What are antibodies?

Antibodies, or immunoglobulin molecules, are glycoproteins produced by the plasma B cells of the adaptive immune system with the aim to recognise and neutralise foreign antigens, such as microorganisms and viruses (e.g. SARS-CoV-2). The specific binding of antibodies to their target molecules activates downstream immune responses leading to the elimination of the intruders. Thus, antibodies play a pivotal role in the immune system’s defence against infection and disease.

Using a powerful and highly sophisticated combination of genetic recombination, somatic hypermutation and clonal selection, the adaptive immune response continuously generates novel antibodies against new antigens, thus making antibodies the most versatile among the currently known classes of binding molecules.

Full-length antibody (lgG)

Figure 1

Figure 1, source: DOI: 10.1039/c8cs00523k–2018-3rd generation antibody discovery methods: in silico rational design. On a structural view, antibodies are tetrameric proteins with a characteristic Y-shaped structure, consisting of two pairs of heavy and light chains. The tips of the Y are formed by the variable antigen-binding fragment (Fab) region, containing the paratopes, which are located within six binding loops, referred to as the complementary determining regions (CDRs). The paratopes mediate the interaction with the target regions of the antigens, also known as epitopes. The base of the antibody, also called the crystallisable fragment (Fc) region, defines the antibody subclass and regulates the communication with parts of the immune system that are important for effector function and serum half-life.

MONOCLONAL ANTIBODIES AS THERAPEUTIC AGENTS

The first use of the term “antibody” occurred by Paul Ehrlich in his article “Experimental Studies on Immunity”, published in October 1891. In 1986, the first therapeutic monoclonal antibody was approved by the US Food and Drug Administration. Since then, more than 100 monoclonal antibodies have been designated as drugs, as they are very effective therapeutic agents. The high specificity of antibodies makes them ideal to reach their intended target and thus is useful to treat many different disease states. Due to their potential for high affinity and specific binding to a large variety of molecular targets, antibodies have been the focus of a wide range of technological developments aiming for the isolation, production and optimisation of these molecules for specific targets of interests in various indications. Antibodies are key tools in research and diagnostics, but also represent the fastest-growing class of biotherapeutics on the market due to the naturally favourable attributes such as specificity, potency and metabolic stability.

HISTORY OF ANTIBODY DRUG DISCOVERY PLATFORMS

Antibody drug discovery refers to the process of identifying new therapeutic antibodies to combat various diseases, such as cancer, autoimmune disease, viral infections, and many others. Over the last four decades, three different generations of antibody discovery technologies have been described. Importantly, the different approaches are not mutually exclusive but are highly complementary to each other.

Figure 2

Figure 2, source: see figure 1. In vivo approaches harness the power of an immune system to generate antibodies, traditionally through animal immunisation and more recently also from human patients. In vitro approaches, on the other hand, rely on the construction of large libraries of antibody sequences mimicking the diversity achieved by the immune system, and thus likely to contain some binding molecules for each given antigen. The third generation covers the in silico attempts of designing and optimising biotherapeutics.

The first major breakthrough in the development of antibodies for research applications was the production of monoclonal antibodies in 1975 by Köhler and Milstein. Their technique involved removing B-cells from the spleen of an animal that had been challenged with an antigen, and subsequently fusing them with an immortal myeloma tumour cell line. This resulted in a single-cell hybrid known as a hybridoma and allowed for the first time the purification of homogenous preparations (monoclonal antibodies; mAbs) of in which every antibody in the product is identical in its protein sequence, and thus every antibody is expected to have the same antigen recognition site, affinity, biological interactions, and downstream biologic effects. More recently, the direct interrogation of single antibody-secreting B cells upon immunisation or virus challenge using technologies, such as fluorescence-activated cell sorting or the more modern ultra-highthroughput (UHT) microfluidic is rapidly evolving to become commonplace in the antibody discovery industry. The importance of this technology was recently highlighted by the real-time isolation of neutralising mAbs from convalescent COVID-19 patient’s memory B cells that may serve as a promising intervention to SARSCoV-2 infection. In parallel to these in vivo (B-cell derived) discovery efforts, in vitro approaches relying on recombinant antibody technologies that can be used for the creation of large libraries of antibody sequences evolved. These antibodies are supposed to mimic the diversity achieved by the immune system, and in combination with the power of in vitro technologies, such as phage display, the selection of antibodies specific for any given target can be achieved. As a third generation technology, the in silico design or engineering (molecular optimisation) of antibodies has emerged more recently. While being highly affine and selective for their targets, antibodies also need to exhibit different biophysical features, such as stability and solubility. Often these traits are conflicting as some of the mutations may cause advantages for some features but correlate at the same time with worsening others. Therefore, simultaneous optimisation of all of these features is required. For this, computational approaches offer a promising avenue, as they could drastically reduce time and costs of antibody discovery.