U.S. patent application number 17/327241 was filed with the patent office on 2022-04-14 for pharmaceutical preparation of carbohydrates for therapeutic use.
This patent application is currently assigned to Glycomine, Inc.. The applicant listed for this patent is Glycomine, Inc.. Invention is credited to Tatyana CHERNENKO, Agnes RAFALKO.
Application Number | 20220110871 17/327241 |
Document ID | / |
Family ID | |
Filed Date | 2022-04-14 |
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United States Patent
Application |
20220110871 |
Kind Code |
A1 |
RAFALKO; Agnes ; et
al. |
April 14, 2022 |
PHARMACEUTICAL PREPARATION OF CARBOHYDRATES FOR THERAPEUTIC USE
Abstract
The disclosure provides methods for preparation of carbohydrate
replacement therapies (CRT) that include nanocarriers of
carbohydrates and glycolipids for pharmaceutical delivery to cell
interior, endoplasmic reticulum, and Golgi for treating CDG type I
and CDG type II diseases as well as other metabolic disorders.
Inventors: |
RAFALKO; Agnes; (San
Francisco, CA) ; CHERNENKO; Tatyana; (San Francisco,
CA) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Glycomine, Inc. |
San Carlos |
CA |
US |
|
|
Assignee: |
Glycomine, Inc.
San Carlos
CA
|
Appl. No.: |
17/327241 |
Filed: |
May 21, 2021 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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16568098 |
Sep 11, 2019 |
11045419 |
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17327241 |
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15788591 |
Oct 19, 2017 |
10449149 |
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16568098 |
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15022194 |
Mar 15, 2016 |
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PCT/US2014/055921 |
Sep 16, 2014 |
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15788591 |
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61878591 |
Sep 16, 2013 |
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International
Class: |
A61K 9/127 20060101
A61K009/127; A61K 31/7028 20060101 A61K031/7028 |
Claims
1-20. (canceled)
21. A method for treating a congenital disorder of glycosylation
(CDG) in a human in need thereof, wherein the CDG is CDG-Ia,
CDG-Ie, CDG-Ii, CDG-Ik, CDG-Io, CDG-Ip, or DPM2-CDG, the method
comprising administering to the human a pharmaceutical composition
comprising: (i) a liposome comprising one or more phospholipids
conjugated to polyethylene glycol (PEG); and (ii) one or more
phosphorylated monosaccharide, phosphorylated disaccharide,
phosphorylated oligosaccharide, or phosphorylated polysaccharide,
or any combination thereof, encapsulated in the liposome.
22. The method of claim 21, wherein the pharmaceutical composition
further comprises at least one pharmaceutically acceptable
carrier.
23. The method of claim 21, wherein the pharmaceutical composition
is administered to the human intravenously.
24. The method of claim 21, wherein the PEG minimizes degradation
of the liposome, enhances retention of the liposome, or makes the
liposome immunotolerant when administered to a human in need
thereof, or any combination thereof.
25. The method of claim 21, wherein the PEG is present at a
molecular weight concentration between about 0.5 molar percent and
about 20 molar percent.
26. The method of claim 21, wherein the PEG is present at a
molecular weight concentration between about 1 molar percent and
about 8 molar percent.
27. The method of claim 21, wherein one or more of the
phospholipids comprise a polar head group.
28. The method of claim 21, wherein one or more of the
phospholipids comprise choline, ethanolamine, serine, glycerol, or
inositol, or any combination thereof.
29. The method of claim 21, wherein one or more of the
phospholipids comprise phosphatidylcholine (PC),
phosphatidylethanolamine (PE), phosphatidylserine (PS),
phosphatidylglycerol (PG), or phosphatidylinositol (PI), or any
combination thereof.
30. The method of claim 21, wherein one or more of the
phospholipids comprise phosphatidylcholine (PC) or
phosphatidylethanolamine (PE), or any combination thereof.
31. The method of claim 21, wherein the liposome further comprises
one or more glycerolipids or sphingolipids, or any combination
thereof.
32. The method of claim 21, wherein the liposome further comprises
cholesterol, a fatty acid, or any combination thereof.
33. The method of claim 21, wherein the pharmaceutical composition
comprises: (ii) one or more phosphorylated monosaccharide
encapsulated in the liposome.
34. The method of claim 21, wherein the pharmaceutical composition
comprises: (ii) one or more phosphorylated mannose, phosphorylated
mannofuranose, or phosphorylated mannopyranose, or any combination
thereof, encapsulated in the liposome.
35. The method of claim 21, wherein the pharmaceutical composition
comprises: (ii) mannose-1-phosphate encapsulated in the
liposome.
36. The method of claim 35, wherein the CDG is CDG-Ia.
37. The method of claim 35, wherein the CDG is CDG-Ie.
38. The method of claim 35, wherein the CDG is CDG-Ii.
39. The method of claim 35, wherein the CDG is CDG-Ik.
40. The method of claim 35, wherein the CDG is CDG-Io.
41. The method of claim 35, wherein the CDG is CDG-Ip.
42. The method of claim 35, wherein the CDG is DPM2-CDG.
43. The method of claim 35, wherein: the PEG is present at a
molecular weight concentration between about 0.5 molar percent and
about 20 molar percent; and one or more of the phospholipids
comprise choline, ethanolamine, serine, glycerol, or inositol, or
any combination thereof.
44. The method of claim 43, wherein the PEG is present at a
molecular weight concentration between about 1 molar percent and
about 8 molar percent.
45. The method of claim 43, wherein one or more of the
phospholipids comprise phosphatidylcholine (PC),
phosphatidylethanolamine (PE), phosphatidylserine (PS),
phosphatidylglycerol (PG), or phosphatidylinositol (PI), or any
combination thereof.
46. The method of claim 43, wherein one or more of the
phospholipids comprise phosphatidylcholine (PC) or
phosphatidylethanolamine (PE), or any combination thereof.
47. A pharmaceutical composition, comprising: (i) a lipid particle
comprising one or more phospholipids conjugated to polyethylene
glycol (PEG); and (ii) a nucleotide sugar, a uridine diphosphate, a
guanine diphosphate, a cytosine monophosphate, fucose, GDP-fucose,
a sialic acid, CMP-sialic acid, N-acetylneuraminic acid (Neu5Ac),
CMP-Neu5Ac or any combination thereof, encapsulated in the lipid
particle.
48. A method for treating a congenital disorder of glycosylation
(CDG) in a human in need thereof, comprising administering to the
human a pharmaceutical composition of claim 47.
49. The method of claim 48, wherein the CDG is CDG-Ia, CDG-Ie,
CDG-Ii, CDG-Ik, CDG-Io, CDG-Ip, or DPM2-CDG.
Description
CROSS REFERENCE TO RELATED APPLICATIONS
[0001] This application claims the benefit of U.S. Provisional
Application No. 61/878,591, filed Sep. 16, 2013, which is hereby
incorporated by reference in its entirety.
FIELD
[0002] The present invention relates to compositions containing
carbohydrates encapsulated in a lipid particle for delivery to the
cell interior, methods of delivering the encapsulated carbohydrates
into the interior of cells, and methods of using such compositions
for treating diseases and disorders, such as congenital disorders
of glycosylation (CDG). For example, the present invention relates
to methods for preparation of carbohydrates and glycolipids for
pharmaceutical delivery to cell interior, endoplasmic reticulum
(ER), and Golgi.
BACKGROUND
[0003] Glycosylation, the enzymatic attachment of carbohydrates
(glycans) to proteins and lipids is a co-translational and
post-translational modification (PTM) that is more common than any
other PTM as it applies to a majority of proteins synthesized in
the rough endoplasmic reticulum (ER). Glycosylation plays a
critical role in a variety of biological processes of membrane and
secreted proteins. In the ER, glycosylation defines protein
structure and folding and acts as a quality control mechanism that
dictates the export of properly folded proteins to Golgi or targets
misfolded ones for degradation. Glycan moieties may also act as
ligands for cell surface receptors to mediate cell attachment or
stimulate signal transduction pathways. Congenital disorders of
glycosylation, also known as CDG syndromes, are a group of rare
genetic diseases where tissue proteins and/or lipids carry
defective glycosylation and/or lack of glycosylation. These
diseases are linked to numerous enzymatic deficiencies and often
times cause severe, sometimes fatal, impairments of the nervous
system, muscles, intestines, and several other organ systems.
[0004] Common clinical symptoms in children with CDG include
hypotonia, developmental delay, failure to thrive, hepatic
dysfunction, coagulopathy, hypothyroidism, esotropia, abnormal fat
pattern and inverted nipples, hypoglycemia, seizure, cerebellar
hypoplasia, and stroke-like episodes in a developmentally delayed
child. At an older age, in adolescence, and adulthood, the
presentation may include ataxia, cognitive impairment, and absence
of puberty in females, small testes in males, retinitis pigmentosa,
scoliosis, joint contractures, and peripheral neuropathy.
[0005] CDG may be classified into two groups: CDG type I and CDG
type II. CDG type I is characterized by defects in the initial
steps of N-linked protein glycosylation, i.e., biosynthesis of
dolichol pyrophosphate linked oligosaccharide (DLO), which occur in
the ER, or transfer of the DLO to asparagine residues of nascent,
polypeptides. CDG type II involves defects in further processing
(synthetic or hydrolytic) of the protein-bound glycan. Currently,
Twenty-two CDG type I and fourteen type IT variants have been
identified. One of the most common subtype of CDG is CDG-Ia
(approximately 70% of all CDG cases), which is characterized by
loss or reduction of phosphomannomutase 2 (PMM) activity leading to
deficiency or insufficiency in intracellular N-glycosylation.
(Jaeken et, al. J. of Inherit. Met. Disease. 2008, 31: 669-672).
PMM is responsible for the conversion of mannose-6-phosphate to
mannose-1-phosphate.
[0006] Although several different approaches to developing
therapies for CDG have been explored, researchers continue their
search for a suitable cure or a therapy for mitigating the disease
itself. Existing treatments for manifestations include, for
example, nutritional supplements, tube feeding, and a wide range of
therapies that attempt to treat gastro-esophageal reflux,
persistent vomiting, developmental delays, ocular abnormalities,
and hypothyroidism. Patients also require intravenous (IV)
hydration and physical therapy for stroke-like episodes. Adults
with orthopedic symptoms often require wheel chairs, transfer
devices, and surgical treatment for scoliosis (Sparks et al.,
Disorders of Glycosylation Overview. 2005 in: Pagon R A, Adam M P,
Bird T D, et al., editors. GeneReviews.TM.. Seattle (Wash.):
University of Washington, Seattle; 1993-2013).
[0007] Currently, CDG-Ib is the only known CDG for which a
relatively effective treatment is available, namely oral D-mannose
administration. However, such therapy may not be as effective in
treating CDG-Ia patients and there are currently limited treatment
options for other CDG type I subtypes and CDG type II diseases. One
of the reasons for a lack in established therapy for CDG-I
disorders may be due to the plethora of heterogeneous clinical
phenotypes presented that do not show a direct correlation to the
PMM enzyme activity.
