U.S. patent application number 11/497914 was filed with the patent office on 2006-12-28 for methods of treating schizophrenia.
This patent application is currently assigned to Titan Pharmaceuticals, Inc.. Invention is credited to Richard C. Allen, Michael Cornfeldt.
Application Number | 20060292128 11/497914 |
Document ID | / |
Family ID | 23112130 |
Filed Date | 2006-12-28 |
United States Patent
Application |
20060292128 |
Kind Code |
A1 |
Allen; Richard C. ; et
al. |
December 28, 2006 |
Methods of treating schizophrenia
Abstract
The invention provides methods for the treatment of abnormal
psychiatric states, particularly the negative symptoms of
schizophrenia and extrapyramidal side effects (EPS) of
antipsychotic drugs. The inventive methods relate to the
administration of therapeutic cells (which produce dopamine or
dopamine precursors) adhered to support matrices to subjects
suffering from the negative symptoms of schizophrenia and/or EPS.
The therapeutic cells may be coadministered with cells which
protect the therapeutic cells from immune rejection and/or cells
which produce neurotrophic factors which improve the viability of
the therapeutic cells.
Inventors: |
Allen; Richard C.;
(Flemington, NJ) ; Cornfeldt; Michael;
(Morristown, NJ) |
Correspondence
Address: |
MORRISON & FOERSTER LLP
755 PAGE MILL RD
PALO ALTO
CA
94304-1018
US
|
Assignee: |
Titan Pharmaceuticals, Inc.
South San Francisco
CA
|
Family ID: |
23112130 |
Appl. No.: |
11/497914 |
Filed: |
August 1, 2006 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
09289576 |
Apr 9, 1999 |
7115256 |
|
|
11497914 |
Aug 1, 2006 |
|
|
|
Current U.S.
Class: |
424/93.7 ;
435/368 |
Current CPC
Class: |
A61K 48/00 20130101;
A61K 35/30 20130101; A61P 25/18 20180101 |
Class at
Publication: |
424/093.7 ;
435/368 |
International
Class: |
A61K 35/30 20060101
A61K035/30; C12N 5/08 20060101 C12N005/08 |
Claims
1. A method for providing dopamine or a dopamine precursor to a
dopamine-deficient brain of a subject exhibiting extrapyramidal
side effects (EPS) produced by antipsychotic drugs, comprising
administering an effective amount of a cell/support complex to the
brain of the subject, wherein said cell/support complex comprises
cells adhered to a support matrix, wherein said cells produce
dopamine or a dopamine precursor, wherein the cells are allogeneic,
non-genetically modified retinal pigment epithelial cells, wherein
said support matrix is made of material selected from the group
consisting of glass, polystyrene, polypropylene, polyethylene,
polyvinylidene fluoride, polyurethane, polyalginate, polysulphone,
polyvinyl alcohol, acrylonitrile polymers, polyacrylamide,
polycarbonate, polypentene, polypentane, nylon, magnetite, natural
polysaccharide, modified polysaccharide, collagen, gelatin and
modified gelatin, and wherein said extrapyramidal side effects
(EPS) produced by antipsychotic drugs are ameliorated.
2. The method of claim 1, wherein said cell/support complex is
administered to the subject by injection.
3. The method of claim 1, wherein said cell/support complex is
administered to the subject by implantation.
4. The method of claim 1, wherein said support matrix is gelatin or
modified gelatin.
5. The method of claim 4 wherein said support matrix is crosslinked
gelatin.
6. The method of claim 1 wherein the cells produce a dopamine
precursor.
7. The method of claim 1 wherein the cells produce dopamine.
8. The method of claim 1 wherein the subject is a human.
9. The method of claim 1 wherein said EPS is tardive
dyskinesia.
10. The method of claim 9 wherein said cell/support matrix is
administered to the striatal area of said subject's brain.
11. The method of claim 10 wherein said cell/support complex is
administered by injection.
12. The method of claim 11 wherein said cell/support complex is
administered by implantation.
13. The method of claim 1 wherein said EPS is tardive dystonia.
14. The method of claim 1 wherein said EPS is tardive
akathisia.
15. The method of claim 1 wherein said cells are administered in a
bilaterally-symmetrically pattern.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application is a divisional of U.S. application Ser.
No. 09/289,576, filed Apr. 9, 1999, which is herein incorporated by
reference in its entirety.
TECHNICAL FIELD
[0002] The invention relates generally to the treatment of
psychiatric disorders, and particularly to methods of treating
negative symptoms of schizophrenia and/or certain extrapyramidal
side effects (EPS) of antipsychotics by administering an effective
number of cells, adhered to a support matrix, which are capable of
producing dopamine or a dopamine precursor to a subject suffering
from schizophrenia.
BACKGROUND
[0003] Disorders of the nervous system comprise a wide variety of
conditions, and can generally be separated into "neurological
disorders" and "psychiatric disorders". Neurological disorders can
be characterized as the product of the death of nervous system
cells. Examples of neurological disorders include Parkinson's
Disease, which involves the death of dopaminergic neurons,
Alzheimer's disease, in which cholinergic neurons are lost, and
multiple sclerosis, in which neurons and glia are killed in an
autoimmune process. Psychiatric disorders, on the other hand, are
the result of dysregulation of neurons, which can result in
abnormal levels of various neurochemicals and/or abnormal
responsiveness to neurochemicals.
[0004] Psychiatric disorders, such as schizophrenia, are major
public health concerns. Schizophrenia, for example, affects
approximately 2 million Americans. At any particular time, about
20% of the hospital beds in the U.S. are occupied by schizophrenic
patients. The illness usually develops between adolescence and age
30 and is characterized by positive symptoms (delusions or
hallucinations), negative symptoms (blunted emotions and lack of
interest) and disorganized symptoms (confused thinking and speech
or disorganized behavior and perception). Additionally, cognitive
deficits are also frequently observed, particularly in elderly
schizophrenia patients (Purohit et al., 1993, Biol. Psychiatry
33(4):255-260). For some patients, the disorder is lifelong, while
others may have periodic episodes of psychosis.
[0005] The causes of schizophrenia are essentially unknown.
Although it is believed to have a genetic component, environmental
factors appear to influence the onset and severity of the disease.
Neuropathological changes in schizophrenics may include enlargement
of the lateral ventricles, cavities in the brain which are part of
the cerebrospinal fluid system. Sometimes, there is a decrease in
overall brain mass.
[0006] A number of animal models have been developed for
schizophrenia, utilizing both non-primate (rat) and primate
(monkey) animals. In one commonly used animal model of
schizophrenia, phencyclidene (PCP) is chronically administered to
the animal subjects, resulting in dysfunctions similar to those
seen in schizophrenic humans (Jentsch et al., 1997, Science
277:953-955; Piercey et al., 1988, Life Sci. 43(4):375-385).
