RPTOR is an Alzheimer’s Disease susceptibility gene associated with Alzheimer’s Disease risk factors BMI and infectious encephalitis

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RPTOR is an Alzheimer’s Disease susceptibility gene associated with Alzheimer’s Disease risk factors BMI and infectious encephalitis

https://doi.org/10.21203/rs.3.rs-3639903/v1

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Background: In comparison to persons who did not have viral encephalitis, people with viral encephalitis had a later-life risk of Alzheimer's disease (AD) that was 31 times higher. In a previous study, we were able to confirm the association of viral encephalitis with AD and suggest that West Nile Virus infection is a significant AD risk factor. A genome wide association study (GWAS) with UK Biobank data revealed that the gene RAR Related Orphan Receptor B (RORB), previously linked to AD, is significantly associated with viral encephalitis.

Methods: In the current study we used data from PheWeb to identify genes other than RORB that might be involved in both infectious encephalitis and AD. We used UK Biobank data to examine gene expression and phenotypic expression.

Results: PheWeb identified additional genes associated with both infectious encephalitis and AD. RPTOR, a gene associated with the mTOR pathway, emerges as significant. Analyses of UK Biobank data reveal the impact of RPTOR on AD risk, with carriers of the minor allele A exhibiting decreased prevalence in subjects under age 55. Further analysis demonstrates that RPTOR genotypes influence Body Mass Index (BMI) in subjects of all ages, with carriers of the minor allele A having lower BMI. Logistic regression analyses confirm the association between reduced BMI and increased AD risk, along with the established factor of age.

Conclusion: RPTOR may represent an early onset AD gene, though mTOR's role in AD and BMI is complex. Nevertheless, RPTOR and mTOR could represent potential therapeutic targets for AD.

Cellular & Molecular Neuroscience

neurodegeneration

encephalitis

body mass index

An increased chance of developing Alzheimer's or Parkinson's disease later in life has been linked to infections with the influenza virus and other common viruses, according to an examination of about 450,000 electronic health records. One of the strongest correlations was found between viral encephalitis, an uncommon brain inflammation that can be brought on by a variety of viruses, and Alzheimer's disease (AD). In comparison to those who did not have encephalitis, those with encephalitis had a later-life risk of Alzheimer's disease that was roughly 31 times higher [1, 2].

West Nile virus (WNV) is the main mosquito-borne disease in the continental United States. The most typical way for WNV to spread to humans is by a mosquito bite. The mosquito season, which begins in the summer and lasts until the fall, is when WNV cases develop. The majority of WNV carriers have no symptoms. One in five infected individuals have fever and other symptoms. One in 150 infected individuals develop encephalitis [3].

In a previous study, we were able to confirm the association of viral encephalitis with AD and suggest that WNV infection is a significant AD risk factor. A genome wide association study (GWAS) with UK Biobank data revealed that the gene RAR Related Orphan Receptor B (RORB) is significantly associated with viral encephalitis. [4]. RORB is an AD vulnerability factor [5] and an epilepsy gene [6]. Results in Finngen indicate that epilepsy increases risk of viral encephalitis, and thus AD, with hazards ratio (HR) 4.92 [7].

In the current study we used data from PheWeb [8] to identify genes other than RORB that might be involved in both infectious encephalitis and AD. We used UK Biobank data to examine gene expression and phenotypic expression.

PheWeb is a user-friendly, open-source web application for sharing, exploring, and visualizing GWAS results [8]. PheWeb includes findings from genome-wide association studies (GWAS) of 1542 broad phenome wide association studies (PheWAS) codes collected from electronic health records (EHRs) for over 51.8 million imputed variations in 51,583 individuals of European ancestry from the Michigan Genomics Initiative (MGI), a biobank of patients at Michigan Medicine. The PheWAS R package (https://github.com/PheWAS/PheWAS) is used to extract diagnosis codes from patient electronic health records and generate phenotypes. The TOPMed reference panel is used to impute genetic data in MGI participants to provide association testing at high-quality genetic variants for each phenotype. Pheome-wide association analysis of individual genetic variants and interactive study of individual GWAS are made possible via the PheWeb interface. The gene regulatory associated protein of MTOR complex 1 (RPTOR) emerged in the GWAS (see results below).

To examine RPTOR in relation to AD and BMI we used UK Biobank data. The UK Biobank (UKBB) is a large prospective observational study comprising approximately 500,000 men and women (N = 229,134 men, N = 273,402 women), more than 90% white, aged 40–69 years at enrollment. Our UK Biobank project: 57245, Lehrer and Rheinstein. We analyzed the RPTOR snp rs12940622, an intron variant, G>A, minor allele (A) frequency 0.43.

