Sarah M. Carpanini

The association of genetic variants in the Human Leukocyte Antigen (HLA) locus with late-onset Alzheimer’s disease has been stringently replicated across several, powerful genome-wide association studies. However, no clear picture has yet emerged of the mechanistic relationship between Alzheimer’s disease and this top genetic hit, despite the fact that the HLA locus is one of the most influential gene loci of the immune system, known to influence antigen presentation, T cell responses and brain plasticity. In this review, we explore this association by outlining five research questions, namely: (i) the association of HLA Class I and Class II genes with Alzheimer’s disease at the allelic and haplotypic levels, (ii) the unconventional role of HLA Class I in the brain, (iii) the infection hypothesis of Alzheimer’s disease in the context of the known role HLA proteins play in immunity, (iv) the possible antigen presentation of Alzheimer’s disease relevant self-antigens and in turn (v) the possibility of T cells existing that are specific for these antigens. Identifying the functional mechanisms underlying this important genetic association with Alzheimer’s disease may hold the key to unravelling new avenues of Alzheimer’s disease immunotherapeutics.

Introduction: Triggering receptor expressed on myeloid cells-2 (TREM2) variants have been identified as risk factors for neurodegenerative disease, including Alzheimer’s disease. TREM2 is a cell surface receptor on microglia that regulates homeostatic and immunomodulatory functions, including phagocytosis of apoptotic debris and the resolution of damage-associated inflammation. It remains unclear how TREM2 may mediate an influence on neurodegenerative disease, particularly in relation to key neuropathological hallmarks such as neuronal loss and proteinopathy.Methods: We used the ME7 prion disease model to assess the role of TREM2 in the progression and pathology of neurodegenerative disease. Prion diseases are characterised by the accumulation of misfolded prion proteins and provide a highly tractable platform to determine if TREM2 has disease-modifying effects.Results: Trem2^−/− and wild-type (WT) mice were inoculated intracerebrally with mouse-passaged ME7 scrapie prions, and their effects on CNS disease pathogenesis were determined. Although the accumulation of prion disease-specific PrP was similar in the brains of mice from each group, the severity of neuropathology was increased in Trem2^−/− mice. Morphometric analysis of the microglia also indicated blunted disease-induced reactivity in the brains of infected Trem2^−/− mice compared to wild-type (WT) controls. Expression of genes involved in myelination was reduced in prion-infected Trem2^−/− mice compared to infected WT mice.Conclusion: We conclude that during brain infection with prions, TREM2 supports microglial reactive changes associated with resilience to neuronal loss independently of affecting misfolded PrP deposition. These data imply that TREM2 status may be an important influence on the downstream response to CNS proteinopathy, which alters the susceptibility of neurons and brain tissue to proteinopathy-induced degenerative changes.

Complement is an important component of innate immune defence against pathogens and crucial for efficient immune complex disposal. These core protective activities are dependent in large part on properly regulated complement-mediated inflammation. Dysregulated complement activation, often driven by persistence of activating triggers, is a cause of pathological inflammation in numerous diseases, including neurological diseases. Increasingly, this has become apparent not only in well-recognized neuroinflammatory diseases like multiple sclerosis but also in neurodegenerative and neuropsychiatric diseases where inflammation was previously either ignored or dismissed as a secondary event. There is now a large and rapidly growing body of evidence implicating complement in neurological diseases that cannot be comprehensively addressed in a brief review. Here, we will focus on neurodegenerative diseases, including not only the ‘classical’ neurodegenerative diseases such as Alzheimer’s disease and Parkinson’s disease, but also two other neurological diseases where neurodegeneration is a neglected feature and complement is implicated, namely, schizophrenia, a neurodevelopmental disorder with many mechanistic features of neurodegeneration, and multiple sclerosis, a demyelinating disorder where neurodegeneration is a major cause of progressive decline. We will discuss the evidence implicating complement as a driver of pathology in these diverse diseases and address briefly the potential and pitfalls of anti-complement drug therapy for neurodegenerative diseases.

Background: Alzheimer’s disease (AD) has been associated with immune dysregulation in biomarker and genome-wide association studies (GWAS). GWAS hits include the genes encoding complement regulators clusterin (CLU) and complement receptor 1 (CR1), recognised as key players in AD pathology, and complement proteins have been proposed as biomarkers. Main body: To address whether changes in plasma complement protein levels in AD relate to AD-associated complement gene variants we first measured relevant plasma complement proteins (clusterin, C1q, C1s, CR1, factor H) in a large cohort comprising early onset AD (EOAD; n = 912), late onset AD (LOAD; n = 492) and control (n = 504) donors. Clusterin and C1q were significantly increased (p < 0.001) and sCR1 and factor H reduced (p < 0.01) in AD plasma versus controls. ROC analyses were performed to assess utility of the measured complement biomarkers, alone or in combination with amyloid beta, in predicting AD. C1q was the most predictive single complement biomarker (AUC 0.655 LOAD, 0.601 EOAD); combining C1q with other complement or neurodegeneration makers through stepAIC-informed models improved predictive values slightly. Effects of GWS SNPs (rs6656401, rs6691117 in CR1; rs11136000, rs9331888 in CLU; rs3919533 in C1S) on protein concentrations were assessed by comparing protein levels in carriers of the minor vs major allele. To identify new associations between SNPs and changes in plasma protein levels, we performed a GWAS combining genotyping data in the cohort with complement protein levels as endophenotype. SNPs in CR1 (rs6656401), C1S (rs3919533) and CFH (rs6664877) reached significance and influenced plasma levels of the corresponding protein, whereas SNPs in CLU did not influence clusterin levels. Conclusion: Complement dysregulation is evident in AD and may contribute to pathology. AD-associated SNPs in CR1, C1S and CFH impact plasma levels of the encoded proteins, suggesting a mechanism for impact on disease risk.

