Personalised management of age-related macular degeneration by the identification of high-risk genetic variants in complement Factor H & Factor I

Abstract

MD is the predominant cause of irreversible blindness in the developed world. There is currently no treatment for ‘dry’ AMD. Several pivotal studies have associated AMD with genetic variants in complement factor I (CFI) and factor H (CFH), which, through the formation of a trimolecular complex (TMC), cleave C3b and control complement activation. To investigate the consequences of rare CFI variants in AMD, I initially assayed plasma FI levels in patients and demonstrated that rare genetic variants in CFI leading to low FI levels were strongly associated with AMD (early or late AMD: odds ratio [OR] 12.05, P = 0.03; early AMD: OR 30.3, P = 0.02; late AMD: OR 10.64, P < 0.01). Measurement of FI in aqueous humour revealed a large FI concentration gradient between systemic circulation and the eye (∼286-fold). Rare CFI variants without quantitative defects were analysed in the context of recombinant, fully processed FI revealing profound (Y459S, R474Q) and more subtle (D403N, E554V) functional defects. To probe these defects in real time, I developed a surface plasmon resonance (SPR) assay of AP regulatory TMC formation using an inactive FI backbone and as proof of concept demonstrated impaired TMC formation with Y459S FI. Analysis of inactive (S525A) FI revealed the potential for dysfunctional variants to have a dominant negative effect. Rare CFH variants identified in AMD patients were produced and functionally analysed in the context of CFH CCP1-4. Established functional tests revealed severe (R166W, R232Q) and subtle (P26S, T91S) impairments, which were confirmed by my real-time AP regulatory TMC building assay. Overall, the data presented herein demonstrates that understanding protein variant dysfunction at a molecular level may enable streamlined clinical trials; to offer personalised therapeutic choices based on an individual’s genetics. My real-time assay of AP regulatory TMC complex formation could provide greater granularity and sensitivity in patient stratification. The impermeability of the BRB and potential dominant negative effects of variants should be considered if replacement therapy is to be used to treat AMD.