[0008] Patients suffering from a reduction in PMM activity have
reduced production of mannose-1-phosphate (Man-1-P), associated
with symptoms of multivisceral impairments. In order to overcome
PMM production deficiency, it is important to supply downstream
enzymes with the required substrate (i.e., Man-1-P). However, the
delivery and maintenance of such a systemic supply of Man-1-P is
problematic, as extracellular enzymes within bodily fluids degrade
Man-1-P when delivered exogenously by oral or intravenous
administration. Another problem with exogenously delivered Man-1-P
is that its high polarity prevents Man-1-P from penetrating into
the cell interior (i.e., cytosol) and thus treating the deficiency
in PMM production.
[0009] Derivatives of the polar Man-1-P can be synthesized to make
Man-1-P more cell-permeable (US Patent Publication No.
2009/0054353). This approach, however, is also problematic, as the
cell-permeable Man-1-P derivative has been shown to be either
unstable for clinical use or cytotoxic via the by-products of the
Man-1-P derivative (Eklund et al., Glycobiology 2005, 15: 10844093;
Rutschow et at Bioorg Med Chem 2002, 10: 4043-4049; and Hardre et
al., Bioorg Med Chem Lett 2007, 17: 152455).
[0010] Other potential therapies have focused on inhibiting enzymes
that catabolize mannose-6-phosphate (Man-6-P), a precursor to
Man-1-P, via inhibition of phosphomannose isomerases (PMI). The
approach focuses on forcing the reaction towards optimizing
homeostasis, which with the use of PMI inhibitors, would have been
skewed toward production of Man-6-P. These approaches, however, are
ineffective as clinical treatment options due to their associated
toxicity, off-target side effects, and poor selective tissue
penetration.
[0011] Accordingly, unmet needs exist for improved compositions and
methods for delivering carbohydrates, such as Man-1-P, to the cell
interior in order to treat disorders, such as a congenital disorder
of glycosylation (CDG), to subjects (including, for example,
humans) in need of such treatment.
BRIEF SUMMARY
[0012] The present disclosure meets the unmet needs described above
by providing carbohydrates encapsulated by nanocarriers that
address the cytotoxicity and stability problems associated with
either permeabilizing cells or using carbohydrate derivatives. Such
carbohydrates may be endogenous carbohydrates, including, for
example, mannose-1-phosphate. By utilizing endocytotic pathways,
nanocarriers can enter the cell interior and deliver the
carbohydrate into the cytosol of a cell. This implementation of
nanocarriers encapsulating carbohydrates, facilitates
simultaneously the ease of therapy, allowing for higher dosages of
the carbohydrate to reach the biochemical glycosylation pathway, as
well as potential easement on administration. In some aspects, the
disclosure provides methods for preparation of carbohydrates and
glycolipids using nanocarriers for pharmaceutical delivery to cell
interior, endoplasmic reticulum, and Golgi. The disclosure provides
carbohydrate replacement therapies for, but not limited to,
treating diseases of CDG type I and II.
[0013] The present disclosure also addresses unmet needs related to
effective therapeutics for diseases, such as diseases of CDG, as
well as issues related to the delivery of such therapeutics. In
some aspects, the disclosure provides a method of delivering
carbohydrates such as mannose-1-phosphate to the cytosol of cells.
Without wishing to be bound by any theory, it is believed that such
method of deliver the carbohydrates (e.g., as part of a medicament)
would bypass the work of genetically defective cytosolic enzymes,
namely phosphomannomutase (PMM) and phosphomannose isomerase (PMI),
which are found prevalent in CDG I disorders.
[0014] In some embodiments, the compositions of the present
disclosure may involve the delivery of, for example,
mannose-1-phosphate into the cytosol of the cell using
nanocarriers. In particular, biodegradable and biocompatible
lipid-sterol-water lamellar nanostructures may be used to
encapsulate the carbohydrate. The disclosed medicament represents
the most potent and viable treatment option for disorders, such as
the CDG Ia disorder.
[0015] In certain embodiments, the present disclosure provides
compositions containing a lipid particle and carbohydrates, such as
endogenous carbohydrates, encapsulated in the lipid particle, and
methods of using such compositions to deliver the carbohydrate to
the interior of cells and to treat diseases and disorders, such as
congenital disorders of glycosylation (CDG).
[0016] In some embodiments, the compositions described herein are
medicaments. Such compositions (e.g., medicaments) may include an
aqueous suspension of liposomes encasing, for example,
mannose-1-phosphate. Liposomes may be prepared from vesicle-forming
lipids, such as but not limited to phosphatidylcholine (PC). The
liposome may also be prepared in conjunction with a sterol, namely
cholesterol. The full liposome may be enriched with
polyethylene-glycol (PEG), or PEGylated, in order to enhance
circulation and retention with the body fluids upon administration,
as well as protect the liposome from premature degradation and
excretion of the medicament from the body of a subject in need of
such treatment.
[0017] Accordingly, certain aspects of the present disclosure
relate to a composition comprising a lipid particle; and an
endogenous carbohydrate encapsulated in the lipid particle, where
the lipid particle contains a molecule that is capable of
minimizing degradation of the lipid particle and/or enhancing
retention of the lipid particle when administered to a subject,
and/or makes the lipid particle immunotolerant when administered to
a subject. In some embodiments, the molecule is a stealth molecule,
such as ethylene oxide, an ethylene oxide oligomer, an ethylene
oxide polymer, polyethylene glycol (PEG), or any combination
thereof; and/or a PEGylated neutral lipid.
[0018] Other aspects of the present disclosure relate to a
composition containing a lipid particle that contains ethylene
oxide, an ethylene oxide oligomer, an ethylene oxide polymer,
polyethylene glycol (PEG), or any combination thereof; and a
carbohydrate encapsulated in the lipid particle. Other aspects of
the present disclosure relate to a composition containing a lipid
particle; and a carbohydrate encapsulated in the lipid particle,
wherein the lipid particle contains choline, ethanolamine,
glycerol, inositol, or any combination thereof. In some
embodiments, the lipid particle further includes a molecule, such
as a stealth molecule, that is capable of minimizing degradation of
the lipid particle and/or enhancing retention of the lipid particle
when administered to a subject, and/or makes the lipid particle
immunotolerant when administered to a subject. In some embodiments,
the lipid particle is selected from a liposome, a micelle, a solid
lipid nanoparticle, and a niosome. In some embodiments, the lipid
particle is a liposome. In some embodiments, the liposome is a
stealth liposome that may be immunotolerant. In some embodiments,
the endogenous carbohydrate and/or carbohydrate is selected from a
monosaccharide, a phosphorylated monosaccharide, a disaccharide, a
phosphorylated disaccharide, an oligosaccharide, a phosphorylated
oligosaccharide, a polysaccharide, a phosphorylated polysaccharide,
mannose, a phosphorylated mannose, a mannofuranose, a
phosphorylated mannofuranose, a mannopyranos, a phosphorylated
mannopyranos, mannose-1-phosphate, a nucleotide sugar, a uridine
diphosphate, a guanine diphosphate, a cytosine monophosphate,
fucose, GDP-fucose, a sialic acid, CMP-sialic acid,
N-acetylneuraminic acid (Neu5Ac), CMP-Neu5Ac, and derivatives
thereof. In some embodiments, the endogenous carbohydrate is
mannose-1-phosphate.
[0019] Other aspects of the present disclosure relate to a
composition containing a liposome; and mannose-1-phosphate
encapsulated in the liposome, wherein the liposome comprises
cholesterol and phosphatidylethanolamine (PE) attached to
polyethylene glycol (PEG).
[0020] Other aspects of the present disclosure relate to a
pharmaceutical composition containing a composition of any of the
preceding embodiments, and a pharmaceutically acceptable
carrier.
[0021] Other aspects of the present disclosure relate to a kit
containing a composition of any of the preceding embodiments for
use in any of the methods described herein.
[0022] Other aspects of the present disclosure relate to a method
for delivering a carbohydrate to a subject in need thereof, by
administering to the subject a composition of any of the preceding
embodiments. In some embodiments, the composition is administered
orally, topically, dermally, nasally, intravenously,
intramuscularly, intraperitoneally, intracerobrospinally,
intracranially, intraspinally, subcutaneously, intraarticularly,
intrasynovialy, or intrathecaly.
[0023] Other aspects of the present disclosure relate to a method
for delivering a carbohydrate to a cell interior of a subject in
need thereof, by administering to the subject a composition of any
of the preceding embodiments. In some embodiments, the composition
is administered orally, topically, dermally, nasally,
intravenously, intramuscularly, intraperitoneally,
intracerobrospinally, intracranially, intraspinally,
subcutaneously, intra-articularly, intrasynovialy, or
intrathecaly.
[0024] Other aspects of the present disclosure relate to a method
for treating a congenital disorder of glycosylation (CDG) in a
subject in need thereof, by administering to the subject a
composition of any of the preceding embodiments. In some
embodiments, the congenital disorder of glycosylation (CDG) is
selected from a CDG type I disorder, a CDG-Ia disorder, a CDG type
II disorder, a CDG-IIc disorder, and a CDG-IIf disorder. In some
embodiments, the congenital disorder of glycosylation (CDG) is a
CDG-Ia disorder. In some embodiments, the composition is
administered orally, topically, dermally, nasally, intravenously,
intramuscularly, intraperitoneally, intracerobrospinally,
intracranially, intraspinally, subcutaneously, intra-articularly,
intrasynovialy, or intrathecaly.
DESCRIPTION OF THE DRAWINGS
[0025] The following enumerated embodiments are representative of
some aspects of the invention.
[0026] FIG. 1A depicts an exemplary structure of
mannose-1-phosphate (M1P). FIG. 1B depicts an exemplary structure
of polyethylene glycol (PEG). FIG. 1C depicts an exemplary
structure of phosphatidylethanolamine (PE) attached to PEG. FIG. 1D
depicts an exemplary structure of phosphatidylcholine (PC).
[0027] FIG. 2 depicts an exemplary the structure of a liposome
composition containing a carbohydrate encapsulated within the core
of the liposome.
DETAILED DESCRIPTION
[0028] Provided herein are pharmaceutical nanocarriers of
carbohydrates and glycolipids (e.g., lipid compositions containing
a lipid particle and one or more carbohydrates) that are capable of
delivering the carbohydrates and/or glycolipids to a cell interior
including, without limitation, the cytoplasm, endoplasmic
reticulum, and Golgi. Suitable nanocarriers include, but are not
limited to, liposomes, micelles, solid lipid nanoparticles, and
niosomes.
[0029] As disclosed herein, the composition of suitable
nanocarriers is not limited to any class and molecular weight of
lipids, polyethylene glycol, or their derivatives. In some
embodiments, the present disclosure relates more specifically, but
is not limited to, liposomal preparations of mannose-1-phosphate,
guanosine 5'-diphospho-beta-L-fucose (GDP-fucose),
cytidine-5'-monophospho-N-acetylneuraminic acid (CMP-sialic acid),
and dolichol pyrophosphate linked oligosaccharide,
Glc3Man9GlcNAc2-PP-Dol, where Glc=glucose, Man=mannose,
GlcNAc=N-Acetylglucosamine, P=phosphate, Dol=dolichol. In some
variations, the dolichol may have chain lengths of at least 10
isoprene units, and in certain variations, 14-18 isoprene
units.
