Combating Alzheimer’s Disease with CRISPR: A Step Forward Towards New Therapeutics

Alzheimer’s disease (AD) is a progressive neurodegenerative disorder, with around 55 million people suffering from it or related dementia worldwide. It is the most common type of dementia, primarily affecting older individuals over the age of 65 years. Alzheimer’s is characterized by memory loss, cognitive decline, behavioral changes and can seriously impact an individual’s ability to perform daily activities at its peak. With no current cure, treatments usually focus on symptom management. Recent advancements such as anti-amyloid drugs like aducanumab, target amyloid plaques in the brain, offering hope for slowing disease progression. Ongoing research is exploring promising approaches, including CRISPR gene editing, tau-targeted therapies, and combination treatments, with the goal of developing more effective treatments for this complex disease.

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Alzheimer’s Disease: Overview

Discovered in 1906 and named after Dr. Alois Alzheimer, Alzheimer disease (AD) is an irreversible and progressive brain disease beginning with mild memory loss which later on causes loss of orientation and other cognitive impairments. It primarily inflicts the part of the brain that is important for memory, thought and language.

While not very common in younger individuals, people over 60 are more prone to getting affected by Alzheimer’s. Changes in the brain can begin years before the first symptoms appear and over time account for accumulation of the protein beta-amyloid (outside neurons) and twisted strands of the protein tau (inside neurons). They are accompanied by the death of neurons and damage to brain tissue. Other accompanying symptoms of Alzheimer’s include Inflammation and atrophy of brain tissue.

Even though there has been ongoing research to study the disease, the scientists haven’t been able to find the cure yet. The present treatments are helpful in slowing the dementia symptoms but none can stop the disease progression. It is estimated that by 2060, the number of patients suffering from Alzheimer may triple in the USA alone if we are not able to find effective treatment soon.

Alzheimer’s symptoms, progression & stages

Alzheimer’s disease develops gradually and affects various brain functions over several years. It is a progressive disease impacting memory, thinking, judgment, language, problem-solving, personality, and movement of the affected individuals. This is also called the Alzheimer’s disease continuum. The disease progresses through five stages: preclinical Alzheimer’s, mild cognitive impairment, mild dementia, moderate dementia, and severe dementia (Alzheimer’s Association). These stages help understand the progression, but each person's experience is unique. Dementia, in general, refers to symptoms affecting intellectual and social abilities enough to interfere with daily life.

The progression rate of Alzheimer’s disease varies widely, with an average life expectancy of three to eleven years after diagnosis, though some individuals may live 20 years or more. The degree of impairment at diagnosis and untreated vascular risk factors like hypertension can influence the progression speed. Common causes of death in Alzheimer’s patients include pneumonia (due to impaired swallowing as well as dehydration), malnutrition, falls, and other infections (Alzheimer’s Association).

Alzheimer’s manifestation: common causes

While its exact cause remains unknown, many risk factors contribute to the likelihood of developing the disorder. It is believed that it could be a combination of genetic, lifestyle, and environmental factors (“What Causes Alzheimer’s Disease?”).

Genetics influence the likelihood of developing Alzheimer’s, though no definitive genetic cause is known till date. It is believed that multiple genes in combination with other factors such as lifestyle and environmental factors can play a role. It is possible that a person may carry more than one genetic variant or group of variants that can either increase or reduce the risk of Alzheimer’s. Also it is not always true that people who develop Alzheimer always have a family history of the disease, whereas having a parent or sibling diagnosed with the disease can lead to a higher risk of Alzheimer’s.

Several gene targets are associated with Alzheimer’s after years of research. Gaining a better understanding of their role in the disease onset and progression is the key to finding the cure. One such target that is well studied in the field is the apolipoprotein E (APOE) gene (Raulin and Doss 2022).The APOE gene is involved in making a protein that helps carry cholesterol and other types of fat in the bloodstream. Problems in this process are thought to contribute to the development of Alzheimer’s.

Other than APOE gene, three rare single-gene variants are known to cause the disease:

• Amyloid precursor protein (APP) on chromosome 21
• Presenilin 1 (PSEN1) on chromosome 14
• Presenilin 2 (PSEN2) on chromosome 1

Changes in these three genes result in the production of abnormal proteins that are associated with the disease (Cacace et al. 2016).

