A Immunotherapy: Lessons From Mice, Monkeys and Men

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Aß ImmunotherapyLessons From Mice, Monkeys and
Men

• Cynthia A. Lemere
• Center for Neurologic Diseases
• Brigham and Womens Hospital
• Harvard Medical School
• Boston, MA USA

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Silver Stained Plaques and Tangles
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Alzheimers Disease
A syndrome arising from a chronic imbalance
between Aß production and Aß clearance that leads
to gradual cerebral accumulation of Aß42. The
imbalance can be caused by numerous distinct
genetic (or environmental) alterations (e.g.,
mutations in APP, PS1 or PS2 lead to increased Aß
production and early onset FAD). Aß aggregates
toxic in vitro and cause impaired memory and
learning, gliosis and neuritic dystrophy in vivo
(APP, PSAPP tg mice)
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Several Therapeutic Approaches Based Upon the
Importance of ß-Amyloid in AD

• inhibit Aß-generating proteases (ß- or
  g-secretase)
• prevent the aggregation of Aß into oligomers
  and fibrils
• prevent the deposition of Aß into plaques
• enhance Aß clearance
• interfere with the toxic response of neurons to
  amyloid
• inhibit the inflammatory process around amyloid
  plaques

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Aß Vaccination Potential Therapy For
Alzheimers Disease
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Mouse Studies

• Active Aß immunization in APP and APP/PS1 Tg
  mice
• Generates anti-Aß titers
• Lowers cerebral Aß levels (if early enough)
  (Schenk et al., Nature 1999 Weiner, Lemere et
  al., Ann Neurol 2000 and many other
  publications)
• B cell epitopes within Aß amino-terminus, Aß1-15
• T cell epitopes within Aß16-42
• Increases peripheral levels of Aß in blood
• (Lemere et al, Neurobiol Dis 2003)
• Improves behavior/cognition on certain tasks
• (Janus et al., Nature 2000 Morgan et al., Nature
  2000 Dodart et al., Nature NS 2002, etc.)

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Passive Transfer of Aß Antibodies in Mice

• Results - direct application of Aß antibodies
  to brain surface clears local plaques (Bacskai
  et al., Nat Med 2001)
• - i.p. injections decreased brain Aß (Bard et
  al., Nat Med 2000) - increased Aß in blood
  (DeMattos et al., PNAS 2001)
  - improved behavior (Dodart et
  al, Nat Neurosci 2002 Kotilinek et al., J
  Neurosci 2002 Wilcock et al., J
  Neuroinflammation 2004)
• HC injections of 2 Aß Mabs in 3xAD-Tg mice
  cleared extracellular and intracellular Aß as
  well as early tau aggregates but not
  hyperphosphorylated tau (Oddo et al., Neuron
  2004)
• Intracerebral ventricular injections of Aß
  antibodies protected APP tg mice from synaptic
  loss and gliosis (Chuahan et al., J Neurosci
  Res, 2002)
• Passive transfer protected neurons from induced
  seizures in APP tg mice (Mohajeri et al., J
  Biol Chem 2002)
• Microhemorrhage observed in very old APP tg
  mice with CAA
• (Pfiefer et al., Science 2002 Wilcock et al., J
  Neuroinflamm 2004 Racke et al., J Neurosci
  2005 )

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Passive immunization in PDAPP Tg mice
Aß Pab by i.p. weekly for 6 months 93 reduction
Bard et al., Nature Medicine, 2000
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Aß Immunization of Caribbean Vervet Monkeys
(Chlorocebus atheiops)

• African Green Monkeys
• Imported to St. Kitts 1670
• Collaboration with BSF

• -- 8 s.c. injections 1 mg Aß40/42 CFA/IFA over
  10 months
• -- 5 vervets immunized, 5 controls (all between
  15 and 30 yr)
• -- B cell epitope in Aß
  N-terminus, reduced cerebral Aß load and
  gliosis, increased plasma Aß, no T or B cells in
  brain
• (Lemere et al., Am J Pathol 2004)

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Aß Immunization Reduced Cerebral Aß
Controls
Immunized
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Proposed Mechanisms

• Aß antibodies prevent Aß aggregation and/or
  dissolve Aß aggregates in vitro (Solomon
  et al., PNAS 1996, 1997)
• Fc-mediated microglial phagocytosis of Aß in
  brain (Bard et al., Nature Med 2000 )
• -- Fc-mediated response not required (Fab
  FcR-/- mice)
  
• (Bacskai et al., J Neurosci 2002
  Das et al., J Neurosci 2003)
• Peripheral Sink Hypothesis shift in gradient
  across the BBB such that there is an increase in
  efflux of Aß from brain to blood (DeMattos et
  al., PNAS 2001 Lemere et al., Neurobiol Dis,
  2003)

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ELAN/AHP/Wyeth Human Clinical Trials