[0007] Several different theories have been developed regarding the
etiology of schizophrenia, including the dopaminergic,
glutamatergic, and cholinergic theories of schizophrenia. The
dopamine hypothesis posits that positive symptoms result from
excess function of the neurotransmitter dopamine in the mesolimbic
area of the brain. This hypothesis is based largely on indirect,
pharmacological evidence that (1) dopamine-antagonizing drugs are
effective antipsychotic agents; (2) dopamine-mimicking drug
exacerbate schizophrenic symptoms and (3) certain symptoms of acute
paranoid schizophrenia can be elicited in non-schizophrenics by
amphetamine, a drug that activates dopamine systems. Conversely,
negative symptoms have been associated with regionally localized
dopamine deficits in the prefrontal cortex.
[0008] The glutamate hypothesis is based, among other things, the
actions of phencyclidine (PCP) in the brains of abusers. PCP, which
induces a number of symptoms also found in schizophrenia, blocks
the N-methyl-D-aspartate (NMDA) receptor, through which glutamate
exerts some of its effects. Additionally, observations from the
phencyclidine (PCP) animal model of schizophrenia, indicate that
glutamate is dramatically increased in the brains of rats
chronically dosed with PCP (Moghaddam et al., 1997, J. Neurosci.
17:2921). Inhibition of the rise in glutamate levels with a
metabotropic glutamate receptor agonist blocked a variety of
schizophrenia symptoms in PCP-dosed rats, including hyperactive
behavior, head turning, and memory deficits (Moghaddan et al.,
1998, Science 281:1349-1352).
[0009] Current treatments for schizophrenia include antipsychotic
drugs; electric shock treatment for severe catatonia, depression,
or elation; and psychotherapy. Antipsychotic drug treatments
generally include both dopamine and acetylcholine antagonists
(anticholinergics), although anticholinergics are generally used to
treat extrapyramidal side effects (EPS) of commonly used dopamine
antagonists, rather than the disorder itself. EPS induced by
dopamine antagonists result from dopamine blockade in the striatum
and include dyskinesias, especially tardive dyskinesia, which are
characterized by uncontrolled movements. In most cases, tardive
dyskinesia resolves following discontinuation of dopamine agonist
drug therapy, but some patients continue to experience dyskinesia
long after termination of dopamine antagonist therapy.
[0010] Clopazine, an antipsychotic that affects several receptor
systems, is currently viewed as the "gold standard" of drug
treatment for schizophrenia. However, although clopazine seems to
ameliorate certain symptoms (e.g., verbal fluency, reaction time
and attention), it appears to have an adverse effect on higher
level executive functions, such as working memory ability and
visual memory. Goldberg et al. (1994) J. Clin. Psychiatry 55:9
(suppl. B). Moreover, many dopamine antagonist antipsychotic drugs
are not as effective at controlling the "negative" symptoms, as
compared to the "positive" symptoms of schizophrenia (Angrist et
al., 1982, Psychopharm. 78:128-130).
[0011] U.S. Pat. No. 5,447,948 asserts that dopamine reuptake
inhibitors are useful in treating schizophrenic patients suffering
from negative symptoms. However, dopamine reuptake inhibitors
oppose the effects of clopazine and other dopamine antagonists,
potentially nullifying the activity of this group of drugs in the
treatment of the positive symptoms of schizophrenia.
[0012] U.S. Pat. No. 5,618,531 describes methods of increasing the
viability of cells which are administered to the brain or spinal
cord of a mammalian subject by attaching the cells to a support
matrix. It is indicated that the cells may be used to "neurological
disorders," but not psychiatric disorders. As commonly understood
in the art, "neurological disorders" are disorders of the nervous
system which involve death or major dysfunction of cells of the
nervous system, while "psychiatric disorders" involve the
misregulation of cells of the nervous system. For example,
Parkinson's disease, which involves the death and dysfunction of
dopaminergic neurons, is a neurological disorder, while
schizophrenia, which involves the misregulation of neurotransmitter
levels, is a psychiatric disorder.
[0013] In sum, there remains a need in the art for an effective
treatment of the symptoms of schizophrenia, particularly the
negative symptoms of schizophrenia and cognitive defects associated
with schizophrenia, which will not interfere with treatments for
the positive symptoms of the disorder. Additionally, there exists a
need in the art for an effective treatment for tardive dyskinesia
and other EPS.
SUMMARY OF THE INVENTION
[0014] The invention relates to methods of treating schizophrenia,
particularly the negative symptoms of schizophrenia, by
administering therapeutic cells which produce dopamine to a subject
having schizophrenia. The therapeutic methods of the invention
provide treatments for the negative symptoms of schizophrenia which
do not interfere with contemporaneous, pharmacological treatments
for the positive symptoms of schizophrenia. The invention also
provides methods for the treatment of EPS, particularly tardive
dyskinesia (TD), also by the administration of therapeutic
cells.
[0015] In one aspect, the invention provides methods for treating
symptoms of schizophrenia, particularly the negative symptoms of
schizophrenia and cognitive deficits associated with schizophrenia,
by administering therapeutic cells adhered to a support matrix
(cell/support complex) to the subject. The therapeutic cells may be
paraneural cells, neural cells, cells engineered by somatic cell
hybridization, cells derived from the adrenal medulla, cells that
have been genetically engineered to produce dopamine, or any other
cells which produce dopamine. The support matrix is a
non-encapsulating particulate support.
[0016] In another aspect, the invention provides methods in which
protective cells (cells which produce an immunologically privileged
site) are administered with the therapeutic cells. Alternately, the
therapeutic cells may have the properties of protective cells.
[0017] In a further aspect, the invention provides methods in which
support cells, which provide factors which improve the viability or
function of the therapeutic cells, are administered with the
therapeutic cells. A combination of therapeutic cells, protective
cells, and support cells for administration to the patient is also
contemplated.
[0018] In another embodiment, the invention provides methods for
improving cognitive deficits associated with schizophrenia,
particularly elderly schizophrenia patients. Cognitive deficits are
treated by the administration of therapeutic cells, or a
combination of therapeutic cells with protective cells and/or
support cells, adhered to a support matrix, to the brain of a
subject suffering from schizophrenia-associated cognitive deficits.
Administration of the therapeutic cells results in an improvement
in cognitive function in the subject.
[0019] In further embodiments, the invention provides methods for
the treatment of EPS such as tardive dyskinesia, tardive dystonia
and tardive akathisia, particularly tardive dyskinesia. In
accordance with the invention, EPS are treated by administration of
therapeutic cells which produce dopamine, adhered to a substrate,
to a subject. The therapeutic cells may be administered with
protective cells, or have the properties of protective cells.
Additionally, support cells may be administered with the
therapeutic cells.
[0020] Also provided are pharmaceutical compositions comprising the
therapeutic cells adhered to a support matrix. The pharmaceutical
compositions may optionally comprise protective cells, support
cells, or both.
[0021] In a further embodiment, the invention provides kits which
may be utilized for practicing the instant inventive methods. The
kits comprise therapeutic cells and a support matrix, and may
additionally include protective cells, support cells, or both
protective cells and support cells.
