TREM2 Variants Research Articles

INVESTIGATING GENETIC VARIATION IN ALZHEIMER’S DISEASE USING WHOLE-EXOME SEQUENCING

Alzheimer's disease (AD) is an incurable neurodegenerative disorder; in which the death of brain cells characteristically result in memory loss and cognitive decline. It is the most common form of dementia, affecting around 850,000 people in the UK. Sporadic late-onset AD (LOAD) accounts for 95% of all cases and is genetically complex in nature. It is believed that combinations of genetic and environmental factors are at play. Recent genome-wide association studies (GWAS) have uncovered over 20 new gene candidates for AD risk, however these exhibit small effect sizes. We are now utilising next generation sequencing (NGS) to explore the contribution made by rare variants (MAF<5%). Recently this approach highlighted the role of TREM2 and SORL1 variants in AD risk and emerging NeuroX chip and NGS data is set to generate more genes of interest. DNA was extracted from post-mortem brain tissue obtained from the BDR for healthy and diseased individuals. Whole-exome sequencing was performed on 292 samples, including 133 AD cases, 53 controls and 106 other phenotypes. Samples were screened for mutations in APP, PSEN1 and PSEN2 to distinguish early-onset AD cases and known LOAD risk genes. Using a combination of bioinformatics and statistical tools variants were tested for association with AD and their functionality assessed in silico. Presently over 340,000 variants have been identified in the whole-exome dataset, with 18% predicted to be novel. Coding variants account for ∼55% of all variants, with >50% of these being missense or frameshift mutations. A total of 653 SNPs were found in GWAS nominated genes. APOEε4 SNP rs429358 reached genome-wide significance (p<5x10-8, OR=6.5) in single SNP association without adjusting for covariates. It is well known that APOEε4 significantly increases AD risk. Although no other SNPs reached genome-wide significance, it is likely that important variants could lie below this threshold. Therefore AD risk gene variants and other neurodegenerative disease risk genes will also be investigated further.

Mutations in TYROBP are not a common cause of dementia in a Turkish cohort

Mutations in TYROBP and TREM2 have been shown to cause polycystic lipomembranous osteodysplasia with sclerosing leukoencephalopathy. Recently, variants in TREM2 were also associated with frontotemporal dementia and Alzheimer's disease. Given the functional proximity between these 2 genes, we investigated the genetic variation of TYROBP in a Turkish cohort of 103 dementia patients. No mutations or copy number variants predicted to be pathogenic were identified. These results indicate that mutations in TYROBP are not a common cause of dementia in this Turkish cohort.

Neocortical and hippocampal TREM2 protein levels during the progression of Alzheimer's disease

Heterozygous triggering receptor expressed on myeloid cells (TREM2) mutations are an Alzheimer's disease (AD) risk factor. Nonmutated TREM2 dysregulation occurs in AD brain. Whether TREM2 is altered in prodromal AD remains unknown. Western blotting was used to determine levels of TREM2 (∼25 kDa) and Iba1 in the frontal cortex and TREM2 in the hippocampus from people who died with an ante-mortem clinical diagnosis of non- and mild-cognitive impairment, mild/moderate AD, and severe AD (sAD). Immunohistochemistry defined the relationship between amyloid and Iba1 profiles. Polymerase chain reaction analysis revealed that all subjects did not carry the most common R47H TREM2 variant. TREM2 was significantly upregulated in sAD frontal cortex but stable in hippocampus. Frontal TREM2 mRNA and protein level patterns were similar but not significantly different. Iba1 immunopositive microglia counts increased significantly in frontal cortex containing plaques in sAD. TREM2 and Iba1 levels were not associated with plaques, tangles, neuropathological criteria, or cognitive performance. Frontal cortex TREM2 upregulation is a late event and may not play a major role early in the pathogenesis of the disease.

