8.1 Alzheimer’s Disease

8.1.1b-Amyloid and Tau

The pathological hallmarks of AD are represented by extracellular senile plaques and intracellular neurofibrillary tangles, which are always present in the brains of AD patients. Both plaques and tangles are the result of aberrant protein accumulation and aggregation, respectively of amyloid b protein (Ab) and of hyperphosphorylated tau protein.4 In addition, a third distinctive feature of the AD brain is a chronic inflammatory status.

Microtubule-associated protein tau is involved in the regulation of microtubule assembly and disassembly, and its activation is dependent on phosphorylation at di erent sites. In AD, tau is in a hyperphosphorylated state that increases its potential to aggregate into filaments that accumulate inside the cell, disrupting the cytoskeleton.5 In addition, the protein undergoes multiple truncations that influence its conformation and ability to polymerize.6 Although much debated, tauopathy is considered to be secondary to Ab dysregulation. In support of this, it is not strictly related to AD but it is common to other forms of dementia.5

Classically, the amyloid cascade hypothesis is the more widely accepted mechanism proposed for AD pathogenesis.7,8 Ab is a short peptide (39–42 amino acids) derived from the proteolytic cleavage of the amyloid precursor protein (APP). APP is endowed with a single transmembrane domain and is cleaved at di erent sites by transmembrane proteolytic complexes known as secretases. Secretase activity yields peptides of di erent lengths and with different tendencies to aggregate. According to the amyloid theory, an imbalance leading to an over-production of the highly aggregation-prone Ab 42 species triggers its accumulation and aggregation first into low-molecular-weight oligomers, then into fibrils and finally into plaques, in specific brain regions. Plaques have long been considered the main cause of AD. Plaques contain Ab as well as degenerating neurites, and evoke strong local inflammatory responses. Activated microglia and astrocytes are attracted to the area around the plaque and release a number of cytokines, chemokines and complement components. Even though microglia and astrocytes are involved in Ab removal mechanisms, the real significance of inflammation around plaques still needs clarification, since it is not yet evident if it indeed represents a protective mechanism against neurodegeneration or rather it is responsible for increased neuronal damage in the areas surrounding the plaques.9,10 In a more recent interpretation of the amyloid hypothesis, based also on the observation that plaque load does not e ectively correlate with cognitive decline, oligomers have been recognized possibly to represent the driving cause of the disease. Recent findings show oligomers to be highly synaptotoxic species able to cause neuronal synaptic dysfunction and degeneration.7,11,12

APP processing implies the alternative activation of three identified secretase complexes, a, b and g as reviewed recently7,13 g-Secretase includes at least five di erent proteins, Presenilin (PS) 1 and 2, APH1, PEN-2 and Nicastrin, and performs a C-terminal cut inside the lipid bilayer of the plasma membrane. Such C-terminal cleavage is coupled to cleavage in the N-terminal ectodomain,

alternatively performed by a or b secretases. a-Secretase activity has been attributed to metalloproteases belonging to the family of A Dysintegrin and Metalloproteinase (ADAM). Three main ADAMs have been assigned a role in APP processing, ADAM10, 9 and 17, with ADAM10 apparently being the main player. a-Secretase activity is considered to be favorable in pathologic conditions, for its site of cleavage is located inside the Ab 42 sequence, thereby precluding the formation of the amyloidogenic isoforms. Release of the Ab peptides depends instead on the coupling of g-secretase with b-secretase activity. b-site APP-cleaving enzyme 1 (BACE1) is the main b-secretase, and its action is the rate-limiting step in Ab 42 formation. It is, however, the precise site of cleavage by g-secretase that determines the length of the amyloid species produced, which ranges from 39 to 42 amino acids, with Ab 40 and Ab 42 being the most represented. As mentioned, Ab 42 is the most insoluble species and has a greater tendency to aggregate compared to Ab 40.

