BACE 1

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BACE 1 Alzheimers Disease

• By Anisha Vora Lindsey Wood

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What is Alzheimers?

• Neurodegenerative disorder characterized by the
  progressive and irreversible loss of nerve cells
  (neurons) located in the specific brain areas
  the hippocampus and polymodal association areas
• 4th leading cause of death in adults (cases rise
  with aging)
• Cortical brain areas heavily filled with two
  different types of lesions amyloid plaques and
  neurofibriallary tangles
• Amyloid plaques spherical extracellular amyloid
  deposits
• Neurofibrillary tangles an intra-neuronal
  accumulation of pathological fibrils called
  Paired Helical Filaments (PHF) basis of these
  tangles is the tau protein

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Why this disease?

• Prevalence is increasing
• United States and Congress has classified the
  treatment of this disease as a major national
  priority
• 100 billion per year is spent on Alzheimer's
  disease in the United States, making it the third
  most costly disease after heart disease and
  cancer
• 7 million Americans may be afflicted by the year
  2010

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Plaques Tangles

• Disrupt all three processes
• Communication (sending messages)
• Metabolism (turning chemicals and nutrients into
  energy to keep neurons working)
• Repair (keeping long-lived neurons in good
  working order)
• Cause nerve cells to stop working, lose
  connections w/ other cells and eventually die
• Destruction causes memory failure, personality
  changes, etc.
• Find abundance of ß-amyloid plaques and
  neurofibrillary tangles, especially in regions
  for memory

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The Changing Brain

• PET scan measures blood and glucose metabolism
  giving map of active brain
• Damage by free radicals increase
• Shrinking of neurons
• Increased levels of tangles plaques develop in
  AD

Normal Brain
Alzheimers Brain
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Neurofibrillary Tangles

• Insoluble twisted fibers found inside of brains
  cells
• Primarily tau, which forms part of structure
  called a microtubule
• Microtubule helps transport nutrients and other
  important substance from one part of nerve cell
  to another
• In AD, tau is abnormal and microtubule structures
  collapse

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Neurofibrillary Tangles
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ß-Amyloid Plaques

• Dense, insoluble deposits made of 42 residue
  ß-amyloid peptides
• Arranged in an anti-parallel arrangement
• Fragments clump together and are mixed w/
  molecules, neurons and non-nerve cells
• Develop in hippocampus (deep in brain, encode
  memories) and later in other areas of cerebral
  cortex (those involved thinking and making
  decisions)

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Amyloid Precursor Protein
APP is associated with the cell membrane, the
thin barrier that encloses the cell. After it is
made, APP sticks through the neuron's membrane,
partly inside and partly outside the cell.
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From APP to ß-amyloid
Enzymes act on the APP and cut it into fragments
of protein, one of which is called beta-amyloid.
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From APP to ß-amyloid
The beta amyloid fragments begin coming together
into clumps outside the cell, then join other
molecules and non-nerve cells to form insoluable
plaques.
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Overall Mechanism of AD formation
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BACE 1

• ß-Secretase (BACE-1 for ß-site APP-cleaving
  enzyme) is a type 1 transmembrane protein
  containing aspartyl protease activity
• 501 amino acids
• Mediates the primary amyloidogenic cleavage of
  APP
• Generates a membrane-bound APP C-terminal
  fragment-immediate precursor for the
  intramembraneous ?-secretase cleavage.
• BACE-1 is the only protease with a well-defined
  ß-secretase activity shown by the homozygous
  knockout of the BACE-1 gene, which does not allow
  any Aß generation
• (Cai et al, 2001 Luo et al, 2001).

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APP Cleavage Sites
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3 proteases

• Three proteases, alpha-, ß- and gamma-secretases,
  are involved in APP processing.
• At the cell surface, APP undergoes proteolysis by
  alpha-secretase releasing a large, soluble
  ectodomain (a-APP sAPP). The C-terminal fragment
  is retained within the cell membrane.
• This fragment can then be cleaved by
  gamma-secretase releasing the p3 peptide.
• In an alternate pathway, ß-secretase cleaves APP
  releasing a large secreted derivative sAPPß and a
  C-terminal fragment CTFß that can be further
  cleaved by gamma-secretase to form Aß which is
  released into the extracellular region.

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Cleavage Diagram
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Structure of BACE 1 complexed with inhibitor
(Hong et al, 2000),

• Resolution found to 1.9 Angstroms
• 4 domains
• Lumenal domain, transmembrane domain, cytosol
  c-terminal tail
• Active site is more open and less hydrophobic
  than other human proteases

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Function of BACE 1

• Responsible for the proteolytic processing of the
  amyloid precursor protein (APP).
• Cleaves at the N-terminus of the A-beta peptide
  sequence, between residues 671 and 672 of APP,
• Leads to the generation and extracellular release
  of beta-cleaved soluble APP, and a corresponding
  cell-associated C-terminal fragment which is
  later released by gamma-secretase.
• BACE-1 expression is significantly enhanced in
  brains from patients with AD
• Reasons for the enhanced expression are currently
  unclear. Speculated that AD-associated stressors
  such as oxidative stress, radicals, unfolded
  proteins, head trauma, and others may induce
  BACE-1 transcription and/or expression/activity
  during aging

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Cleavage of APP, APLP1, and APLP2 by BACE 1
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Drug Targeting for AD

• BACE-1 is an obvious target for Aß-lowering
  drugs.
• In contrast to ?-secretase, BACE-1 can be fully
  knocked out in mice without causing any bad
  phenotypes. Interfering with BACE-1 is unlikely
  to result in unwanted side effects.
• The three-dimensional structure of BACE-1 has
  been solved and peptidomimetic and
  nonpeptidomimetic inhibitors have already been
  generated (Hong et al, 2000 Vassar, 2001).
• Developed further to improve their affinity with
  the rather large active site cleft of BACE-1
  before they can be used in human trials.

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Inhibitors of BACE 1 invitro
Table 1. In vitro values Cp P3 P1' X BACE FRET
HEK293 Aß inhibition at P3 IC50(uM)a
IC50(uM)a 1 CHMe2 Me N 0.082 (0.023)
2.831 (0.129) 2 H H 9.704 (1.413)
gt100 3 Me H 0.285 (0.016)
gt100 4 Me N 4.925 (0.169)
gt100 5 Me H 2.397 (0.089)
gt100 6 CHMe2 Me N 0.260(0.002) 5.305
(0.191) 7 (S)-MeCHEt(Ile) MeN 0.065
(0.039) 0.880 (0.042) a Values are means
of three experiments, standard deviation is given
in parentheses.
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BACE-1 Inhibitors

• Macrocycles derived from a hydroxyethylene core
  structure
• Molecule inhibits production of Aß peptide in
  HEK293 cells overexpressing APP751sw
• Improve potency, Val residue at the P3 position
  replaced with Ile (compound 7)

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BACE-1 Inhibitory Complex

• Compound 7 co-crystallized with BACE-1
• X-ray structure of complex elucidated at 1.6Å
  resolution to an R-factor of 0.211 using data
  from the Advanced Photon Source

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Future Exploration with BACE 1

• Try to modify inhibitors to have a higher
  affinity for BACE 1 active site
• Decrease formation of AB42 and aggregate creation
• Further cleavage of AB42 to eliminate plaques
• Breakdown of beta amyloid plaques formed in the
  brain

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Questions?