Identification of a rare coding variant in TREM2 in a Chinese individual with Alzheimer’s disease

ABSTRACTRare variation in the TREM2 gene is associated with a broad spectrum of neurodegenerative disorders including Alzheimer’s disease (AD). TREM2 encodes a receptor expressed in microglia which is thought to influence neurodegeneration by sensing damage signals and regulating neuroinflammation. Many of the variants reported to be associated with AD, including the rare R47H variant, were discovered in populations of European ancestry and have not replicated in diverse populations from other genetic backgrounds. We utilized a cohort of elderly Chinese individuals diagnosed as cognitively normal, or with mild cognitive impairment or AD to identify a rare variant, A192T, present in a single patient diagnosed with AD. We characterized this variant using biochemical cell surface expression assays and found that it significantly altered cell surface expression of the TREM2 protein. Together these data provide evidence that the A192T variant in TREM2 could contribute risk for AD. This study underscores the increasingly recognized role of immune-related processes in AD and highlights the importance of including diverse populations in research to identify genetic variation that contributes risk for AD and other neurodegenerative disorders.

Rare coding variants in PLCG2, ABI3, and TREM2 implicate microglial-mediated innate immunity in Alzheimer's disease

Rare coding variants in PLCG2, ABI3, and TREM2 implicate microglial-mediated innate immunity in Alzheimer's disease

Neurodegenerative disease mutations in TREM2 reveal a functional surface and distinct loss-of-function mechanisms.

Genetic variations in the myeloid immune receptor TREM2 are linked to several neurodegenerative diseases. To determine how TREM2 variants contribute to these diseases, we performed structural and functional studies of wild-type and variant proteins. Our 3.1 Å TREM2 crystal structure revealed that mutations found in Nasu-Hakola disease are buried whereas Alzheimer's disease risk variants are found on the surface, suggesting that these mutations have distinct effects on TREM2 function. Biophysical and cellular methods indicate that Nasu-Hakola mutations impact protein stability and decrease folded TREM2 surface expression, whereas Alzheimer's risk variants impact binding to a TREM2 ligand. Additionally, the Alzheimer's risk variants appear to epitope map a functional surface on TREM2 that is unique within the larger TREM family. These findings provide a guide to structural and functional differences among genetic variants of TREM2, indicating that therapies targeting the TREM2 pathway should be tailored to these genetic and functional differences with patient-specific medicine approaches for neurodegenerative disorders.

Rare TREM2 variants associated with Alzheimer's disease display reduced cell surface expression.

Rare variation in TREM2 has been associated with greater risk for Alzheimer’s disease (AD). TREM2 encodes a cell surface receptor expressed on microglia and related cells, and the R47H variant associated with AD appears to affect the ability of TREM2 to bind extracellular ligands. In addition, other rare TREM2 mutations causing early-onset neurodegeneration are thought to impair cell surface expression. Using a sequence kernel association (SKAT) analysis in two independent AD cohorts, we found significant enrichment of rare TREM2 variants not previously characterized at the protein level. Heterologous expression of the identified variants showed that novel variants S31F and R47C displayed significantly reduced cell surface expression. In addition, we identified rare variant R136Q in a patient with language-predominant AD that also showed impaired surface expression. The results suggest rare TREM2 variants enriched in AD may be associated with altered TREM2 function and that AD risk may be conferred, in part, from altered TREM2 surface expression.Electronic supplementary materialThe online version of this article (doi:10.1186/s40478-016-0367-7) contains supplementary material, which is available to authorized users.

Alzheimer's disease-associated TREM2 variants exhibit either decreased or increased ligand-dependent activation

IntroductionTREM2 is a lipid-sensing activating receptor on microglia known to be important for Alzheimer's disease (AD), but whether it plays a beneficial or detrimental role in disease pathogenesis is controversial. MethodsWe analyzed AD risk of TREM2 variants in the NIMH AD Genetics Initiative Study and AD Sequencing Project. We compared each variant's risk and functional impact by a reporter assay. Finally, we analyzed expression of TREM2 on human monocytes. ResultsWe provide more evidence for increased AD risk associated with several TREM2 variants, and show that these variants decreased or markedly increased binding to TREM2 ligands. We identify HDL and LDL as novel TREM2 ligands. We also show that TREM2 expression in human monocytes is minimal compared to monocyte-derived dendritic cells. DiscussionOur results suggest that TREM2 signaling helps protect against AD but can cause harm in excess, supporting the idea that proper TREM2 function is important to counteract disease progression.

