U.S. patent application number 12/187896 was filed with the patent office on 2009-05-14 for slow intraventricular delivery.
This patent application is currently assigned to GENZYME CORPORATION. Invention is credited to Seng H. Cheng, James Dodge, Marco A. Passini, Lamya Shihabuddin.
Application Number | 20090123451 12/187896 |
Document ID | / |
Family ID | 38372004 |
Filed Date | 2009-05-14 |
United States Patent
Application |
20090123451 |
Kind Code |
A1 |
Dodge; James ; et
al. |
May 14, 2009 |
SLOW INTRAVENTRICULAR DELIVERY
Abstract
Neurological diseases, including lysosomal storage diseases, can
be successfully treated using intraventricular delivery of the
therapeutic agents to bypass the blood-brain barrier. Similarly,
diagnostic agents and anesthetic agents can be delivered to the
brain in this manner. The administration can be performed slowly to
achieve maximum effect. Such administration permits greater
penetration of distal portions of the brain.
Inventors: |
Dodge; James; (Worcester,
MA) ; Passini; Marco A.; (Shrewsbury, MA) ;
Shihabuddin; Lamya; (Brighton, MA) ; Cheng; Seng
H.; (Natick, MA) |
Correspondence
Address: |
GENZYME CORPORATION;LEGAL DEPARTMENT
15 PLEASANT ST CONNECTOR
FRAMINGHAM
MA
01701-9322
US
|
Assignee: |
GENZYME CORPORATION
Cambridge
MA
|
Family ID: |
38372004 |
Appl. No.: |
12/187896 |
Filed: |
August 7, 2008 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
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PCT/US2007/003382 |
Feb 8, 2007 |
|
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12187896 |
|
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60771451 |
Feb 9, 2006 |
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Current U.S.
Class: |
424/94.61 ;
424/94.1 |
Current CPC
Class: |
A61P 1/16 20180101; A61P
25/08 20180101; A61K 9/0024 20130101; A61P 25/28 20180101; A61P
3/10 20180101; A61K 31/00 20130101; A61P 3/00 20180101; A61P 25/14
20180101; A61P 43/00 20180101; A61P 3/08 20180101; A61P 25/00
20180101; A61P 25/16 20180101; A61P 23/00 20180101; A61P 11/00
20180101; A61P 13/12 20180101 |
Class at
Publication: |
424/94.61 ;
424/94.1 |
International
Class: |
A61K 38/43 20060101
A61K038/43; A61P 43/00 20060101 A61P043/00 |
Claims
1. A method of delivering an agent to a patient's brain, the method
comprising: administering the agent to the patient via a lateral
ventricle of the brain at a rate such that the administration of a
single dose consumes more than two hours.
2. A method of delivering an agent to a patient's brain, the method
comprising: administering the agent to the patient via a lateral
ventricle of the brain at a rate such that the administration of a
single dose consumes at least 50% of the turn-over time of the
cerebrospinal fluid in the patient.
3. A method for delivering an agent to a patient's brain, the
method comprising: estimating turn-over time of cerebrospinal fluid
of the patient; selecting a rate and a total delivery time for an
agent via a lateral ventricle of the brain based on the turn-over
time; setting a pump to deliver the agent at said selected rate for
said total delivery time.
4. A method of delivering an agent to a patient's brain, the method
comprising: estimating turn-over time of cerebrospinal fluid of the
patient; selecting a rate and a total delivery time for an agent
via a lateral ventricle of the brain based on the turn-over time;
delivering the agent to the patient at said selected rate for said
total delivery time.
5. A method of delivering an agent to a patient's brain, the method
comprising: administering the agent to the patient via a lateral
ventricle of the brain at a rate such that the administration of a
single dose continues at least until the agent is detectable in
serum of the patient.
6. The method of claim 3 wherein the rate delivers a single dose of
the agent for a time greater than or equal to 50% of the estimated
turn-over time.
7. The method of claim 4 wherein the rate delivers a single dose of
the agent for a time greater than or equal to 50% of the estimated
turn-over time.
8. The method of claim 3 wherein the rate delivers a single dose of
the agent for a time greater than or equal to 100% of the estimated
turn-over time.
9. The method of claim 4 wherein the rate delivers a single dose of
the agent for a time greater than or equal to 100% of the estimated
turn-over time.
10. The method of claim 3 wherein the rate delivers a single dose
of the agent for a time greater than or equal to 150% of the
estimated turn-over time.
11. The method of claim 4 wherein the rate delivers a single dose
of the agent for a time greater than or equal to 150% of the
estimated turn-over time.
12. The method of claim 2 wherein the administration consumes at
least 100% of the turn-over time.
13. The method of claim 2 wherein the administration consumes at
least 150% of the turn-over time.
14. The method of claim 2 wherein the administration consumes at
least 200% of the turn-over time.
15. The method of claim 2 wherein the administration consumes at
least 250% of the turn-over time.
16. The method of claim 2 wherein the agent accesses the third
ventricle.
17. The method of claim 2 wherein the agent accesses the Aqueduct
of Sylvius.
18. The method of claim 2 wherein the agent accesses the fourth
ventricle.
19. The method of claim 2 wherein the agent accesses Foramina of
Lushka.
20. The method of claim 2 wherein the agent accesses the Foramina
of Magendie.
21. The method of claim 2 wherein the agent accesses the spinal
cord.
22. The method of claim 2, wherein the agent accesses the
subarachnoid space.
23. The method of claim 2 wherein the agent accesses the serum.
24-26. (canceled)
27. The method of claim 1, 2, 3, 4, or 5 wherein the agent is an
enzyme.
28. The method of claim 1, 2, 3, 4, or 5 wherein the agent is an
enzyme that is deficient in a lysosomal storage disease.
29-31. (canceled)
32. The method of claim 2 wherein the agent is
sphingomyelinase.
33. The method of claim 2 wherein the patient has Niemann-Pick B
disease.
34. The method of claim 2 wherein the agent is
alpha-L-iduronidase.
35. The method of claim 2 wherein the patient has Hurler
syndrome.
36. The method of claim 2 wherein the patient has Gaucher's
disease.
37. The method of claim 2 wherein the agent is
glucocerebrosidase.
38. The method of claim 2, wherein the patient has Fabry
disease
39. The method of claim 2, wherein the agent is alpha-galactosidase
A.
