Enzolytics Inc (ENZC)

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Therapeutic monoclonal antibodies

The biopharmaceutical market, like the global pharmaceutical market, has been expanding every year and is expected to reach approximately 230 billion dollars in 2018, twice the sum in 2010 [1]. It is forecasted to be >380 billion dollars in 2024 [1]. Recombinant proteins, followed by therapeutic antibodies, have been the main contributors to the biopharmaceutical market in the past. However, in 2017 [2] the sales of antibody-based medicines took on the leading position, with expected sales of 172.8 billion dollars in 2022, about 20% of the global pharmaceutical market [2].

It is noteworthy that the range of therapeutic monoclonal antibodies has been remarkably increasing since the late 1990s, the total number approved by the US FDA reaching 64 in 2018 [2,3] (see Table 1 list for those from 2015 to 2017). In 2017, a total of 11 therapeutic monoclonal antibodies were approved. There are interesting changes in the characteristics of the antibodies attracting attention. As shown in Figure 1, the percentage of human and humanized antibodies in the total has dramatically increased. The reasons for continuous expansion of the market for therapeutic antibodies include an increase in the number of approvals, efforts to explore other target diseases for the already approved therapeutic antibodies, as well as improvements in formulations and dosage forms. Finding new target antigens will be of critical importance for further development of antibody-based medicines in the future.64 therapeutic monoclonal antibodies have been approved by the US FDA. The ratio of human and humanized monoclonal antibodies to the total sum of therapeutic monoclonal antibodies is increasing year by year.

Bispecific antibodies [6–10], antibody–drug conjugates [11], sugar chain-modified antibodies [12–15] and low molecular weight antibodies [6,16] are now being emphasized as next-generation products. Some bispecific antibodies and antibody–drug conjugates are already on the market, with emicizumab [17] as an example of the former, and gemtuzumab ozogamicin [18], inotuzumab ozogamicin [19] and brentuximab vedotin [20] featuring in the latter.

There is another important point regarding further development of therapeutic monoclonal antibodies. A critical reason for their employment is their high specificity and affinity for targeted antigens. It is known that monoclonal antibodies can specifically recognize two types of epitopes (Figure 2). One is linear in the primary structures of proteins. The other is conformational, dependent on secondary and tertiary structures. In addition to their primary, secondary and tertiary structures, proteins may also exhibit quarterly structures formed by hetero- or homosubunits, which provide unique interfacial geometries on their complexes. Intact native proteins mainly feature secondary and tertiary structures, but various kinds of monoclonal antibodies do not necessarily have specificity for conformational structures, although they have been raised against a large number of proteinous antigens.
Epitope recognition by antibodies is generally divided into two types. One is linear epitope-specific recognition and the other is conformational or discontinuous epitope specific.

Such specific recognition of conformational structures of proteins would be expected to give more strict selectivity and affinity for therapeutic purposes. Notably, stereo-specific monoclonal antibodies recognizing 3D configurations of molecules, offer advantages over linear epitope-specific monoclonal antibodies, which only recognize 2D configuration. Again we should stress that target antigens for therapy predominantly feature secondary and tertiary structures. Monoclonal antibodies recognizing primary structures can have affinity for only restricted regions in intact antigens, because linear epitopes can be masked by conformational folding. Thus, the employment of stereo-specific monoclonal antibodies should be our goal.

Production of stereo-specific monoclonal antibodies against different proteins
The next point is how to produce stereo-specific monoclonal antibodies. Until very recently, no practical technologies have been available for their generation, because of difficulties like how to immunize a mouse maintaining the structure of the antigen intact in the presence of adjuvant. While adjuvants generally allow more effective sensitization, they usually disrupt the original protein native structure. If an adjuvant is not used, immunization efficiency may be very low. Another difficulty is strict selection of sensitized B lymphocytes secreting stereo-specific monoclonal antibodies. Even when immunization is successful with a native intact antigen, the number of desired sensitized B lymphocytes is usually extremely small, accounting for only a few percent of the total spleen cells after repeated immunization.

However, we have now established an original stereo-specific targeting (SST) technique [21–25]. The protocol promises efficient generation of stereo-specific monoclonal antibodies on the basis of hybridoma technology, as illustrated in Figure 3. One of the critical points is strict selection of the required sensitized B lymphocytes by intact antigens expressed on myeloma cells through B-cell receptors (BCRs). This precise selection also enables efficient formation of B-cell and myeloma-cell complexes for generating hybridoma cells secreting the aimed for antibodies.
https://www.futuremedicine.com/doi/full/10.2217/imt-2018-0130