Selectivity of C3-opsonin targeted complement inhibitors: A distinct advantage in the protection of erythrocytes from paroxysmal nocturnal hemoglobinuria patients

Abstract

Paroxysmal nocturnal hemoglobinuria (PNH) is characterized by complement-mediated cell lysis due to deficiency of GPI-anchored complement regulators. Blockage of the lytic pathway by eculizumab is the only available therapy for PNH patients and shows remarkable benefits, but regularly yields PNH erythrocytes opsonized with fragments of complement protein C3, rendering such erythrocytes prone to extravascular hemolysis. This effect is associated with insufficient responsiveness seen in a subgroup of PNH patients. Novel C3-opsonin targeted complement inhibitors act earlier in the cascade, at the level of activated C3 and are engineered from parts of the natural complement regulator Factor H (FH) or complement receptor 2 (CR2). This inhibitor class comprises three variants of “miniFH” and the clinically developed “FH-CR2” fusion-protein (TT30). We show that the approach of FH-CR2 to target C3-opsonins was more efficient in preventing complement activation induced by foreign surfaces, whereas the miniFH variants were substantially more active in controlling complement on PNH erythrocytes. Subtle differences were noted in the ability of each version of miniFH to protect human PNH cells. Importantly, miniFH and FH-CR2 interfered only minimally with complement-mediated serum killing of bacteria when compared to untargeted inhibition of all complement pathways by eculizumab. Thus, the molecular design of each C3-opsonin targeted complement inhibitor determines its potency in respect to the nature of the activator/surface providing potential functionality in PNH.

Incomplete inhibition by eculizumab: mechanistic evidence for residual C5 activity during strong complement activation

“…This is consistent with previous data showing that the C3-opsonintargeting inhibitors mini-FH and TT30 efficiently control AP activation in several assays. 38,44,45,49,50 As observed for the double inhibition of C5 in NHS (above), addition of coversin or PAS-coversin to the patientderived, eculizumab-containing serum completely stopped the residual hemolysis if compared with eculizumab alone. We verified that the patient sera indeed contained an excess amount of eculizumab over C5 ( Figure 1F).…”

“…[42][43][44] Detailed descriptions are given in supplemental Methods, available on the Blood Web site.…”

Therapeutic complement inhibition in complement-mediated hemolytic anemias: Past, present and future

“…Initial PK and PD data demonstrate that TT30 treatment may result in pharmacologically relevant CAP inhibition in PNH, as supported by LDH decrease; nevertheless, further clinical development is hampered by the short half-life of this compound [174]. A different approach aiming to improve surface targeting of CFH exploits an engineered miniaturized version of CFH, named mini-FH; different versions of mini-FH have been described [175][176][177], all aiming to maximize the CAP inhibitory effect of CFH (C3 convertase decay and co-factor activities) at sites of complement activation [175]. Mini-FH (Amyndas, AMY-201) is a small (43 kDa) protein consisting of the regulatory complement control protein (CCP) 1-4 domains of CFH attached to its CCP 19-20 domains (which harbor a C3 binding-site) [175].…”

From orphan drugs to adopted therapies: Advancing C3-targeted intervention to the clinical stage

“…This new class of surface-targeted inhibitors (e.g., AMY-201, Amyndas) has shown promising therapeutic results in an AP-driven disease model, protecting the erythrocytes of PNH patients from C3 opsonization and intravascular hemolysis (Schmidt et al , 2013). Notably, a recent study guided by molecular design has revealed distinct selectivity criteria that determine the relative inhibitory potency of surface-targeted C3 inhibitors on different C3-opsonized surfaces (Schmidt et al , 2016). In this context, mini-FH showed greater efficacy in abrogating complement activation on PNH erythrocytes, whereas other targeted regulator constructs showed distinct activities on different surfaces (Schmidt et al, 2016).…”

“…Notably, a recent study guided by molecular design has revealed distinct selectivity criteria that determine the relative inhibitory potency of surface-targeted C3 inhibitors on different C3-opsonized surfaces (Schmidt et al , 2016). In this context, mini-FH showed greater efficacy in abrogating complement activation on PNH erythrocytes, whereas other targeted regulator constructs showed distinct activities on different surfaces (Schmidt et al, 2016). These findings may better guide C3 therapeutic design by allowing us to tailor the application of surface-targeted inhibitors to appropriate pathologies.…”