[0022] As will become apparent, preferred features and
characteristics of one aspect of the invention are applicable to
any other aspect of the invention.
DETAILED DESCRIPTION OF THE INVENTION
[0023] The present invention provides methods for treating the
negative symptoms of schizophrenia by administration of cells into
one or more sites in the central nervous system (CNS) of a subject
suffering from the symptoms of schizophrenia, particularly negative
symptoms of schizophrenia or cognitive deficits associated with
schizophrenia. Cognitive deficits associated with schizophrenia may
also be treated in accordance with the invention by the
administration of therapeutic cells. The invention also provides
methods for the treatment of EPS, particularly tardive dyskinesia,
by administration of cells into one or more sites in the CNS of
subject suffering from the symptoms of EPS such as tardive
dyskinesia (TD). In accordance with the methods of the invention,
cells which produce dopamine ("therapeutic cells") are introduced
into the brain of a subject suffering from the negative symptoms of
schizophrenia, cognitive deficits associated with schizophrenia,
and/or EPS. The therapeutic cells are normally administered
adherent to a microcarrier (cell/support complex) when administered
to the subject, and can be administered with other cells. The
instant invention is advantageous for the treatment of
schizophrenia, particularly for the treatment of the negative
symptoms of schizophrenia, cognitive deficits associated with
schizophrenia, particularly in elderly schizophrenics, and EPS
(e.g., TD) associated with the administration of antipsychotic
drugs.
[0024] The instant methods may be used as a "stand alone" therapy,
but are also contemplated for use in combination with other
therapies. In particular, it is contemplated that the instant
methods may be utilized in combination with standard schizophrenia
therapies, such as clozapine. It is further contemplated that
treatment of a subject in accordance with the invention will result
in "cross-talk" within the brain of the subject, resulting in an
improvement in positive symptoms suffered by the subject,
independent of any improvement in positive symptoms due to
treatment with antipsychotic drugs. Accordingly, the invention also
provides methods for alleviating negative and positive symptoms of
Schizophrenia.
[0025] The practice of the present invention will employ, unless
otherwise indicated, conventional techniques of molecular biology,
microbiology, cell biology and recombinant DNA, which are within
the skill of the art. See, e.g., Sambrook et al., MOLECULAR
CLONING: A LABORATORY MANUAL, 2nd edition (1989); CURRENT PROTOCOLS
IN MOLECULAR BIOLOGY, (F. M. Ausubel et al. eds., 1987); the series
METHODS IN ENZYMOLOGY (Academic Press, Inc.); PCR 2: A PRACTICAL
APPROACH (M. J. McPherson, B. D. Hames and G. R. Taylor eds.,
1995); ANIMAL CELL CULTURE (R. I. Freshney. Ed., 1987); and
ANTIBODIES: A LABORATORY MANUAL (Harlow et al. eds., 1987).
Definitions
[0026] As used herein, the term "schizophrenia" is intended to
include the group of mental disorders characterized by disruptions
in thinking and perception. In a clinical evaluation, schizophrenia
is commonly marked by "positive symptoms" such as auditory
hallucinations (especially hearing voices), disorganized thought
processes and delusions as well as "negative symptoms" which
include affective flattening, alogia, avolition, and anhedonia.
[0027] As used herein, "the negative symptoms of schizophrenia"
refer to a class of symptoms of schizophrenia which can be
considered to reflect a `loss` in functional, directed thought or
activity. Negative symptoms of schizophrenia are well known in the
art, and include affective flattening (characterized by, for
example, an immobile and/or unresponsive facial expression, poor
eye contact and reduced body language), alogia (`poverty of speech`
or brief, laconic and/or empty replies), avolition (characterized
by a reduced or absent ability to initiate and carry out
goal-directed activities), anhedonia (loss of interest or
pleasure), social withdrawal, apathy and other negative symptoms
known to those of skill in the art. The negative symptoms of
schizophrenia may be assessed using any methodology known in the
art including, but not limited to, the Brief Psychiatric Rating
Scale (BPRS), the Positive and Negative Symptom Scale (PANSS), the
Rorschach Schizophrenia Index (SCZI), and the Scale for the
Assessment of Negative Symptoms (SANS). Some of these methods may
also be used to assess positive symptoms (e.g., BPRS, PANSS and
SCZI), although methods for specifically assessing positive
symptoms are also available (e.g., the Scale for the Assessment of
Positive Symptoms, or PANS).
[0028] As used herein "cognitive deficits associated with
schizophrenia" refers to cognitive deficits in schizophrenia
patients. Cognitive deficits include, but are not limited to
deficits of working memory, visuospatial memory, and low
performance on `frontal` neuropsychological tests such as the
Wisconsin Card Sorting Test (WCST).
[0029] The term "support matrix" includes any material to which
cells adhere following in vitro incubation, and on which cells can
grow, and which can be implanted into a mammalian brain without
producing a toxic reaction, or an inflammatory or gliosis reaction
which would destroy the implanted cells or otherwise interfere with
their biological or therapeutic activity. Such materials may be
synthetic or natural chemical substances or substances having a
biological origin. The matrix materials include, but are not
limited to, glass and other silicon oxides, polystyrene,
polypropylene, polyethylene, polyvinylidene fluoride, polyurethane,
polyalginate, polysulphone, polyvinyl alcohol, acrylonitrile
polymers, polyacrylamide, polycarbonate, polypentene, polypentane,
nylon, amylases, gelatin, modified (e.g., crosslinked) gelatin,
collagen, natural and modified polysaccharides, including dextrans
and celluloses (e.g. nitrocellulose), agar, and magnetite.
[0030] Either resorbable or non-resorbable materials may be used.
Also intended are extracellular matrix materials, which are
well-known in the art (see below). Extracellular matrix materials
may be obtained commercially or prepared by growing cells which
secrete such a matrix, removing the secreting cells, and allowing
the cells which are to be transplanted to interact with and adhere
to the matrix.
[0031] As used herein, the term "therapeutic cell" refers to a cell
which produces dopamine or a precursor of dopamine (e.g., L-DOPA).
Therapeutic cells may be of neural origin, paraneural cells such as
retinal pigmented epithelium (RPE) cells and chromaffin cells,
cells engineered by somatic cell hybridization, cells derived from
the adrenal medulla, cells that have been genetically engineered to
produce biologically active factors, and any other cells which
produce dopamine or a dopamine precursor. In some cases,
therapeutic cells will also have the attributes of protective cells
(e.g., RPE cells). Therapeutic cells are preferably "species
matched" (derived from the same species as the subject), but may be
derived from any species, preferably a mammalian species.
[0032] A "paraneural cell" is a cell which is derived from the
embryonic neural crest. Examples of paraneural cells include
retinal pigmented epithelium (RPE) cells and cells derived from the
adrenal medulla, such as adrenal chromaffin cells.
