Compstatin: a C3-targeted complement inhibitor reaching its prime for bedside intervention

Abstract

There is a growing awareness that complement plays an integral role in human physiology and disease, transcending its traditional perception as an accessory system for pathogen clearance and opsonic cell killing. As the list of pathologies linked to dysregulated complement activation grows longer, it has become clear that targeted modulation of this innate immune system opens new windows of therapeutic opportunity for anti-inflammatory drug design. Indeed, the introduction of the first complement-targeting drugs has reignited a vibrant interest in the clinical translation of complement-based inhibitors. Compstatin was discovered as a cyclic peptide that inhibits complement activation by binding C3 and interfering with convertase formation and C3 cleavage. As the convergence point of all activation pathways and a molecular hub for crosstalk with multiple pathogenic pathways, C3 represents an attractive target for therapeutic modulation of the complement cascade. A multidisciplinary drug optimization effort encompassing rational 'wet' and in silico synthetic approaches and an array of biophysical, structural and analytical tools has culminated in an impressive structure-function refinement of compstatin, yielding a series of analogues that show promise for a wide spectrum of clinical applications. These new derivatives have improved inhibitory potency and pharmacokinetic profiles and show efficacy in clinically relevant primate models of disease. This review provides an up-to-date survey of the drug design effort placed on the compstatin family of C3 inhibitors, highlighting the most promising drug candidates. It also discusses translational challenges in complement drug discovery and peptide drug development and reviews concerns related to systemic C3 interception.

Keywords: Cp40; clinical translation; complement-based drug design; compstatin; nonhuman primate models; peptidic C3 inhibitors.

© 2015 Stichting European Society for Clinical Investigation Journal Foundation.

Figures

Figure 1. Overview of the complement cascade illustrating its multiple interconnections to human physiology and disease, with an emphasis on key aspects of complement-targeted therapeutic design

A. Complement activation on diseased host surfaces involves diverse recognition molecules and initiation mechanisms that underlie three distinct activation pathways operating either in parallel or independently of each other. Briefly, binding of C1 to antigen-antibody complexes (e.g. in autoimmune pathologies) triggers the classical pathway (CP), while binding of MBL/ficolin or properdin to target surfaces, via recognition of distinct molecular patterns, initiates the lectin pathway (LP) and alternative pathway (AP), respectively. Irrespective of their initiating trigger, all pathways converge at the proteolytic activation of C3, the central component of the cascade. Assembly of C3 convertases via any of the three aforementioned activation pathways promotes surface opsonization by C3 fragment deposition mainly through an AP-mediated amplification loop, and also leads to effector generation (e.g. C3a anaphylatoxin release). C3 activation is the prerequisite for the downstream activation of the lytic pathway that begins with the cleavage of C5 and culminates in the assembly of the cell-perforating MAC. Both fluid-phase and surface-bound complement regulators (e.g. factor H, CR1, MCP) promote the factor I-mediated degradation of C3b into bioactive fragments that mediate a plethora of immunomodulatory functions in health and disease (including inflammatory signaling, phagocytic activation and modulation of adaptive immune responses). Cleavage of the central complement components C3 and C5 by their respective convertases results in the generation of the anaphylatoxins C3a and C5a that, through interaction with their cognate receptors (C3aR, C5aR1), trigger potent inflammatory signaling and chemotactic responses in a variety of myeloid and non-myeloid cell types. B. Therapeutic modulation of complement using the peptidic C3-targeted inhibitor compstatin: Compstatin acts as a highly selective protein-protein interaction inhibitor. Binding of compstatin to native C3 and also to C3b-containing convertases results in potent and broad complement inhibition through abrogation of C3 activation and AP-mediated amplification of C3 cleavage. The lack of C3b deposition also prevents C5 convertase assembly and downstream effector generation (e.g. C5a, MAC). The complement inhibitory action of compstatin is exerted regardless of the upstream trigger or complement pathway (CP, LP, AP) involved. C. Therapeutic intervention at different steps of the complement cascade affords distinct benefits and limitations to each targeting approach. Intervention at the early stages of complement initiation, such as inhibition of C1s and MASPs by C1-INH, affords effective blockade of the CP and LP, while letting the alternative pathway operate and preserve its homeostatic and immune surveillance functions. However, upstream inhibition may also still allow amplification of potentially harmful activation on diseased tissue. C3 interception using peptidic inhibitors of the compstatin family results in abrogation of C3 activation by the convertases, inhibition of AP-mediated amplification and downstream effector generation. However, compstatin does not affect initial activation of the AP through the ‘C3 tick-over’ mechanism and also allows for C4b generation through upstream activation of the CP and LP. Therefore compstatin treatment may still allow for residual opsonic activity and complement-mediated immune surveillance. Finally, C5 blockade selectively targets the lytic pathway of complement abrogating C5 activation and generation of detrimental downstream effectors (i.e. C5a, MAC), while leaving intact upstream activation pathways including C3 activation and the AP-mediated amplification loop. Abbreviations: BCR, B-cell receptor; CR1 complement receptor type 1; CR2, complement receptor type 2; CP, classical pathway, AP, alternative pathway; LP, lectin pathway; MBL, mannose-binding lectin; MASP, MBL-associated serine protease; MCP, membrane cofactor protein; DAF, decay-accelerating factor; CR’s complement receptors;

Figure 2. The unique binding and inhibitory modes of compstatin on human C3

A: The resolution of the crystal structure of the C3c-compstatin complex revealed that the binding of compstatin to C3 occurs at the macroglobulin (MG) ring of the β-chain, in a shallow pocket formed between the MG4 and MG5 domains of the β-chain (colored blue and green respectively). Open circles depict additional contact sites on C3 that have been explored through targeted modifications of compstatin, i.e. via N-terminus extension in analog Cp40 (red) or by adding albumin-binding tags (magenta). B: Schematic illustration of the key protein interactions leading to the formation of the alternative pathway (AP) C3 convertase on a target surface, displaying the subsequent binding of native C3 to the nascent convertase and its cleavage into C3a and C3b. C: Compstatin acts as a protein-protein interaction inhibitor. It binds both native C3 and C3b and sterically inhibits the binding and cleavage of native C3 by C3 convertases.

Figure 3. Milestones of the preclinical and clinical development of compstatin and its most potent derivatives

A comprehensive timeline of the molecular characterization, structure-guided optimization, preclinical and clinical evaluation of the most promising compstatin derivatives, including milestones that have illuminated key structural elements of the interaction with C3 and have propelled the drug design effort placed on this family of peptidic C3 inhibitors (see text for more details).

Figure 4. The pharmacokinetic profile of compstatin and structure-guided modifications aimed at improving its plasma residence

A: A ‘target-driven’ model of compstatin’s elimination from plasma: A first rapid phase of clearance of the excess of free (unbound) compstatin is followed by a second, slower phase of plasma elimination that is driven by the tight binding of compstatin to its target protein, C3. B: Schematic overview of N- or C-terminal modifications for the development of compstatin derivatives with increased plasma residence. These compstatin derivatives are designed to display longer plasma retention via binding to carrier proteins (i.e. albumin) or via reducing renal filtration by conjugation to high molecular weight-PEG moieties. More details on the various peptide modification strategies employed to improve the pharmacokinetic profile and plasma residence of compstatin can be found in the text.