40. The method of claim 2 wherein the patient has Pompe
disease.
41. The method of claim 2 wherein the agent is acid maltase.
42. The method of claim 2, wherein the patient has Tay-Sachs
disease.
43. The method of claim 2 wherein the agent is hexosaminidase.
44. The method of claim 2 wherein the patient has Glycogen storage
disease type II.
45. The method of claim 2 wherein the agent is
alpha-glucosidase.
46. The method of claim 1 wherein the rate is such that the
administration of a single dose consumes more than four hours.
47. The method of claim 1 wherein the rate is such that the
administration of a single dose consumes more than six hours.
48. The method of claim 1 wherein the rate is such that the
administration of a single dose consumes more than eight hours.
49. The method of claim 1 wherein the rate is such that the
administration of a single dose consumes more than ten hours.
50. The method of claim 2 wherein the agent is delivered using a
catheter.
51. The method of claim 2 wherein the agent is delivered using a
pump.
52. The method of claim 2 wherein the agent is delivered using an
implantable pump.
53. (canceled)
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application is a continuation of PCT Application No.
PCT/US2007/03382, filed Feb. 8, 2007, which claims priority under
35 U.S.C. .sctn. 119 (e) to U.S. Provisional Application Ser. No.
60/771,451 filed Feb. 9, 2006 the contents of which are hereby
incorporated by reference into the present disclosure in their
entirety.
TECHNICAL FIELD OF THE INVENTION
[0002] This invention is related to delivery of agents to the
brain. In particular it relates to brain diagnosis, treatment, and
imaging.
[0003] A group of metabolic disorders known as lysosomal storage
diseases (LSD) includes over forty genetic disorders, many of which
involve genetic defects in various lysosomal hydrolases.
Representative lysosomal storage diseases and the associated
defective enzymes are listed in Table 1.
TABLE-US-00001 TABLE 1 Lysosomal storage disease Defective enzyme
Aspartylglucosaminuria Aspartylglucosaminidase Fabry
.alpha.-Galactosidase A Infantile Batten Disease* (CNL1) Palmitoyl
Protein Thioesterase Classic Late Infantile Batten Disease* (CNL2)
Tripeptidyl Peptidase Juvenile Batten Disease* (CNL3) Lysosomal
Transmembrane Protein Batten, other forms* (CNL4- CNL8) Multiple
gene products Cystinosis Cysteine transporter Farber Acid
ceramidase Fucosidosis Acid .alpha.-L-fucosidase
Galactosidosialidosis Protective protein/cathepsin A Gaucher types
1, 2*, and 3* Acid .beta.-glucosidase, or G.sub.M1 gangliosidosis*
Acid .beta.-galactosidase Hunter* Iduronate-2-sulfatase
Hurler-Scheie* .alpha.-L-Iduronidase Krabbe* Galactocerebrosidase.
alpha.-Mannosidosis* Acid .alpha.-mannosidase. beta.-Mannosidosis*
Acid .beta.-mannosidase Maroteaux-Lamy Arylsulfatase B
Metachromatic leukodystrophy* Arylsulfatase A Morquio A
N-Acetylgalactosamine-6-sulfate Morquio B Acid .beta.-galactosidase
Mucolipidosis II/III* N-Acetylglucosamine-1- Niemann-Pick A*, B
Acid sphingomyelinase Niemann-Pick C* NPC-1 Pompe* Acid
.alpha.-glucosidase Sandhoff* .beta.-Hexosaminidase B Sanfilippo A*
Heparan N-sulfatase Sanfilippo B* .alpha.-N-Acetylglucosaminidase
Sanfilippo C* Acetyl-CoA: alpha.-glucosaminide Sanfilippo D*
N-Acetylglucosamine-6-sulfate Schindler Disease*
.alpha.-N-Acetylgalactosaminidase Schindler-Kanzaki.
alpha.-N-Acetylgalactosaminidase Sialidosis .alpha.-Neuramidase
Sly* .beta.-Glucuronidase Tay-Sachs* .beta.-Hexosaminidase A
Wolman* Acid Lipase *CNS involvement
[0004] The hallmark feature of LSD is the abnormal accumulation of
metabolites in the lysosomes which leads to the formation of large
numbers of distended lysosomes in the perikaryon. A major challenge
to treating LSD (as opposed to treating a liver-specific
enzymopathy) is the need to reverse lysosomal storage pathology in
multiple separate tissues. Some LSDs can be effectively treated by
intravenous infusion of the missing enzyme, known as enzyme
replacement therapy (ERT). For example, Gaucher type 1 patients
have only visceral disease and respond favorably to ERT with
recombinant glucocerebrosidase (Cerezyme.TM., Genzyme Corp.).
However, patients with metabolic disease that affects the CNS
(e.g., type 2 or 3 Gaucher disease) partially respond to
intravenous ERT because the replacement enzyme is prevented from
entering the brain by the blood brain barrier (BBB). Furthermore,
attempts to introduce a replacement enzyme into the brain by direct
injection have been limited in part due to enzyme cytotoxicity at
high local concentrations and limited parenchymal diffusion rates
in the brain (Partridge, Peptide Drug Delivery to the Brain, Raven
Press, 1991).
[0005] According to UniProtKB/Swiss-Prot entry P17405, defects in
the SMPD1 gene, located on chromosome 11, (11p 15.4-p 15.1), are
the cause of Niemann-Pick disease type A (NPA), also referred to as
the classical infantile form. Niemann-Pick disease is a clinically
and genetically heterogeneous recessive disorder. It is caused by
the accumulation of sphingomyelin and other metabolically related
lipids in the lysosomes, resulting in neurodegeneration starting
from early life. Patients may show xanthomas, pigmentation,
hepatosplenomegaly, lymphadenopathy and mental retardation.
Niemann-Pick disease occurs more frequently among individuals of
Ashkenazi Jewish ancestry than in the general population. NPA is
characterized by very early onset in infancy and a rapidly
progressive course leading to death by three years. The acid
sphingomyelinase enzyme (aSM) converts sphingomyelin to ceramide.
aSM also has phospholipase C activities toward
1,2-diacylglycerolphosphocholine and
1,2-diacylglycerolphosphoglycerol. The enzyme converts
Sphingomyelin+H.sub.2O.fwdarw.N-acylsphingosine+choline
phosphate.