Interestingly, all identified mutations (more than 200) linked to FAD are

located in the APP or PS genes and are responsible for the increased ratio of Ab 42 vs. Ab 40.7,14,15 Also, a variety of mouse models of AD have been obtained

by inserting into their genomes the human APP and PS1 genes carrying one or multiple mutations. Although these animals do not fully reproduce the human pathology, they are characterized by abnormal production of Ab 42, which aggregates in hippocampal and cortical plaques.16,17

Sporadic AD is associated with dysregulation of Ab synthesis, leading to accumulation of Ab 42. The e4 allele of the apoE gene is the prevalent risk factor for this form of the disease, although its exact role remains to be elucidated. ApoE is a lipid carrier protein that binds to amyloid peptides and influences transport, aggregation and metabolism of Ab.18,19 The apoEe4 isoform has been persistently associated with increased cerebral levels of Ab.19

Another factor influencing Ab synthesis is linked to tra cking of APP and secretases, all transmembrane proteins. Tra cking represents a crucial regulatory event that may a ect their location and interaction, and thus

the chance of Ab formation. In this regard, lipoprotein receptors have been shown to play a role in regulating APP endocytic tra cking.20,21

Recently, an important role has emerged also for intracellular accumulation of Ab, linked to development of synaptic pathology. Ab has been recently shown to be taken up and concentrated inside acidic intracellular compartments such as lysosomes, where it is cleared. In pathologic conditions, the load of intracellular Ab 42 likely becomes excessive. In these conditions, it begins to accumulate and aggregate inside the cell, eventually leading to neuritic

degeneration. This causes aggregated Ab to be released in the extracellular space, where it may in turn give rise to plaque formation.15,22,23

Although attention has been primarily focused on the biochemistry of Ab production, the importance of Ab clearance mechanisms has recently emerged. The brain is endowed with Ab-degrading systems mainly related to the activation of microglial and astroglial cells. In addition, recent evidence suggests the possibility that circulating monocytes could infiltrate the AD brain and contribute to Ab degradation with a distinct functional role.24 It has been shown

that astrocytes are in a reactive state in the areas surrounding plaques, where they release Ab-degrading enzymes.25–27 Several proteases have been identified with Ab-degrading action. The most e cient has been shown to be neprylisin,28 belonging to a family of zinc-metallopeptidase that includes at least two other enzymes endowed with Ab-degrading activity, endothelin-converting enzymes 1 and 2.29 Matrix metalloproteinases (MMP), mainly MMP2 and MMP9, have been demonstrated to possess the same ability.30 Notably, the insulin-degrading enzyme (IDE) is another primary enzyme responsible for Ab degradation,31 and accordingly insulin signaling has been shown to be involved in APP processing and Ab clearance. While hyperinsulinemia has been indicated as a risk factor for AD, levels of insulin in the central nervous system (CNS) are inversely related to the levels of Ab 42 accumulation.

8.1.2Target for AD Therapy

Current therapy for AD and related dementias includes only symptomatic treatments that delay progression of the disease during its early stages (Table 8.1). Based on pre-clinical studies and on the available knowledge of AD pathogenesis, alternative approaches should be used only at very early stages of the disease, even when no clinical evidence is present, in order to realize a

preventive strategy. According to current approaches, research is now trying to prove the e ectiveness of b- and g-secretase inhibitors32,33 or alternatively a-

secretase activators. This kind of intervention would a ect the processing of APP leading to Ab generation. It is also possible to modulate the activity of g- secretase, thus reducing the production of Ab 1–42 in favor of shorter, less toxic fragments.34 Other potential interventions are aimed at inhibiting plaque formation or favoring plaque removal. In this regard, major attention should be reserved to Ab passive immunotherapy that is at present being actively tested in humans and appears promising, despite some serious adverse e ects that have

Table 8.1 AD therapy.

Current treatments

Acetylcholinesterase inhibitors

Donepezil Rivastigmine Galantamine

NMDA antagonist

Memantine

Potential future pharmacological targets a-, b-, g-secretases

Tau kinase Tau clearance

Ab and tau aggregation Immunotherapy