TREM2 Binds to Apolipoproteins, Including APOE and CLU/APOJ, and Thereby Facilitates Uptake of Amyloid-Beta by Microglia

Genetic variants of TREM2, a protein expressed selectively by microglia in the brain, are associated with Alzheimer's disease (AD). Starting from an unbiased protein microarray screen, we identified a set of lipoprotein particles (including LDL) and apolipoproteins (including CLU/APOJ and APOE) as ligands ofTREM2. Binding of these ligands by TREM2 was abolished or reduced by disease-associated mutations. Overexpression of wild-type TREM2 was sufficient to enhance uptake of LDL, CLU, and APOE in heterologous cells, whereas TREM2 disease variants were impaired in this activity. Trem2 knockout microglia showed reduced internalization of LDL and CLU. β-amyloid (Aβ) binds to lipoproteins and this complex is efficiently taken up by microglia in a TREM2-dependent fashion. Uptake of Aβ-lipoprotein complexes was reduced in macrophages from human subjects carrying a TREM2 AD variant. These data link three genetic risk factors for AD and reveal apossible mechanism by which mutant TREM2 increases risk of AD. VIDEO ABSTRACT.

AD-ASSOCIATED TREM2 VARIANTS LEAD TO SOME SUBPOPULATIONS OF MICROGLIA TO BE LESS ABUNDANT BUT MORE ACTIVATED

AD-ASSOCIATED TREM2 VARIANTS LEAD TO SOME SUBPOPULATIONS OF MICROGLIA TO BE LESS ABUNDANT BUT MORE ACTIVATED

Mutation analysis of the MS4A and TREM gene clusters in a case-control Alzheimer's disease data set

Genome wide association studies have identified an association between Alzheimer's disease (AD) and common polymorphisms in the MS4A and TREM loci (each containing a cluster of homologous genes) and should be thoroughly investigated for the presence of potentially functional variations. We conducted a mutation analysis by next generation sequencing of 15 genes within the MS4A and TREM gene clusters; and catalogued rare coding variants detected in a North American data set of 210 cases and 233 controls. Investigation of the 5 homologues genes in the TREM locus revealed potentially damaging rare variants in TREM2, TREML1, TREML2, and TREML4. In agreement with a previous report, we observed a significant enrichment of TREM2-damaging missense substitutions in cases (N = 9; 4.2%) compared with controls (N=2; 0.9%; p = 0.010; after Yates' correction p = 0.022). Among known AD-associated TREM2 substitutions, we detected p.R47H, p.D87N, and p.H157Y affecting both TREM2 isoforms (NM_018965 and NM_001271821). In addition, we identified 2 cases with novel TREM2 variants (p.L205P and p.G219C), which mapped only to the isoform NM_001271821 at the C-terminus. Investigation of the MS4A gene cluster revealed that potentially damaging missense substitutions and loss-of-function variants were twice as frequent in controls (N = 19; 8.2%) than cases (N = 9; 4.3%), generating a nominally significant result (p = 0.047; after Yates' correction p = 0.07). Validation of our observation in large data sets might address the question whether such variants could contribute to the protective effect of the minor alleles of Genome wide association study–significant single nucleotide polymorphisms at the MS4A locus.

TREM2 Function in Alzheimer’s Disease and Neurodegeneration

Alzheimer's disease (AD), the most common cause of dementia in the elderly, is a complex neurodegenerative disease marked by the appearance of amyloid-β (Aβ) plaques and hyperphosphorylated tau tangles. Alzheimer's disease has a strong genetic component, and recent advances in genome technology have unearthed novel variants in several genes, which could provide insight into the pathogenic mechanisms that contribute to AD. Particularly interesting are variants in the microglial-expressed receptor TREM2 which are associated with a 2-4-fold increased risk of developing AD. Since the discovery of a link between TREM2 and AD, multiple studies have emerged testing whether partial or complete loss of TREM2 function contributed to Aβ deposition or Aβ-associated microgliosis. Although some confounding conflicting data have emerged from these studies regarding the role of TREM2 in regulating Aβ deposition within the hippocampus, the most consistent and striking observation is a strong decrease in microgliosis surrounding Aβ plaques in TREM2 haploinsufficient and TREM2 deficient mice. Interestingly, a similar impairment in microgliosis has been reported in mouse models of prion disease, stroke, and multiple sclerosis, suggesting a critical role for TREM2 in supporting microgliosis in response to pathology in the central nervous system. In this Review, we summarize recent reports on the role of TREM2 in AD pathology and hypothesized mechanisms by which TREM2 function could influence AD-induced microgliosis.

