scaffolds offer over MAbs include small size (12–50 kDa), high thermal stability and solubility, better tissue penetration and accumulation, low production or synthesis costs, and intellectual property protection and freedom to operate around high value MAb-directed targets ( 7, 8, 9). Both have yielded smaller molecules that possess most of the desirable features of MAbs (e.g., antigen specificity and binding) but lack many of the limitations mentioned above. Such platforms can be divided into two major groups: antibody-derived and nonantibody-derived protein scaffolds. Finally, the complex molecular structure of MAbs typically renders them thermally unstable, which requires that they be stored at refrigeration temperatures to ensure stability and clinical efficacy ( 5).īecause of such limitations, scientists over the past 10–15 years began to develop novel and alternative MAb engineering platforms. In some cases, MAbs are unable to bind to epitopes on the surface of protein targets that are accessible only by smaller-sized molecules ( 8). That can reduce their effectiveness in treating some types of cancers (e.g, solid tumors). Third, most MAbs (because of their large size) are limited in their ability to penetrate and accumulate in tissues. Second, their large size (150–160 kDa) makes them unsuitable for most intracellular therapeutic targets, so MAbs must be delivered to patients by injection or infusion ( 7). That process can be extremely labor intensive and costly ( 6). First, their complex molecular structure (four polypeptide chains - two light and two heavy), glycosylation of heavy chains, and at least one disulfide bond in each of several immunoglobulin domains require that full-size antibodies be manufactured in eukaryotic or mammalian expression systems (e.g., CHO cells). Some Clinical Limitations of Therapeutic MAbsĪlthough MAbs are one of the biotechnology industry’s greatest commercial successes to date, they possess several characteristics that can limit their utility and clinical effectiveness ( 4, 5). Currently, ~70 chimeric and humanized MAbs and 30 fully human MAbs are in various stages of clinical development worldwide ( 2). To that point, Humira (Abbott) - one of the first fully human MAbs to be marketed as a treatment for rheumatoid arthritis and other chronic inflammatory indications (Table 1) - is one of the world’s top-selling drugs with sale projected to exceed $9.3 billion in 2012 alone ( 3). Such investments - coupled with major advances in MAb engineering technology - led to development of fully-human MAbs (molecules that contain only human antibody protein sequences).įully human MAbs generally exhibit improved therapeutic efficacy and possess fewer side effects (better safety profiles) than chimeric or humanized MAbs. The commercial success of those early products prompted huge investments by pharmaceutical and biotechnology companies in therapeutic MAb drug development. KEYWORDS: ANTIBODY FRAGMENT ANALYSIS FLOW CYTOMETRY, IMMUNOHISTOCHEMISTRY RADIOLABELING PROTEIN FUSION WHO SHOULD READ: PRODUCT DEVELOPMENT, ANALYTICAL PRODUCT FOCUS: MONOCLONAL ANTIBODIES, FRAGMENTS, AND OTHER PROTEINS Since their introduction, many of those products - including Remicade, Enbrel, and other drugs - have attained blockbuster status. The first MAbs to be commercialized in the early 2000s were chimeric MAbs (genetically engineered hybrid molecules containing murine and human sequences) and “humanized” MAbs (containing 90–95% human antibody protein sequences). Table 1: Commercialized monoclonal antibodies in the United States and European Union ()
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