Title: Non-transgenic rodent model of alzheimer's
United States Patent: 6,172,277
Inventors: Tate; Barbara A. (Sharon, MA); Majocha; Ronald
(late of Sharon, MA); Newton; Julie L. (Pawtucket, RI)
Assignee: The Miriam Hospital (Providence, RI)
Appl. No.: 959148
Filed: October 28, 1997
The invention features a method of inducing amyloid plaque deposition
in a mammal by infusing into the brain of the mammal an amyloid peptide at
a basic pH, a nontransgenic animal model for Alzheimer's disease, and
methods of identifying compounds to treat Alzheimer's disease.
SUMMARY OF THE INVENTION
The invention features a nontransgenic animal model (e.g., a rat model) of
human Alzheimer's disease and a method useful for evaluating strategies to
prevent or treat the development of the disease. In addition to the use of
the model in evaluating the efficacy of candidate compounds or other
therapeutic interventions, the etiology of the disease may be determined
using the model.
A method of inducing amyloid plaque deposition in a mammal is carried out
by infusing into the brain of the mammal an amyloid peptide at a basic pH,
e.g., a pH greater than 8.0 such as a pH greater than 9.0 but less than
11. Preferably, the pH is 9.5.
An amyloid peptide is a peptide derived from human amyloid .beta. protein
that causes amyloid plaque deposition in mammalian brain tissue.
Preferably, the peptide has an amino acid sequence corresponding to the
first 40 amino acids of human amyloid .beta. protein, i.e., the amino acid
ly-Val-Val (SEQ ID NO:1).
Amyloid peptide is continuously infused into the brain for at least one
week, preferably two weeks, more preferably four weeks, and most
preferably eight weeks. Peptides may be infused continuously for a year or
more with regular changes of the pump as the contents becomes depleted.
Preferably, the amyloid peptide is infused at a concentration of at least
1 mg/ml (preferably in the range of about 1-10 mg/ml, more preferably in
the range of about 1-5 mg/ml, and most preferably at about 2 mg/ml). The
rate of infusion is about 0.5 .mu.l/hour for about 2 weeks. Preferably,
the rate is at least 100 mg per week, more preferably the peptide is
infused at a rate of about 200 mg per week. Amyloid peptide is
administered to an animal e.g., a rat, at a dose within the range of 0.1
mg/kg to 50 mg/kg of body weight. Preferably, the dose is in the range of
1 to 1.5 mg/kg of body weight.
The invention also features a nontransgenic mammalian model for human
Alzheimer's disease. Amyloid plaques are induced in the brain tissue of a
nontransgenic mammal, e.g., a rodent such as a rat, mouse, hamster, guinea
pig, or degu, by infusing into the brain of the mammal an amyloid peptide
at a basic pH for an extended period of time, e.g., weeks to months. In
animals that are long-lived (e.g., the degu which has an average life span
of about 10 years), the amyloid peptide may be infused for a period of
months to years to induce or accelerate plaque deposition and/or other
Alzheimer's type pathology.
Animals infused with a peptide having the amino acid sequence of SEQ ID
NO:1 develop a plurality of amyloid plaques in their brain tissue. Control
animals infused with vehicle alone or a non-toxic control peptide, e.g, a
peptide which differs from SEQ ID NO:1 by several amino acid
substitutions, develop few or no plaques. Preferably, the sequence of the
control peptide is
al-Ser-Glu (SEQ ID NO:2).
The nontransgenic model is characterized by Alzheimer's associated
disrupted sleep and circadian activity patterns compared to a control
mammal (receiving vehicle alone or the control peptide). Control mammals
typically develop few or no plaques, whereas mammals infused with the
amyloid peptide are characterized by at least a 50% increase in the number
of amyloid plaques compared to the number in a control mammal. Preferably,
the mammal is characterized by at least a 100% increase in the number of
amyloid plaques compared to the number in a control mammal. More
preferably, the mammal is characterized by at least a 200% or greater
increase in the number of amyloid plaques compared to the number in a
Also within the invention is an in vivo screening assay to determine
whether a compound reduces deposition of amyloid plaques. The screening
assay is carried out by (a) providing a first and second nontransgenic
mammal, e.g., a rat, each of which is characterized as having amyloid
plaques in a brain tissue (the plaques having been induced by infusing
into the brain of each mammal an amyloid peptide at a basic pH); (b)
contacting the first mammal with a candidate compound; (c) maintaining the
second mammal in the absence of the candidate compound; and (d) comparing
the degree of amyloid plaque deposition in the brain of the first mammal
with the degree of amyloid plaque deposition in the brain of the second
mammal. A lesser degree of deposition in the first mammal compared to that
in the second mammal is an indication that the candidate compound reduces
amyloid plaque deposition associated with Alzheimer's disease pathology.
In addition to quantifying plaque deposition, the screening assay may be
used to determine whether a compound reduces an Alzheimer's
disease-associated disruption in behavioral abnormalities, e.g., sleep and
circadian activity disruptions. In the latter case, the last step of the
assay requires comparing a sleep and circadian activity pattern of the
first mammal with the sleep and circadian activity pattern of the second
mammal. A lesser degree of disruption of a normal or control pattern
(e.g., a pattern obtained from an animal infused with vehicle alone or
control peptide) in the first mammal compared to that in the second mammal
is an indication that the candidate compound reduces Alzheimer's
disease-associated disruption in sleep and circadian activity.
The methods of the invention have several advantages over known methods of
inducing amyloid plaques to simulate Alzheimer's type pathology, e.g.,
reduced mechanical tissue damage from administration procedures, reduced
neurotoxicity of vehicle, increased solubility of the peptide over long
periods of time (e.g., over the weeks required for continuous infusion
protocols), and increased delivery (total load and efficiency of transfer
from infusion apparatus) of peptide to brain tissue.
In contrast to prior art control peptides, the control peptide described
herein produces few or no amyloid plaques. In addition, amyloid deposition
in amyloid peptide-infused animals is associated with a pronounced
cellular immune response, wherease infusion of identical amounts of
control peptide does not elicit a detectable immune response. This immune
response is analogous to that observed in brain tissue derived from human
Alzheimer's disease patients.
In contrast to transgenic models of Alzheimer's disease which are harbor
DNA encoding amyloid peptide (a significant percentage of which fail to
express the recombinant peptide), most or all of the animals chronically
infused with amyloid peptide according to the invention develop
Alzheimer's type pathology such as amyloid plaque deposition. The amyloid-infused
animals also display behavioral symptoms such as disruptions in sleep and
circadian rhythm patterns not observed in other rodent models.
Claim 1 of 32 Claims
What is claimed is:
1. A method of inducing amyloid plaque deposition in a rodent, comprising
infusing into the brain of said rodent a solution comprising a .beta.-amyloid
peptide at a basic pH, wherein the infusion of said peptide results in the
formation of amyloid plaques in the brain of said rodent in a greater
number than in a control rodent infused with buffer alone or receiving a
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