The H Factor
Complement-regulating proteins play a major role in dry AMD pathology
Simon J. Clark | | Longer Read
Age-related macular degeneration (AMD) is a major cause of blindness worldwide – and there are no effective treatments for this condition. There are, however, a plethora of gene variants (45 common and 7 rare) associated with modifying AMD risk. Many of these variants are linked directly to the complement pathway, which regulates the immune system and subsequent inflammation. This is important, because inflammation at the back of the eye is well known to play a role in AMD pathology – and it is clear that failure to regulate complement in AMD is a factor in disease onset and progression.
This information, along with our previous research, led us to investigate the role of complement pathway proteins on AMD – in particular, the factor H protein family that regulates complement activation. Our approach uses mass spectrometry to investigate AMD pathogenesis at the protein level, using the circulatory blood as a window to disease progression.
What we’ve discovered
We now know that genetic risk markers for AMD lead to increased factor H-related (FHR) proteins in the blood; that some FHR proteins accumulate in the back of the eye and cause an unwanted immune-inflammatory reaction; and, finally, that these proteins can be measured and tracked using a new technique that requires a very small (~100 µL) blood sample (1).
Where did this project come from? Last year, we stumbled across an unexpected FHR-4 elevation in AMD patients’ blood (2) and identified a link between the FHR family of proteins and AMD. Although there are seven members of the factor H protein family, we could only detect two of them, and the family as a whole was poorly understood. Once we developed a technique capable of measuring all seven proteins of the factor H family in human blood, we immediately tested it on the AMD blood samples from the previous study. Not only did we confirm the original FHR-4 discovery, but we also showed that almost all of the FHR proteins were elevated in AMD patients’ blood. In fact, the pattern in which they were elevated could be even more informative about a patient’s disease state or speed of progression.
A happy coincidence
As we drew closer to finalizing our study, we heard about another research group – led by Anneke den Hollander in Nijmegen, the Netherlands – who had set out to answer the same question. In their study, they used blood from a different, independent patient cohort and a different detection method that measured six of the seven factor H family proteins (3). Remarkably, their results were almost identical to ours, confirming the role of elevated circulating FHR proteins in driving AMD. We agreed to publish our two studies “back-to-back” in the same journal (1,3), each effectively providing independent validation data for the other original study.
The detection of all seven factor H family members simultaneously (FH, FHL-1, FHR-1, -2, -3, -4, and -5) had never previously been achieved. This new measurement technique opens up the possibility of examining the proteins’ role in a wide range of diseases in which complement activation is believed to play a role. Also, we know now that most or all of the FHR proteins are involved in AMD, giving us a better idea of what needs to be targeted therapeutically to try to prevent FHR-mediated risk of AMD.
This process has not been without barriers. Our biggest challenge was the one that has eluded scientists for years – how to distinguish all seven members of the factor H protein family from one another. They are so similar that most methods have failed – but now, by using mass spectrometry to make highly accurate protein measurements, we can differentiate them. A limitation of our approach is that we can’t specify an exact tissue location for FHRs. For that, we rely on the old antibody methods, which are tricky and specific for only two or three of the proteins. That is the next challenge – to visualize all seven of the proteins in eye tissue (instead of blood samples).
Our findings should impact the wider dry AMD research field. We suspect that there is a portion of dry AMD patients whose condition is driven by elevated FHR proteins accumulating in their macula and causing inflammation. If we’re right, targeting these FHR proteins so they don’t get to the eye could slow these patients’ disease progression. FHR proteins aren’t made in the eye; they are almost exclusively made in the liver, so lowering their levels in blood may prove effective. Because we can now measure the proteins in blood, we can easily identify patients who may benefit from such a therapeutic strategy and monitor their progress while targeting their FHR protein levels. Admittedly, we don’t yet have a FHR protein-targeting therapeutic but, now that we know we need one, it may be developed in the future. Importantly, we now have a way of measuring all of the FHR proteins in a single low-volume blood sample.
Looking to the future
We intend to progress this research further in several ways. First, we’re developing antibodies that can be used to visualize these FHR proteins in tissues. This will help us understand where they are accumulating and what they’re interacting with. This is not limited to eye research, but can also be used in organs affected by complement overactivation (for instance, the kidney, brain, or liver). Second, we are exploring ways of reducing FHR protein levels to try to help patients with elevated levels.
- V Cipriani et al., “Beyond factor H: The impact of genetic-risk variants for age-related macular degeneration on circulating factor-H-like 1 and factor-H-related protein concentrations,” Am J Hum Genet, 108, 1385 (2021). PMID: 34260948.
- V Cipriani et al., “Increased circulating levels of Factor H-Related Protein 4 are strongly associated with age-related macular degeneration,” Nat Commun, 11, 778 (2020). PMID: 32034129.
- L Lorés-Motta et al., “Common haplotypes at the CFH locus and low-frequency variants in CFHR2 and CFHR5 associate with systemic FHR concentrations and age-related macular degeneration” Am J Hum Genet, 108, 1367 (2021). PMID: 34260947.