Cerebrospinal fluid soluble TREM2 is higher in Alzheimer disease and associated with mutation status.

Low frequency coding variants in TREM2 are associated with increased Alzheimer disease (AD) risk, while loss of functions mutations in the gene lead to an autosomal recessive early-onset dementia, named Nasu-Hakola disease (NHD). TREM2 can be detected as a soluble protein in cerebrospinal fluid (CSF) and plasma, and its CSF levels are elevated in inflammatory CNS diseases. We measured soluble TREM2 (sTREM2) in the CSF of a large AD case-control dataset (n=180) and 40 TREM2 risk variant carriers to determine whether CSF sTREM2 levels are associated with AD status or mutation status. We also performed genetic studies to identify genetic variants associated with CSF sTREM2 levels. CSF, but not plasma, sTREM2 was highly correlated with CSF total tau and phosphorylated-tau levels (r=0.35, P<1×10(-4); r=0.40, P<1×10(-4), respectively), but not with CSF Aβ42. AD cases presented higher CSF sTREM2 levels than controls (P=0.01). Carriers of NHD-associated TREM2 variants presented significantly lower CSF sTREM2 levels, supporting the hypothesis that these mutations lead to reduced protein production/function (R136Q, D87N, Q33X or T66M; P=1×10(-3)). In contrast, CSF sTREM2 levels were significantly higher in R47H carriers compared to non-carriers (P=6×10(-3)), suggesting that this variant does not impact protein expression and increases AD risk through a different pathogenic mechanism than NHD variants. In GWAS analyses for CSF sTREM2 levels the most significant signal was located on the MS4A gene locus (P=5.45×10(-07)) corresponding to one of the SNPs reported to be associated with AD risk in this locus. Furthermore, SNPs involved in pathways related to virus cellular entry and vesicular trafficking were overrepresented, suggesting that CSF sTREM2 levels could be an informative phenotype for AD.

Targeted sequencing approach to identify genetic mutations in Nasu-Hakola disease.

Nasu-Hakola disease (NHD) is a rare autosomal recessive disorder characterized by sclerosing leukoencephalopathy and multifocal bone cysts, caused by a loss-of-function mutation of either TYROBP (DAP12) or TREM2. TREM2 and DAP12 constitute a receptor/adaptor signaling complex expressed exclusively on osteoclasts, dendritic cells, macrophages, and microglia. Premortem molecular diagnosis of NHD requires genetic analysis of both TYROBP and TREM2, in which 20 distinct NHD-causing mutations have been reported. Due to genetic heterogeneity, it is often difficult to identify the exact mutation responsible for NHD. Recently, the revolution of the next-generation sequencing (NGS) technology has greatly advanced the field of genome research. A targeted sequencing approach allows us to investigate a selected set of disease-causing genes and mutations in a number of samples within several days. By targeted sequencing using the TruSight One Sequencing Panel, we resequenced genetic mutations of seven NHD cases with known molecular diagnosis and two control subjects. We identified homozygous variants of TYROBP or TREM2 in all NHD cases, composed of a frameshift mutation of c.141delG in exon 3 of TYROBP in four cases, a missense mutation of c.2T>C in exon 1 of TYROBP in two cases, or a splicing mutation of c.482+2T>C in intron 3 of TREM2 in one case. The results of targeted resequencing corresponded to those of Sanger sequencing. In contrast, causative variants were not detected in control subjects. These results indicate that targeted sequencing is a useful approach to precisely identify genetic mutations responsible for NHD in a comprehensive manner.