[0006] There is a continuing need in the art for methods to treat
LSDs that have both cerebral and visceral disease pathologies.
There is a continuing need in the art for methods to access
portions of the brain with diagnostic and therapeutic agents that
do not readily cross the blood-brain barrier.
SUMMARY OF THE INVENTION
[0007] According to one embodiment of the invention a method is
provided of delivering an agent to a patient's brain. The agent is
administered to the patient via a lateral ventricle of the brain at
a rate such that the administration of a single dose consumes more
than two hours.
[0008] According to another embodiment of the invention a method is
provided of delivering an agent to a patient's brain. The agent is
administered to the patient via a lateral ventricle of the brain at
a rate such that the administration of a single dose consumes at
least 50% of the turn-over time of the cerebrospinal fluid in the
patient.
[0009] According to still another embodiment of the invention a
method is provided of delivering an agent to a patient's brain.
Turn-over time of cerebrospinal fluid of the patient is estimated.
A rate for delivery and a total delivery time of the agent via a
lateral ventricle of the brain is selected based on the turn-over
time. A pump is set to deliver the agent at said selected rate for
said total delivery time.
[0010] According to yet another embodiment of the invention a
method is provided of delivering an agent to a patient's brain.
Turn-over time of cerebrospinal fluid of the patient is estimated.
A rate and a total delivery time is selected for delivery of the
agent via a lateral ventricle of the brain based on the turn-over
time. The agent is delivered to the patient at said selected rate
for said total delivery time.
[0011] According to another aspect of the invention a method is
provided of delivering an agent to a patient's brain. The agent is
administered to the patient via a lateral ventricle of the brain at
a rate such that the administration of a single dose continues at
least until the agent is detectable in serum of the patient.
[0012] According to one embodiment of the invention a patient with
Niemann-Pick A or B disease is treated. An acid sphingomyelinase is
administered to the patient via intraventricular delivery to the
brain in an amount sufficient to reduce sphingomyelin levels in
said brain.
[0013] Another aspect of the invention is a kit for treating a
patient with Niemann-Pick A or B disease. The kit comprises an acid
sphingomyelinase, and a catheter for delivery of said acid
sphingomyelinase to the patient's brain ventricles.
[0014] Yet another aspect of the invention is a kit for treating a
patient with Niemann-Pick A or B disease. The kit comprises an acid
sphingomyelinase and a pump for delivery of said acid
sphingomyelinase to the patient's brain ventricles.
[0015] According to the invention a patient can be treated who has
a lysosomal storage disease which is caused by an enzyme deficiency
which leads to accumulation of the enzyme's substrate. The enzyme
is administered to the patient via intraventricular delivery to the
brain. The rate of administration is such that the administration
of a single dose consumes more than four hours. Substrate levels in
said brain are thereby reduced.
[0016] These and other embodiments which will be apparent to those
of skill in the art upon reading the specification provide the art
with methods for delivering agents to hard-to-reach portions of the
brain.
BRIEF DESCRIPTION OF THE DRAWINGS
[0017] FIG. 1 shows diagram of sections of brain that were analyzed
for sphingomyelin. S1 is at the front of brain and S5 is at the
back of brain.
[0018] FIG. 2 shows that intraventricular administration of rhASM
reduces SPM levels in the ASMKO mouse brain.
[0019] FIG. 3 shows intraventricular administration of rhASM
reduces SPM levels in the ASMKO liver, spleen, and lung.
[0020] FIG. 4 shows hASM staining in the brain following
intraventricular infusion.
[0021] FIG. 5 shows that intraventricular infusion of rhASM over a
6 h period reduces SPM levels in the ASMKO mouse brain.
[0022] FIG. 6 shows that intraventricular infusion of rhASM over a
6 h period reduces SPM levels in ASMKO liver, serum, and lung.
[0023] FIG. 7 shows documented hASM variants and their relationship
to disease or enzyme activity.
[0024] FIG. 8 shows the ventricular system which bathes the entire
brain and spinal cord with cerebrospinal fluid.
DETAILED DESCRIPTION OF THE INVENTION
[0025] The inventors have discovered that intraventricular delivery
of agents to patients at a slow rate, rather than in a bolus
delivery, increases the effective penetration of the agents to
distal portions of the brain from the site of introduction. Agents
which can be administered in this manner are any, but include
diagnostic agents, imaging agents, anesthetic agents, and
therapeutic agents. This mode of delivery is particularly useful
for agents which cannot cross the blood-brain barrier.
[0026] Applicants have observed that bolus intraventricular
administration is not very effective, whereas slow infusion is very
effective. While applicants do not wish to be bound by any
particular theory of operation, it is believed that the slow
infusion is effective due to the turn-over of the cerebrospinal
fluid (CSF). While estimates and calculations in the literature
vary, the adult human cerebrospinal fluid is believed to turn over
within about 4, 5, 6, 7, or 8 hours. The turn-over rate may vary
depending on the size of the individual and the volume of
cerebrospinal fluid in the individual. Thus for example, children
have less cerebrospinal fluid than adults and therefore have a
shorter turn-over time. The slow infusion of the invention can be
metered so that the delivery time is about equal to or greater than
the turn-over time of the CSF. The metering can be a fixed time,
for example, greater than 2, 4, 6, 8, or 10 hours, or it can be set
to be a fraction of the estimated turn-over time, for example
greater than 50%, 75%, 100%, 150%, 200%, 300%, or 400%. The CSF
empties into the venous blood system. The delivery can be performed
for a time until the delivered agent is detectable in the serum of
the patient. One can also detect and/or measure delivered agent in
other parts of the CNS such as in the spinal cord and the
subarachnoid space. These, too, can be used as endpoints for
delivery.
[0027] CSF is secreted at a rate of about 430 to 600 ml/day or
about 0.35 to 0.4 per minute in adults and the volume at any given
moment is approximately 80 to 150 ml, with the entire volume being
replaced every six to eight hours. Infants are estimated to produce
0.15 ml per minute. The choroid plexuses of the lateral ventricals
are the largest and produce most of the CSF. The fluid flows
through the intraventricular foramina in the third ventricle, is
augmented by fluid formed in the choroid plexus of that ventricle,
and passes through the cerebral aqueduct of Sylvius to the fourth
ventricle. CSF flows from 4th ventricle to foramen of Magendie to
the sub-arachnoid space that surrounds the spinal cord; CSF flows
from 4th ventricle to the foramen of Lushka to the sub-arachnoid
space that surrounds the brain. The arachnoid membrane lines the
sub-arachnoid space; arachnoid villi are part of the membrane.