Genome wide association studies (GWAS) have highlighted the importance of the complement cascade in pathogenesis of Alzheimer's disease (AD). Complement receptor 1 (CR1; CD35) is among the top GWAS hits. The long variant of CR1 is associated with increased risk for AD; however, roles of CR1 in brain health and disease are poorly understood. A critical confounder is that brain expression of CR1 is controversial; failure to demonstrate brain expression has provoked the suggestion that peripherally expressed CR1 influences AD risk. We took a multi‐pronged approach to establish whether CR1 is expressed in brain. Expression of CR1 at the protein and mRNA level was assessed in human microglial lines, induced pluripotent stem cell (iPSC)‐derived microglia from two sources and brain tissue from AD and control donors. CR1 protein was detected in microglial lines and iPSC‐derived microglia expressing different CR1 variants when immunostained with a validated panel of CR1‐specific antibodies; cell extracts were positive for CR1 protein and mRNA. CR1 protein was detected in control and AD brains, co‐localizing with astrocytes and microglia, and expression was significantly increased in AD compared to controls. CR1 mRNA expression was detected in all AD and control brain samples tested; expression was significantly increased in AD. The data unequivocally demonstrate that the CR1 transcript and protein are expressed in human microglia ex vivo and on microglia and astrocytes in situ in the human brain; the findings support the hypothesis that CR1 variants affect AD risk by directly impacting glial functions.

The presence of complement activation products at sites of pathology in post-mortem Alzheimer’s disease (AD) brains is well known. Recent evidence from genome-wide association studies (GWAS), combined with the demonstration that complement activation is pivotal in synapse loss in AD, strongly implicates complement in disease aetiology. Genetic variations in complement genes are widespread. While most variants individually have only minor effects on complement homeostasis, the combined effects of variants in multiple complement genes, referred to as the “complotype”, can have major effects. In some diseases, the complotype highlights specific parts of the complement pathway involved in disease, thereby pointing towards a mechanism; however, this is not the case with AD. Here we review the complement GWAS hits; CR1 encoding complement receptor 1 (CR1), CLU encoding clusterin, and a suggestive association of C1S encoding the enzyme C1s, and discuss difficulties in attributing the AD association in these genes to complement function. A better understanding of complement genetics in AD might facilitate predictive genetic screening tests and enable the development of simple diagnostic tools and guide the future use of anti-complement drugs, of which several are currently in development for central nervous system disorders.

Late-onset Alzheimer’s disease (LOAD), the most common cause of dementia, and a huge global health challenge, is a neurodegenerative disease of uncertain aetiology. To deliver effective diagnostics and therapeutics, understanding the molecular basis of the disease is essential. Contemporary large genome-wide association studies (GWAS) have identified over seventy novel genetic susceptibility loci for LOAD. Most are implicated in microglial or inflammatory pathways, bringing inflammation to the fore as a candidate pathological pathway. Among the most significant GWAS hits are three complement genes: CLU, encoding the fluid-phase complement inhibitor clusterin; CR1 encoding complement receptor 1 (CR1); and recently, C1S encoding the complement enzyme C1s. Complement activation is a critical driver of inflammation; changes in complement genes may impact risk by altering the inflammatory status in the brain. To assess complement gene association with LOAD risk, we manually created a comprehensive complement gene list and tested these in gene-set analysis with LOAD summary statistics. We confirmed associations of CLU and CR1 genes with LOAD but showed no significant associations for the complement gene-set when excluding CLU and CR1. No significant association with other complement genes, including C1S, was seen in the IGAP dataset; however, these may emerge from larger datasets.

The complement system plays critical roles in development, homeostasis, and regeneration in the central nervous system (CNS) throughout life; however, complement dysregulation in the CNS can lead to damage and disease. Complement proteins, regulators, and receptors are widely expressed throughout the CNS and, in many cases, are upregulated in disease. Genetic and epidemiological studies, cerebrospinal fluid (CSF) and plasma biomarker measurements and pathological analysis of post-mortem tissues have all implicated complement in multiple CNS diseases including multiple sclerosis (MS), neuromyelitis optica (NMO), neurotrauma, stroke, amyotrophic lateral sclerosis (ALS), Alzheimer's disease (AD), Parkinson's disease (PD), and Huntington's disease (HD). Given this body of evidence implicating complement in diverse brain diseases, manipulating complement in the brain is an attractive prospect; however, the blood-brain barrier (BBB), critical to protect the brain from potentially harmful agents in the circulation, is also impermeable to current complement-targeting therapeutics, making drug design much more challenging. For example, antibody therapeutics administered systemically are essentially excluded from the brain. Recent protocols have utilized “Trojan horse” techniques to transport therapeutics across the BBB or used osmotic shock or ultrasound to temporarily disrupt the BBB. Most research to date exploring the impact of complement inhibition on CNS diseases has been in animal models, and some of these studies have generated convincing data; for example, in models of MS, NMO, and stroke. There have been a few recent clinical trials of available anti-complement drugs in CNS diseases associated with BBB impairment, for example the use of the anti-C5 monoclonal antibody (mAb) eculizumab in NMO, but for most CNS diseases there have been no human trials of anti-complement therapies. Here we will review the evidence implicating complement in diverse CNS disorders, from acute, such as traumatic brain or spine injury, to chronic, including demyelinating, neuroinflammatory, and neurodegenerative diseases. We will discuss the particular problems of drug access into the CNS and explore ways in which anti-complement therapies might be tailored for CNS disease.