Characterization of alternative complement pathway regulator Factor H and Factor H related proteins

Factor H (FH) is an abundant plasma glycoprotein, and is the main regulator of the alternative complement pathway. FH serves as a cofactor for Factor I (FI) in the inactivation of C3b to iC3b and has decay-accelerating activity towards C3 convertases. FH binds to host cell surfaces, by binding to host specific glycosaminoglycans (GAGs) on the cell surface following initial complement activation and targets C3b to inhibit further complement activation. Dysfunctional FH may lead to severe inflammatory disease due to uncontrolled complement activation on the endothelial cells of the vasculature. As the major inhibitor of the alternative pathway of complement, FH may potentially be used as therapeutic agent for several complement mediated human diseases.

We have developed a purification strategy to isolate functional FH from human plasma. In addition, in close collaboration with Dr. T. Rispens (Dept of IP and Prof. T.W. Kuijpers (AMC-UMC) we have developed several antibodies against FH and Factor H Related proteins (FHRs). We investigate FH and FHRs levels and function in health and disease. In addition, we have developed a novel specific monoclonal antibody that potentiates the function of FH on human cell surfaces.
 

Complement activation and regulation on red blood cells

All healthy human cells are equipped with several surface complement regulators to prevent damage through unwanted complement activation. Endothelial cells and blood cells are particularly well protected, as these are constantly exposed to the complement system in plasma. Red blood cells (RBCs) bear all membrane complement regulators, except CD46, which is only expressed on nucleated cells

Various diseases are associated with complement mediated RBC destruction, which may be antibody mediated (autoimmune hemolytic anemia (AIHA), allo-immunization) or the result of defective complement regulation (atypical hemolytic syndrome (aHUS), paroxysmal nocturnal hemoglobinuria (PNH)). Disease severity and response to existing complement inhibiting therapeutics is variable in these patients. This illustrates the urgent need to better understand both the triggering mechanisms of complement activation in these diseases and how the balance of complement activation and regulation on different cell surfaces is maintained.

We are investigating how RBCs are protected from unwanted complement activation, both in normal and pathological conditions, e.g. in AIHA and PNH. To answer our research questions we make use of a human cell line (HAP1) that is genetically engineered by CRISPR/Cas9 technology to lose one or several complement regulators (collaboration with Dr. R. Spaapen, dept. of IP). Furthermore, we use isolated RBCs from healthy donors and PNH patients and plasma samples of AIHA patients.

Key publications

Pouw RB, Brouwer MC, de Gast M, van Beek AE, van den Heuvel L, Schmidt CQ, van der Ende A, Sanchez-Corral P, Kuijpers TW and Wouters D. Potentiation of complement regulator factor H protects human endothelial cells from complement attack in aHUS sera. Blood Adv. 2019 Feb 26;3(4):621-632. doi: 10.1182/bloodadvances.2018025692.

van Beek AE, Kamp A, Kruithof S, Nieuwenhuys EJ, Wouters D, Jongerius I, Rispens T, Kuijpers TW, Gelderman KA. Reference Intervals of Factor H and Factor H-Related Proteins in Healthy Children. Front Immunol. 2018 doi, 10.3389/fimmu.2018.01727.

van Beek AE, Pouw RB, Brouwer MC, van Mierlo G, Geissler J, Ooijevaar-de Heer P, de Boer M, van Leeuwen K, Rispens T, Wouters D, Kuijpers TW. Factor H-Related (FHR)-1 and FHR-2 Form Homo- and Heterodimers, while FHR-5 Circulates Only As Homodimer in Human Plasma. Front Immunol. 2017 Oct 18;8:1328. doi: 10.3389/fimmu.2017.01328.

Hovingh, E.S., van den Broek, B., Pinelli, E., Kuipers, B., Rooijakkers, S.H.M., and Jongerius, I. Acquisition of C1 inhibitor by Bordetella pertussis virulence associated gene 8 results in C2 and C4 consumption away from the bacterial surface. Plos Pathogens. 2017, doi, 10.1371/journal.ppat.1006531.

Hovingh ES, de Maat S, Cloherty APM, Johnson S, Pinelli E, Maas C, Jongerius I. Virulence Associated Gene 8 of Bordetella pertussis Enhances Contact System Activity by Inhibiting the Regulatory Function of Complement Regulator C1 Inhibitor. Front Immunol. 2018 Jun 4;9:1172. doi: 10.3389/fimmu.2018.01172. eCollection 2018.