[0030] As disclosed herein in some embodiments, carbohydrates and
glycolipids of the present disclosure can be integrated into
liposomes designed for specific targeting to cell interior, ER, or
Golgi. A chemoenzymatic approach to synthesis of dolichol
pyrophosphate linked oligosaccharide may be utilized (Wang Z. et
al., Science, 2013, 341, 379-383 and Weerapana E. et al., J. Am.
Chem. Soc., 2005, 127, 13766-13767). In some embodiments, liposomal
preparations may be optimized for each targeted cell compartment as
described in Torchilin V. et al., Curr. Protein Pept. Sci., 2003,
4, 133-140; Pollock S. et al., FASEB, 2010, 24, 1866-1878; Fujiwara
T. et al., Int. J. Pharm., 2010, 386, 122-130; and WO 2009/118658
A2.
[0031] In other embodiments, architectural modification to a
liposome may include, but not limited to, cholesteryl hemisuccinate
(CH) and polyethylene glycol (PEG) chains. As disclosed herein,
such modifications can shield carbohydrate and glycolipid moieties
of the customized nanoparticle from degradation and will enhance
circulation and/or retention time of the pharmaceutical within the
body.
[0032] Accordingly, the present disclosure provides lipid
compositions containing a lipid particle and one or more
carbohydrates, such as endogenous carbohydrates, encapsulated in
the lipid particle, where the lipid compositions are capable of
delivering the carbohydrate into the interior of a cell. In some
embodiments, such carbohydrate-carrying lipid compositions are
useful for treating, preventing, or reducing risk of a congenital
disorder of glycosylation (CDG) in a subject in need thereof. In
some embodiments, the lipid particle may be a liposome, a micelle,
a solid lipid nanoparticle, or a noisome. In some embodiments, the
one or more carbohydrates may be mannose-1-phosphate, guanosine
5'-diphospho-beta-L-fucose (GDP-fucose), and/or
cytidine-5'-monophospho-N-acetylneuraminic acid (CMP-sialic acid).
In some embodiments, the lipid particle contains a molecule that is
capable of minimizing degradation of the lipid particle and/or
enhancing retention of the lipid particle when administered to a
subject, and/or makes the lipid particle immunotolerant when
administered to a subject. As disclosed herein, such molecules may
shield, or otherwise protect, the lipid particle and encapsulated
carbohydrate of the present disclosure from degradation, and may
also enhance cell permeability, circulation and/or retention time
of the lipid particle when the lipid particle is administered to a
subject in need thereof. Additionally, such molecules can allow the
lipid particle to avoid detection by the immune system of a subject
in need thereof that is administered the lipid particle, and as
such are considered stealth molecules. Accordingly, in some
embodiments, the molecule is a stealth molecule that is capable of
inducing immunotolerance when administered to a subject in need
thereof. Examples of such suitable molecules include, without
limitation, ethylene oxide, an ethylene oxide oligomer, an ethylene
oxide polymer, polyethylene glycol (PEG), or any combination
thereof; and/or a PEGylated neutral lipid.
[0033] Thus, in certain aspects, the present disclosure provides a
composition containing a lipid particle; and an endogenous
carbohydrate encapsulated in the lipid particle, where the lipid
particle contains a molecule that is capable of minimizing
degradation of the lipid particle and/or enhancing retention of the
lipid particle when administered to a subject, and/or makes the
lipid particle immunotolerant when administered to a subject. In
some embodiments the molecule is a stealth molecule, such as
ethylene oxide, an ethylene oxide oligomer, an ethylene oxide
polymer, polyethylene glycol (PEG), or any combination thereof;
and/or a PEGylated neutral lipid.
[0034] In certain aspects, the present disclosure provides a
composition containing a lipid particle comprising ethylene oxide,
an ethylene oxide oligomer, an ethylene oxide polymer, polyethylene
glycol (PEG), or any combination thereof; and a carbohydrate
encapsulated in the lipid particle.
[0035] In certain aspects, the present disclosure provides a
composition containing a lipid particle; and a carbohydrate
encapsulated in the lipid particle, wherein the lipid particle
comprises choline, ethanolamine, glycerol, inositol, or any
combination thereof. In some embodiments, the lipid particle
contains a molecule that is capable of minimizing degradation of
the lipid particle and/or enhancing retention of the lipid particle
when administered to a subject, and/or makes the lipid particle
immunotolerant when administered to a subject. In some embodiments
the molecule is a stealth molecule, such as ethylene oxide, an
ethylene oxide oligomer, an ethylene oxide polymer, polyethylene
glycol (PEG), or any combination thereof; and/or a PEGylated
neutral lipid.
[0036] In certain aspects, the present disclosure provides a
composition containing a liposome; and mannose-1-phosphate
encapsulated in the liposome, wherein the liposome comprises
cholesterol and phosphatidylethanolamine (PE) attached to
polyethylene glycol (PEG).
[0037] Additionally, pharmaceutical compositions are provided
containing a carbohydrate-containing lipid composition of the
present disclosure in combination with pharmaceutically acceptable
carriers. The present disclosure also provides a kit containing a
carbohydrate-containing lipid composition of the present disclosure
for use in any of the methods described herein.
[0038] The present disclosure further provides methods of using a
carbohydrate-containing lipid composition of the present disclosure
to deliver a carbohydrate to a subject in need thereof. The present
disclosure further provides methods of using a
carbohydrate-containing lipid composition of the present disclosure
to deliver a carbohydrate to a cell interior to a subject in need
thereof. The present disclosure further provides methods of using a
carbohydrate-containing lipid composition of the present disclosure
to treat, prevent, or reduce risk of a congenital disorder of
glycosylation (CDG) in a subject in need thereof.
Lipid Particles
[0039] Certain aspects of the present disclosure relate to
compositions containing lipid particles that contain carbohydrates
encapsulated within the lipid particle.
[0040] As used herein, the term "lipid particle" refers to
particles formed by lipids in an aqueous solution. Suitable
examples of lipid particles include, without limitation, liposomes,
micelles, solid lipid nanoparticles, niosome, lipospheres,
emulsomes, and emulsions.
[0041] As used herein, a carbohydrate that is "encapsulated" in a
lipid particle, refers to a lipid particle that provides an active
agent or therapeutic agent, such as a carbohydrate of the present
disclosure, with full encapsulation, partial encapsulation, or
both. In some variations, at least a portion of the carbohydrate
may be "encapsulated" by a lipid particle and localized within the
core of a lipid particle and/or within the inner surface (e.g., the
membrane) of a lipid particle. Alternatively, in other variations,
the entire carbohydrate may be "encapsulated" by a lipid particle
and localized within the core of a lipid particle and/or within the
inner surface (e.g., the membrane) of a lipid particle.
[0042] Any lipid particle known in the art suitable for delivering
an encapsulated carbohydrate of the present disclosure to the
interior of a cell may be used. Examples of suitable lipid
particles include, without limitation, liposomes, micelles, solid
lipid nanoparticles, and niosomes.
[0043] In some embodiments, lipid particles of the present
disclosure have an average particle size that ranges from about
0.02 microns in diameter to about 0.5 microns in diameter. In
certain embodiments, lipid particles of the present disclosure have
an average particle size of about 0.02 microns in diameter, about
0.03 microns in diameter, about 0.04 microns in diameter, about
0.05 microns in diameter, about 0.06 microns in diameter, about
0.07 microns in diameter, about 0.08 microns in diameter, about
0.09 microns in diameter, about 0.10 microns in diameter, about
0.15 microns in diameter, about 0.20 microns in diameter, about
0.25 microns in diameter, about 0.30 microns in diameter, about
0.35 microns in diameter, about 0.40 microns in diameter, about
0.45 microns in diameter, or about 0.50 microns in diameter.
[0044] In some embodiments, lipid particles of the present
disclosure are capable of delivering a carbohydrate of the present
disclosure to the interior of a cell, such as the cytoplasm,
endoplasmic reticulum, or Golgi.
[0045] In some embodiments, lipid particles of the present
disclosure may be optimized for targeting to particular organelles
within a cell, including without limitation, the endoplasmic
reticulum (ER), Golgi, and lysosome. In some embodiments, lipid
particles of the present disclosure may be optimized for targeting
to particular tissues and/or organs in a subject. Methods of
optimizing lipid particles for targeting specific organelles,
tissues, and organs are well known in the art (e.g., Torchilin V et
al., Curr. Protein Pept. Sci., 2003, 4, 133-140; Torchilin V et
al., J. Drug Target., 2011, 19, 606-614; Huwyler J. et al., J.
Pharmacol. Exp. Ther., 1997, 282: 1541-1546; Pollock S. et al.,
FASEB, 2010, 24, 1866-1878; Fujiwara T. et al., Int. J. Pharm.,
2010, 386, 122-130; and WO 2009/118658).
[0046] In some embodiments, lipid particles of the present
disclosure contain a carbohydrate of the present disclosure, that
when delivered to the interior of a cell of a subject (including,
for example, a human) in need thereof can induce at least a
0.05-fold to at least a 10-fold increase in cellular production of
higher-order lipid-linked oligosaccharides, as compared to cellular
production of higher-order lipid-linked oligosaccharides in the
absence of lipid particles of the present disclosure containing a
carbohydrate of the present disclosure. In some embodiments, the
lipid particles provided herein, when administered to a subject
(including, for example, a human) in need thereof, may induce at
least a 0.05-fold, at least a 0.1-fold, at least a 0.2-fold, at
least a 0.3-fold, at least a 0.4-fold, at least a 0.5-fold, at
least a 0.6-fold, at least a 0.7-fold, at least a 0.8-fold, at
least a 0.9-fold, at least a 1-fold, at least a 1.5-fold, at least
a 2-fold, at least a 2.5-fold, at least a 3-fold, at least a
3.5-fold, at least a 4-fold, at least a 4.5-fold, at least a
5-fold, at least a 5.5-fold, at least a 6-fold, at least a
6.5-fold, at least a 7-fold, at least a 7.5-fold, at least an
8-fold, at least an 8.5-fold, at least a 9-fold, at least a
9.5-fold, or at least a 10-fold increase in cellular production of
higher-order lipid-linked oligosaccharides in the subject, as
compared to cellular production of higher-order lipid-linked
oligosaccharides in the subject in the absence of administering
such lipid particles to the subject.
[0047] As used herein a "higher-order lipid-linked oligosaccharide"
refers to an oligosaccharide having at least five monosaccharide
subunits and that is linked to a lipid. For example, a higher-order
linked oligosaccharide may refer to an oligosaccharide containing
at least four mannose subunits (Man4), five mannose subunits
(Man5), six mannose subunits (Man6), seven mannose subunits (Man7),
eight mannose subunits (Man8), or nine mannose subunits (Man9) and
an N-acetylglucosamine disaccharide (GlcNAc2), where the
oligosaccharide is linked to a dolichol. In some embodiments, the
higher-order lipid linked oligosaccharides may include an
oligosaccharide portion that includes Man4GlcNAc2, Man5GlcNAc2,
Man6GlcNAc2, Man7GlcNAc2, Man8GlcNAc2, Man8GlcNAc2, or any
combination thereof.
