The complement pathway in geographic atrophy explained

Bios Dr. Alekseev is a Heed fellow in medical retina at Duke University, Durham, North Carolina. Dr. Lad is an associate professor at Duke University. Dr. Steinle is a partner at California Retina Consultants in Southern California. DISCLOSURES: Dr. Alekseev has no relevant relationships to disclose. Dr. Lad disclosed relationships with Apellis, Roche, Novartis, Allegro, Gallimedix, Retrotope, Gemini Therapeutics and LumiThera. Dr. Steinle disclosed relationships with Alimera Sciences, Apellis, Carl Zeiss Meditec, Genentech, Notal Vision, Novartis, Regenerative Patch Technologies, Regeneron and RegenxBio, and is an investor in Vortex Surgical.

The complement pathway, a key component of innate immunity, maintains immune homeostasis throughout the body, including in ocular tissues. It’s composed of three pathways —classical, alternative and lectin—­all eventually converging on C3 cleavage (Figure). 

Endogenous inhibitory factors suppress activated complement to maintain immune equilibrium. Thus, autologous damage can arise due to overaction, whether through inappropriate activation or inadequate inhibition. 

In the retina, the complement system, particularly the alternative pathway, has been implicated in the development of age-related macular degeneration and geographic atrophy. In all, six members of the complement cascade—CFB, CFH, CFI, C2, C3 and C9—are estimated to account for 40 to 60 percent of AMD heritability.¹  

Complement activation levels differ significantly between AMD disease stages, with higher activation corresponding with more advanced AMD and central GA.² Many complement components and regulators deposit within drusen, where they act as inflammatory foci.³ 

This review examines the role of complement factors in the pathogenesis of GA.

Complement factor B

CFB is an activating factor in the alternative pathway. It’s converted by CFD into its active subunit Bb, which then becomes an integral part of the C3 convertase. Activation products of CFB (Ba and Bb) are elevated in the serum of AMD patients.³  

Two protective variants of CFB have been described: R32Q and L9H.⁴ The R32Q polymorphism also appears to slow GA progression.⁵ These polymorphisms may be protective due to the impaired activity of the resultant C3 convertase. 

Complement factor D 

CFD is directly upstream of CFB and also activates the alternative pathway. A CFD-/- knockout mouse model of AMD showed decreased photoreceptor damage compared to wild type.⁶ One polymorphism (rs3826945) has been associated with AMD.⁷ In addition, serum levels of CFD have been shown to be elevated in AMD patients,⁷ a noteworthy finding since CFD is considered to be rate-limiting in the activation of the alternative pathway.

Complement factor H 

CFH is a key regulatory factor responsible for complement cascade inactivation.  CFH is the first and most researched complement factor to be linked to AMD.³ It prevents damage to healthy bystander cells, but not pathogens, by selective inhibition of complement activity on self-surfaces.⁸  

Genome-wide association studies have identified numerous risk-imparting, as well as disease-protective, CFH variants. Of these, Y402H is the most notable common variant; it’s present in 50 percent of AMD patients⁹ and correlates with the presence of GA.^10,11 This polymorphism compromises glycosaminoglycan-mediated interaction between CFH and the damaged retina, thereby attenuating inactivation of the complement cascade and allowing for chronic low-level inflammation.¹²  

In contrast, R1210C is an example of a rare but highly penetrant variant that imparts a more severe disease course, including earlier disease onset by approximately six years.¹³ Interestingly, CFH accumulates in drusen at the choroid-RPE interface, where it’s thought to expose retinal pigment epithelium cells to continuous membrane attack complex (MAC) damage.⁹ In addition to conferring general AMD risk, CFH variants impart higher risk of conversion from large drusen to GA.^14,15

Three complement pathways—classical, lectin and alternative—converge on the common terminal pathway, eventually leading to formation of the membrane attack complex. Complement factors with known genetic associations with age-related macular degeneration are marked with A (CFB, CFD, CFH, CFI, C1 inhibitor, C2, C3 and C9). Those with known genetic associations specifically with geographic atrophy are marked with G (CFB, CFH and C3). Complement regulators appear in orange boxes. Complement-targeting medications under current or former clinical trial investigation (Table, page 33) appear in dashed boxes. (Adapted from Khandhadia S, Cipriani V, Yates JR, Lotery AJ. Age-related macular degeneration and the complement system. Immunobiology. 2012;217:127-146.)

Complement factor I 

CFI is a C3b/C4b inactivator and the rate-limiting enzyme in complement termination. Interestingly, amyloid-beta, a component of drusen, binds to CFI and reduces its activity.¹⁶ While several high-risk CFI variants have been proposed, a meta-analysis has confirmed a strong association between AMD and CFI variants rs10033900T>C (protective) and rs2285714C>T (high-risk).¹⁷  

Subsequently, a large systematic functional testing study revealed numerous rare CFI variants that likely contribute to AMD.¹⁸ A rare but particularly pathogenic polymorphism is G119R, which results in a hypoactive version of CFI and imparts a 20-fold increased risk of AMD.¹⁹

Complement factor P

CFP (properdin) is the only known positive regulator of the complement cascade, wherein it stabilizes the C3 and C5 convertases. While no properdin polymorphisms are known to associate with AMD, it presents an appealing therapeutic target (Table).