Arachnoid villi are pumps that take in the CSF and return it to the
venous circulation. The CSF is reabsorbed into the blood through
the arachnoid villi.
[0028] Slower-than-bolus delivery according to the invention has
the advantage of delivering agents to portions of the brain that
are not reached with a bolus. Bolus-delivered agent accumulates in
the ependymal layer or in the parenchyma adjacent to the injection
site. In contrast, slow-delivered agents are found to access distal
regions of the parenchyma from the injection site (widespread
delivery across the anterior-to-posterior axis of the brain; in
addition widespread delivery dorsally and ventrally to the
ependymal layer), the third ventricle, the Aqueduct of Sylvius, the
fourth ventricle, the Foramina of Lushka, the Foramina of Magendie,
the spinal cord, the subarachnoid space, and the serum. From the
serum, peripheral organs can also be reached.
[0029] The CSF empties into the blood via the arachnoid villi and
intracranial vascular sinuses, thereby delivering the enzymes to
the visceral organs. The visceral organs which are often affected
in Niemann-Pick disease are the lungs, spleen, kidney, and liver.
The slow intraventricular infusion provides diminished amounts of
substrate in at least these visceral organs.
[0030] The reduction in substrate accumulated in the brain, lungs,
spleen, kidney, and/or liver is dramatic. Reductions of greater
that 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% can be achieved.
The reduction achieved is not necessarily uniform from patient to
patient or even from organ to organ within a single patient.
[0031] Agents for delivery can be any that are known in the art for
treatment and imaging of brains. Imaging agents can be radioactive,
radio-opaque, fluorescent, etc. Therapeutic agents can be any that
are useful for treating neurological or other brain diseases.
Anesthetics can be for treating chronic or acute pain, for example
lidocaine hydrochloride and morphine. Examples of therapeutic
agents include the enzymes that are deficient in lysosomal storage
diseases. Other possible agents for use include nucleic acid
vectors, such as plasmid and viral vectors, siRNA, anti-sense RNAs,
etc. Other therapeutic agents include those which increase or
decrease excitation of neurons in the brain. These include agonists
or antagonists of glutamate, GABA, and dopamine. Specific examples
include cycloserine, carboxyphenylglycine, glutamic acid,
dizocilpine, ketamaine, dextromethorphan, baclofen, muscinol,
gabazine, saclofen, haloperidol, and methane sulfonate. Additional
agents which can be used are anti-inflammatory agents, in
particular non-steroidal anti-inflammatory agents such as
indomethacin and cyclooxygenase inhibitors.
[0032] Nucleic acids can be delivered in any desired vector. These
include viral or non-viral vectors, including adenovirus vectors,
adeno-associated virus vectors, retrovirus vectors, lentivirus
vectors, and plasmid vectors. Exemplary types of viruses include
HSV (herpes simplex virus), AAV (adeno associated virus), HIV
(human immunodeficiency virus), BW (bovine immunodeficiency virus),
and MLV (murine leukemia virus). Nucleic acids can be administered
in any desired format that provides sufficiently efficient delivery
levels, including in virus particles, in liposomes, in
nanoparticles, and complexed to polymers.
[0033] Adenovirus is a non-enveloped, nuclear DNA virus with a
genome of about 36 kb, which has been well-characterized through
studies in classical genetics and molecular biology (Hurwitz, M.
S., Adenoviruses Virology, 3.sup.rd edition, Fields et al., eds.,
Raven Press, New York, 1996; Hitt, M. M. et al., Adenovirus
Vectors, The Development of Human Gene Therapy, Friedman, T. ed.,
Cold Spring Harbor Laboratory Press, New York 1999). The viral
genes are classified into early (designated E1-E4) and late
(designated L1-L5) transcriptional units, referring to the
generation of two temporal classes of viral proteins. The
demarcation of these events is viral DNA replication. The human
adenoviruses are divided into numerous serotypes (approximately 47,
numbered accordingly and classified into 6 groups: A, B, C, D, E
and F), based upon properties including hemaglutination of red
blood cells, oncogenicity, DNA and protein amino acid compositions
and homologies, and antigenic relationships.
[0034] Recombinant adenoviral vectors have several advantages for
use as gene delivery vehicles, including tropism for both dividing
and non-dividing cells, minimal pathogenic potential, ability to
replicate to high titer for preparation of vector stocks, and the
potential to carry large inserts (Berkner, K. L., Curr. Top. Micro.
Immunol. 158:39-66, 1992; Jolly, D., Cancer Gene Therapy 1:51-64
1994). Adenoviral vectors with deletions of various adenoviral gene
sequences, such as pseudoadenoviral vectors (PAVs) and
partially-deleted adenoviral (termed "DeAd"), have been designed to
take advantage of the desirable features of adenovirus which render
it a suitable vehicle for delivery of nucleic acids to recipient
cells.
[0035] In particular, pseudoadenoviral vectors (PAVs), also known
as `gutless adenovirus` or mini-adenoviral vectors, are adenoviral
vectors derived from the genome of an adenovirus that contain
minimal cis-acting nucleotide sequences required for the
replication and packaging of the vector genome and which can
contain one or more transgenes (See, U.S. Pat. No. 5,882,877 which
covers pseudoadenoviral vectors (PAV) and methods for producing
PAV, incorporated herein by reference). PAVs have been designed to
take advantage of the desirable features of adenovirus which render
it a suitable vehicle for gene delivery. While adenoviral vectors
can generally carry inserts of up to 8 kb in size by the deletion
of regions which are dispensable for viral growth, maximal carrying
capacity can be achieved with the use of adenoviral vectors
containing deletions of most viral coding sequences, including
PAVs. See U.S. Pat. No. 5,882,877 of Gregory et al.; Kochanek et
al., Proc. Natl. Acad. Sci. USA 93:5731-5736, 1996; Parks et al.,
Proc. Natl. Acad. Sci. USA 93:13565-13570, 1996; Lieber et al., J.