Genetic analysis of TREM2 variants in Chinese Han patients with sporadic Parkinson’s disease

Parkinson’s disease (PD) is the second most common neurodegenerative disorder after Alzheimer’s disease and is characterized by the degeneration of dopaminergic neurons in substantia nigra. Recently, rs75932628 (p.R47H) of the triggering receptor expressed on myeloid cells 2 gene (TREM2) was identified to be associated with PD in American, Spanish, Irish, and Polish population. To explore whether TREM2 variants are related to susceptibility of sporadic PD in Chinese Han population, we designed a case-control comparison study and studied two variants rs75932628 (p.R47H) and rs2234253 (p.T96K) of the TREM2 gene in 512 Chinese Han patients with sporadic PD and 512 age, gender and ethnicity matched normal controls from Mainland China. No variant for either rs75932628 or rs2234253 was found in both PD and control cohorts. Our data suggest that neither variant rs75932628 nor rs2234253 be a major susceptibility factor of sporadic PD in Chinese Han population from Mainland China.

Convergent Genetic and Expression Datasets Highlight TREM2 in Parkinson's Disease Susceptibility.

A rare TREM2 missense mutation (rs75932628-T) was reported to confer a significant Alzheimer's disease (AD) risk. A recent study indicated no evidence of the involvement of this variant in Parkinson's disease (PD). Here, we used the genetic and expression data to reinvestigate the potential association between TREM2 and PD susceptibility. In stage 1, using 10 independent studies (N = 89,157; 8787 cases and 80,370 controls), we conducted a subgroup meta-analysis. We identified a significant association between rs75932628 and PD (P = 3.10E-03, odds ratio (OR) = 3.88, 95% confidence interval (CI) 1.58-9.54) in No-Northern Europe subgroup, and significantly increased PD risks (P = 0.01 for Mann-Whitney test) in No-Northern Europe subgroup than in Northern Europe subgroup. In stage 2, we used the summary results from a large-scale PD genome-wide association study (GWAS; N = 108,990; 13,708 cases and 95,282 controls) to search for other TREM2 variants contributing to PD susceptibility. We identified 14 single-nucleotide polymorphisms (SNPs) associated with PD within 50-kb upstream and downstream range of TREM2. In stage 3, using two brain expression GWAS datasets (N = 773), we identified 6 of the 14 SNPs regulating increased expression of TREM2. In stage 4, using the whole human genome microarray data (N = 50), we further identified significantly increased expression of TREM2 in PD cases compared with controls in human prefrontal cortex. In summary, convergent genetic and expression datasets demonstrate that TREM2 is a potent risk factor for PD and may be a therapeutic target in PD and other neurodegenerative diseases.

Personalized medicine for effective Alzheimer disease treatment.

Treatments for Alzheimer disease—even treatments that minimally modify the disease—have remained stubbornly elusive. Although hypotheses concerning a disease mechanism have resulted in target-engaging drugs that have been tested in large clinical trials, the results of all trials to date have been negative. Most of the rethinking prompted by these clinical trial failures has driven researchers to reexamine their hypotheses concerning the disease mechanism and to initiate trials earlier in the course of the disease. 1 However, another reason for the apparent failure of large clinical trials to identify effective treatments may be the erroneous assumption that Alzheimer disease is a single clinical entity. A complete risk profile of a patient with Alzheimer disease reveals vast variation in risk markers that may serve to guide treatment. Associating diseases that require different treatment strategies may obscure small responsive subgroups. The data set that defines a person’s risk of developing Alzheimer disease is becoming increasingly rich. These data include numerous genetic variants, some of which are rare, while others are more frequent. The most well-recognized variant is APOE e4, which exerts its effect in a dose-dependent manner. Each APOE e4 allele increases a person’s likelihood of developing Alzheimer disease by about a decade compared with the general population. Risk variants with smaller effect size than APOE e4 include noncoding changes in ABCA7, BIN1, CASS4, CD33, CD2AP, CELF1, CLU, CR1, DSG2, EPHA1, FERMT2, HLA-DRB5-DBR1, INPP5D, MS4A, MEF2C, NME8, PICALM, PTK2B, SLC24H4-RIN3, SORL1, and ZCWPW1 and coding variants in PLD3 and TREM2. 2 Risk variants are not uniformly distributed in the population; therefore, the likelihood of carrying a specific genetic variant depends on one’s genetic background. Ascertaining the presence of these variants may shape the choice of treatment strategies because some of these genes may result in opportunities for effective drug treatments for carriers of the variant. For example, variants in genes related to inflammation may suggest a treatment cocktail that includes medications that suppress