Liposomes
[0048] In some embodiments, a lipid particle of the present
disclosure may be a liposome. As used herein, a "liposome" refers
to a vesicle composed of a lamellar phase lipid bilayer. Any
suitable liposome known in the art may be used. In some
embodiments, the liposome has a lamellar nanostructure. As used
herein, the term "lamellar nanostructure" refers to a
nanostructure, such as a lipid particle, that includes parallel
amphiphilic bilayers separated by a lumen.
[0049] Liposomes of the present disclosure may be prepared by any
suitable method known in the art and disclosed herein. Examples of
suitable methods for preparing liposomes include, without
limitation, disrupting biological membranes, such as by mechanical
dispersion including sonication, thin-film hydration, emulsions,
french pressure cell, extrusion, and reconstitution of dried
vesicles; solvent dispersion including ethanol injection, ether
injection, double emulsion, reverse phase, and vaporization; and
detergent removal methods.
[0050] In certain embodiments, the liposome is a stealth liposome
that may be immunotolerant. As used herein, the term "stealth
liposome" refers to liposomes that are capable of avoiding
detection by a subject's immune system. As such, a stealth liposome
may be immunotolerant. For example, the subject may be a human.
Micelles
[0051] In some embodiments, a lipid particle of the present
disclosure may be a micelle. As used herein, a "micelle" refers to
an aggregate of surfactant molecules (e.g., soaps, detergents,
fatty acids, lipids, phospholipids, etc.) that are dispersed in a
liquid colloid. A micelle in aqueous solution may form an aggregate
with hydrophilic head regions in contact with surrounding solvent,
sequestering the hydrophobic tail regions in the interior of the
micelle. Any suitable micelle known in the art may be used. In some
embodiments, micelles may be spherical. Micelles of the present
disclosure may be prepared by any suitable method known in the art.
Examples of suitable methods for preparing micelles include,
without limitation, direct dissolution, and direct or
microemulsification dialysis, which may encompass preparation by
detergent or water-miscible solvent removal methods.
Solid Lipid Nanoparticles
[0052] In some embodiments, a lipid particle of the present
disclosure may be a solid lipid nanoparticle. As used herein, a
"solid lipid nanoparticle" (SLN) refers to lipid in water emulsions
composed of lipids that are generally solid at temperatures of at
least 50.degree. C., and typically contain a solid lipid core
matrix that can solubilize lipophilic molecules. In some
variations, solid lipid nanoparticles have a diameter in the range
of 10 to 1000 nanometers. Solid lipid nanoparticles may protect
incorporated active compounds, such as carbohydrates of the present
disclosure, against chemical degradation and can also demonstrate
flexibility in modulating the release of such compounds. The lipid
core of solid lipid nanoparticles may be stabilized by surfactants
(e.g., emulsifiers). The lipid may typically include triglycerides,
diglycerides, monoglycerides, fatty acids, steroids, and/or waxes.
Any suitable solid lipid nanoparticle known in the art may be used.
Solid lipid nanoparticles of the present disclosure may be prepared
by any suitable method known in the art. Examples of suitable
methods for preparing solid lipid nanoparticles include, without
limitation, microemulsification, high-pressure homogenization,
precipitation, and film ultrasound dispersion.
Niosomes
[0053] In some embodiments, a lipid particle of the present
disclosure may be a niosome. As used herein, a "niosome" refers to
a vesicular structure formed of a bilayer of non-ionic surfactant
molecules and that contains an aqueous core. Niosomes are
structurally similar to liposomes in having a lamellar structure,
however, the materials used to prepare niosomes make them more
stable against hydrolytic degradation. Examples of suitable noisome
preparation materials include, without limitation, sterols and one
or more non-ionic surfactants. Any suitable niosome known in the
art may be used. Niosomes of the present disclosure may be prepared
by any suitable method known in the art. Examples of suitable
methods for preparing micelles include, without limitation, ether
injection, agitation, bubble method, reverse phase evaporation,
sonication, multiple membrane extrusion, and microfluidigation.
Components of the Lipid Particles
[0054] Lipid particles of the present disclosure (e.g., liposomes,
micelles, solid lipid nanoparticles, and niosomes) contain one or
more lipids. As used herein, the term "lipid" refers to a substance
of biological or synthetic origin that is soluble or partially
soluble in organic solvents or which partitions into a hydrophobic
environment when present in aqueous phase. In some variations,
lipids may be divided into at least three classes: (1) "simple
lipids," which include, without limitation, fats, oils, and waxes;
(2) "compound lipids," which include, without limitation,
phospholipids and glycolipids; and (3) "derived lipids," which
include, without limitation, steroids.
[0055] In some variations, the lipid may be a neutral lipid or an
amphiphilic lipid. As used herein, the term "neutral lipid" refers
to a lipid that exists either in an uncharged or neutral
zwitterionic form at a selected pH. At physiological pH, such
lipids may include, for example, diacylphosphatidylcholine,
diacylphosphatidylethanolamine, ceramide, sphingomyelin, cephalin,
cholesterol, cerebrosides, and diacylglycerols. As used herein, the
term "amphiphilic lipid" refers to a lipid that contains both
polar, water-soluble groups and non-polar, water-insoluble
groups.
[0056] Examples of suitable lipids include, without limitation,
bilayer-forming lipids, non-bilayer-forming lipids, amphiphilic
lipids, naturally-occurring lipids, phospholipids, glycerolipids,
sphingolipids, phosphatidylglycerol, phosphatidic acid,
lyso-lipids, fatty acids, sphingomyelin, glycosphingolipids,
glucolipids, glycolipids, sulphatides, lipids with ether-linked and
ester-linked fatty acids, polymerizable lipids, synthetic lipids,
and semi-synthetic lipids. Synthetic or semi-synthetic lipids may
be produced via deacylation or reacylation of natural lipids.
Suitable features of synthetic and semi-synthetic lipids include,
without limitation, myristoyl, palmitoyl, and stearoyl fatty acids.
In some embodiments, lipid particles of the present disclosure may
contain a mixture of two or more types of lipids. Such mixture may
be present at any ratio that is suitable for encapsulating a
carbohydrate of the present disclosure and delivering such
carbohydrate to a cell interior. In some embodiments, lipid
panicles of the present disclosure may contain a lipid selected
from phospholipid, a glycerolipid, a sphingolipid, and any
combination thereof. As disclosed herein, such a lipid has a polar
head group and a fatty acid tail that may be linked by, for
example, an ester linkage or an ether linkage.
[0057] In some embodiments, lipid particles of the present
disclosure (e.g., liposomes, micelles, solid lipid nanoparticles,
and niosomes) may contain one or more lipids having polar head
groups. The lipids may contain any suitable polar head group known
in the art. Examples of suitable polar head groups include, without
limitation, choline, ethanolamine, serine, glycerol, inositol, and
any combination thereof.
[0058] In some embodiments, lipid particles of the present
disclosure (e.g., liposomes, micelles, solid lipid nanoparticles,
and niosomes) may be, without limitation, lamellar nanostructures
that contain an amphiphilic lipid. Examples of suitable amphiphilic
lipids include, without limitation, phospholipids, cholesterol,
glycolipids, fatty acids, bile acids, saponins, and
surfactants.
[0059] In some embodiments, lipid particles of the present
disclosure (e.g., liposomes, micelles, solid lipid nanoparticles,
and niosomes) may contain one or more phospholipids. Lipid
particles of the present disclosure may contain any suitable
phospholipid known in the art. Examples of suitable phospholipids
include, without limitation, phosphatidylcholine (PC),
phosphatidylethanolamine (PE), phosphatidylserine (PS),
phosphatidylglycerol (PG), phosphatidylinositol (PI), and any
combination thereof. In some embodiments, lipid particles of the
present disclosure may contain phosphatidylcholine (PC). In some
embodiments, lipid particles of the present disclosure may contain
phosphatidylethanolamine (PE). In certain embodiments, lipid
particles of the present disclosure may contain phosphatidylcholine
(PC) and phosphatidylethanolamine (PE).
[0060] In some embodiments, a phospholipid species of the present
disclosure may be present in a lipid particle of the present
disclosure (e.g., liposomes, micelles, solid lipid nanoparticles,
and niosomes) at a concentration of up to 80 molar percent. In
certain embodiments, a phospholipid of the present disclosure may
be present in a lipid particle of the present disclosure at a
concentration of up to 5 molar percent, up to 10 molar percent, up
to 20 molar percent, up to 30 molar percent, up to 40 molar
percent, up to 50 molar percent, up to 60 molar percent, up to 70
molar percent, or up to 80 molar percent. In certain embodiments, a
phospholipid of the present disclosure may be present in a lipid
particle of the present disclosure at a concentration of about 70
molar percent.
[0061] In some embodiments, a lipid particle of the present
disclosure (e.g., liposomes, micelles, solid lipid nanoparticles,
and niosomes) may be modified to, for example, include a molecule,
such as a stealth molecule, that shields, or otherwise protects, an
encapsulated carbohydrate of the present disclosure from
degradation, and that enhances cell permeability, circulation,
and/or retention time of the carbohydrate-containing lipid
composition within a cell, tissue, organ and/or body of a subject
that is administered the carbohydrate-containing lipid composition.
Without wishing to be bound by theory, a lipid particle of the
present disclosure dramatically enhances cell permeability of
carbohydrates, which are otherwise incapable of traversing the
plasma membrane. Accordingly, in some embodiments, a lipid particle
of the present disclosure includes a molecule that is capable of
minimizing degradation of the lipid particle and/or enhancing
retention of the lipid particle when administered to a subject,
and/or makes the lipid particle immunotolerant when administered to
a subject. In some embodiments, the molecule is one or more of
ethylene oxide, an ethylene oxide oligomer, an ethylene oxide
polymer, polyethylene glycol (PEG), or any combination thereof;
and/or a PEGylated neutral lipid. As used herein, the term
"PEGylated neutral lipid" refers to a neutral lipid, at a selected
pH, to which a polyethylene glycol has been conjugated.
[0062] As used herein, the terms "ethylene oxide," "oxirane,"
"epoxyethane," "dimethylene oxide," and "oxacyclopropane" may be
used interchangeably and refer to a cyclic ether having the formula
C.sub.2H.sub.4O.
[0063] As used herein, the terms "polyethylene glycol," "PEG,"
"polyethylene oxide," "PEO," "polyoxyethylene," and "POE" may be
used interchangeably and refer to a polyether compound that is
composed of two or more ethylene oxide subunits. "Polyethylene
glycol" may be composed of ethylene oxide oligomers (e.g., having
from two to nine ethylene oxide monomer subunits) or ethylene oxide
polymers (e.g., having ten or more nine ethylene oxide monomer
subunits).
