Generation of polyclonal antibodies is relatively simple and cheap as
the use of animals (such as chicken, horses, goats, and rabbits) enables
the recovery of large quantity of antibodies from serum or egg Yolk
(Yakhkeshi et al., 2022 ).
Polyclonal antibodies arising
from diverse B-cell clones constitute a heterogeneous mixture that
manifests varying binding affinities. In contrast, monoclonal antibodies
are homogeneous population originated from a single B-cell clone,
resulting in well-defined binding specificities and affinities. However,
at some point a fresh batch will be sought as the original stock
diminishes, which inevitably leads to a batch-to-batch variation in
terms of the specificity, sensitivity, and reproducibility
(Bradbury & Plückthun, 2015 ). This
might include differences in antibody reactivity and titre, and thus
polyclonal reagents often lack reproducibility. In comparison, the
continuous culture of B cell hybridomas overs a reproducible and
potentially inexhaustible supply of antibody with exquisite specificity.
Consequently, monoclonal antibodies enable the development of
standardised and secure immunoassay system.
Genetically engineered antibody fragments are alternative to full-length
antibody in diagnostics and therapeutics for a variety of diseases due
to their various advantages
(Holliger & Hudson, 2005 ).
Firstly, their small size and simpler structure make them ideal for
large-scale production in eukaryotic systems, such as mammalian and
insect cells. The antibody fragments have known sequences and are
reproducible, verifiable, and manufacturable.
The single-chain variable
fragment (scFv) is a successful example of a genetically engineered
antibody fragment. As the variable light chain and variable heavy chain
coding sequences are genetically linked in a single transcript, there is
no need to balance the expression of the light chain and heavy chain.
Engineered single domain
antibodies (VHH) enable the rapid generation of antibody fragments at
higher yields and lower cost compared to full-size
monoclonal antibodies (mAbs) that
typically require mammalian expression systems. Their small size also
facilitates tissue penetration and access to cryptic epitopes, making
them particularly useful for tumour penetration in cancer immunotherapy
(Zinn et al., 2023 ). Moreover, the
lack of an Fc region removes the risk of bystander immune cell
activation and antibody effector functions such as antibody-dependent
cellular cytotoxicity (ADCC), antibody-dependent cellular phagocytosis
(ADCP), or complement-dependent cytotoxicity (CDC), allowing the
molecule to bind its target without activating the host’s immune system
(Gravina et al., 2023 ). However,
the absence of an Fc region is a double-edged sword. The disadvantage is
that it not only reduces thermostability and enhances propensity for
aggregation, thereby increasing the risk of immunogenicity, but also
shortens half-life due to a lack of FcRn-mediated recycling
(Park et al., 2016 ). This can lead
to the need for higher and more frequent dosing. Fusion of the scFv to
albumin or polyethylene glycol (PEG) can be used to improve half-life
(Albrecht et al., 2004 ;Vazquez-Lombardi et al., 2015 ).
However, such fusions can offset the advantages that an scFv holds over
a mAb due to cost and the increase in size
(Bradbury & Plückthun, 2015 ).
The fourth generation of antibody engineering: antibody mimetics is not
generated by the body immune system naturally but share a common feature
with natural antibody:
they
shared characteristic of complementary shape at the binding site with
the antigen. Similar to enzyme and substrate, the binding interactions
between antibody mimetics and antigens can be explained by three models,
including the classical lock and key theory, the induced fit mode, and
the conformational selection model. Typically, the classical lock and
key theory reflects the initial recognition and specifically between
antibody mimetics and antigens. The induced fit model suggests that both
antibody mimetics and antigens undergo conformational changes upon
binding, in which the dynamic interaction allows the active sites of
antibody mimetics to adapt to their shape, optimizing the fit with the
antigen. Additionally, the conformational selection model proposes a
conformational change occurs prior to the binding of antigens, in which
the antigen seems to select and stabilize a higher-energy conformation
of an antibody mimetic for binding. Of note, these models are not
mutually exclusive, but rather represent different aspects of
interactions between antibody mimetics and antigens. Antibody mimetics
represent a class of engineered molecules mimic the functions and
properties of natural antibodies while offering advantages such as
enhanced stability, smaller size, improved production yields. These
mimetics can manifest in various forms, like smaller antibody fragments
synthesized based on the functional regions of antibody fragments such
as scFv or Fab. It is possible to design novel proteins with binding
properties through site-directed mutagenesis and random mutagenesis. We
summarized and compared the features of all the four generations of in
terms of stability, specificity, affinity, and so forth (Table 1).
Now it may be the right time to review and analyse the field of antibody
mimetics retrospectively and prospectively. Given that scientometric
analysis can quantitively evaluate and investigate on all aspects of the
literature at different development stages
(Mooghali et al., 2011 ), this
article aims to provide a vigorous roadmap for antibody mimetics
research through this method to identify the major players and their
cooperation networks, including countries, academic groups, and
individuals; to analyse research status and hotspots, especially the key
study findings; to discuss the potentially valuable research directions.
Table 1 Antibody production & diversified antibody generation
platforms