[0033] A "cell which produces an immunologically privileged site"
is a cell which produces a locally immunosuppressive environment. A
cell which produces an immunologically privileged site may also be
known as a "protective cell". A locally immunosuppressive
environment may be produced by the expression of anti-inflammatory
or immunosuppressive molecules such as Fas ligand (Fas L),
transforming growth factor beta (TGF-.beta.) and/or other molecules
known in the art. Cells which produce an immunologically privileged
site include cells which naturally produce an immunologically
privileged site, such as RPE cells and Sertoli cells, and cells
which have been genetically modified to produce an immunologically
privileged site by, for example, expressing Fas L or TGF-.beta..
Protective cells are preferably species matched, but may be derived
from any species, although preferably a mammalian species.
[0034] The term "support cell", as used herein, refers to a cell
which produces factors that improve the viability of the
therapeutic cells. Support cells may produce soluble factors, such
as neurotrophic factors or other factors (e.g., growth factors), or
they may produce extracellular matrix or other insoluble factors
which improve the viability of the therapeutic cells. Suitable
support cells for use in the instant invention include glial cells,
which naturally `support` neurons, and cells genetically engineered
to act as support cells by, for example, producing a beneficial
neurotrophic factor, such as NGF.
[0035] As used herein, "treatment" is an approach for obtaining
beneficial or desired clinical results. For purposes of this
invention, beneficial or desired clinical results include, but are
not limited to, alleviation of symptoms associated with the
psychiatric disorder, diminishment of the extent of the disorder,
stabilized (i.e. not worsening) state of disorder, delay or slowing
of disorder progression.
[0036] An "effective amount" or "effective number" is an amount or
number sufficient to effect beneficial or desired therapeutic
results. An effective amount can be administered in one or more
administrations. For purposes of this invention, an effective
amount is that amount which achieves the desired result, namely
amelioration, stabilization, palliation or slowing of the mental
disorder.
[0037] As used herein, the term "comprising" and its cognates are
used in their inclusive sense; that is, equivalent to the term
"including" and its corresponding cognates.
[0038] The term "subject" refers to a mammal, preferably a
human.
[0039] An "expression vector" is a DNA construct which (due to the
presence of appropriate transcriptional and/or translational
control sequences) is capable of expressing a DNA (or cDNA)
molecule which has been cloned into the vector and of thereby
producing a polypeptide or protein in the therapeutic cell.
Expression of the cloned sequences occurs when the expression
vector is introduced into an appropriate host cell. An appropriate
mammalian host cell would be any an cell capable of expressing the
cloned sequences. Procedures for preparing cDNA and for producing a
genomic library are disclosed by Sambrook et al. (supra).
[0040] Two DNA sequences (such as a promoter region sequence and a
coding sequence) are said to be operably linked if the nature of
the linkage between the two DNA sequences does not (1) result in
the introduction of a frame-shift mutation, (2) interfere with the
ability of the promoter region sequence to direct the transcription
of the coding sequence, or (3) interfere with the ability of the
coding sequence to be transcribed by the promoter region sequence.
A promoter region would be operably linked to a DNA sequence if the
promoter were capable of effecting transcription of that DNA
sequence.
[0041] It should be noted that, as used herein, the singular form
"a", "an" and "the" include plural references unless the context
clearly dictates otherwise.
[0042] The present invention provides, in one aspect, methods for
treating the negative symptoms of schizophrenia. In accordance with
the instant invention, the negative symptoms of schizophrenia are
treated by administering a cell/support complex comprising
therapeutic cells that supply dopamine or a dopamine precursor
adhered to a support matrix (cell/support complex) to the subject.
The cell/support complex is generally administered directly into
the brain by injection or implantation. The cell/support complex
may be administered at one or more sites in the subject, and may be
administered on a single administration or in multiple
administrations. After completion of administration, the negative
symptoms of schizophrenia are alleviated, ameliorated or
eliminated. Alleviation, amelioration, or elimination of the
negative symptoms of schizophrenia can be measured using any of the
tests, scales, or indexes known in the art, such as the BPRS, SANS
or PANSS.
[0043] In another aspect, the instant invention provides methods
for treating the negative symptoms of schizophrenia by
administering a cell/support complex comprising therapeutic cells
in combination with cells which produce an immunologically
privileged site ("protective cells"), although in certain other
embodiments the therapeutic cells have the properties of protective
cells. If the therapeutic cells are combined with separate
protective cells, both types of cells are normally administered as
a cell/support complex and are generally administered directly into
the subject brain by injection or implantation.
[0044] The therapeutic cells, optionally with accompanying
protective cells, are administered to subjects suffering from the
negative symptoms of schizophrenia. Administration of the
therapeutic cells (with or without protective cells) results in
alleviation of the symptoms of the negative symptoms of
schizophrenia.
[0045] Preferably, the cells are administered directly to the brain
by injection or implantation. The cells are preferably administered
to one or more sites in the prefrontal cortex. Preferably, the
cells are administered to the prefrontal cortex in a
bilaterally-symmetrical pattern (i.e., at matching sites in the
left and right prefrontal cortex). The administration of the
therapeutic cells results in alleviation of the symptoms of
schizophrenia, particularly negative symptoms.
[0046] In another aspect, the invention provides methods of
alleviating cognitive deficits associated with schizophrenia,
particularly in elderly schizophrenia patients. Therapeutic cells
are administered in a cell/support complex by injection or
implantation into the prefrontal cortex of the subject's brain,
particularly the dorsolateral prefrontal cortex. As with the
instant methods for the treatment of the negative symptoms of
schizophrenia, cognitive deficits may be treated by the
administration of therapeutic cells in a cell/support complex with
protective cells and/or support cells. Administration of
therapeutic cells in a cell/support complex results in improvements
in cognitive function.
[0047] The invention also provides methods for the treatment of
extrapyramidal side effects (EPS). In accordance with the
invention, a cell/support complex comprising therapeutic cells
which produce dopamine or a dopamine precursor adhered to a support
matrix are administered to a subject suffering from EPS (such as
tardive dyskinesia, tardive dystonia and/or tardive akathisia). The
therapeutic cells may optionally be administered in combination
with protective cells. Preferably, the cells are administered by
implantation or injection into one or more sites in the striatal
area of the subject's brain. Preferably, the cells are administered
in a bilaterally-symmetrically pattern.
Cells
[0048] Therapeutic cells, in accordance with the instant invention,
produce dopamine or a dopamine precursor (e.g., L-DOPA). Cells
useful in the practice of various aspects of this invention include
cells of neural origin, paraneural cells such as RPE cells and
chromaffin cells, cells engineered by somatic cell hybridization,
cells derived from the adrenal medulla and cells that have been
genetically engineered to produce biologically active factors.
Neural and paraneural cells are preferred cells for use in the
instant invention.
[0049] Typically, a cell used in the instant invention will be
selected for its ability to produce dopamine or a dopamine
precursor. Alternately, a cell may be selected for other properties
(such as ease of propagation in vitro) and genetically engineered
to produce the desired biologically active compound.
