Preclinical Development and In Vivo Evaluation Of Next-Generation Compstatin Analogs With Improved Systemic Profiles: A Novel Option For The Treatment Of Paroxysmal Nocturnal Hemoglobinuria

Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired orphan disease that is characterized by complement-mediated lysis of clonal populations of erythrocytes that lack glycosyl-phosphatidyl-inositol (GPI)-anchored complement regulators. Hemolysis and uncontrolled complement activation on affected platelets may contribute to the thrombophilia typical of PNH, which largely accounts for the morbidity and mortality of the disease. The introduction of the therapeutic antibody Eculizumab, which blocks activation of complement component C5, to the clinic has dramatically improved the treatment of PNH as it protects from hemolysis and possibly thromboembolisms; yet it also imposes high costs to the health care system. Moreover, while C5-targeted therapy efficiently impairs intravascular lysis of erythrocytes, it does not prevent their ongoing opsonization with activation fragments of complement component C3, thereby increasing the potential for extravascular lysis by immune cells. As a consequence, there is an unmet clinical need, and intervention at the level of C3 is expected to lead to a more comprehensive protection of PNH erythrocytes. Compstatin is a 13-residue disulfide-bridged peptide that selectively binds to human and primate forms of the central complement component C3 and its active fragment, thereby preventing the conversion of C3 to C3b which is essential for the functioning of all initiation, amplification, and terminal pathways of complement. Owing to their potent inhibitory activity of C3 activation, and due the comparatively low production costs of these peptidic drugs, compstatin analogs present an attractive option for the development of improved PNH therapeutics. Whereas early compstatin analogs are in clinical development for the local treatment of age-related macular degeneration, a long-term systemic use of compstatin for treating PNH places higher demands on this drug class. In this study, we therefore evaluated next-generation compstatin analogs with distinct pharmacokinetic properties concerning their efficacy and suitability for systemic administration. For this purpose, the lead analog Cp40 and a PEGylated derivative thereof were evaluated in complement activation assays, binding studies, and in a well-established ex vivo model of PNH using patient-derived erythrocytes. In addition, compstatin analogs were injected to non-human primates (cynomolgus monkeys) in order to determine administration strategies suitable for the treatment of PNH. Both Cp40 and its PEGylated derivatives prevented complement activation equally well, thereby indicating that N-terminal PEGylation does not substantially interfere with the activity of the peptide. Importantly, both compounds impaired deposition of C3 activation fragments on the surface of PNH erythrocytes as determined by FACS analysis and prevented their lysis with an IC 50 of ∼4 µM. Pharmacokinetic profiling showed that PEGylation substantially improved the plasma half-life of Cp40 to more than 5 days whereas the non-PEGylated compound still exerted a value (t 1/2 > 12 hours) that surpasses most peptide drugs. We therefore also evaluated repetitive subcutaneous injection of non-PEGylated Cp40 and could show that doses of 1 mg/kg every 12 hours were sufficient to maintain a therapeutic plasma level of compstatin in cynomolgus monkeys. Both strategies appear highly promising, and further evaluation of the in vivo efficacy and consideration of aspects such as production cost, treatment flexibility/control, and long-term plasma profiles will help determining the best route. Hence, our study provides critical evidence for the efficacy of compstatin analogs in the treatment of PNH, aiming to achieve a better control of all complement-mediated disease manifestations, reveals two suitable options for systemic administration of such peptidic complement inhibitors, and paves the way for a clinical application of this compound class in PNH. Disclosures: Ricklin: University of Pennsylvania: Patent application on Cp40, Patent application on Cp40 Patents & Royalties. Risitano: Alexion Pharmaceuticals: Honoraria, Membership on an entity’s Board of Directors or advisory committees, Research Funding. Lambris: Amyndas Pharmaceuticals: Equity Ownership; University of Pennsylvania: Patent application on Cp40 Patents & Royalties.