Complement component 1 

The trigger of the classical pathway, C1 becomes activated by binding to antigen-antibody complexes. While no known AMD-causing variants of C1 have been identified, potential associations between AMD and variants in the C1-inhibitor gene, SERPING1,²⁰ have been noted, although these findings remain contested.

Complement component 2 

C2 is activated via the classical and lectin pathways to become a part of the C3 convertase. Two major protective variants of C2 are E318D and rs547154. However, since C2 and CFB are in extensive linkage disequilibrium, CFB/C2 is considered a single risk-modifying allele, wherein C2 variants likely do not have an independent molecular role in AMD.²¹

Complement component 3 

C3 is the convergence point of the three complement pathways. It’s activated by C3 convertase to become C3a, which then becomes a core subunit of C5 convertase.  

Activated C3 is often measured as a reflection of overall complement cascade activity. C3 is found deposited within drusen and elevated in plasma of AMD patients.³  C3 overexpression in mice induces retinal pathology that in many aspects recapitulates AMD, including photoreceptor and RPE atrophy.²² The most common AMD-linked C3 variants are R80G and R102G,⁴ whereas K155Q is a rare variant. Variant R102G is strongly associated specifically with GA.¹¹  

As a common relay point in the complement cascade, C3 is an attractive therapeutic target. A C3-neutralizing therapeutic (APL-2, also known as pegcetacoplan, Apellis Pharmaceuticals) has shown promising clinical results in a Phase II trial²³ and is the subject of two fully enrolled ongoing Phase III trials^24,25 (Table). 

Additional agents, including a non-PEGylated compstatin (AMY-106)²⁶ and a novel protease that cleaves and degrades C3 (CB 2782-PEG),²⁷ are currently in preclinical development.

To date, only three clinical trials have shown reduction in geographic atrophy growth: MAHALO, FILLY and GATHER1. Results of the Phase II MAHALO study were subsequently invalidated by the larger CHROMA and SPECTRI trials. Based on positive results of the Phase II FILLY and Phase II/III GATHER1 trials, both APL-2 and avacincaptad pegol are in Phase III clinical trials.

Complement component 5 

C5 is activated by C5 convertase to become C5a, a pro-inflammatory mediator that acts as a nidus for MAC formation. Plasma levels of C5a are elevated in AMD patients¹¹ and stimulation of the choroid C5a receptor causes ICAM-1 overexpression, which likely leads to monocyte recruitment.²⁸ Nevertheless, no consistent and replicable associations between polymorphisms in C5 or C5a receptors and AMD have been reported.^28,29 A C5-based therapeutic (avacincaptad pegol, also known as Zimura, Iveric bio) has already shown promising Phase II/III results,³⁰ with a second Phase III trial ongoing³¹ (Table).

Complement component 9 

C9 participates in the final stage of MAC formation, whereby polymerization of 12-18 molecules of C9 forms a cytolytic transmembrane pore in the target cell. A high-risk C9 variant (P167S) has been reported in a mixed population of GA and/or nAMD,³² although this study didn’t stratify specifically for correlation with GA.  

Another variant, R95S, has only been reported in nAMD.³³ While C9 is currently not being targeted for therapeutic purposes, an endogenous inhibitor of C9 polymerization, CD59, has inspired a gene-therapy-based MAC-inhibitory agent now in a Phase II trial (Table).

Outside the pathway

While most of the research into the role of complement in AMD focuses on polymorphisms that cause complement  dysfunction, it’s important to recognize that the inciting pathology may lie outside the complement genes themselves. Inflammation, reactive oxygen species and complement-expressing macrophages all contribute to complement activation in AMD.  

Particularly interesting is the role of iron homeostasis, critical for retinal and RPE health. Directly relevant to the pathogenesis of GA, cultured RPE cells exposed to elevated iron levels upregulate C3 gene expression, protein levels and protein secretion.³⁴ Mice with elevated iron in the RPE develop activated C3 deposits in Bruch’s membrane.³⁵  

Moreover, iron increases retinal production and deposition of amyloid-beta,³⁶ which is known to inhibit CFI activity.¹⁶ These findings align with the elevated iron levels in the RPE of GA-affected eyes of AMD patients³⁷ and highlight iron-chelation as a potential approach to suppress complement activity for treatment of GA.

Bottom line

When considering the role of complement factors in GA, it’s important to recognize the broad range of outcome measures from the gene-association studies. While most of these studies examine cross-sectional AMD populations of pooled disease stages, the underlying molecular pathobiology is undoubtedly more complex. The genetic and environmental risk signature is likely quite different between the onset of early AMD, progression to GA, enlargement of GA and development of nAMD. 

Relatively few studies have focused on identifying complement gene variants specifically pertinent to GA. Given that this knowledge is of utmost prognostic value and could inform the development of novel therapeutics, a focused characterization of a GA-specific genetic signature would be highly useful academically and clinically.

Phase III trials are ongoing for complement inhibition at the C3 and C5 levels.^24,25,31 The results of these large trials will continue to shape how we approach GA. RS

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