Data processing was performed on Minerva, a Linux mainframe with Centos 7.6, at the Icahn School of Medicine at Mount Sinai. We used PLINK, a whole-genome association analysis toolset, to analyze the UKB chromosome files [9] and the UK Biobank Data Parser (ukbb parser), a python-based package that allows easy interfacing with the large UK Biobank dataset [10]. PheWeb uses LocusZoom for the Manhattan and q-q plots and calculation of lambda, the genetic control inflation factor [11].

Figure 1A displays PheWeb GWAS results for 134 cases of encephalitis and 114 cases of non-infectious encephalitis, Fig. 1B. RANBP3L and RPTOR are significantly associated with encephalitis, but RANBP3L is associated with non-infectious encephalitis. Most encephalitis cases that have been diagnosed in the United States are infectious, caused by enteroviruses, arboviruses (including the West Nile Virus), and herpes simplex virus types 1 and 2, which are spread from infected animals to humans through the bite of an infected tick, mosquito, or other bloodsucking parasite [12]. We have excluded RANBP3L from our analysis to eliminate cases of non-infectious encephalitis.

Figure 2A shows the QQ plot of p values from encephalitis data, Fig. 1A. Note that the leftmost p-values observed follow a uniform distribution (lower segment of line) but those that are in linkage disequilibrium with causal polymorphisms produce significant p-values (right segment of line). The genomic control inflation factor lambda, calculation based on the 50th percentile (median), is 0.205. Values up to 1.1 are generally considered acceptable for GWAS and suggest no systematic biases. Figure 2B is the QQ plot of p values from non-infectious encephalitis data, Fig. 1B, lambda 0.188.

Table 1 shows RPTOR genotype versus AD in 187,297 subjects age < 55. Note that subjects homozygous for the major allele G had increased prevalence of AD, while subjects heterozygous or homozygous for the minor allele A had decreased prevalence of AD (p = 0.05, two tail Fisher exact test).

Table 2 shows logistic regression with 95% confidence intervals, AD in subjects < age 55 dependent variable, sex, age, and RPTOR genotype independent variables. Increasing age was significantly associated with more AD (odds ratio [O.R.] 1.240). Subjects heterozygous or homozygous for the RPTOR minor allele A had decreased risk of AD (O.R. 0.517).

RPTOR genotypes had a significant effect on BMI. 153,249 subjects homozygous for major allele G had higher BMI 27.4702 ± 4.81742 (mean ± SD). 330,564 subjects heterozygous or homozygous for the minor allele A had lower BMI 27.3953 ± 4.77236. This difference was significant (p < 0.001, two tailed t test).

Table 3 shows logistic regression with 95% confidence intervals, AD in subjects all ages dependent variable, sex, age, and BMI independent variables. Reduced BMI was significantly associated with more AD (O.R. 0.982). Increasing age was significantly associated with more AD (O.R. 1.213). Male sex was significantly associated with more AD (O.R. 1.198). In another study of UK Biobank data, men had a higher risk of all-cause, young-onset and late-onset dementias than women [13].

The RPTOR gene is associated with the mTOR pathway [14]. Mammalian Target of Rapamycin, or mTOR, is a protein kinase that plays a crucial role in the regulation of cell growth, proliferation, and survival. It is involved in cellular processes, including protein synthesis, autophagy, and metabolism. Dysregulation of mTOR signaling has been implicated in the pathogenesis of several diseases, including neurodegenerative disorders and AD.

AD is characterized by the accumulation of abnormal protein aggregates in the brain, including beta-amyloid plaques and tau tangles. There is evidence to suggest that mTOR signaling may be involved in the development and progression of AD [15]:

1. Regulation of Protein Synthesis: mTOR is a central regulator of protein synthesis, and excessive protein synthesis can contribute to the accumulation of abnormal proteins, such as beta-amyloid, in the brain. Abnormal levels of beta-amyloid are a hallmark of AD.

2. Autophagy: mTOR regulates autophagy, a cellular process responsible for the degradation and recycling of damaged or dysfunctional cellular components. Dysfunctional autophagy may contribute to the accumulation of protein aggregates in neurodegenerative diseases.

3. Neuronal Function: mTOR is involved in the regulation of synaptic plasticity and neuronal function. Disruptions in these processes can contribute to cognitive decline, a characteristic feature of AD.

The RPTOR gene is related to Body Mass Index (BMI) [16-18]. BMI is a measure of body fat based on an individual's height and weight. It is widely used as a screening tool to categorize individuals into different weight categories, such as underweight, normal weight, overweight, and obesity. We were able to confirm the RPTOR-BMI relationship in UKBB data above, although the effect size was small [19].