Virol 70:8944-8960, 1996; Fisher et al., Virology 217:11-22, 1996;
U.S. Pat. No. 5,670,488; PCT Publication No. WO96/33280, published
Oct. 24, 1996; PCT Publication No. WO96/40955, published Dec. 19,
1996; PCT Publication No. WO97/25446, published Jul. 19, 1997; PCT
Publication No. WO95/29993, published Nov. 9, 1995; PCT Publication
No. WO97/00326, published Jan. 3, 1997; Morral et al., Hum. Gene
Ther. 10:2709-2716, 1998. Such PAVs, which can accommodate up to
about 36 kb of foreign nucleic acid, are advantageous because the
carrying capacity of the vector is optimized, while the potential
for host immune responses to the vector or the generation of
replication-competent viruses is reduced. PAV vectors contain the
5' inverted terminal repeat (ITR) and the 3' ITR nucleotide
sequences that contain the origin of replication, and the
cis-acting nucleotide sequence required for packaging of the PAV
genome, and can accommodate one or more transgenes with appropriate
regulatory elements, e.g. promoter, enhancers, etc.
[0036] Other, partially deleted adenoviral vectors provide a
partially-deleted adenoviral (termed "DeAd") vector in which the
majority of adenoviral early genes required for virus replication
are deleted from the vector and placed within a producer cell
chromosome under the control of a conditional promoter. The
deletable adenoviral genes that are placed in the producer cell may
include E1A/E1B, E2, E4 (only ORF6 and ORF6/7 need be placed into
the cell), pIX and pIVa2. E3 may also be deleted from the vector,
but since it is not required for vector production, it can be
omitted from the producer cell. The adenoviral late genes, normally
under the control of the major late promoter (MLP), are present in
the vector, but the MLP may be replaced by a conditional
promoter.
[0037] Conditional promoters suitable for use in DeAd vectors and
producer cell lines include those with the following
characteristics: low basal expression in the uninduced state, such
that cytotoxic or cytostatic adenovirus genes are not expressed at
levels harmful to the cell; and high level expression in the
induced state, such that sufficient amounts of viral proteins are
produced to support vector replication and assembly. Preferred
conditional promoters suitable for use in DeAd vectors and producer
cell lines include the dimerizer gene control system, based on the
immunosuppressive agents FK506 and rapamycin, the ecdysone gene
control system and the tetracycline gene control system. Also
useful in the present invention may be the GeneSwitch.TM.
technology (Valentis, Inc., Woodlands, Tex.) described in Abruzzese
et al., Hum. Gene Ther. 1999 10:1499-507, the disclosure of which
is hereby incorporated herein by reference. The partially deleted
adenoviral expression system is further described in WO99/57296,
the disclosure of which is hereby incorporated by reference
herein.
[0038] Adeno-associated virus (AAV) is a single-stranded human DNA
parvovirus whose genome has a size of 4.6 kb. The AAV genome
contains two major genes: the rep gene, which codes for the rep
proteins (Rep 76, Rep 68, Rep 52, and Rep 40) and the cap gene,
which codes for AAV replication, rescue, transcription and
integration, while the cap proteins form the AAV viral particle.
AAV derives its name from its dependence on an adenovirus or other
helper virus (e.g., herpesvirus) to supply essential gene products
that allow AAV to undergo a productive infection, i.e., reproduce
itself in the host cell. In the absence of helper virus, AAV
integrates as a provirus into the host cell's chromosome, until it
is rescued by superinfection of the host cell with a helper virus,
usually adenovirus (Muzyczka, Curr. Top. Micor. Immunol.
158:97-127, 1992).
[0039] Interest in AAV as a gene transfer vector results from
several unique features of its biology. At both ends of the AAV
genome is a nucleotide sequence known as an inverted terminal
repeat (ITR), which contains the cis-acting nucleotide sequences
required for virus replication, rescue, packaging and integration.
The integration function of the ITR mediated by the rep protein in
trans permits the AAV genome to integrate into a cellular
chromosome after infection, in the absence of helper virus. This
unique property of the virus has relevance to the use of AAV in
gene transfer, as it allows for a integration of a recombinant AAV
containing a gene of interest into the cellular genome. Therefore,
stable genetic transformation, ideal for many of the goals of gene
transfer, may be achieved by use of rAAV vectors. Furthermore, the
site of integration for AAV is well-established and has been
localized to chromosome 19 of humans (Kotin et al., Proc. Natl.
Acad. Sci. 87:2211-2215, 1990). This predictability of integration
site reduces the danger of random insertional events into the
cellular genome that may activate or inactivate host genes or
interrupt coding sequences, consequences that can limit the use of
vectors whose integration of AAV, removal of this gene in the
design of rAAV vectors may result in the altered integration
patterns that have been observed with rAAV vectors (Ponnazhagan et
al., Hum Gene Ther. 8:275-284, 1997).
[0040] There are other advantages to the use of AAV for gene
transfer. The host range of AAV is broad. Moreover, unlike
retroviruses, AAV can infect both quiescent and dividing cells. In
addition, AAV has not been associated with human disease, obviating
many of the concerns that have been raised with retrovirus-derived
gene transfer vectors.
[0041] Standard approaches to the generation of recombinant rAAV
vectors have required the coordination of a series of intracellular
events: transfection of the host cell with an rAAV vector genome
containing a transgene of interest flanked by the AAV ITR
sequences, transfection of the host cell by a plasmid encoding the
genes for the AAV rep and cap proteins which are required in trans,
and infection of the transfected cell with a helper virus to supply
the non-AAV helper functions required in trans (Muzyczka, N., Curr.
Top. Micor. Immunol. 158:97-129, 1992). The adenoviral (or other
helper virus) proteins activate transcription of the AAV rep gene,
and the rep proteins then activate transcription of the AAV cap
genes. The cap proteins then utilize the ITR sequences to package
the rAAV genome into an rAAV viral particle. Therefore, the
efficiency of packaging is determined, in part, by the availability
of adequate amounts of the structural proteins, as well as the
accessibility of any cis-acting packaging sequences required in the
rAAV vector genome.
[0042] Retrovirus vectors are a common tool for gene delivery
(Miller, Nature (1992) 357:455-460). The ability of retrovirus
vectors to deliver an unrearranged, single copy gene into a broad
range of rodent, primate and human somatic cells makes retroviral
vectors well suited for transferring genes to a cell.
