ADAPTED

Summary

ADAPTED investigated the APOE gene, a well-known risk factor for developing Alzheimer’s disease (AD), in order to better understood why people who carry the APOE4 variant have a considerably higher risk of developing AD. The main outputs of the project are the high-quality cellular models derived from human stem cells that express the different APOE genotypes, including those expressing the risk variant, but also variants that seem to confer neutral or even reduced risk to carriers, and cells where the APOE gene had been deleted.

During the project, the ADAPTED partners amassed a large and important collection of samples of cerebrospinal fluid, blood serum and saliva that are still generating data and are now stored at a Barcelona biobank, as well as troves of data from their various findings about APOE. These findings have done a lot to help advance our understanding of Alzheimer’s risk, including the identification of a rare gene variant that slows AD progression and seems to counteract the effects of APOE4, thus suggesting a new drug target, as well as potential protein blood biomarkers for pre-symptomatic AD, and links between heart problems and later AD diagnosis. Their models, data and biobank samples are all available for the research community to access and use. In addition, an SME partner in the project developed a model of the human blood–brain barrier (BBB) that can be used to assess the passage of large antibodies across the BBB and so could be of enormous value in the drug screening process.

Background

Alzheimer’s disease (AD) affects over 35 million people worldwide, yet there is a severe lack of medicines that can slow down the progress of the disease. The ADAPTED project aimed to boost the development of new medicines by investigating an area of AD research that has previously received little attention – the APOE gene. The APOE gene is well known for conferring varying levels of risk for developing the disease, but it is not known precisely why this is so. People who carry the APOE4 version of the gene have a considerably higher risk of developing AD, and tend to develop the disease much earlier in life.

To APOE models

Apolipoprotein E (APOE) is a protein involved in the transport of cholesterol in the bloodstream, and research suggests that it helps clear beta-amyloid, a type of plaque that is one of the principal hallmarks of AD. The APOE gene is involved in giving instructions for the creation of this protein, and everybody has two copies of it. There are three possible variants that each of these copies can be: e2, e3 and e4. Some people have two copies of the e4 variant, and some people have one copy. Others have none. For those with one or two copies, their risk of developing late-onset AD, which is about 95% of AD cases, is significantly higher than the non-e4 carrying people. APOE e4 carriers also have an earlier disease age of onset, and the disease progresses faster. At least one copy of this variant is found in 60% of AD cases, with one e4 variant conferring a threefold increased risk and two e4 variants conferring a twelvefold increased risk of developing the disease. Conversely, APOE e2 protects against the disease.

To be able to systematically study the biology of these variants, ADAPTED reprogrammed and edited the genomes of cells taken from AD patients as well as healthy controls and created a set of stem cell lines that express the different variations, called genotypes. The stem cells are a type called induced pluripotent stem cells (iPSCs), which means they are derived from cells that can give rise to a number of different cells in the body, and they are pluripotent, meaning they can regenerate indefinitely, allowing other researchers to use them in the lab to study the biology of the APOE gene.

Three sets of iPSC lines carrying one of the following APOE genotypes were generated: APOE-e2/e2, APOE-e3/e3, APOE-e4/e4, APOE-e3/e4, as well as APOE-KO (knock out, meaning the DNA was edited to erase the gene’s expression permanently). The cell lines are a unique resource and the project has made them available to the scientific community via the European Collection of Authenticated Cell Cultures (ECACC).

A trove of data

The ADAPTED researchers used the stem cells to obtain cells representing important cell types for APOE biology - neurons, astrocytes, and cells of the monocyte macrophage lineage and microglial cells. They then integrated and compared results obtained from analyses of these cells with data obtained from analysis of biological samples from the biobank of the project. The project partners carried out a number of investigations using the APOE models, and published their findings in high-impact journals.

Among their findings, they discovered that a rare coding variant in the PLCG2 gene slows AD progression and maintains cognitive function, apparently counteracting the negative effects of the APOE-e4 variant. This highlights the PLCG2 enzyme as a potential target for interventions. They also found that levels of the proteins CDH6 and HAGH are significantly higher in AD patients who also carry the APOE-e4 gene compared to healthy controls, which suggests that CDH6 and HAGH could be used as new blood-based biomarkers for the detection of pre-symptomatic AD. They also showed an unexpected potential genetic link between echocardiographic (ECG) measures in healthy populations and cognitive decline in later life, which could prove important in designing preventative strategies to combat AD.

Another study resulted in the development of a novel model of the human blood–brain barrier (BBB) that can be used to measure the passage of large molecules, such as therapeutic antibodies, across the BBB for drug screening purposes. The in vitro model, developed by SME partner Mimetas, shows sufficient barrier function to study the passage of large molecules and is sensitive to differences in antibody penetration, which could help in the discovery and engineering of new technologies that can overcome limitations inherent in getting molecules into the brain. 

Data for the research community

ADAPTED generated a large quantity of valuable data, having combined and analysed information from publicly available multi-omics studies and genome-wide association studies, from thousands of AD cases and controls, with the ADAPTED isogenic iPSCs. The data, together with the samples described below, are available for the use of the scientific community through European repositories.

Biobank in Barcelona

The research centre at Fundació ACE research centre in Barcelona now manages an extensive and high-quality biobank of samples taken during the lifetime of the project with the patients’ permission. Cerebrospinal fluid samples were collected, stored and frozen, to be analysed by other researchers upon request. The ADAPTED partners also gathered blood serum and saliva collected on the same day as the CSF samples, which they measured using state-of-the-art analytical techniques. There are samples from thousands of donors and the number is still growing. The samples will continue to generate data as the sample size increases, constantly contributing new insights. It is the biggest single-site biobank of its kind in the world, and the only one that is fully open to researchers with an approved protocol.