While there is evidence that RPTOR can influence susceptibility to obesity, multiple genes are likely involved, and the interplay between genetic and environmental factors is complex. Numerous genetic variants have been associated with obesity-related traits, but the specific contribution of each gene and how they interact is still an area of active research [20].

Studies of the association between Body Mass Index (BMI) and Alzheimer's disease (AD) have yielded mixed findings [21].

There may be a link between being underweight or having low BMI and an increased risk of developing Alzheimer's disease [22]. Malnutrition or other factors related to low BMI may influence cognitive function negatively.

Obesity is a well-known risk factor for various health conditions, including cardiovascular diseases and type 2 diabetes. A possible link may exist between obesity and an increased risk of developing Alzheimer's disease [21]. Indeed, both low and high BMI may be associated with an increased risk of Alzheimer's disease, indicating a potential U-shaped relationship, the obesity paradox: obesity in midlife, low BMI in old age [23]. Any relationship between BMI and Alzheimer's is complex, and additional factors such as age, genetics, and overall health contribute to the risk.

Early-onset AD (onset < 65 years) and late-onset AD (onset > 65 years) are the two primary kinds of AD. According to data from twin and family studies, genetic factors are thought to be involved in at least 80% of instances of AD. Linkage analysis has identified the APP, PSEN1, and PSEN2 gene variants as responsible for some early onset AD. However, only a small percentage of early onset AD families may be explained by harmful mutations in these three genes [24]. Our analysis above suggests that RPTOR may represent a fourth early onset AD gene.

A weakness in our study: Effects on the blood brain barrier could not be evaluated. Infectious encephalitis disrupts the blood brain barrier [25, 26], and derangements of the blood brain barrier are an important part of AD [27].

Nevertheless, mTOR and RPTOR may represent potential therapeutic targets for AD, due to their involvement in various cellular processes implicated in AD. Modulating RPTOR and mTOR activity may have the potential to regulate protein synthesis, enhance autophagy, and improve neuronal function in people with AD. The role of RPTOR and mTOR in Alzheimer's disease is complex, and both excessive and insufficient mTOR activity may have negative effects. Balancing mTOR activity to achieve therapeutic benefits without causing harmful side effects will be a challenge.

Data sources described in the article are publicly available or can be accessed after approved application to UK Biobank.

Conflicts of interest: none

Competing interests: The authors declare no competing interests.

Dr. Lehrer and Dr. Rheinstein contributed equally to the conception, writing, and data analysis of this study.

This work was supported in part through the computational resources and staff expertise provided by Scientific Computing at the Icahn School of Medicine at Mount Sinai. Research reported in this paper was also supported by the Office of Research Infrastructure of the National Institutes of Health under award numbers S10OD018522 and S10OD026880. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

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Table 1. RPTOR genotype versus AD in 187,297 subjects age < 55. Note that subjects homozygous for the major allele G had increased prevalence of AD, while subjects heterozygous or homozygous for the minor allele A had decreased prevalence of AD (p = 0.05, two tail Fisher exact test).

		no AD	AD	total
GG	Count	59478	17	59495
	% within ALZ	31.80%	47.20%	31.80%
GA + AA	Count	127783	19	127802
	% within ALZ	68.20%	52.80%	68.20%
total	Count	187261	36	187297
	% within ALZ	100.00%	100.00%	100.00%

Table 2. Logistic regression with 95% confidence intervals, AD in subjects < age 55 dependent variable, sex, age, and RPTOR genotype independent variables, L.B. lower bound, O.R. odds ratio, U.B. upper bound. Increasing age was significantly associated with more AD (O.R. 1.240). Subjects heterozygous or homozygous for the RPTOR minor allele A had decreased risk of AD (O.R. 0.517).

	95% L.B.	O.R.	95% U.B.	p value
sex	0.743	1.430	2.753	0.284
age	1.120	1.240	1.373	<0.001
RPTOR	0.269	0.517	0.996	0.048

Table 3. Logistic regression with 95% confidence intervals, AD in subjects all ages dependent variable, sex, age, and BMI independent variables. Reduced BMI was significantly associated with more AD (O.R. 0.982). Increasing age was significantly associated with more AD (O.R. 1.213). Male sex was significantly associated with more AD (O.R. 1.198).

	95% L.B.	O.R.	95% U.B.	p value
sex	1.053	1.198	1.364	0.006
age	1.196	1.213	1.230	<0.001
BMI	0.967	0.982	0.998	0.020

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RPTOR is an Alzheimer’s Disease susceptibility gene associated with Alzheimer’s Disease risk factors BMI and infectious encephalitis

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