BioPharm

18 BioPharm International eBook April 2019 www.biopharminternational.com Outsourcing Resources Manufacturing cells. Emicizumab, a bsAb engi- neered on a full-length immuno- globulin G4 (IgG4) and approved in 2017, binds to the blood clotting factors IX and X, resulting in the same function as factor VIII but easier to manufacture (4). Antibody constructs, for example Amgen's BiTE technology that uses two dif- ferent Fab fragments with a linker, "pair up" tumor cells with endog- enous T- cells, w ith the goal of enhancing effector functions. A n t i b o d y d r u g c o n j u g a t e s (ADCs), although not new to the arsenal, are seeing a revival as a potential solution to some of med- icine's great unsolved problems, i nc lud i ng c rossi ng t he blo o d- brain barrier or treatment of solid tumors. Originally conceived to carry a toxic small-molecule drug to cancer cells, several groups are now focusing on other payloads, including nano-particles, for tar- geted radiotherapy or antibiotics that target latent infection (5). The number of ADCs in clinical trials more than tripled from 2013 to 2017 (6), and the global market for next-generation antibodies is projected to generate $11.6 billion in sales in 2020 (7). BETTER MABS BY DESIGN The development focus is not just about structure in this new "fam- ily" of mAb formats. Drugmakers can also influence the functional- ity and manufacturability of these next-gen mAbs by designing at the protein sequence level, under- scor ing the need for advanced development a nd ma nu fac t u r- ing know-how. Achieving the cor- rect manufacturing capabilities at the early stages of development reduces the risk of meeting prob- lems in manufacturing or patient toxicity later on in the lifecycle. Aggregation of a mAb, either dur- ing manufacture or upon storage, is a common problem because it can affect both the efficacy and safety of the drug. Today, mAb protein sequences are able to be screened in silico to classify molecules based upon their propensity to aggregate using descriptors relevant to biopro- cessing and manufacturing (8). Immunogenic reactions to a bio- logic drug can appear as allergic reactions or generation of antibod- ies that reduce the drug's efficacy or mediate unwanted immuno - genic reactions. Immunogenicity assessments in the early stages of drug development using a com- bination of in-silico, in-vitro, and in-vivo tools facilitate candidate desig n a nd selec t ion w it h low immunogenicity risks (9). During the past 10 years, anti- body engineering has focused on development of "fit-for-purpose" m A bs w it h i nc re a se d ha l f-l i fe and/or modulated effector func- tions. These include increased or muted antibody- dependent cell med iated c y totox ic it y (A DCC), a n t i b o d y - d e p e n d e n t c e l l u l a r phagocytosis (ADCP), and com- plement-dependent cytotoxicit y (CDC). Tuning effector functions c a n b e ac h ie ve d b y e it he r Fc e n g i ne e r i n g — mo d i f ic at io n o f the amino acid sequence of the Fc reg ion to create novel com- bi nat ions of ef fe c tor f u nc t ion (e.g., oc rel i z u mab) — or by gly- coengineering, typically used to enhance ADCC activity by reduc- ing or eliminating fucose residues on t he m Ab. Cu r rent ly, seven m A b s w it h mo d i f ie d e f f e c tor function are approved for thera- peutic use, and there are more in late-stage clinical trials. CREATING THE RIGHT TOOL BOX The task at hand now is to develop protein engineering and manu- facturing processes that solve the diff ic ulties associated w ith the complexity of the emerging second generation of mAbs and proteins. Scalable production requires a cus- tomizable "toolkit" of synthetic biology capabilities, rather than a "one-size-fits-all" approach. Vectors While standard mAbs contain two different polypeptides, many next- generation biologics, especially bsAbs based on a full-length IgG format, are formed of three or four different poly peptides. Mak ing a bsAb in a single cell requires expression of the four polypep- tides in an optimal ratio, best done by putting the four genes into the same genomic locus using a single gene expression vector. To overcome the challenges associ- ated with high levels of sequence homolog y in the bsAb, a more advanced vector architecture than that typically used with standard mAbs is required. Lonza has devel- oped architectures that allow the insertion of three or four polypep- tides into a single vector, which facilitates selection, results in a more homogeneous product, and simplifies the route from discovery to market. For bioconjugates, a new genera- tion is emerging with a focus on improved pharmacok inetics, in particular controlling the number of "payload" molecules bound to the mAb via linkers. This approach reduces variability in toxin load and therefore results in a more homogenous product that has more consistent efficacy. Site-specific conjugation (SSC) vectors allow the sub-cloning of the variable regions needed for specificity with constant regions that have precisely defined bioconjugate attachment points. Cell lines The dominance of Chinese ham- ster ova r y (CHO) cel ls i n t he

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