Alzheimer's Flashcards
What is the amyloid cascade hypothesis?
The original hypothesis was the result of genetic evidence relating to “early onset” forms of Alzheimer’s disease. Mutations in the amyloid precursor protein (APP) and the presenilins (PS1 and PS2 - form part of the protease complexes responsible for forming Abeta peptides) genes lead to accumulation of Abeta42 peptides Soluble oligomers are toxic via wide ranging impacts on protein function and membrane integrity, leading to disrupted ionic homeostasis (calcium levels) Tau disruption and tangles may be central to cell death pathways.
What is the evidence for the amyloid cascade hypothesis?
The observation of amyloid plaques and tangles in patients with “AD” (Alois Alzheimer 1880s). The identification of the protein within the plaques (Glenner, 1980s) and tangles. The observation of point mutations in APP and PS genes for early-onset AD cases (1990s). The observation of AD pathology in animal models with mutations in APP (requires human Tau) (1990/2010s). Observation of Abeta (amyloid precursor) amyloid causing tangles in cell culture (Takashima et al, 1993). Toxicity of protein only samples in both animal and cell culture models. In order for the model to be expanded to sporadic disease, there needs to be a way that an imbalance in Abeta or Tau can happen spontaneously.
Could be correlation not causation - could be a downstream effect of something else. Is there evidence for causation in that it is only these become mutated?
Do amyloids form simultaneously?
All proteins seem to be able to form amyloid in vitro (given the correct conditions). Proteins are close to their solubility limit in vivo. Propensity to aggregate is correlated with age of onset (most obvious examples from other diseases such as Huntington’s).
Can the environment favour amyloid formation?
Amyloid formation is a “nucleated” process. It occurs by “heterogeneous nucleation” (surfaces will catalyse the formation of nuclei which then grow to form toxic assemblies then amyloid).
What is the biggest factor affecting amyloid formation?
The biggest factor affecting the process is concentration of the protein as well as changes in the environment. Ageing, obesity and smoking all cause fluctuations in proteostasis / lipid levels etc.
What does infection lead to?
An inflammatory response which induces processing of APP to Abeta peptide. Abeta acts as an “AMP” (antimicrobial peptide) and induces break-up of bacteria – DNA and membrane fragments lead to aggregation of Abeta (Soscia et al (2010). Imbalance in Abeta concentration leads to toxicity and inflammatory response (Su et al, 2016). This leads to breakdown in blood brain barrier and permeation to infection. Infection then exacerbates things.
How is Abeta toxicity enhanced?
Toxicity of Abeta enhanced by higher cholesterol: phospholipid levels. Abeta causes membrane damage leading to neuronal dysfunction. Processing of Abeta enhanced by high cholesterol:phospholipid levels. ApoE is the main genetic risk factor for late-onset Alzheimer’s and is a major cholesterol carrier in the brain. Cholesterol may compete with Aβ peptide for its carrier – e.g. 1. ApoE, 2. serum albumin (HSA) in the cerebral spinal fluid and blood, 3. membrane transporters for removal of Abeta to the blood (via the BBB: Blood Brain Barrier).
What are positives of immunisation against Abeta?
Immunisation against Aβ: The first demonstration that removal of amyloid leads to recovery in an animal model (Schenk et al 1999) Passive immunisation using an antibody based drug: the example of aducanumab (Sevigny et al 2016)
What are negatives of immunisation for Abeta?
Immunisation in humans led to death in some patients due to encephalitis (inflammation of the brain) (Gilman et al (2005). Efficacy questioned for passive immunisation despite evidence for amyloid clearance.
Why is amyloid clearance not curing Alzheimers?
- The Amyloid Cascade Hypothesis is wrong (see previous discussion).
- The “domino effect”: we need to treat downstream problems too (inflammation, Tau) – see an example in Live session 2 (UPR targeting).
- We need to target the toxic forms of Abeta better (the “right” oligomers).
- The problem of “co-morbidities”… Are anti-amyloids too specific?
What are the three features identified in Abeta fibrilisation (Linse et al, 2020)?
- Primary nucleation: kn formation of a small assembly, beyond which addition of further monomers is favourable
- Elongation: k+ Growth of the fibril by monomer addition
- Secondary nucleation: k2 an autocatalytic feedback mechanism that accelerates assembly. Nucleation is catalysed on the surface of existing fibres. This is calculated to be the largest contributor to small (presumably toxic) oligomer formation.
How are drugs screened for Alzheimer’s?
Tunicamycin is added to induce UPR in a C. elegans developmental assay. This stalls larval development due to impact on neurogenesis. Screen: >1,000 compounds added to C. elegans developmental assays (microplates) and 20 identified to inhibit this “stalling” of neurogenesis.
20 compounds added to UPR cell culture assay where “CHOP” promoter is engineered upstream of a luciferase reporter gene. Transcription of CHOP is part of PERK downstream response, here induced by tunicamycin (Tm). Luciferase catalyses the formation of a bioluminescent product which can readily be quantified. Only compounds with a partial effect were selected (GSK26… is too strong and causes pancreatic failure).
Animal work showed that administration of trazodone and DBM prevents neurodegeneration in mice with prion disease as well as frontotemporal dementia (Tau aggregates).
Treated animals maintain a very high load of amyloid, which is now no longer causing toxic downstream responses.
How are anti amyloids specific?
The anti-amyloids in the last section are raised against a specific protein target (e.g. Abeta peptide and its assemblies). Many patients will have Tau and even α-synuclein pathology as well as Abeta plaques. Identification of a “general amyloid interaction motif (GAIM)”, able to “remodel” all amyloids, was discovered on the surface of a bacteriophage protein – M13. This has shown promise in animal models and is currently in clinical trials.
Phage display used to scan peptide libraries for amyloid binding candidates: Control phage highest affinity!
What is the M13 phage?
M13 Phage binds fibrillar amyloids including fAβ with nM affinity. M13 remodels amyloids of a range of different proteins. After 3 days, remodelled fibrils can be washed through a 200nm cellulose acetate membrane filter with PBS.
M13 “G3P” (Gene 3 protein) is key to amyloid binding and remodelling. Sedimentation assays with fAβ show the nature of the capsid component of M13 responsible for binding.
What are positive contributions of UPR?
- UPR is central to regulation of (MAP) kinase pathways, inflammatory responses, apoptosis, autophagy and innate immunity (complex mechanisms).