[0050] Generally, cells for use in accordance with the invention
are post-mitotic or of very low or controlled mitotic potential
when administered to the subject. As will be understood by one of
skill in the art, the introduction of actively dividing cells into
the cranium should be avoided. Introduction of actively dividing
cells into the cranium of a subject can lead to the formation of a
tumor, which can damage or destroy structures in the subject's
brain due to compressive damage.
[0051] A preferred cell for use in the instant invention is a
paraneural cell. Retinal pigmented epithelial (RPE) cells, which
are found as a monolayer between the retina and uvea and which
produce dopamine and other factors, are a preferred paraneural cell
(Song et al., 1990, i J. Cell. Physiol. 148:196-203). Methods of
isolating and culturing human RPE cells are known in the art (for
example, as described in Liu et al., 1988, Exp. Eye Res.
47:911-917; Lopez et al., 1989, Invest. Ophthamol. Vis. Sci.
30:586-588; and Lui et al., 1990, Investigative Ophthamol., ARVO
Supplement). RPE cells can be isolated for any source known in the
art, and are preferably human.
[0052] Another example of a cell of paraneural origin is an adrenal
medullary chromaffin cell. The cells which form the mammalian
adrenal medulla are derived from the neural crest and possess the
potential to develop along either neuronal or endocrine lines of
differentiation (Bohn et al., 1981, supra, and 1982, Devel. Biol.
89:299-308; Unsicker, 1985, Develop. Biol. 108:259-268). Mammalian
chromaffin cells from the adrenal medulla, when removed from
adrenal cortical influences and exposed to nerve growth factor
(NGF), change from an endocrine to a neuronal phenotype (Notter et
al., 1986, Cell Tiss. Res. 244:69-70; Stromberg et al., 1985, Exp.
Brain Res. 60:335-349). When co-grafted with cerebral cortical or
hippocampal tissue into the anterior chamber of the rat eye,
adrenal chromaffin cells form nerve fibers which innervate the
adjacent co-grafted brain tissue (Olson et al., 1980, Exp. Neurol.
70:414-426). It has also been shown that transplanted adrenal
medulla tissue can correct functional deficits resulting from
nigrostriatal dopamine depletion in rats (see, for example, Freed
et al., 1981, Nature 292(5821):351-352), although dopamine
secretion, and the functional improvement by the implant, decreases
by about three to six months after implantation. However, NGF
treatment of the transplanted cells induces fiber outgrowth from
the transplant into the host and induces a longer lasting
behavioral recovery (at least a year) and few implant cells survive
without NGF treatment. Accordingly, in embodiments utilizing
adrenal medulla chromaffin cells as the therapeutic cells, the
therapeutic cells are preferably treated with a neurotrophic
factor, preferably NGF, prior to implantation, and may be
optionally implanted in the presence of a neurotrophic factor such
as NGF.
[0053] Another source of therapeutic cells is established neural
cell lines. Many neuronal clones exist which have been used
extensively as model systems of development since they are
electrically active with appropriate surface receptors, specific
neurotransmitters, synapse forming properties and the ability to
differentiate morphologically and biochemically into normal
neurons. Such cells are described, for example, in the following
references: Kimhi et al., 1976, Proc. Natl. Acad. Sci. USA
73:462-466; In: Excitable Cells in Tissue Culture, Nelson, P. G. et
al., eds., Plenum Press, N.Y. 1977, pp. 173-245; Prasad et al., In.
Control of proliferation of Animal Cells, Clarkson, B. et al.,
eds., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.
1974, pp. 581-594; Puro et al., 1976, Proc. Natl. Acad. Sci.USA
73:3544-3548; Notter et al., 1986,Devel. Brain Res. 26:59-68;
Schubert et al., 1970, Proc. Natl. Acad. Sci. USA 67:247-254;
Kaplan et al., In: Basic and Clinical Aspects of molecular
Neurobiology, Guffrida-Stella, A. M. et al., eds., Milano
Fondozione International Manarini, 1982.
[0054] Neural cell lines have a number of advantageous qualities
which may be useful in the instant invention. Generally, these
cells can be propagated in vitro indefinitely, which simplifies
genetic manipulation of the cells. Additionally, the phenotype of
differentiated cells from most neural cell lines can be manipulated
by altering the culture environment. It should be noted that, due
to the potential for such cell lines to form benign or malignant
tumors, therapeutic cells derived from neural cell lines are
rendered amitotic prior to administration in accordance with the
instant invention. Therefore, in one embodiment of the present
invention, cell line cells are modulated in vitro with the
appropriate growth or differentiation factor and with an amitotic
agent before transplantation in order to promote cell survival and
prevent expression of the tumorigenic potential.
[0055] For example, human neuroblastoma cells from the IMR 32 cell
line can survive and express cholinergic markers in primate brain
nine months after transplantation (Gash et al., 1986, Science
233:1420-22). As noted above, these cells are preferably treated to
render them morphologically and biochemically differentiated in
vitro and must be rendered permanently amitotic before
administration, which further aids in their survival (Gash et al.,
supra; Gupta et al., 1985, Dev. Brain Res. 19:21-29). Cells from
the pheochromocytoma/neuroblastoma cell line, PC12, can be
stimulated to differentiate in vitro by the addition of NGF and
anti-mitotic agents to the culture medium. The resulting
differentiated cells produce catecholamine neurotransmitters.
Neural cell lines useful as therapeutic cells in accordance with
the instant invention are those neural cells lines which produce
dopamine naturally, through growth factor/hormone manipulation of
the cells, or through genetic engineering of the cells.
[0056] Somatic cell hybrids may also be used as a source of
therapeutic cells. Somatic cell hybrids can be formed which retain
the properties of differentiated cells. For example, hybrids
derived from fusion of sympathetic ganglia and neuroblastoma cells
can synthesize dopamine (Greene et al., 1975, Proc. Natl. Acad.
Sci. USA 82:4923-4927). Embryonic precursors to dopaminergic
neurons from the CNS can be fused with mitotic cells to incorporate
both genomes into a single one that loses extra chromosomes over
time and results in a new hybrid line. Methods for creating somatic
cell hybrids are well known in the art and it is within the skill
of the art to produce such hybrid neural or paraneural cells
without undue experimentation, screen them for the desired traits
(including dopamine secretion), and select those having the best
potential for transplantation.
[0057] Cells which have been genetically engineered to produce a
substance of neurological interest may also be used as therapeutic
cells. A preferred cell type for genetic engineering is a human
foreskin fibroblast, which is easily obtained and cultured. For use
in the present invention, the cells are genetically altered, using
methods known in the art, to express the enzymes necessary for the
production of dopamine or dopamine precursors (e.g., tyrosine
hydroxylase; Wolff et al., 1989, Proc. Natl. Acad. Sci. USA
86(22):9011-9014, and/or L-DOPA decarboxylase, Scherer et al.,
1992, Genomics 13(2), 469-471). See, for example. Gage et al.,
1987, Neuroscience 23:795-807; Rosenberg et al., 1988, Science
242:1575-1578; Shimohama et al., 1989, Mol. Brain Res.
5:271-278.