[0043] Retroviruses are RNA viruses wherein the viral genome is
RNA. When a host cell is infected with a retrovirus, the genomic
RNA is reverse transcribed into a DNA intermediate which is
integrated very efficiently into the chromosomal DNA of infected
cells. This integrated DNA intermediate is referred to as a
provirus. Transcription of the provirus and assembly into
infectious virus occurs in the presence of an appropriate helper
virus or in a cell line containing appropriate sequences enabling
encapsidation without coincident production of a contaminating
helper virus. A helper virus is not required for the production of
the recombinant retrovirus if the sequences for encapsidation are
provided by co-transfection with appropriate vectors.
[0044] The retroviral genome and the proviral DNA have three genes:
the gag, the pol, and the env, which are flanked by two long
terminal repeat (LTR) sequences. The gag gene encodes the internal
structural (matrix, capsid, and nucleocapsid) proteins; the pol
gene encodes the RNA-directed DNA polymerase (reverse
transcriptase) and the env gene encodes viral envelope
glycoproteins. The 5' and 3' LTRs serve to promote transcription
and polyadenylation of the virion RNAs. The LTR contains all other
cis-acting sequences necessary for viral replication. Lentiviruses
have additional genes including vit vpr, tat, rev, vpu, nef, and
vpx (in HIV-1, HIV-2 and/or SIV). Adjacent to the 5' LTR are
sequences necessary for reverse transcription of the genome (the
tRNA primer binding site) and for efficient encapsidation of viral
RNA into particles (the Psi site). If the sequences necessary for
encapsidation (or packaging of retroviral RNA into infectious
virions) are missing from the viral genome, the result is a cis
defect which prevents encapsidation of genomic RNA. However, the
resulting mutant is still capable of directing the synthesis of all
varion proteins.
[0045] Lentiviruses are complex retroviruses which, in addition to
the common retroviral genes gag, pol and env, contain other genes
with regulatory or structural function. The higher complexity
enables the lentivirus to modulate the life cycle thereof, as in
the course of latent infection. A typical lentivirus is the human
immunodeficiency virus (HIV), the etiologic agent of AIDS. In vivo,
HIV can infect terminally differentiated cells that rarely divide,
such as lymphocytes and macrophages. In vitro, HIV can infect
primary cultures of monocyte-derived macrophages (MDM) as well as
HeLa-Cd4 or T lymphoid cells arrested in the cell cycle by
treatment with aphidicolin or gamma irradiation. Infection of cells
is dependent on the active nuclear import of HIV preintegration
complexes through the nuclear pores of the target cells. That
occurs by the interaction of multiple, partly redundant, molecular
determinants in the complex with the nuclear import machinery of
the target cell. Identified determinants include a functional
nuclear localization signal (NLS) in the gag matrix (MA) protein,
the karyophilic virion-associated protein, vpr, and a C-terminal
phosphotyrosine residue in the gag MA protein. The use of
retroviruses for gene therapy is described, for example, in U.S.
Pat. No. 6,013,516; and U.S. Pat. No. 5,994,136, the disclosures of
which are hereby incorporated herein by reference.
[0046] The inventors have discovered that intraventricular delivery
of lysosomal hydrolase enzymes to patients who are deficient in the
enzymes, leads to improved metabolic status of both the brain and
the affected visceral (non-CNS) organs. This is particularly true
when the delivery rate is slow, relative to a bolus delivery. One
particularly useful enzyme for treating Niemann-Pick A, B, or D is
acid sphingomyelinase (aSM), such as that shown in SEQ ID NO:
1..sup.1 .sup.1Residues 1-46 constitute the signal sequence which
is cleaved upon secretion.
[0047] Although a particular amino acid sequence is shown in SEQ ID
NO: 1, normal variants in the human population which retain
activity can be used as well. Typically these normal variants
differ by just one or two residues from the sequence shown in SEQ
ID NO: 1. The variants to be used should be at least 95%, 96%, 97%,
98%, or 99% identical to SEQ ID NO: 1. Variants which are
associated with disease or reduced activity should not be used.
Typically the mature form of the enzyme will be delivered. This
will begin with residue 47 as shown in SEQ ID NO: 1. Variants which
are associated with disease are shown in FIG. 7.
[0048] Kits according to the present invention are assemblages of
separate components. While they can be packaged in a single
container, they can be subpackaged separately. Even a single
container can be divided into compartments. Typically a set of
instructions will accompany the kit and provide instructions for
delivering the diagnostic, therapeutic, or anesthetic agents, such
as lysosomal hydrolase enzymes, intraventricularly. The
instructions may be in printed form, in electronic form, as an
instructional video or DVD, on a compact disc, on a floppy disc, on
the internet with an address provided in the package, or a
combination of these means. Other components, such as diluents,
buffers, solvents, tape, screws, and maintenance tools can be
provided in addition to the agent, one or more cannulae or
catheters, and/or a pump. Printed matter or other instructional
materials may correlate volume of CSF, turn-over time of CSF,
patient weight, patient age, delivery rate, delivery time, and/or
other parameters. Pumps may be calibrated to deliver at specified
rates based on CSF volume and/or turn-over time and/or patient age
and/or patient weight.
[0049] The populations treated by the methods of the invention
include, but are not limited to, patients having or at risk for
developing a neurometabolic disorder, e.g., an LSD, such as
diseases listed in Table 1, particularly, if such a disease affects
the CNS and visceral organs. In an illustrative embodiment, the
disease is type A Niemann-Pick disease. If genetic propensity for
the disease has been determined, treatment may begin prenatally.
Other diseases or conditions which may be treated include but are
not limited to neurosurgical patients, stroke patients,
Huntington's disease, epilepsy, Parkinson's disease, Lou Gehrig's
disease, Alzheimer's disease.