[0064] In some embodiments, a lipid particle of the present
disclosure may be modified to include one or more lipids conjugated
to polyethylene glycol (PEG) in order to allow the lipid particle
to avoid detection by the immune system of a subject that is
administered the lipid particle. In some embodiments, a lipid
particle of the present disclosure (e.g., liposomes, micelles,
solid lipid nanoparticles, and niosomes) increases cell
permeability for hydrophobic carbohydrates, such as endogenous
carbohydrates of the present disclosure, that are otherwise unable
to get through the cell membrane on their own.
[0065] Accordingly, in some embodiments, lipid particles of the
present disclosure (e.g., liposomes, micelles, solid lipid
nanoparticles, and niosomes) contain ethylene oxide, oligomers of
ethylene oxide and/or polymers of ethylene oxide. In certain
embodiments, lipid particles of the present disclosure contain
polyethylene glycol (PEG). Alternatively, in some embodiments,
lipid particles of the present disclosure (e.g., liposomes,
micelles, solid lipid nanoparticles, and niosomes) may contain,
without limitation, polyvinyl pyrrolidone (PVP), polyacryl amide
(PAA), G.sub.M1 ganglioside, cerebroside, and/or sulfates. Without
wishing to be bound by theory, such modifications can shield lipid
particles from opsonization.
[0066] In some embodiments, polyethylene glycol (PEG) may be
present in a lipid particle of the present disclosure (e.g.,
liposomes, micelles, solid lipid nanoparticles, and niosomes) at a
concentration that ranges from about 0.5 molar percent to about 20
molar percent. In some embodiments, PEG may be present in a lipid
particle of the present disclosure (e.g., liposomes, micelles,
solid lipid nanoparticles, and niosomes) at a concentration of
about 0.5 molar percent, about 1 molar percent, about 2 molar
percent, about 3 molar percent, about 4 molar percent, about 5
molar percent, about 6 molar percent, about 7 molar percent, about
8 molar percent, about 9 molar percent, about 10 molar percent,
about 11 molar percent, about 12 molar percent, about 13 molar
percent, about 14 molar percent, about 15 molar percent, about 16
molar percent, about 17 molar percent, about 18 molar percent,
about 19 molar percent, or about 20 molar percent.
[0067] In some embodiments, polyethylene glycol (PEG) may be
present in a lipid particle of the present disclosure (e.g.,
liposomes, micelles, solid lipid nanoparticles, and niosomes) at a
molecular weight concentration that ranges from about 200 Da to
about 10,000 Da. In some embodiments, polyethylene glycol (PEG) may
be present in a lipid particle of the present disclosure (e.g.,
liposomes, micelles, solid lipid nanoparticles, and niosomes) at a
molecular weight concentration of about 200 Da, about 300 Da, about
400 Da, about 500 Da, about 600 Da, about 700 Da, about 800 Da,
about 900 Da, about 1,000 Da, about 1,500 Da, about 2,000 Da, about
2,500 Da, about 3,000 Da, about 3,500 Da, about 4,000 Da, about
4,500 Da, about 5,000 Da, about 5,500 Da, about 6,000 Da, about
6,500 Da, about 7,000 Da, about 7,500 Da, about 8,000 Da, about
8,500 Da, about 9,000 Da, about 9,500 Da, or about 10,000 Da.
[0068] In some embodiments, lipid particles of the present
disclosure (e.g., liposomes, micelles, solid lipid nanoparticles,
and niosomes) may contain PEG conjugated to a phospholipid of the
present disclosure. The PEG may be conjugated to one or more of
phosphatidylcholine (PC), phosphatidylethanolamine (PE),
phosphatidylserine (PS), phosphatidylglycerol (PG),
phosphatidylinositol (PI), and any combination thereof. In certain
embodiments, lipid particles of the present disclosure may contain
PEG conjugated to phosphatidylcholine (PC). In certain embodiments,
lipid particles of the present disclosure may contain PEG
conjugated to phosphatidylethanolamine (PE).
[0069] In some embodiments, a phospholipid conjugated to
polyethylene glycol (PEG) may be present in a lipid particle of the
present disclosure (e.g., liposomes, micelles, solid lipid
nanoparticles, and niosomes) at a concentration that ranges from
about 0.5 molar percent to about 20 molar percent. In some
embodiments, a phospholipid conjugated to polyethylene glycol (PEG)
may be present in a lipid particle of the present disclosure (e.g.,
liposomes, micelles, solid lipid nanoparticles, and niosomes) at a
concentration of about 0.5 molar percent, about 1 molar percent,
about 2 molar percent, about 3 molar percent, about 4 molar
percent, about 5 molar percent, about 6 molar percent, about 7
molar percent, about 8 molar percent, about 9 molar percent, about
10 molar percent, about 11 molar percent, about 12 molar percent,
about 13 molar percent, about 14 molar percent, about 15 molar
percent, about 16 molar percent, about 17 molar percent, about 18
molar percent, about 19 molar percent, or about 20 molar percent.
In certain embodiments, a phospholipid conjugated to polyethylene
glycol (PEG) may be present in a lipid particle of the present
disclosure at a concentration of about 2 molar percent.
[0070] In some embodiments, lipid particles of the present
disclosure (e.g., liposomes, micelles, solid lipid nanoparticles,
and niosomes) may contain one or more sterols. Examples of suitable
phospholipids include, without limitation, cholesterol and
cholesteryl hemisuccinate (CH), dicetyl phosphate, and Solulan
C24.
[0071] In some embodiments, a sterol of the present disclosure,
such as cholesterol, may be present in a lipid particle of the
present disclosure (e.g., liposomes, micelles, solid lipid
nanoparticles, and niosomes) at a concentration of up to 40 molar
percent. In some embodiments, a sterol of the present disclosure,
such as cholesterol, may be present in a lipid particle of the
present disclosure (e.g., liposomes, micelles, solid lipid
nanoparticles, and niosomes) at a concentration of up to 5 molar
percent, up to 10 molar percent, up to 15 molar percent, up to 20
molar percent, up to 30 molar percent, or up to 40 molar percent.
In certain embodiments, a sterol of the present disclosure, such as
cholesterol, may be present in a lipid particle of the present
disclosure at a concentration of at most 15 molar percent. In some
embodiments, a sterol of the present disclosure, such as
cholesterol, may be present in a lipid particle of the present
disclosure (e.g., liposomes, micelles, solid lipid nanoparticles,
and niosomes) at a molar ratio of 1:1 or 2:1.
[0072] In some embodiments, lipid particles of the present
disclosure (e.g., liposomes, micelles, solid lipid nanoparticles,
and niosomes) may contain one or more fatty acids. In some
embodiments, fatty acids of the present disclosure may have a
carbon chain that ranges in length from about 14 carbon atoms to
about 18 carbon atoms.
[0073] In some embodiments, lipid particles of the present
disclosure (e.g., liposomes, micelles, solid lipid nanoparticles,
and niosomes) may contain one or more neutral lipids. In certain
embodiments, a neutral lipid of the present disclosure may be
PEGylated (Le conjugated to PEG). In some embodiments, a PEGylated
neutral lipid of the present disclosure may be present in a lipid
particle of the present disclosure at a concentration that ranges
from about 0.2 molar percent to about 20 molar percent. In some
embodiments, a neutral lipid of the present disclosure may be
PEGylated (i.e., conjugated to PEG). In some embodiments, a
PEGylated neutral lipid of the present disclosure may be present in
a lipid particle of the present disclosure at a concentration of
about 0.5 molar percent, about 1 molar percent, about 2 molar
percent, about 3 molar percent, about 4 molar percent, about 5
molar percent, about 6 molar percent, about 7 molar percent, about
8 molar percent, about 9 molar percent, about 10 molar percent,
about 11 molar percent, about 12 molar percent, about 13 molar
percent, about 14 molar percent, about 15 molar percent, about 16
molar percent, about 17 molar percent, about 18 molar percent,
about 19 molar percent, or about 20 molar percent.
[0074] In some embodiments, lipid particles of the present
disclosure (e.g., liposomes, micelles, solid lipid nanoparticles,
and niosomes) may be modified to target specific organelles and/or
tissues. Any suitable method known in the art may be used to target
a lipid particle to an organelle and/or tissue. Suitable examples
include, without limitation, modification of the material used for
the preparation of the lipid particle itself (e.g., by utilizing
distinct mixtures and molar ratios of phospholipids such as PC, PS,
PI, and PE), covalent attachment of a linker (e.g., peptide chains
such as poly-arginine chains and octadecyl-rhodamine B) to the
lipid particle surface, or an antibody modification. For example,
octadecyl-rhodamine B modification can target a lipid particle to
the lysosome of cells. In some embodiments, lipid particles of the
present disclosure (e.g., liposomes, micelles, solid lipid
nanoparticles, and niosomes) may be antibody-conjugated liposomes,
or immunoliposomes, that may be used to direct encapsulated
carbohydrates of the present disclosure to diseased tissues and/or
organs.
Carbohydrates
[0075] Other aspects of the present disclosure relate to
compositions containing carbohydrates encapsulated within a lipid
particle of the present disclosure (e.g., liposomes, micelles,
solid lipid nanoparticles, and niosomes). Any suitable carbohydrate
known in the art may be used. Examples of suitable carbohydrates
that may be used include, without limitation, monosaccharides,
phosphorylated monosaccharides, disaccharides, phosphorylated
disaccharides, oligosaccharides, phosphorylated oligosaccharides,
polysaccharides, phosphorylated polysaccharides, nucleotide sugars,
endogenous carbohydrates, and phosphorylated endogenous
carbohydrates.
[0076] In some embodiments, the carbohydrate may be present within
a lipid particle of the present disclosure at a concentration that
ranges from about 0.10 mg per mL of lipid particle to about 10 mg
per mL of lipid particle. In some embodiments, the carbohydrate may
be present within a lipid particle of the present disclosure at a
concentration of about 0.10 mg per mL of lipid particle, about 0.20
mg per mL of lipid particle, about 0.30 mg per mL of lipid
particle, about 0.40 mg per mL of lipid particle, about 0.50 mg per
mL of lipid particle, about 0.60 mg per mL of lipid particle, about
0.70 mg per mL of lipid particle, about 0.80 mg per mL of lipid
particle, about 0.90 mg per mL of lipid particle, about 1 mg per mL
of lipid particle, about 1.50 mg per mL of lipid particle, about 2
mg per mL of lipid particle, about 2.50 mg per mL of lipid
particle, about 3 mg per mL of lipid particle, about 3.50 mg per mL
of lipid particle, about 4 mg per mL of lipid particle, about 4.50
mg per mL of lipid particle, about 5 mg per mL of lipid particle,
5.50 mg per mL of lipid particle, about 6 mg per mL of lipid
particle, 6.50 mg per mL of lipid particle, about 7 mg per mL of
lipid particle, 7.50 mg per mL of lipid particle, about 8 mg per mL
of lipid particle, 8.50 mg per mL of lipid particle, about 9 mg per
mL of lipid particle, about 9.50 mg per mL of lipid particle, or
about 10 mg per mL of lipid particle. In certain embodiments, a
carbohydrate of the present disclosure may be present within a
lipid particle of the present disclosure at a concentration of
about 1 mg per mL of lipid particle.