[0058] The recombinant DNA molecules useful for the methods of the
present invention can be produced through any of a variety of
means, such as, for example, DNA or RNA synthesis, or more
preferably, by application of recombinant DNA techniques.
Techniques for synthesizing such molecules are disclosed by, for
example, Wu et al. (1978, Prog. Nucl. Acid. Res. Molec. Biol.
21:101-141). Procedures for constructing recombinant molecules are
well known in the art and are disclosed in detail by, for example,
Sambrook et al. (supra).
[0059] Dopamine is synthesized from the amino acid tyrosine in a
two step process. Tyrosine is first hydroxylated by tyrosine
hydroxylase to produce L-DOPA. L-DOPA, which is utilized as a
therapy in neurological disorders involving the loss of
dopaminergic neurons, such as Parkinson's Disease, is converted to
dopamine by L-DOPA decarboxylase. Tyrosine hydroxylase and L-DOPA
decarboxylase genes are known from a number of different species,
including humans.
[0060] Preferably, the cells are genetically engineered to produce
"species matched" enzymes; that is, if the subject is a human, the
genetically engineered dopamine pathway enzymes are also human.
However, since the brain is an immunologically privileged site, it
is not necessary that the genetically engineered dopamine pathway
enzymes are species matched.
[0061] The gene(s) for genetic engineering may be conveniently
amplified from DNA or RNA which contains the gene(s) of interest
using the polymerase chain reaction or any other template-dependent
amplification method, as is well known in the art. The amplified
DNA is then isolated using standard techniques (such as agarose gel
electrophoresis). Alternately, the gene(s) may be isolated from a
DNA, typically a cDNA, library which is known or believed to
contain the gene(s) of interest. The gene(s) is isolated using
techniques known in the art (including nucleic hybridization
screening, phage display, expression screening using soluble
receptors or antibodies which bind to the desired gene product, and
DNA sequencing).
[0062] Once isolated, the gene(s) is subcloned into an "expression
vector" (a DNA construct which comprises the appropriate
transcriptional and translational regulatory sequences to allow
transcription of the gene and translation of the resulting mRNA in
the therapeutic cell) such that the gene(s) is operably linked to
the transcriptional and translational regulatory sequences. If more
than one gene is genetically engineered into the cells, the genes
are preferably linked to separate transcriptional/translational
regulatory sequences.
[0063] Expression constructs for use in eukaryotic cells are well
known in the art. Generally, the expression construct will comprise
an enhancer, a promoter including transcriptional and translational
initiation/regulatory sequences, one or more introns (which may be
located at the 5' end of the gene, within the gene sequence, or at
the 3' end of the gene), the gene of interest, a termination codon,
3' untranslated sequence (3'UTR), and a poly-A addition site. The
precise details of the expression construct will vary according to
the species and cell type of the therapeutic or support cell that
is the subject of the genetic engineering, as is known in the
art.
[0064] As will be apparent to one of skill in the art, the
promoter, enhancer, introns, 3' UTR, and poly-A addition site may
be native to the gene or may be heterologous, depending on the gene
and the therapeutic cell. In most cases, the promoter will be a
heterologous promoter, such as the mouse metallothionein I promoter
(Hamer et al., 1982, J. Mol. Appl. Gen. 1:273-288); the TK promoter
of Herpes virus (McKnight, 1982, Cell 31:355-365); the SV40 early
promoter (Benoist et al., 1981, Nature (London) 290:304-310) and
the collagen promoter (Prockop et al., 1979, New Eng. J. Med.
301:13-23, 77-85; Eyre, 1980, Science 207:1315-1322; Martin et al.,
1985, Trends Bioch. Sci. 10:285-287). Useful heterologous enhancers
include the SV40 enhancer.
[0065] It will be apparent to one of ordinary skill in the art that
cell surface markers may be utilized to select the therapeutic
cells prior to administration. Specific markers can be used to
select fully differentiated cells from a population of cells, or to
select genetically modified cells. A number of CNS-specific markers
are known which may be useful for selection of cells, including
CNS-specific gangliosides, cell surface receptors for
neurotransmitters and neuropeptides, and other markers as are known
in the art, as are methods for utilizing these markers to select
desired cells.
[0066] If the therapeutic cells are heterologous to the subject
(i.e., derived from another individual and/or species), then the
possibility of an immune reaction against the therapeutic cells
must be considered. As discussed above, the therapeutic cells are
preferably species matched. If the therapeutic cells are species
matched, the risk of hyperacute rejection is of very low order.
However, acute and chronic graft rejection reactions are possible.
Accordingly, in certain embodiments of the invention, particularly
where the therapeutic cells are not autologous, the therapeutic
cells have the properties of protective cells (e.g., RPE cells) or
are administered with protective cells (e.g., Sertoli cells). If
the therapeutic cells are administered with separate protective
cells, the two cell types may be mixed prior to or during
attachment to the support matrix, or they may be separately
attached to support matrices and combined immediately before or
during administration to the subject. If the therapeutic cells and
the protective cells are separately adhered to support matrices,
the support matrices for the therapeutic cells and the protective
cells may be the same or different. Generally, approximately
10.sup.2-10.sup.7 protective cells are administered with the
therapeutic cells per site of administration, preferably
10.sup.3-10.sup.6 protective cells, although the exact number will
vary depending on the properties of the protective cells (e.g., the
level of expression of the immunosuppressive molecule), as will be
apparent to one of skill in the art.
[0067] Cells transplanted into the mammalian brain according to the
present invention can survive in the absence of added growth
factors. However, in certain embodiments of the invention, the
therapeutic cells are treated with a neurotrophic factor (e.g.,
NGF) before, during or after attachment to the support matrix,
and/or may be treated with a neurotrophic factor during or after
administration to the subject.
[0068] Therapeutic cells of the invention may also be implanted or
injected with support cells. Glial cells normally act to support
neurons in the nervous system, and grafted glial cells may play an
important role in functional recovery of neurons by, for example,
supplying important neurotrophic factors (Doering et al., 1984, J.
Neurolog. Sci. 63:183-196; Gumple et al., 1984, Neurosci. Lett.
37:307-311). Additionally any cell of choice may be genetically
engineered to supply neurotrophic factors. Accordingly, in certain
embodiments of the invention, the therapeutic cells are
administered to the subject in combination with support cells. The
therapeutic cells may be combined with the support cells prior to
or during attachment to the support matrix, or they may be
separately attached to support matrices and combined immediately
before or during administration to the subject. If the therapeutic
cells and the support cells are separately adhered to support
matrices, the support matrices for the therapeutic cells and the
support cells may be the same or different. Additionally, support
cells may be combined with therapeutic cells and protective
cells.
Support Matrices
[0069] The support matrix is made of material which is preferably
non-toxic, for example, glass, polystyrene, polypropylene,
polyethylene, polycarbonate, polypentane, acrylonitrile polymer,
nylon, magnetite, natural polysaccharide, a modified
polysaccharide, collagen, gelatin and modified gelatin such as
crosslinked gelatin.