Another genetic correlation that is known to cause Alzheimer’s disease is presence of an extra copy of chromosome 21 in people with Down syndrome, which carries the APP gene, and a higher risk of developing early-onset Alzheimer’s. It is estimated that 50% or more of people living with Down syndrome will develop Alzheimer’s with symptoms appearing in their 50s and 60s.

Scientists looking at Alzheimer’s disease in humans have observed that certain genetic variations in brain cells called microglia show a strong correlation with their risk of getting AD. One of the strongest associations is between AD and a microglia associated gene called TREM2 (Huang et al. 2023) .

Alzheimer’s is broadly defined as either late-onset or early-onset, affected by different genetic components. Late-onset, typically affecting those in their mid-60s and older, is linked to the APOE, Clusterin (CLU), and Bridging Instigator 1 (BIN1) genes, possible risk factors but not a definitive cause. Early-onset, affecting fewer than 10% of cases, occurs between the mid-30s and mid-60s and is often associated with mutations in the APP, PSEN1, and PSEN2 genes.

Whatever the original trigger(s) may have been, the symptoms of Alzheimer’s develop because of beta-amyloid plaques and tau tangles in brain neurons. Beta-amyloid, a fragment of the amyloid precursor protein (APP), clumps together into plaques that disrupt cell communication and trigger inflammatory responses, leading to neuron death. Tau proteins, which normally support cellular transport, become misfolded and form tau tangles in Alzheimer’s. This disruption further contributes to brain cell death. The exact functions of APP and the reasons for these protein malfunctions remain unclear. Each of these mutations contributes to the breakdown of APP, a protein whose function isn’t completely understood. The breakdown of APP is part of a process that makes harmful forms of sticky amyloid fragments. These fragments cluster to form plaques in the brain, which is a hallmark of Alzheimer’s.

Alzheimer’s Disease Treatments & Cure

Alzheimer treatment primarily focuses on symptom management rather than treatment or cure. Several prescription drugs are approved by the U.S. Food and Drug Administration (FDA) for Alzheimer’s disease that work best for people in the early or middle stages of Alzheimer’s. Even after a century of disease discovery, disease-modifying drug treatments (DMTs) are still not available. The increasing life expectancy in today’s world is leading to increased prevalence of Alzheimer’s disease and other types of dementia. This highlights the urgent need for alternate strategies that can give a new hope for AD treatment and cure.

Pharmacological therapy for Alzheimer’s symptom management

Even though several pharmacological drugs exist to alleviate symptoms associated with Alzheimer’s disease, they don't cure the disease. Most of these approved drugs such as Galantamine, rivastigmine, and donepezil are cholinesterase inhibitors that are prescribed for mild to moderate Alzheimer’s and may help reduce or control some cognitive and behavioral symptoms. Similarly, Memantine, which is a NMDA receptor antagonist works by blocking the effects of an excessive amount of glutamate in the brain, which can kill neurons and lead to memory loss. Memantine is used for moderate or severe Alzheimer’s disease. It's suitable for those who cannot take or are unable to tolerate AChE inhibitors.

According to a recent review highlighting data from clinicaltrials.gov, in 2024, there are 164 clinical trials assessing 127 drugs in the Alzheimer’s disease drug development pipeline. In comparison to 2023, there are fewer trials, fewer drugs, and fewer new chemical entities being tested. The range of drugs being targeted for the AD drug development pipeline include neurotransmitter receptors, inflammation, amyloid, and synaptic plasticity to name a few.

Immunotherapies

Recently, Immunotherapies targeting AD have made significant progress after years of research setbacks. The approval of aducanumab in 2021 ended a 17-year drought in new AD treatments. This was followed by the accelerated and then standard approval of lecanemab in 2023, based on evidence linking amyloid plaque reduction to slower cognitive decline. The latest to join the list is Donanemab, approved in July 2024 as an injection for the treatment of Alzheimer’s in the patients with mild cognitive or mild dementia stage of disease. In 2023, brexpiprazole became the first approved treatment for agitation in AD-associated dementia.

While this is promising, and there have been recent advancements in the drug development pipeline targeting amyloid plaques in the brain, there are several doubts and concerns regarding their efficacy in improving cognitive functions and restoring mental health of the patients. This leads to constant need for exploring new therapies for this disease.