[0077] In some embodiments, a carbohydrate of the present
disclosure is an endogenous carbohydrate. As used herein,
"endogenous carbohydrate" refers to a carbohydrate that is found as
a natural product in a subject (including, for example, a human).
It should be understood, however, that an endogenous carbohydrate
may be either (i) naturally produced by a subject (including, for
example, a human) and extracted from the living cells of such
subject, or (ii) synthetically made. In some embodiments, the
endogenous carbohydrate is produced in vivo by a subject
(including, for example, human). In other embodiments, the
endogenous carbohydrate is naturally produced by a cell derived
from a subject (including, for example, human), such as a cultured
cell line. Thus, the source of such endogenous carbohydrates may
include, without limitation, a synthetic source (e.g., chemical
synthesis) or a natural source (e.g., extraction, isolation, or
purification from a subject or cell that naturally produces the
endogenous carbohydrate or a recombinant cell, such as a bacterial
cell, that has been genetically engineered to produce the
endogenous carbohydrate). Examples of endogenous carbohydrates may
include, without limitation, carbohydrates involved in protein and
lipid glycosylation.
[0078] Examples of suitable endogenous carbohydrates include,
without limitation, a monosaccharide, a phosphorylated
monosaccharide, a disaccharide, a phosphorylated disaccharide, an
oligosaccharide, a phosphorylated oligosaccharide, a
polysaccharide, a phosphorylated polysaccharide, mannose, a
phosphorylated mannose, a mannofuranose, a phosphorylated
mannofuranose, a mannopyranos, a phosphorylated mannopyranos,
mannose-1-phosphate, a nucleotide sugar, a uridine diphosphate, a
guanine diphosphate, a cytosine monophosphate, fucose, GDP-fucose,
a sialic acid, CMP-sialic acid, N-acetylneuraminic acid (Neu5Ac),
and CMP-Neu5Ac.
[0079] In other embodiments, the carbohydrate is a dolichol
pyrophosphate linked oligosaccharide, Glc3Man9GlcNAc2-PP-Dol, where
Glc=glucose, Man=mannose, GlcNAc=N-Acetylglucosamine, P=phosphate,
and Dol=dolichol with a chain length that includes, but not limited
to, 14-18 isoprene units. Any suitable method known in the art for
preparing dolichol pyrophosphate linked oligosaccharides may be
used. In one example, a chemoenzymatic approach may be used to
synthesize dolichol pyrophosphate linked oligosaccharide (e.g.,
Wang Z. et al., Science, 2013, 341, 379-383 and Weerapana E. et
al., J. Am. Chem. Soc., 2005, 127, 13766-13767).
[0080] Other aspects of the present disclosure relate to
compositions containing glycolipids encapsulated within a lipid
particle of the present disclosure. Any suitable glycolipid known
in the art may be used. Glycolipids of the present disclosure
include, without limitation, any class and molecular weight of
lipids and carbohydrates.
[0081] In some embodiments, carbohydrates and glycolipids of the
present disclosure may be integrated into lipid particles of the
present disclosure, such as liposomes, that may be designed for
specific targeting to a cell interior, the ER of a cell, or the
Golgi of a cell.
Pharmaceutical Compositions
[0082] Compositions of the present disclosure containing a lipid
particle of the present disclosure and a carbohydrate of the
present disclosure encapsulated in the lipid particle can be
incorporated into a variety of formulations for therapeutic use
(e.g., by administration) or in the manufacture of a medicament
(e.g., for delivering a carbohydrate of the present disclosure to a
subject in need thereof and/or cell interior of a subject in need
thereof and/or for treating or preventing a disease or disorder
such as a congenital disorder of glycosylation (CDG) in a subject
in need thereof) by combining the composition with appropriate
carriers (including, for example, pharmaceutically acceptable
carriers or diluents), and may be formulated, without limitation,
into preparations in liquid, aerosolized, semisolid, or powder
forms.
[0083] "Carriers" as used herein include pharmaceutically
acceptable carriers, excipients, or stabilizers that are nontoxic
to the cell or subject being exposed thereto at the dosages and
concentrations employed. Often the physiologically acceptable
carrier is an aqueous pH buffered solution. Examples of
physiologically acceptable carriers include, without limitation,
buffers such as phosphate, citrate, and other organic acids;
antioxidants including ascorbic acid; low molecular weight (less
than about 10 residues) polypeptide; proteins, such as serum
albumin, gelatin, or immunoglobulins; hydrophilic polymers such as
polyvinylpyrrolidone; amino acids such as glycine, glutamine,
asparagine, arginine or lysine; monosaccharides, disaccharides, and
other carbohydrates including glucose, mannose, or dextrins;
chelating agents such as EDTA; sugar alcohols such as mannitol or
sorbitol; salt-forming counterions such as sodium; and/or nonionic
surfactants such as TWEEN.TM., polyethylene glycol (PEG), and
PLURONICS.TM..
[0084] Examples of suitable formulations include, without
limitation, solutions, injections, inhalants, microspheres,
aerosols, gels, ointments, creams, lotions, powders, dry vescicular
powders, tablets, and capsules. Pharmaceutical compositions can
include, depending on the formulation desired,
pharmaceutically-acceptable, non-toxic carriers of diluents, which
are vehicles commonly used to formulate pharmaceutical compositions
for animal or human administration. The diluent is selected so as
not to affect the biological activity of the combination. Examples
of such diluents include, without limitation, distilled water,
buffered water, physiological saline, PBS, Ringer's solution,
dextrose solution, and Hank's solution. A pharmaceutical
composition or formulation of the present disclosure can further
include, without limitation, other carriers or non-toxic,
nontherapeutic, nonimmunogenic stabilizers, and excipients. The
compositions can also include additional substances to approximate
physiological conditions, such as pH adjusting and buffering
agents, toxicity adjusting agents, wetting agents and detergents. A
pharmaceutical composition of the present disclosure can also
include any of a variety of stabilizing agents, such as an
antioxidant for example.
[0085] For oral administration, the active ingredient can be
administered in solid dosage forms, such as capsules, tablets, and
powders, or in liquid dosage forms, such as elixirs, syrups, and
suspensions. The active component(s) can be encapsulated in gelatin
capsules together with inactive ingredients and powdered carriers,
such as glucose, lactose, sucrose, mannitol, starch, cellulose or
cellulose derivatives, magnesium stearate, stearic acid, sodium
saccharin, talcum, magnesium carbonate. Examples of additional
inactive ingredients that may be added to provide desirable color,
taste, stability, buffering capacity, dispersion or other known
desirable features are red iron oxide, silica gel, sodium lauryl
sulfate, titanium dioxide, and edible white ink. Similar diluents
can be used to make compressed tablets. Both tablets and capsules
can be manufactured as sustained release products to provide for
continuous release of medication over a period of hours. Compressed
tablets can be sugar coated or film coated to mask any unpleasant
taste and protect the tablet from the atmosphere, or enteric-coated
for selective disintegration in the gastrointestinal tract. Liquid
dosage forms for oral administration can contain coloring and
flavoring to increase patient acceptance.
[0086] Formulations suitable for parenteral administration include
aqueous and non-aqueous, isotonic sterile injection solutions,
which can contain antioxidants, buffers, bacteriostats, and solutes
that render the formulation isotonic with the blood of the intended
recipient, and aqueous and non-aqueous sterile suspensions that can
include suspending agents, solubilizers, thickening agents,
stabilizers, and preservatives.
[0087] The components used to formulate the pharmaceutical
compositions are preferably of high purity and are substantially
free of potentially harmful contaminants (e.g., at least National
Food (NF) grade, generally at least analytical grade, and more
typically at least pharmaceutical grade). Moreover, compositions
intended for in vivo use are usually sterile. To the extent that a
given compound must be synthesized prior to use, the resulting
product is typically substantially free of any potentially toxic
agents, particularly any endotoxins, which may be present during
the synthesis or purification process. Compositions for parental
administration are also sterile, substantially isotonic and made
under GMP conditions.
Pharmaceutical Dosages
[0088] Pharmaceutical compositions of the present disclosure
containing a composition containing a lipid particle of the present
disclosure and a carbohydrate of the present disclosure
encapsulated by the lipid particle may be used (e.g., administered
to a subject in need of treatment with a carbohydrate of the
present disclosure, such as a human individual) in accord with
known methods, such as oral administration, intravenous
administration as a bolus or by continuous infusion over a period
of time, by intramuscular, intraperitoneal, intracerobrospinal,
intracranial, intraspinal, subcutaneous, intra-articular,
intrasynovial, intrathecal, oral, topical, or inhalation
routes.
[0089] Dosages and desired concentration of pharmaceutical
compositions of the present disclosure may vary depending on the
particular use envisioned. The determination of the appropriate
dosage or route of administration is well within the skill of an
ordinary artisan. Animal experiments provide reliable guidance for
the determination of effective doses for human therapy.
Interspecies scaling of effective doses can be performed following
the principles described in Mordenti, J. and Chappell, W. "The Use
of Interspecies Scaling in Toxicokinetics," In Toxicokinetics and
New Drug Development, Yacobi et al., Eds, Pergamon Press, New York
1989, pp. 42-46.
[0090] For in vivo administration of any of the compositions of the
present disclosure containing a lipid particle of the present
disclosure and a carbohydrate of the present disclosure
encapsulated by the lipid particle, normal dosage amounts may vary
from 10 ng/kg up to 100 mg/kg of a subject's body weight per
day.
[0091] Administration of a composition of the present disclosure
containing a lipid particle of the present disclosure and a
carbohydrate of the present disclosure encapsulated by the lipid
particle can be continuous or intermittent, depending, for example,
on the recipient's physiological condition, whether the purpose of
the administration is therapeutic or prophylactic, and other
factors known to skilled practitioners.
[0092] It is within the scope of the present disclosure that
different formulations will be effective for different treatments
and different disorders, and that administration intended to treat
a specific organ or tissue may necessitate delivery in a manner
different from that to another organ or tissue. Moreover, dosages
may be administered by one or more separate administrations, or by
continuous infusion. For repeated administrations over several days
or longer, depending on the condition, the treatment is sustained
until a desired suppression of disease symptoms occurs. However,
other dosage regimens may be useful. The progress of this therapy
is easily monitored by conventional techniques and assays.
[0093] Thus, in some variations, the compositions provided herein
may be chronically or intermittently administered to a subject
(including, for example, a human) in need thereof. "Chronic"
administration refers to administration of the medicament(s) in a
continuous as opposed to acute mode, so as to maintain the initial
therapeutic effect (activity) for an extended period of time.
"Intermittent" administration refers to treatment that is not
consecutively done without interruption, but rather is cyclic in
nature.
Therapeutic Uses
[0094] The present disclosure provides compositions containing a
lipid particle of the present disclosure and a carbohydrate of the
present disclosure encapsulated by the lipid particle that are
capable of delivering the carbohydrate into the interior of a cell.
These compositions are useful for delivering carbohydrates of the
present disclosure to a subject in need of such carbohydrates.