[0070] A preferred support matrix is a microcarrier of gelatin, as
described for example in U.S. Pat. No. 4,935,365. Suitable gelatin
microcarriers are commercially available as Cultispher.RTM. porous
microcarriers. These gelatin microcarriers have diameters ranging
from 80 .mu.m to 330 .mu.m. Particularly preferred in the practice
of the claimed invention are Cultispher-S.RTM., porous
microcarriers of crosslinked gelatin having a diameter between 80
.mu.m to 170 .mu.m (mean of 120 .mu.m). When using human RPE cells,
it has been determined that these gelatin microcarriers adhere, on
average, 24 cells per microcarrier, as compared to glass or
collagen carriers (e.g., Cytodex) which adhere approximately 5-7
cells per microcarrier.
[0071] It should be noted that the support matrix of the present
invention is not an encapsulating matrix or material. According to
the present invention, the cells are attached to or coating the
surface of the support; they are not encapsulated within a closed
compartment. When attached to the support according to the present
invention, the cells used for transplantation are generally on the
"outer surface" of the support. The support may be solid or porous.
However, even in a porous support, the cells are in direct contact
with the external milieu without an intervening membrane or other
barrier. Thus, according to the present invention, the cells are
considered to be on the "outer surface" of the support even though
the surface to which they adhere may be in the form of internal
folds or convolutions of the porous support material which are not
at the exterior of the particle or bead itself. It should be
further noted that the support matrix of the present invention
presents no requirement that the material have particular
permeability properties, such as the particular molecular weigh
"cut off".
[0072] The configuration of the support is preferably spherical, as
in a bead, but may be cylindrical, elliptical a flat sheet or
strip, a needle or pin shape, and the like. Bead sizes may range
from about 10 .mu.m to 200 .mu.m in diameter, preferably from about
90 to about 150 .mu.m, even more preferably around 100 .mu.m. For a
description of various microcarrier beads, see, for example. Fisher
Biotech Source 87-88. Fisher Scientific, Co., 1987, pp. 72-75;
Sigma Cell Culture Catalog, Sigma Chemical Co., St. Louis, 1991,
pp. 162-163; Ventrex Product Catalog, Ventrex Laboratories, 1989;
U.S. Pat. No. 4,935,365; these references are hereby incorporated
by reference.
[0073] To improve cell adhesion, survival and function, the solid
matrix may optionally be coated on its external surface with
factors known in the art to promote cell adhesion, growth or
survival. Such factors include cell adhesion molecules,
extracellular matrix, such as, for example, fibronectin, laminin,
collagen, elastin, glycosaminoglycans, or proteoglycans (see:
Albers, B. supra, pp. 802-834) or growth factors, such as, for
example, NGF. Alternatively, if the solid matrix to which the
implanted cells are attached is constructed of porous material, the
growth- or survival-promoting factor or factors may be incorporated
into the matrix material, from which they would be slowly released
after implantation in vivo.
Adhering Cells to a Support Matrix
[0074] The cells useful in the practice of the present invention
can be adhered to the support matrix by any methods known in the
art. Typically, cells can be adhered simply by incubating them with
the support matrix. Relative amounts and concentrations of cells
and microcarriers can be readily determined by those of skill in
the art, but in one preferred embodiment, cells and microcarriers
are mixed at a ratio of from about 5 to about 50 cells per
microcarrier.
[0075] In a preferred embodiment, Cultispher.RTM. microcarriers
(crosslinked gelatin) are incubated with therapeutic cells,
protective cells, support cells, or any combination thereof, under
conditions which promote binding of the cells to the microcarriers,
for example for at least approximately 4 hours while mixing or at
least approximately 15 hours without mixing, forming a cell-support
complex (therapeutic cells, protective cells, support cells, or any
combination thereof adhered to a support matrix). It should be
noted that microcarriers should be sterilized before incubation
with the cells, using a sterilization technique appropriate to the
microcarrier, as will be apparent to one of skill in the art.
[0076] The cell-support complex is preferably formulated as a
liquid suspension. The liquid may simply comprise a physiologically
and pharmaceutically acceptable isotonic buffer or may contain
additional components, including, but not limited to, nutrients,
vitamins, anti-oxidants, growth factors and neurotrophic factors.
In one preferred embodiment, the cell-support complex is formulated
in Hank's buffered saline solution (HBSS).
Administration of Cell/Support Complex to Subjects
[0077] Methods of delivering cells to a particular site in the
brain will be known to those of skill in the art. Preferably, the
cell-support complex is administered directly into a particular
site in the brain, for example, by injection or by implantation. To
treat the negative symptoms of schizophrenia, for example, the
cell-support complex may be administered into the prefrontal
cortex. For improvement of cognitive deficits associated with
schizophrenia, the cell-support complex is also administered to the
prefrontal cortex, particularly the dorsolateral prefrontal cortex.
EPS (e.g., tardive dyskinesia, tardive dystonia and/or tardive
akathisia) is treated by administration of a therapeutic
cell/support matrix complex to the striatal area of the subject's
brain.
[0078] Typically, the site(s) of interest is identified and, using
standard stereotaxic Atlas coordinates, or alternately using
coordinates developed from a magnetic resonance imaging (MRI) scan,
and the cell-support complex is injected. As will be apparent to
one of skill in the art, needles used for injection should be
selected to minimize clogging of the needle and minimize damage to
the tissue at the site of injection. Accordingly, the needle size
will depend on the size and shape of the support matrix.
[0079] The needle used for injection may be straight, beveled or
have a bent tip. If a beveled or bent tip needle is utilized, care
should be taken to correct the stereotaxic coordinates to
compensate for the tip of the needle.
[0080] Alternately, the cell-support complex may be implanted. If
implantation is used, the cell-support complex is preferably
formulated as a slurry or a pellet. The site for implantation is
identified, and the cell-support slurry or pellet is deposited at
the implantation site using standard surgical techniques.
[0081] The number of cells which are administered to a particular
subject will depend on a number of individual variables, as will be
apparent to one of skill in the art. The severity of the disorder,
the psychiatric and medical history of the subject, and the
properties of the therapeutic cells are all considered when
selecting the number and type of cells to administer to the
subject. Generally, about 10.sup.3-10.sup.7 therapeutic cells are
administered per site, preferably 10.sup.5-10.sup.6 cells per site,
in a total of from 1 to 25 sites, more preferably 5 to 20 sites for
treatment of the negative symptoms of schizophrenia, cognitive
deficits associated with schizophrenia or tardive dyskinesia.
[0082] Preferably, the cell-support complex is administered to a
subject suffering from the negative symptoms of schizophrenia,
cognitive deficits associated with schizophrenia, or EPS on a
single occasion (although the subject may receive the cell-support
complex at multiple sites on that single occasion), to minimize
trauma to the subject. However, as will be apparent to one of skill
in the art, the cell-support matrix may be delivered to the subject
in multiple doses. Preferably, any cell-support complex
administered subsequent to the first occasion is administered at
least one week, and preferably four weeks, after the previous
occasion of administration, to allow for assessment of the subject
after each administration.