Stem Cell therapy

One of the upcoming therapies with immense potential for success in the treatment of neurodegenerative diseases is the use of stem cells in tissue regeneration. Stem cell therapies can replace diseased cells with healthy ones, addressing the root of the disease. Some Alzheimer’s studies have shown that stem cell therapies can:

1. Regenerate neurons i.e. transplanted neural stem cells (NSCs) can regenerate damaged neurons, and NSC-derived neurons can form connections with nearby neurons (Chen et al. 2023).
2. Improve cognitive function as seen in studies using mesenchymal stem cells (MSCs) where they can improve cognitive function in mouse models and a subset of people with AD (Hernández and García 2021).
3. Reduce inflammation as a part of MSC replacement treatment with reduction of beta-amyloid (Aβ) levels, in AD-transgenic mice (Yokokawa,, Iwahara, and Shimohama 2019)
4. Protect against memory loss (Mishra, n.d., 2019)

Additionally, there is extensive ongoing research using induced Pluripotent stem cells (iPSCs) for the treatment of AD and related neurological disorders as they retain the reprogramming capabilities that allows them to be differentiated into different cell types .

The biggest challenge associated with bringing these experimental therapies into actual clinical practices is the need for pre-differentiation of these stem cells in vitro into various neuroblasts.

Gene Therapy

Given the success of gene therapy for several diseases (eg cystic fibrosis, alpha-1 antitrypsin deficiency, hemophilia), it is only fair to consider it as one the strongest contender as the treatment course for AD. Gene therapy for Alzheimer’s disease aims to deliver modified genes to help correct the altered protein activity. Specifically, gene therapy can help slow progression, reduce symptoms and delay onset the Alzheimer’s if not present a sure shot cure for now. For example gene therapy directly targeting CD33 in an attempt to reduce neuroinflammation and Aβ pathology in a mouse model of AD (). Researchers are also manipulating the APOE variants for finding potential therapies. There is an interest in negating the risk factor APOE4 by administering virus-vector mediated delivery of APOE2 to the brain. Work is also ongoing on another potential target, PGC-1α, which is implicated in improved mitochondrial dysfunction and oxidative stress in AD.

This is really promising path but continued research is crucial for refining gene therapy approaches and improving their effectiveness in managing Alzheimer’s symptoms and slowing disease progression.

Utilizing CRISPR for Alzheimer’s Research

Since the traditionally used AD treatments only treat the symptoms and not the disease itself there has been a need to look for alternate strategies. With the advent of gene editing techniques and rising impact of CRISPR in diseases research such as sickle cell anemia and β-thalassaemia, is giving new hopes to the researchers. At the 2023 Alzheimer’s Association International Conference (AAIC) in Amsterdam, researchers introduced two groundbreaking CRISPR-based therapies for Alzheimer’s disease. One approach is designed to counteract the effects of the APOE-e4 gene, a major genetic risk factor for Alzheimer’s. The other strategy aims to decrease the production of beta-amyloid, a protein linked to the disease's progression. These advancements offer promising new avenues for treatment and renewed hope for those impacted by Alzheimer’s. Some other CRISPR initiatives that are driving the AD disease research forward are worth mentioning and are listed below:

Disease modeling of Alzheimer’s using CRISPR

Till date numerous animal models have been created to study AD pathology and disease progression, but few have been seen as fully replicating the key pathological features of AD. Out of the known genetic targets, Aβ-based mouse and rats models are very popular even though they are unable to capture all possible AD lead pathology and cell death. Recent development of a chimeric The novel APP^hu/hu is an animal model that was generated using the CRISPR-Cas9 tool to produce a humanized Aβ sequence by generating F681Y, G676R, and R684H mutations in the rodent APP gene. By introducing a single chimeric App gene through a knock-in strategy, scientists successfully developed a rat model that replicates multiple Alzheimer’s disease (AD) pathologies, mirroring many of the abnormalities observed in the human AD brain thus creating a tool for studying the interactions between different disease mechanisms. This model also enables direct comparisons of various therapeutic molecules targeting different pathways, helping to prioritize candidates for clinical testing.

Several cell lines are also routinely used in AD research including human neuroblastoma cells SH-SY5Y and SK-N-SH, mouse hippocampal neuron cell lines HT22 and glial cells BV2, and mouse glioblastoma cells N2a. Also utilized commonly are the iPSCs that are getting the increasing attention as a great cell model. Specifically, patient-derived iPSCs have unique physiological profiles and can act as great tools to understand the relationships of genetic targets and their roles in the disease pathway by allowing the easy manipulations and downstream functional analysis readouts.

