Evolution of Therapeutic Antibodies, Influenza Virus Biology, Influenza, and Influenza Immunotherapy

Table 1

Comparison on some attributions of the conventional four-chain antibodies and engineered antibodies.

Attribution	Conventional four-chain antibodies	Engineered antibodies (four-chain and fragments)
Selection	In vivo or achieved via hybridoma technology which need repeated animal immunization; Hybridoma technology requires tissue culture facility and hand on experience; Antigen: must be immunogenic with appropriate dose, route, etc.; Host factors to be considered: genetics, MHC and immune status	In vitro: antibody coding genes can be selected from bacteria, yeast, and mammalian display systems; Animal-free system (alleviates animal welfare concern). Antigen: no restriction; Free from influence of host status
Generating time	Relatively long process	Relatively short-time (less than 4 weeks to get antigen binding clones from the display systems)
Production	Hybrodomas require tissue culture facility and expensive culture medium	Various and flexible expression systems including bacterial, yeast, and mammalian; High yield can be obtained, for example, from optimal mammalian expression system
Reproducibility	In vivo: animal-to-animal and batch-to-batch variation in quality	Low batch-to-batch variation; No life-time limit
Genetic stability	Genetic drift (hybridomas)	Relatively more stable
Molecular structure	Mostly unknown	Known DNA sequence information, defined structure (CDRs and FRs)
Format	Four chains with strict species, isotypes, subisotypes	Can be four-chain or engineered at genetic level to preferable formats (to suit the purpose of use): chimeric, humanized, fully human, F(ab)′2, Fab, scFv, sdAb, multi-valent, multimeric, and many other possibility
Purity	Antibodies from in vivo immunization and hybridoma culture can be contaminated with the host proteins, and disease causative and adventitious agents from animal derived raw material	Can be purified to be free from adventitious agents with high purity (up to 99.8% at GMP level); Animal-free raw material
Affinity	Usually high but cannot be improved or modulated	Can be improved and modulated by in vitro affinity maturation, point mutation(s) or resurfacing of the antigen binding site
Cell penetrating ability	No; inaccessible to intracellular target	Yes, by linking molecularly to a cell penetrating peptide; thus, can be accessible to the intracellular target
Half-life in vivo	Can be several weeks (isotype-matched)	Can be many hours to several weeks depending on the designed format; increased longevity and pharmacokinetics can be done, such as by PEGylation, multimerization, or modulating IgG/FcRn interaction; The cell-penetrating antibody (superantibody) can stay in vivo for relatively long period of time as they can cross the membrane of all cells but get accumulated intracellularly only where the target antigen is present. Thus, disappearance of the superantibodies from the blood circulation does not imply that they were eliminated from the body
Fc fragment	The antibody has functional capabilities that are mediated by the Fc including complement dependent cytotoxicity (CDC), antibody-dependent cell-mediated cytotoxicity (ACDD), opsonization, and immune complex removal; nevertheless, the Fc function is derived by chance	Antibody fragments devoid of Fc usually do not cause Fc-mediated inflammation. They cannot mediate CDC, ADCC, opsonization and immune complex removal.; Engineered four-chain antibodies can be designed for appropriate immunological functions such that their Fc can fix properly to receptors, either activating receptors, such as FcRI, FcRIIa, FcRIIIa (CD16) or inhibitory receptor (FcIIb)
Tissue penetrating ability	Relatively low, mostly depends on their interaction to FcRs; Tend to comigrate with FcR-bearing immune cells	Relatively high, due to small size and no Fc restriction; they can freely migrate to the site of infection/affected areas (high tissue penetration)
Antibody-dependent enhancement (ADE) of viral infection	Frequent for many viral infections, such as Dengue, influenza, Zika, Chikungunya, West Nile, and HIV-1	Relatively safe for use in treatment of various viral infections as the antibody fragments devoid of Fc do not have ADE ability, while Fc fragments of intact four-chain engineered antibodies can be modified to abrogate Fc receptor binding ability
Side effects	Uncontrolled binding site, affinity, and Fc function; Can cause adverse effects such as serum sickness, tumor lysis syndrome, cytokine release syndrome, and anaphylaxis	Minimized potential for causing adverse effects can be achieved through modulation of binding site and affinity, humanization, and Fc engineering