[0095] As used herein, the term "subject" refers to a mammal, such
as a human, domestic animal, such as a feline or canine subject,
farm animal (e.g., bovine, equine, caprine, ovine, and porcine
subject), wild animal (whether in the wild or in a zoological
garden), research animal, such as mouse, rat, rabbit, goat, sheep,
pig, dog, and cat, and birds. In one embodiment, the subject is a
human.
[0096] In some variations, the subject may be at risk. For example,
in one variation, the subject is an at risk human. As used herein,
a subject "at risk" of developing a particular disease, disorder,
or condition, such as a congenital disorder of glycosylation, may
or may not have detectable disease or symptoms of disease, and may
or may not have displayed detectable disease or symptoms of disease
prior to the treatment methods described herein. "At risk" denotes
that an individual has risk factors, which are measurable
parameters that correlate with development of a particular disease,
disorder, or condition, as known in the art. A subject having one
or more of these risk factors has a higher probability of
developing a particular disease, disorder, or condition such as a
congenital disorder of glycosylation, than a subject without one or
more of these risk factors.
[0097] In some embodiments, compositions containing a lipid
particle of the present disclosure and a carbohydrate of the
present disclosure encapsulated by the lipid particle may also be
used for delivering carbohydrates of the present disclosure to the
cell interior of a subject in need thereof. In certain embodiments,
compositions containing a lipid particle of the present disclosure
and a carbohydrate of the present disclosure encapsulated by the
lipid particle may be used for preventing, reducing risk, or
treating a disease or disorder, such as a congenital disorder of
glycosylation (CDG) in a subject in need thereof.
[0098] Accordingly, as disclosed herein, compositions of the
present disclosure containing a lipid particle of the present
disclosure and a carbohydrate of the present disclosure
encapsulated by the lipid particle may be used for delivering a
carbohydrate of the present disclosure to a subject in need
thereof, for delivering a carbohydrate of the present disclosure to
the cell interior of a subject in need thereof, and for treating,
preventing, or reducing risk of a disease or disorder, such as a
congenital disorder of glycosylation (CDG) in a subject in need
thereof. In some embodiments, the subject has such a disease or
disorder. In some embodiments, the subject is a human having such a
disease or disorder.
[0099] As used herein, the term "congenital disorders of
glycosylation" (CDG) refers to a group of genetic disorders that
result in errors of metabolism in which glycosylation of a variety
of tissue proteins and/or lipids is deficient or defective.
Congenital disorders of glycosylation may also be known as CDG
syndromes. CDG syndromes may often cause serious, occasionally
fatal, malfunction of several different organ systems, such as the
nervous system, brain, muscles, and intestines, in affected
infants. Manifestations of CDG syndromes may range from severe
developmental delay and hypotonia beginning in infancy, to
hypoglycemia and protein-losing enteropathy with normal
development. Developmental delay can be a common initial indication
for a CDG diagnosis. One of the most common subtype of CDG
syndromes is CDG-Ia (also known as PMM2-CDG) where the genetic
defect leads to the loss of phosphomannomutase 2, which is the
enzyme responsible for the conversion of mannose-6-phosphate into
mannose-1-phosphate.
[0100] CDG syndromes may be classified as type I (CDG-I) and type
II (CDG-II). Such classification may depend on the nature and
location of the biochemical defect in the metabolic pathway
relative to the action of oligosaccharyltransferase. Methods for
screening for CDG subtype may include the analysis of transferrin
glycosylation status by, for example, isoelectric focusing or
ESI-MS. Examples of CDG type I include, without limitation, Ia
(PMM2-CDG), Ib (MPI-CDG), Ic (ALG6-CDG), Id (ALG3-CDG), Ie
(DPM1-CDG), If (MPDU1-CDG), Ig (ALG12-CDG), Ih (ALG8-CDG), Ii
(ALG2-CDG), Ij (DPAGT1-CDG), Ik (ALG1-CDG), 1L (ALG9-CDG), Im
(DOLK-CDG), In (RFT1-CDG), Io (DPM3-CDG), Ip (ALG11-CDG), Ig
(SRD5A3-CDG), Ir (DDOST-CDG), DPM2-CDG, TUSC3-CDG, MAGT1-CDG,
DHDDS-CDG, and I/IIx. Examples of CDG type II include, without
limitation, IIa (MGAT2-CDG), IIb (GCS1-CDG), IIc (SLC335C1-CDG),
IId (B4GALT1-CDG), IIe (COG7-CDG), IIf (SLC35A1-CDG), IIg
(COG1-CDG), IIh (COG8-CDG), IIi (COG5-CDG), IIj (COG4-CDG), IIL
(COG6-CDG), ATP6V0A2-CDG, MAN1B1-CDG, and ST3GAL3-CDG.
[0101] Congenital disorders of glycosylation (CDG) that may be
treated with compositions of the present disclosure containing a
lipid particle of the present disclosure and a carbohydrate of the
present disclosure encapsulated by the lipid particle include,
without limitation, Ia (PMM2-CDG), Ib (MPI-CDG), Ic (ALG6-CDG), Id
(ALG3-CDG), Ie (DPM1-CDG), If (MPDU1-CDG), Ig (ALG12-CDG), Ih
(ALG8-CDG), Ii (ALG2-CDG), Ij (DPAGT1-CDG), Ik (ALG1-CDG), 1L
(ALG9-CDG), Im (DOLK-CDG), In (RFT1-CDG), Io (DPM3-CDG), Ip
(ALG11-CDG), Iq (SRD5A3-CDG), Ir (DDOST-CDG), DPM2-CDG, TUSC3-CDG,
MAGT1-CDG, DHDDS-CDG, I/IIx, IIa (MGAT2-CDG), IIb (GCS1-CDG), IIc
(SLC335C1-CDG), IId (B4GALT1-CDG), IIe (COG7-CDG), IIf
(SLC35A1-CDG), IIg (COG1-CDG), IIh (COG8-CDG), Ili (COG5-CDG), IIj
(COG4-CDG), IIL (COG6-CDG), ATP6V0A2-CDG, MAN1B1-CDG, and
ST3GAL3-CDG.
[0102] Accordingly, the compositions of the present disclosure
containing a lipid particle of the present disclosure and a
carbohydrate of the present disclosure encapsulated by the lipid
particle may be used to treat, prevent, or improve one or more
symptoms of a congenital disorder of glycosylation (CDG) of the
present disclosure. In some embodiments, the present disclosure
provides methods of treating, preventing, or improving one or more
symptoms in subjects having a congenital disorder of glycosylation
(CDG) disorder of the present disclosure by administering to the
subject a composition of the present disclosure containing a lipid
particle of the present disclosure and a carbohydrate of the
present disclosure encapsulated by the lipid particle to, for
example, deliver a carbohydrate to the subject to correct, or
otherwise improve protein and/or lipid glycosylation in the
subject.
[0103] In some embodiments, the present disclosure provides methods
for treating, preventing, or improving one or more symptoms of a
congenital disorder of glycosylation (CDG) to a subject in need
thereof, by administering to the subject a therapeutically
effective amount of a composition comprising a lipid particle; and
an endogenous carbohydrate encapsulated in the lipid particle. In
some embodiments, the present disclosure provides methods for
treating, preventing, or improving one or more symptoms of a
congenital disorder of glycosylation (CDG) to a subject in need
thereof, by administering to the subject a therapeutically
effective amount of a composition comprising a lipid particle
comprising ethylene oxide; and a carbohydrate encapsulated in the
lipid particle. In some embodiments, the present disclosure
provides methods for treating, preventing, or improving one or more
symptoms of a congenital disorder of glycosylation (CDG) to a
subject in need thereof, by administering to the subject a
therapeutically effective amount of a composition comprising a
lipid particle; and a carbohydrate encapsulated in the lipid
particle, wherein the lipid particle comprises choline,
ethanolamine, glycerol, inositol, or any combination thereof. In
some embodiments, the present disclosure provides methods for
treating, preventing, or improving one or more symptoms of a
congenital disorder of glycosylation (CDG) to a subject in need
thereof, by administering to the subject a therapeutically
effective amount of a composition comprising a liposome; and
mannose-1-phosphate encapsulated in the liposome, wherein the
liposome comprises cholesterol and phosphatidylethanolamine (PE)
attached to polyethylene glycol (PEG). In some embodiments, the
congenital disorder of glycosylation (CDG) may be a CDG type I
disorder, a CDG-Ia disorder, a CDG type II disorder, a CDG-IIc
disorder, or a CDG-IIf disorder. In certain embodiments, the
congenital disorder of glycosylation (CDG) is a CDG-Ia disorder. In
some embodiments, the composition is administered orally, dermally,
nasally, or intravenously.
[0104] As used herein, the term "treatment" or "treating" is an
approach for obtaining beneficial or desired results including
clinical results. Beneficial or desired clinical results may
include one or more of the following: a) inhibiting the disease or
condition (e.g., decreasing one or more symptoms resulting from the
disease or condition, and/or diminishing the extent of the disease
or condition); b) slowing or arresting the development of one or
more clinical symptoms associated with the disease or condition
(e.g., stabilizing the disease or condition, preventing or delaying
the worsening or progression of the disease or condition, and/or
preventing or delaying the spread of the disease or condition);
and/or c) relieving the disease, that is, causing the regression of
clinical symptoms (e.g., ameliorating the disease state, providing
partial or total remission of the disease or condition, enhancing
effect of another medication, delaying the progression of the
disease, increasing the quality of life, and/or prolonging
survival.
[0105] As used herein, the term "prevention" or "preventing" refers
to any treatment of a disease or condition that causes the clinical
symptoms of the disease or condition not to develop. Compounds may,
in some embodiments, be administered to a subject (including a
human) who is at risk or has a family history of the disease or
condition.
[0106] An "effective amount" refers to at least an amount
effective, at dosages and for periods of time necessary, to achieve
the desired therapeutic or prophylactic result. An effective amount
can be provided in one or more administrations.
[0107] A "therapeutically effective amount" is at least the minimum
concentration required to effect a measurable improvement of a
particular disease, disorder, or condition, such as a congenital
disorder of glycosylation. A therapeutically effective amount
herein may vary according to factors such as the disease state,
age, sex, and weight of the subject, and the ability of the lipid
compositions of the present disclosure to elicit a desired response
in the subject. A therapeutically effective amount is also one in
which any toxic or detrimental effects of the lipid compositions of
the present disclosure are outweighed by the therapeutically
beneficial effects.
