Targeting the terminal pathway in complement – driven disease.

Zelek, Wioleta 2019. Targeting the terminal pathway in complement – driven disease. PhD Thesis, Cardiff University. Item availability restricted.

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Abstract

Objectives; In the last decade there has been an explosion of interest in inhibiting complement for therapy of disease; the principle target has been complement component C5, activation of which generates the pro-inflammatory and cytotoxic products C5a and the membrane attack complex. The anti-C5 monoclonal antibody Eculizumab is a blockbuster drug, currently only approved for two ultra-rare diseases. There is abundant evidence that complement, and specifically the membrane attack complex, is involved in more common diseases for example in age related macular degeneration, myasthenia gravis and neurological disorders like neuromyelitis optica spectrum disorder and multiple sclerosis. Hence, there is an urgent need to develop better, cheaper drugs targeting membrane attack complex. Here I describe novel monoclonal antibodies targeting membrane attack complex downstream of C5 that are efficient inhibitors of its formation in humans and rodents. Methods; A large panel of anti-membrane attack complex monoclonals was generated from spleens of terminal pathway deficient mice hyperimmunised with human or rat C5b6 and/or C5b67. Antibody clones were screened using immunochemical and novel functional (haemolytic) assays to identify those with binding and blocking activities. Biophysical methods, including surface plasmon resonance were used to characterise monoclonal antibodies of interest, and selected antibodies were tested in rodents to characterise pharmacokinetics and therapeutic capacity in complement-mediated disease models. Results; Several function inhibitory anti-membrane attack complex antibodies were identified and characterised in terms of binding to membrane attack complex intermediates and cross-species lytic inhibitory capacity in comparison to current therapeutic anti-C5 monoclonal antibody. Blocking antibody selected include three against C5, two against C6 and five against C7; these efficiently inhibited lysis by human, rat, or mouse serum in vitro and were further characterised for binding specificity and affinity by enzyme linked immunosorbent assay, western blot and surface plasmone resonance, confirming strong and stable binding. One antibody has so far been tested in vivo; administration of this antibody intraperitoneally in rats efficiently inhibited serum lytic activity and blocked the disease and protected muscle endplates from destruction in a rat model of myasthenia gravis. Conclusions; I report novel blocking antibodies that target membrane attack complex formation at different stages of the pathway to efficiently inhibit complement terminal pathway across species (human, rat and mouse). Rodents treated with selected antibody were protected in a complement-driven disease model. These antibodies offer an alternative to C5 targeting for membrane attack complex-driven pathologies in animal models and man and add new data on membrane attack complex assembly and function.

Item Type:	Thesis (PhD)
Date Type:	Completion
Status:	Unpublished
Schools:	Medicine
Funders:	HEFCW, Welsh Government, Life Sciences Research network Wales
Last Modified:	25 Oct 2020 03:05
URI:	https://orca.cardiff.ac.uk/id/eprint/126319

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