• Trial halted (January 2002) 6 of
  Aß-immunized patients (18/300) developed
  meningoencephalitis anti-Aß antibodies detected
  in serum in subset (Schenk, Nature Rev 2002
  Orgogozo et al, Neurology 2003 Gilman et al.,
  Neurology 2005)
• Anti-Aß antibodies recognize AD plaques and CAA
  but not soluble Aß42 nor APP (Hock et al.,
  Nature Med 2002) epitopes against free
  N-terminus of Aß (Lee et al., Ann Neurol 2005)
• Some slowing of cognitive decline (Hock et al.,
  Neuron 2003 Gilman et al., Neurology 2005)
• Reduced Aß deposition in brain (Case Studies
  Nicoll et al., Nature Med 2003 Ferrer et al.,
  Brain Pathol 2004 Masliah et al., Neurology
  2005)
• Reduced tau levels in CSF (Gilman et al.,
  Neurology 2005)
• Cortical shrinkage by MRI in patients who
  generated Aß antibodies even though some clinical
  improvement observed (Fox et al., Neurology 2005)

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Reduced Plaque Burden in Immunized Pt
From Nicoll et al., Nature Medicine 2003
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Possible explanations for the adverse effects?

• The immunogen, Aß1-42, may have been recognized
  as self- antigen by some, leading to an
  autoimmune, toxic T cell response in the CNS
• The adjuvant, QS-21, purified saponin, is known
  to have
• prodominantly Th1/CD4 cellular immune
  effects and can induce CTL against antigens
  (Kenney et al., Vaccine 2002 Cribbs et al., Int
  Immunol 2003)
• Increased T cell response to Aß in aged humans
  (Monsonego et al., JCI 2003)
• Addition of polysorbate 80 to vaccine
  formulation
• (Nicholl et al., Nature Med 2003)
• Thus, development of an active Aß vaccine that
  would produce a strong humoral response but
  avoid an Aß-specific T cell response may be
  beneficial.

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Novel Immunogens to Short Aß PeptidesPublicatio
ns in Preparation(cannot show immunogens yet)
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Intranasal Aß ImmunizationOur previous work

• Intranasal (i.n.) Aß immunization without
  adjuvant resulted in a modest humoral immune
  response that lowered cerebral A? in PDAPP tg
  mice (Weiner et al., Ann Neurol, 2000 Lemere et
  al., NY Acad Sci, 2000).
• Adjuvant LT(R192G), a mutant form of E. coli heat
  labile toxin that is less toxic than its native
  form, enhanced the generation of Aß antibodies
  when administered intranasally with Aß peptides
  (Lemere et al., Neurobiol Aging, 2002).

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Immunization of J20 APP tg mice with 2 novel Aß
immunogens

• J20 APP-tg mice (hAPPSw, Ind)/PDGF-promoter on a
  mixed C57BL/6 x DBA background (L. Mucke, UCSF)
• 6 month-old mice received intranasal immunization
  weekly
• (n 7 mice/ group)
• Treatment Groups
• 100?g Aß Immunogen 1 5?g LT(R192G)
• 50 µg Aß Immunogen 2 5 µg LT(R192G)
• 5 µg LT(R192G)
• H2O
• Treated for 24 weeks at end, 5 mice per group
  except vehicle (6).

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New Aß Immunogens Summary

• Intranasal immunization using our novel short Aß
  immunogens led to a robust humoral response in
  both WT and J20 APP tg mice.
• Ig isotypes were mainly of the Th2 type B cell
  epitopes were located within the amino-terminus
  of Aß.
• Splenocytes proliferated against the immunizing
  peptides but not full-length Aß.
• Immunization resulted in lowering of plaque
  burden and plaque-associated pathology in APP tg
  mice.
• No toxicity or untoward affects were observed. B
  and T cells were absent from brain.

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Aß Immunotherapy Conclusions

• Active Aß vaccination prevents or reduces AD-like
  pathology and improves cognition but needs to be
  made safer.
• Generating a humoral immune response while
  avoiding an Aß-specific cellular response may
  improve safety.
• Passive transfer of Aß antibodies is also
  effective at lowering cerebral Aß and improving
  cognition but has been associated with
  microhemorrhage in the presence of abundant CAA
  in old APP tg mice. Human trials are underway.
• Ultimately, an active vaccine would be less
  costly and would allow coverage of a much larger
  population compared to passive transfer.
• Our novel immunogens appear to be safe. Further
  testing is required but first an animal model is
  needed.

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All truth passes through three stages. First,
it is ridiculed. Second, it is violently opposed.
Third, it is accepted as being self-evident. Art
hur Schopenhauer (1788-1860) German Philosopher
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Acknowledgements
Center for Neurologic Diseases Lemere Lab
Others Tim Seabrook Gal Bitan Marcel
Maier Weiming Xia Liying Jiang Jackie Sears
Katelyn Thomas Dominic Walsh Ying Peng
Noel Lazo Amy Huynh Former Lab Members Edward
Spooner Jeanne Bloom Diana Li
Dr. Lennart Mucke (J20 APP tg mice) --
Gladstone Institute, SF, CA Dr. John Clements
LT(R192G) -- Tulane U. Medical School,
LA Elan Pharmaceuticals (ELISA Mabs) -- So.
San Francisco, CA Drs. Todd Golde and Pritam Das
(ELISA) -- Mayo Clinic, Jacksonville,
FA Funding Alzheimers Association, NIH
(AG20159) to CAL, and a special thanks to the
Brunozzi Family.