The role of TREM2 R47H as a risk factor for Alzheimer's disease, frontotemporal lobar degeneration, amyotrophic lateral sclerosis, and Parkinson's disease

A rare variant in TREM2 (p.R47H, rs75932628) was recently reported to increase the risk of Alzheimer's disease (AD) and, subsequently, other neurodegenerative diseases, i.e. frontotemporal lobar degeneration (FTLD), amyotrophic lateral sclerosis (ALS), and Parkinson's disease (PD). Here we comprehensively assessed TREM2 rs75932628 for association with these diseases in a total of 19,940 previously untyped subjects of European descent. These data were combined with those from 28 published data sets by meta-analysis. Furthermore, we tested whether rs75932628 shows association with amyloid beta (Aβ42) and total-tau protein levels in the cerebrospinal fluid (CSF) of 828 individuals with AD or mild cognitive impairment. Our data show that rs75932628 is highly significantly associated with the risk of AD across 24,086 AD cases and 148,993 controls of European descent (odds ratio or OR = 2.71, P = 4.67 × 10−25). No consistent evidence for association was found between this marker and the risk of FTLD (OR = 2.24, P = .0113 across 2673 cases/9283 controls), PD (OR = 1.36, P = .0767 across 8311 cases/79,938 controls) and ALS (OR = 1.41, P = .198 across 5544 cases/7072 controls). Furthermore, carriers of the rs75932628 risk allele showed significantly increased levels of CSF-total-tau (P = .0110) but not Aβ42 suggesting that TREM2's role in AD may involve tau dysfunction.

R47H TREM2 variant increases risk of typical early-onset Alzheimer's disease but not of prion or frontotemporal dementia

BackgroundRare TREM2 variants are significant risk factors for Alzheimer's disease (AD). MethodsWe used next generation sequencing of the whole gene (n = 700), exon 2 Sanger sequencing (n = 2634), p.R47H genotyping (n = 3518), and genome wide association study imputation (n = 13,048) to determine whether TREM2 variants are risk factors or phenotypic modifiers in patients with AD (n = 1002), frontotemporal dementia (n = 358), sporadic (n = 2500), and variant (n = 115) Creutzfeldt-Jakob disease (CJD). ResultsWe confirm only p.R47H as a risk factor for AD (odds ratio or OR = 2.19; 95% confidence interval or CI = 1.04-4.51; P = .03). p.R47H does not significantly alter risk for frontotemporal dementia (OR = 0.81), variant or sporadic CJD (OR = 1.06 95%CI = 0.66-1.69) in our cohorts. Individuals with p.R47H associated AD (n = 12) had significantly earlier symptom onset than individuals with no TREM2 variants (n = 551) (55.2 years vs. 61.7 years, P = .02). We note that heterozygous p.R47H AD is memory led and otherwise indistinguishable from “typical” sporadic AD. ConclusionWe find p.R47H is a risk factor for AD, but not frontotemporal dementia or prion disease.

Lack of genetic association between TREM2 and late-onset Alzheimer's disease in a Japanese population.

Rare non-synonymous variants of TREM2 have recently been shown to be associated with Alzheimer's disease (AD) in Caucasians. We here conducted a replication study using a well-characterized Japanese sample set, comprising 2,190 late-onset AD (LOAD) cases and 2,498 controls. We genotyped 10 non-synonymous variants (Q33X, Y38C, R47H, T66M, N68K, D87N, T96K, R98W, H157Y, and L211P) of TREM2 reported by Guerreiro et al. (2013) by means of the TaqMan and dideoxy sequencing methods. Only three variants, R47H, H157Y, and L211P, were polymorphic (range of minor allele frequency [MAF], 0.0002-0.0059); however, no significant association with LOAD was observed in these variants. Considering low MAF of variants examined and our study sample size, further genetic analysis with a larger sample set is needed to firmly evaluate whether or not TREM2 is associated with LOAD in Japanese.