[0083] Treatment of a subject suffering from the negative symptoms
of schizophrenia by administration of therapeutic cells to the
subject results in an improvement in the negative symptoms by, for
example, alleviation or elimination of one or more of the negative
symptoms, diminishment of the extent of the negative symptoms,
stabilization of the subject's negative symptoms or a delay or
slowing of progression in the negative symptoms. The subject's
negative symptoms of schizophrenia can be measured (before, during
or after the administration of therapeutic cells) using any
relevant measurement device, scale or method known in the art, such
as the PANSS, the SAPS, the SANS or the SCZI, preferably the
PANSS.
[0084] Treatment of a subject suffering from cognitive deficit(s)
associated with schizophrenia by administration of therapeutic
cells to the subject results in an alleviation of the cognitive
deficit(s) by, for example, improvement of cognitive function, or a
delay or slowing of increased cognitive deficit(s). Cognitive
function can be measured before and after treatment in accordance
with the invention, using any relevant test or scale known in the
art, including standard neuropsychological tests such as the
Wisconsin Card Sorting Test (WCST), the Mini-Mental State
Examination (MMSE), the Alzheimer's Disease Assessment Scale (ADAS)
and the Clinical Global Impression (CGI), but more preferably pen
and paper versions of tasks that activate Walker area 56 in
non-human primates such as the working memory test described by
Keefe et al. (1995, Schizophr. Res. 17(1):25-33).
[0085] Treatment of a subject suffering from an EPS such as tardive
dyskinesia, tardive dystonia and/or tardive akathisia by
administration of therapeutic cells to the subject results in an
improvement in the symptoms of the EPS by, for example, reduction
or elimination of undirected movements, reduction, elimination or
stabilization of dystonia at any particular affected site,
reduction, elimination or stabilization of akathisia-related
symptoms, stabilization of the subject's dyskinesia symptoms or a
delay or slowing of progression in the dyskinesia symptoms. The
subject's EPS can be measured (before, during or after the
administration of therapeutic cells) using any relevant measurement
device, scale or method known in the art, such as the Abnormal
Involuntary Movement Scale (AIMS), the Barnes Akathisia Scale
(BAS), the Modified Simpson Dyskinesia Scale (MSDS), and the
Simpson-Angus Extrapyramidal Effects Scale (S-AS), preferably the
AIMS.
[0086] The publications, patents, patent applications, and
published patent specifications referenced in this application are
hereby incorporated by reference in their entirety into the present
disclosure to more fully describe the state of the art to which
this invention pertains.
EXAMPLES
Example 1
Treatment of the Negative Symptoms of Schizophrenia by
Administration of RPE cells Adhered to a Support Matrix
[0087] Humans who have ingested PCP on a long term basis exhibit a
large number of symptoms associated with schizophrenia, such as
withdrawal, affective blunting, paranoia, delusions and
hallucinations. This observation is the basis of an art accepted
model of schizophrenia, which utilizes long term administration of
phencyclidine (PCP) to vervet monkeys (Jentsch et al., supra)
[0088] Vervet monkeys (Cercopithecus aethiops sabaeus) are divided
into three groups. The first group ("Control") are given
intramuscular (IM) saline injections bid, for 14 days. The second
and third groups ("PCP-" and "PCP+") are given 0.3 mg/kg PCP by IM
injection, bid, for 14 days.
[0089] Human RPE cells are prepared according to the method of Lui
et al. (1990, Investigative Ophthamol., ARVO Supplement). The cells
are adhered to crosslinked gelatin microspheres (Cultispher-S.RTM.,
mean diameter 120.mu.m) by mixing microspheres with cells suspended
in culture medium (Dulbecco's modified essential medium (DMEM) with
10% fetal calf serum (FCS)) then incubating the mixture overnight
at 37.degree. C. in a 5% CO.sub.2 atmosphere to form the
cell-support complex. The cell-support complex is rinsed and
resuspended in HBSS.
[0090] Cell-support complex is administered to PCP+ monkeys at 10
sites in the prefrontal cortex by stereotaxic injection.
Approximately 5.times.10.sup.5 cells are administered per site.
Control and PCP- monkeys receive injections of an equal volume of
the support matrix suspended HBSS at 10 sites in the prefrontal
cortex by stereotactic injection.
[0091] After allowing the animals to recover from the surgery for
one to two weeks, all monkeys are evaluated by testing prefrontal
cognitive function using "object retrieval with detour task"
testing. In such testing, the monkeys are trained to recover a
reward from a transparent box with a single opening. The monkeys
are scored on first try success rate (recovering the reward through
the opening on the first try), barrier reaches (number of reaches
toward the reward on a closed side of the box), and perserveration
(number of repeat attempts to reach the reward through a closed
side).
Example 2
Treatment of Tardive Dyskinesia by Administration of RPE Cells
Adhered to a Support Matrix
[0092] Long term administration of Cebus monkeys with certain
antipsychotic drugs provides a model for human EPS associated with
antipsychotic drug therapy, particularly TD (Kovacic et al., 1982,
J. Clin. Psychopharm. 2(5):305-307). Cebus monkeys treated with
long term administration of fluphenazine exhibit classic TD
symptoms, including rhythmic finger movements, movements analogous
to human pacing, and oral symptoms such as rhythmic up and down
motions of the jaw and tongue movements.
[0093] Cebus apella monkeys are treated to produce a model of TD,
by long term administration of depot fluphenazine enanthate.
Fluphenazine is administered biweekly, progressing from a low dose
(0.1 mg/kg) to 3.2 mg/kg by six months. Treatment is continued for
at least one year.
[0094] Activity of the monkeys is observed, with particularly
attention to locomotor activity and signs of TD, to establish
baseline levels of EPS. The monkeys are split into two groups:
control and treatment.
[0095] Human RPE cells are prepared according to the method of Lui
et al. (1990, Investigative Ophthamol., ARVO Supplement). The cells
are adhered to crosslinked gelatin microspheres (Cultispher-S.RTM.,
mean diameter 120 .mu.m) by mixing microspheres with cells
suspended in culture medium (Dulbecco's modified essential medium
(DMEM) with 10% fetal calf serum (FCS)) then incubating the mixture
overnight at 37.degree. C. in a 5% CO.sub.2 atmosphere to form the
cell-support complex. The cell-support complex is rinsed and
resuspended in HBSS.
[0096] The cell-support complex is administered by stereotaxic
injection to the striatum of treatment group monkeys, administering
approximately 5.times.10.sup.5 cells are administered per site at a
total of 10 sites. Control group monkeys receive injections of an
equal volume of the support matrix suspended HBSS at 10 sites in
the striatum by stereotactic injection.
[0097] Following a one to two week recovery period, control and
treatment group monkeys are evaluated for locomotor activity and
signs of TD.
[0098] The present invention has been detailed both by direct
description and by example. Equivalents and modifications of the
present invention will be apparent to those skilled in the art, and
are encompassed within the scope of the invention.
* * * * *