Structural changes of interfacially adsorbed antibody molecules

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Cancer treatments have undergone three revolutionary stages over the past few decades: chemotherapies, biomarkers targeted at mutated genes, and combined treatments of biomarker targeting and immune process mediation. Each stage of these treatments was facilitated by advances in our understanding of the behaviour of cancers, especially at the molecular and cell levels. Our early understanding of the high growth rates of cancerous cells led to the development of chemical/drug therapies together with radiation treatments to kill cancerous cells. However, such medical treatments must undergo rigorous clinical trials and meet regulatory requirements, e.g., FDA approval, before clinical deployment. This process plus further improvement to perfect the treatments can take more than a decade. Currently, chemotherapy is still the mainstream cancer treatment for patients, but their toxicity remains a major limiting factor to survival rates. Over the past 5-8 years, the ability to moderate immune processes has been realised, and a number of pioneering treatments based on this new line of thinking have very recently achieved clinical success. This prospect has driven a major global effort to develop new treatments based on antibody technologies, covering not only oncology but also other major diseases including cardiovascular, respiratory, autoimmunity and infectious diseases. Antibodies used for cancer or other disease treatments must be designed, manufactured, separated, purified and eventually formulated into medical products ready for clinical use. A popular means of administration is to apply via intravenous injection. This option requires the antibody drugs to be formulated as a stable protein solution in a bottle (glass or plastic) or a ready-to-inject syringe set, with a shelf-life between 1-2 years. Because these bioengineered antibodies have to be equipped with two or more biological functions, their amino acid sequences (so called primary sequences) must be altered. As a result, we do not know how stable their folded domains are and how instability from the modified domains will affect the stability of the whole antibody.All proteins are amphiphilic due to the presence of both polar and apolar amino acids on their surfaces. This amphiphilic character drives proteins to adsorb and desorb at different interfaces spontaneously. During these interfacial processes, proteins interact with the substrate surface and with themselves, and depending on the nature of the substrate surface and the close proximity between them once adsorbed, deformation of the globular structures and even local unfolding can occur, causing exposure of hydrophobic patches that may induce aggregation and precipitation, compromising the bioactivity of such antibody drugs. Newly bioengineered antibodies are often unstable, and adsorption can accelerate their instability.Using a series of bioengineered antibodies with well-controlled sequence modifications in Fab and Fc domains, this LINK project forges a new collaborative team involving MedImmune, Manchester University and Imperial College London, with the aim to develop new understanding by combining neutron reflection experiments with molecular dynamics simulation. We will examine how certain well-controlled sequence modifications in Fab and Fc domains affect their adsorbed globular structures and how instability in these domains affects the structure of the whole mAb. Another line of work will be to examine how representative substrate surfaces affect structural deformation and unfolding. These studies will lead to new results that will be of great value to the biopharmaceutical industry and to academic research. The successful delivery of this project will lead to new primary structure-stability relationships that will assist MedImmune and other protein drug developers to improve their antibody stability in their biotherapeutics. The outcome will ultimately benefit the general public.