[0050] An agent, such as a lysosomal hydrolase enzyme, can be
incorporated into a pharmaceutical composition. The composition can
be useful to diagnose, anesthetize, or treat, e.g., inhibit,
attenuate, prevent, or ameliorate, a condition characterized by an
insufficient level of a lysosomal hydrolase activity. The
pharmaceutical composition can be administered to a subject
suffering from a lysosomal hydrolase deficiency or someone who is
at risk of developing said deficiency. The compositions should
contain a an effective diagnostic, anesthetic, therapeutic or
prophylactic amount of the agent, in a pharmaceutically-acceptable
carrier. The pharmaceutical carrier can be any compatible,
non-toxic substance suitable to deliver the polypeptides to the
patient. Sterile water, alcohol, fats, and waxes may be used as the
carrier. Pharmaceutically-acceptable adjuvants, buffering agents,
dispersing agents, and the like, may also be incorporated into the
pharmaceutical compositions. The carrier can be combined with the
agent in any form suitable for administration by intraventricular
injection or infusion (also possibly intravenous or intrathecal) or
otherwise. Suitable carriers include, for example, physiological
saline, bacteriostatic water, Cremophor EL.TM. (BASF, Parsippany,
N.J.) or phosphate buffered saline (PBS), other saline solutions,
dextrose solutions, glycerol solutions, water and oils emulsions
such as those made with oils of petroleum, animal, vegetable, or
synthetic origin (peanut oil, soybean oil, mineral oil, or sesame
oil). An artificial CSF can be used as a carrier. The carrier will
preferably be sterile and free of pyrogens. The concentration of
the agent in the pharmaceutical composition can vary widely, i.e.,
from at least about 0.01% by weight, to 0.1% by weight, to about 1%
weight, to as much as 20% by weight or more.
[0051] For intraventricular administration, the composition must be
sterile and should be fluid. It must be stable under the conditions
of manufacture and storage and must be preserved against the
contaminating action of microorganisms such as bacteria and fungi.
Prevention of the action of microorganisms can be achieved by
various antibacterial and antifungal agents, for example, parabens,
chlorobutanol, phenol, ascorbic acid, thimerosal, and the like. In
many cases, it will be preferable to include isotonic agents, for
example, sugars, polyalcohols such as manitol, sorbitol, sodium
chloride in the composition.
[0052] Dosage of any agent, whether aSM or other lysosomal
hydrolase enzyme, may vary somewhat from individual to individual,
depending on the particular agent or enzyme and its specific in
vivo activity, the route of administration, the medical condition,
age, weight or sex of the patient, the patient's sensitivities to
the aSM agent or components of vehicle, and other factors which the
attending physician will be capable of readily taking into account.
While dosages may vary depending on the disease and the patient,
the enzyme is generally administered to the patient in amounts of
from about 0.1 to about 1000 milligrams per 50 kg of patient per
month, preferably from about 1 to about 500 milligrams per 50 kg of
patient per month.
[0053] One way for delivering a slow infusion is to use a pump.
Such pumps are commercially available, for example, from Alzet
(Cupertino, Calif.) or Medtronic (Minneapolis, Minn.). The pump may
be implantable or external. Another convenient way to administer
the enzymes, is to use a cannula or a catheter. The cannula or
catheter may be used for multiple administrations separated in
time. Cannulae and catheters can be implanted stereotaxically. It
is contemplated that multiple administrations will be used to treat
the typical patient with a lysosomal storage disease. Catheters and
pumps can be used separately or in combination. Catheters can be
inserted surgically, as is known in the art. Kits can comprise an
agent and a catheter and/or a pump. The pump may have settings
suitable for delivery rates based on the volume of CSF in an
individual.
[0054] The above disclosure generally describes the present
invention. All references disclosed herein are expressly
incorporated by reference. A more complete understanding can be
obtained by reference to the following specific examples which are
provided herein for purposes of illustration only, and are not
intended to limit the scope of the invention.
EXAMPLE 1
Animal Model
[0055] ASMKO mice are an accepted model of types A and B
Niemann-Pick disease (Horinouchi et al. (1995) Nat. Genetics,
10:288-293; Jin et al. (2002) J. Clin. Invest., 109:1183-1191; and
Otterbach (1995) Cell, 81:1053-1061). Niemann-Pick disease (NPD) is
classified as a lysosomal storage disease and is an inherited
neurometabolic disorder characterized by a genetic deficiency in
acid sphingomyelinase (ASM; sphingomyelin cholinephosphohydrolase,
EC 3.1.3.12). The lack of functional ASM protein results in the
accumulation of sphingomyelin substrate within the lysosomes of
neurons and glia throughout the brain. This leads to the formation
of large numbers of distended lysosomes in the perikaryon, which
are a hallmark feature and the primary cellular phenotype of type A
NPD. The presence of distended lysosomes correlates with the loss
of normal cellular function and a progressive neurodegenerative
course that leads to death of the affected individual in early
childhood (The Metabolic and Molecular Bases of Inherited Diseases,
eds. Scriver et al., McGraw-Hill, New York, 2001, pp. 3589-3610).
Secondary cellular phenotypes (e.g., additional metabolic
abnormalities) are also associated with this disease, notably the
high level accumulation of cholesterol in the lysosomal
compartment. Sphingomyelin has strong affinity for cholesterol,
which results in the sequestering of large amounts of cholesterol
in the lysosomes of ASMKO mice and human patients (Leventhal et al.
(2001) J. Biol. Chem., 276:44976-44983; Slotte (1997) Subcell.
Biochem., 28:277-293; and Viana et al. (1990) J. Med. Genet.,
27:499-504.)
EXAMPLE 2
"Intraventricular Infusion of rhASM in the ASMKO Mouse II"
[0056] Goal: To determine what effect intraventricular infusion of
rhASM has on storage pathology (i.e., sphingomyelin and cholesterol
storage) in the ASMKO mouse brain
[0057] Methods: ASMKO mice were stereotaxically implanted with an
indwelling guide cannula between 12 and 13 weeks of age. At 14
weeks of age mice were infused with 0.250 mg of hASM (n=5) over a
24 h period (.about.0.01 mg/h) for four straight days (1 mg total
was administered) using an infusion probe (fits inside the guide
cannula) which is connected to a pump. Lyophilized hASM was
dissolved in artificial cerebral spinal fluid (aCSF) prior to
infusion. Mice were sacrificed 3 days post infusion. At sacrifice
mice were overdosed with euthasol (>150 mg/kg) and then perfused
with PBS or 4% parformaldehyde. Brain, liver, lung and spleen were
removed and analyzed for sphingomyelin (SPM) levels. Brain tissue
was divided into 5 sections before SPM analysis (S1=front of brain,
S5=back of the brain; see FIG. 1)
TABLE-US-00002 TABLE 2 Group Treatment n ASMKO .250 mg/24 h (1 mg
total) 5 ASMKO None 4 WT None 4
[0058] Results summary: Intraventricular infusion of hASM at 0.250
mg/24 h for 4 continuous days (1 mg total) resulted in hASM
staining and filipin (i.e., cholesterol storage) clearance
throughout the ASMKO brain. Biochemical analysis showed that
intraventricular infusion of hASM also led to a global reduction in
SPM levels throughout the brain. SPM levels were reduced to that of
wild type (WT) levels. A significant reduction in SPM was also
observed in the liver and spleen (a downward trend was seen in the
lung).