A great example on how these tools can be used for AD research is the NIH led project. Approximately 50 genes are linked to Alzheimer’s disease and related dementias (ADRD), but their interactions remain poorly understood. To tackle this challenge, the NIH launched the Inducible Pluripotent Stem Cell Neurodegeneration Initiative (iNDI). This project aims to create a standardized collection of isogenic iPSC lines, each carrying over 100 mutations associated with ADRD. Using CRISPR technology, iNDI has engineered multiple single nucleotide variants (SNVs) per gene across various targets in induced pluripotent stem cells.

Identification of new targets and biomarkers using CRISPR

CRISPR-Cas9 in vitro and in vivo models allow the identification of new therapeutic targets. Reliable biomarkers are crucial for any condition, as they provide measurable indicators of a disease's presence, progression, or risk and their levels can act as a measure of therapeutic effectiveness. For AD, utilizing the CRISPR screening techniques, researchers were able to identify calcium, and integrin-binding protein 1 (CIB1) as the new protein target playing a role in causing AD pathology. STIM (Stromal Interaction Molecule 1- an endoplasmic reticulum protein) was identified as another key player when its expression was found to be downregulated in the brain tissue of AD patients. Furthermore, using CRISPR based knock out of STIM-1 gene in SH-SY5Y cells, they were able to establish its role in neurodegeneration, discovering that eliminating STIM1 expression led to reduced calcium ion transport through the plasma membrane, ultimately resulting in cell death. Identification of biomarkers like this gives hope in creating precise disease models that can mirror brain chemistry of the majority of AD patients.

Other CRISPR based possibilities for Alzheimer research

Extensive research has been conducted on Alzheimer’s disease, focusing on identifying its causes and potential treatments or cures. Numerous genetic variants and pharmacological agents have been examined for their roles in disease onset, progression, and pathology. Researchers are employing high-throughput screens to investigate interactions between known targets and to identify potential drug candidates. A lot of research is published trying to study the co-occurrences in these genetic risk factors and target proteins and enzymes which is very important for tailored treatments to the relevant pharmacological pathway, drug development, developing preventive care in absence of symptoms and creating more effective therapies and treatment strategies.

The advances in CRISPR technology along with high throughput genome wide sequencing, is going to play a major role in Alzheimer research and cure. Some recent studies conducted using CRISPR/Cas9 for the screening of Sporadic AD pathogenic genes have identified TREM2 playing role pathogenesis of AD as the knock-out of TREM2 in iPSCs impacted the survival of microglia, the clearance of APOE and SDF-1α/CXCR4-mediated chemotaxis was seriously impacted and led to impaired response to beta-amyloid plaques. Further mutations in TREM2 (R47H) have been correlated with increased AD risk. Similarly, introducing a homozygous mutation (rs377155188, C > G, p.S1038C) in TTC3 may increase the risk of AD. For the known AD targets such PSEN2, mutation correction led to normalization of cells’ electrophysiological function and Aβ secretion in iPSC derived neurons. Genetic variant of APP gene A673T was found to reduce the AD biomarker beta-amyloid and likelihood of developing Alzheimer by a factor of 4. Other protective mutation in the 3’UTR of the APP gene is mice has been identified using CRISPR that can protect against cognitive decline and amyloid plaque accumulation. While there is continuous research needed till we reach the optimum results, all these targets being studied provide a platform to explore AD biology and potential drug development possibilities.

Future of CRISPR in Alzheimer’s Research

CRISPR-Cas9 mediated gene editing to identify novel mechanistic insights into disease pathogenesis and to mediate accurate gene therapy is routinely leveraged by researchers in different fields. There are several publications highlighting the experiments aimed at generating novel dementia models to showcase proof-of-concept studies in preclinical animal models. While the benefits are clear and many, some hurdles do prevent CRISPR from being the frontrunner of the therapeutic bandwagon. Off-target concerns, mode of delivery, biocompatibility, safety and tissue specificity of CRISPR components still remain points of concern. There is definitely hope owing to the successful clinical trials using CRISPR based therapies in treating certain cancers and blood disorders, it’s only a matter of time when CRISPR based clinical trials would get approval for brain disorders such as Alzheimer’s.

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