EXAMPLE 3
"Intraventricular Delivery of hASM in ASMKO Mice III"
[0059] Goal: to determine lowest efficacious dose over a 6 h
infusion period.
[0060] Methods: ASMKO mice were stereotaxically implanted with an
indwelling guide cannula between 12 and 13 weeks of age. At 14
weeks of age mice were infused over a 6 period at one of the
following doses of hASM: 10 mg/kg (0.250 mg; n=12), 3 mg/kg (0.075
mg; n=7), 1 mg/kg (0.025 mg; n=7), 0.3 mg/kg (0.0075 mg; n=7), or
aCSF (artificial cerebral spinal fluid; n=7). Two mice from each
dose level were perfused with 4% parformaldehyde immediately
following the 6 h infusion to assess enzyme distribution in the
brain (blood was also collected from these to determine serum hASM
levels). The remaining mice from each group were sacrificed 1 week
post infusion. Brain, liver, and lung tissue from these mice was
analyzed for SPM levels as in study 05-0208.
TABLE-US-00003 TABLE 3 Group Treatment n ASMKO 0.250 mg (10 mg/kg)
12 ASMKO 0.075 mg (3 mg/kg) 7 ASMKO 0.025 mg (1 mg/kg) 7 ASMKO
0.0075 mg (.3 mg/kg) 7 ASMKO aCSF 7 WT None 7
[0061] Results summary: Intraventricular hASM over a 6 h period led
to a significant reduction in SPM levels throughout the brain
regardless of does. Brains SPM levels in mice treated with
doses>0.025 mg were reduced to WT levels. Visceral organ SPM
levels were also significantly reduced (but not to WT levels) in a
dose dependent manner. In support of this finding hASM protein was
also detected in the serum of ASMKO mice infused with hASM protein.
Histological analysis showed that hASM protein was widely
distributed throughout the brain (from S1 to S5) after
intraventricular administration of hASM.
EXAMPLE 4
"Intraventricular Infusion of rhASM in ASMKO Mice IV"
[0062] Goal: To determine (1) the time it takes for SPM to
reaccumulate within the brain (and spinal cord) after a 6 h
infusion of hASM (dose=0.025 mg); (2) if there are sex differences
in response to intraventricular hASM administration (pervious
experiments demonstrate that there are sex differences in substrate
accumulation in the liver, whether or not this occurs in the brain
is unknown).
[0063] Methods: ASMKO mice were stereotaxically implanted with an
indwelling guide cannula between 12 and 13 weeks of age. At 14
weeks of age mice were infused over a 6 period with 0.025 mg of
hASM. After intraventricular delivery of hASM mice were sacrificed
either at 1 week post infusion (n=7 males, 7 females), or at 2
weeks post infusion (n=7 males, 7 females) or at 3 weeks post
infusion (n=7 males, 7 females). At sacrifice the brain, spinal
cord, liver and lung were removed for SPM analysis.
TABLE-US-00004 Group Treatment n Sacrifice male ASMKO .025 mg 7 1
week post infusion Female ASMKO .025 mg 7 1 week post infusion male
ASMKO .025 mg 7 2 weeks post infusion Female ASMKO .025 mg 7 2
weeks post infusion male ASMKO .025 mg 7 3 weeks post infusion
Female ASMKO .025 mg 7 3 weeks post infusion male ASMKO aCSF 7 1
week post infusion Female ASMKO aCSF 7 1 week post infusion male WT
None 7 1 week post infusion Female WT None 7 1 week post
infusion
[0064] Tissue samples are prepared for SPM analysis.
EXAMPLE 5
"Effect of Intraventricular Infusion of rhASM on Cognitive Function
in ASMKO Mice"
[0065] Goal: to determine if intraventricular infusion of rhASM
alleviates diseased induced cognitive deficits in ASMKO mice
[0066] Methods: ASMKO mice will be stereotaxically implanted with
an indwelling guide cannula between 9 and 10 weeks of age. At 13
weeks of age mice will be infused over a 6 period with 0.025 mg of
hASM. At 14 and 16 weeks of age mice will undergo cognitive testing
using the Barnes maze.
EXAMPLE 6
"hASM Protein Distribution within the ASMKO CNS After
Intraventricular Infusion"
[0067] Goal: to determine the distribution of hASM protein (as
function of time) within the brain and spinal cord of ASMKO mice
after intraventricular infusion
[0068] Methods: ASMKO mice will be stereotaxically implanted with
an indwelling guide cannula between 12 and 13 weeks of age. At 14
weeks of age mice will be infused over a 6 period with 0.025 mg of
hASM. Following infusion procedure mice will either be sacrificed
immediately or 1 week or 2 weeks or 3 weeks later.
REFERENCES
[0069] The disclosure of each reference cited is expressly
incorporated herein. [0070] 1) Belichenko P V, Dickson P I, Passage
M, Jungles S, Mobley W C, Kakkis E D. Penetration, diffusion, and
uptake of recombinant human alpha-1-iduronidase after
intraventricular injection into the rat brain. Mol Genet Metab.
2005; 86(1-2):141-9. [0071] 2) Kakkis E, McEntee M, Vogler C, Le S,
Levy B, Belichenko P, Mobley W, Dickson P, Hanson S, Passage M.
Intrathecal enzyme replacement therapy reduces lysosomal storage in
the brain and meninges of the canine model of MPS I. Mol Genet
Metab. 2004; 83(1-2): 163-74. [0072] 3) Bembi B, Clana G, Zanatta
M, et al. Cerebrospinal-fluid infusion of alglucerase in the
treatment for acute neuronopathic Gaucher's disease. Pediatr Res
1995; 38:A425. [0073] 4) Lonser R R, Walbridge S, Murray G J,
Aizenberg M R, Vortmeyer A O, Aerts J M, Brady R O, Oldfield E H.
Convection perfusion of glucocerebrosidase for neuronopathic
Gaucher's disease. Ann Neurol. 2005 April; 57(4):542-8.
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