[0108] In other aspects of the present disclosure, compositions of
the present disclosure containing a lipid particle of the present
disclosure and a carbohydrate of the present disclosure
encapsulated by the lipid particle may be used for delivering a
carbohydrate to a subject in need thereof. Accordingly, in some
embodiments, the present disclosure provides methods for delivering
a carbohydrate to a subject in need thereof, by administering to
the subject a therapeutically effective amount of a composition
comprising a lipid particle; and an endogenous carbohydrate
encapsulated in the lipid particle. In some embodiments, the
present disclosure provides methods for delivering a carbohydrate
to a subject in need thereof, by administering to the subject a
therapeutically effective amount of a composition comprising a
lipid particle comprising ethylene oxide; and a carbohydrate
encapsulated in the lipid particle. In some embodiments, the
present disclosure provides methods for delivering a carbohydrate
to a subject in need thereof, by administering to the subject a
therapeutically effective amount of a composition comprising a
lipid particle; and a carbohydrate encapsulated in the lipid
particle, wherein the lipid particle comprises choline,
ethanolamine, glycerol, inositol, or any combination thereof. In
some embodiments, the present disclosure provides methods for
delivering a carbohydrate to a subject in need thereof, by
administering to the subject a therapeutically effective amount of
a composition comprising a liposome; and mannose-1-phosphate
encapsulated in the liposome, wherein the liposome comprises
cholesterol and phosphatidylethanolamine (PE) attached to
polyethylene glycol (PEG). In some embodiments, the subject has a
congenital disorder of glycosylation (CDG). In some embodiments,
the congenital disorder of glycosylation (CDG) may be a CDG type I
disorder, a CDG-Ia disorder, a CDG type II disorder, a CDG-IIc
disorder, and a CDG-IIf disorder. In certain embodiments, the
congenital disorder of glycosylation (CDG) is a CDG-Ia
disorder.
[0109] In other aspects of the present disclosure, compositions of
the present disclosure containing a lipid particle of the present
disclosure and a carbohydrate of the present disclosure
encapsulated by the lipid particle may be used for delivering a
carbohydrate to a cell interior of a subject in need thereof.
Accordingly, in some embodiments, the present disclosure provides
methods for delivering a carbohydrate to a cell interior of a
subject in need thereof, by administering to the subject a
therapeutically effective amount of a composition comprising a
lipid particle; and an endogenous carbohydrate encapsulated in the
lipid particle, wherein the administered composition traverses the
cell plasma membrane thereby delivering the carbohydrate to the
cell interior. In some embodiments, the present disclosure provides
methods for delivering a carbohydrate to a cell interior of a
subject in need thereof, by administering to the subject a
therapeutically effective amount of a composition comprising a
lipid particle comprising ethylene oxide; and a carbohydrate
encapsulated in the lipid particle, wherein the administered
composition traverses the cell plasma membrane thereby delivering
the carbohydrate to the cell interior. In some embodiments, the
present disclosure provides methods for delivering a carbohydrate
to a cell interior of a subject in need thereof, by administering
to the subject a therapeutically effective amount of a composition
comprising a lipid particle; and a carbohydrate encapsulated in the
lipid particle, wherein the lipid particle comprises choline,
ethanolamine, glycerol, inositol, or any combination thereof, and
wherein the administered composition traverses the cell plasma
membrane thereby delivering the carbohydrate to the cell interior.
In some embodiments, the present disclosure provides methods for
delivering a carbohydrate to a cell interior of a subject in need
thereof, by administering to the subject a therapeutically
effective amount of a composition comprising a liposome; and
mannose-1-phosphate encapsulated in the liposome, wherein the
liposome comprises cholesterol and phosphatidylethanolamine (PE)
attached to polyethylene glycol (PEG), and wherein the administered
composition traverses the cell plasma membrane thereby delivering
the carbohydrate to the cell interior. In some embodiments, the
subject has a congenital disorder of glycosylation (CDG). In some
embodiments, the congenital disorder of glycosylation (CDG) may be
a CDG type I disorder, a CDG-Ia disorder, a CDG type II disorder, a
CDG-IIc disorder, and a CDG-IIf disorder. In certain embodiments,
the congenital disorder of glycosylation (CDG) is a CDG-Ia
disorder. In some embodiments, administration of the composition
induces at least a 0.05-fold to at least a 2-fold increase in
cellular production of higher-order lipid-linked oligosaccharides
in the subject, as compared to cellular production of higher-order
lipid-linked oligosaccharides in the subject in the absence of the
composition. In certain embodiments, the higher-order lipid-linked
oligosaccharides comprise Man4GlcNAc2, Man5GlcNAc2, Man6GlcNAc2,
Man7GlcNAc2, Man8GlcNAc2, Man9GlcNAc2, or any combination thereof.
In some embodiments, the composition is administered orally,
topically, dermally, nasally, intravenously, intramuscularly,
intraperitoneally, intracerobrospinally, intracranially,
intraspinally, subcutaneously, intra-articularly, intrasynovialy,
or intrathecaly.
Articles of Manufacture and Kits
[0110] The present disclosure also provides articles of manufacture
and/or kits containing a composition of the present disclosure
containing a lipid particle of the present disclosure and a
carbohydrate of the present disclosure encapsulated by the lipid
particle. Articles of manufacture and/or kits of the present
disclosure may include one or more containers comprising a purified
composition of the present disclosure. Suitable containers may
include, without limitation, bottles, vials, syringes, and IV
solution bags. The containers may be formed from a variety of
materials such as glass or plastic. In some embodiments, the
articles of manufacture and/or kits further include instructions
for use in accordance with any of the methods of the present
disclosure. In some embodiments, these instructions comprise a
description of administration of the composition containing a lipid
particle of the present disclosure and a carbohydrate of the
present disclosure encapsulated by the lipid particle to deliver
the carbohydrate to a subject in need thereof, to deliver the
carbohydrate to a cell interior of a subject in need thereof, or to
treat a congenital disorder of glycosylation (CDG) to a subject in
need thereof, according to any of the methods of the present
disclosure. In some embodiments, the instructions comprise a
description of how to detect a congenital disorder of glycosylation
(CDG), for example in a subject, in a tissue sample, or in a cell.
The article of manufacture and/or kit may further comprise a
description of selecting a subject suitable for treatment based on
identifying whether that subject has the disease and the stage of
the disease.
[0111] The instructions generally include information as to dosage,
dosing schedule, and route of administration for the intended
treatment. The containers may be unit doses, bulk packages (e.g.,
multi-dose packages) or sub-unit doses. Instructions supplied in
the articles of manufacture and/or kits of the present disclosure
are typically written instructions on a label or package insert
(e.g., a paper sheet included in the article of manufacture and/or
kit), but machine-readable instructions (e.g., instructions carried
on a magnetic or optical storage disk) are also acceptable.
[0112] The label or package insert indicates that the composition
is used for delivering a carbohydrate and/or treating, e.g., a
congenital disorder of glycosylation (CDG). Instructions may be
provided for practicing any of the methods described herein.
[0113] The articles of manufacture and/or kits of the present
disclosure may be in suitable packaging. Suitable packaging
includes, without limitation, vials, bottles, jars, and flexible
packaging (e.g., sealed Mylar or plastic bags). Also contemplated
are packages for use in combination with a specific device, such as
an inhaler, nasal administration device (e.g., an atomizer) or an
infusion device such as a minipump. An article of manufacture
and/or kit may have a sterile access port (for example the
container may be an intravenous solution bag or a vial having a
stopper pierceable by a hypodermic injection needle). The container
may also have a sterile access port (e.g., the container may be an
intravenous solution bag or a vial having a stopper pierceable by a
hypodermic injection needle). At least one active agent in the
composition is a carbohydrate capable of treating a congenital
disorder of glycosylation (CDG) and/or improving one or more
symptoms thereof. The container may further comprise a second
pharmaceutically active agent.
[0114] Articles of manufacture and/or kits may optionally provide
additional components such as buffers and interpretive information.
Normally, the article of manufacture and/or kit comprises a
container and a label or package insert(s) on or associated with
the container.
[0115] Unless defined otherwise, all scientific and technical terms
are understood to have the same meaning as commonly used in the art
to which they pertain. For the purpose of the present disclosure,
the following terms are defined.
[0116] The term "about" as used herein refers to the usual error
range for the respective value readily known to the skilled person
in this technical field. Reference to "about" a value or parameter
herein includes (and describes) embodiments that are directed to
that value or parameter per se. For example, "about x" includes and
describes "x" per se. In some embodiments, the term "about" when
used in association with a measurement, or used to modify a value,
a unit, a constant, or a range of values, refers to variations of
+/-2%.
[0117] As used herein and in the appended claims, the singular
forms "a," "an," and "the" include plural reference unless the
context clearly indicates otherwise. For example, reference to a
"lipid particle" is a reference to from one to many lipid
particles, such as molar amounts, and includes equivalents thereof
known to those skilled in the art, and so forth.
Enumerated Embodiments
[0118] The following enumerated embodiments are representative of
some aspects of the invention.
1. A composition comprising:
[0119] a lipid particle; and
[0120] an endogenous carbohydrate encapsulated in the lipid
particle.
2. The composition of embodiment 1, wherein the lipid particle
comprises a molecule that is capable of minimizing degradation of
the lipid particle and/or enhancing retention of the lipid particle
when administered to a subject, and/or makes the lipid particle
immunotolerant when administered to a subject. 3. The composition
of embodiment 2, wherein the molecule is ethylene oxide, an
ethylene oxide oligomer, an ethylene oxide polymer, polyethylene
glycol (PEG), or any combination thereof. 4. The composition of
embodiment 2, wherein the molecule is ethylene oxide. 5. The
composition of embodiment 2, wherein the molecule is an ethylene
oxide oligomer or an ethylene oxide polymer. 6. The composition of
embodiment 2, wherein the molecule is polyethylene glycol (PEG). 7.
The composition of any one of embodiments 1-6, wherein the lipid
particle further comprises a lipid selected from the group
consisting of a phospholipid, a glycerolipid, a sphingolipid, and
any combination thereof; optionally wherein the lipid comprises
choline, ethanolamine, serine, glycerol, inositol, or any
combination thereof. 8. The composition of any one of embodiments
1-7, wherein the lipid particle further comprises a lipid having a
polar head group. 9. The composition of embodiment 8, wherein the
polar head group is selected from the group consisting of choline,
ethanolamine, serine, glycerol, inositol, and any combination
thereof. 10. The composition of any one of embodiments 1-7, wherein
the lipid particle further comprises a phospholipid. 11. The
composition of embodiment 10, wherein the phospholipid comprises
phosphatidylcholine (PC), phosphatidylethanolamine (PE),
phosphatidylserine (PS), phosphatidylglycerol (PG),
phosphatidylinositol (PI), or any combination thereof. 12. The
composition of any one of embodiments 1-11, wherein the lipid
particle further comprises a sterol. 13. The composition of
embodiment 12, wherein the sterol is cholesterol. 14. The
composition of embodiment 13, wherein the cholesterol is present at
a concentration of at most 15 molar percent. 15. The composition of
any one of embodiments 1-14, wherein the lipid particle further
comprises a fatty acid. 16. The composition of any one of
embodiments 1-15, wherein the lipid particle further comprises a
PEGylated neutral lipid. 17. The composition of any one of
embodiments 1-16, wherein the lipid particle is a lamellar
nanostructure comprising an amphiphilic lipid. 18. The composition
of any one of embodiments 1-17, wherein the lipid particle is
selected from the group consisting of a liposome, a micelle, a
solid lipid nanoparticle, and a niosome. 19. The composition of any
one of embodiments 1-17, wherein the lipid particle is a liposome.
20. The composition of embodiment 17, wherein the liposome
comprises polyethylene-glycol (PEG). 21. The composition of
embodiment 20, wherein the liposome is a stealth liposome. 22. The
composition of embodiment 21, wherein the stealth liposome is
immunotolerant. 23. The composition of any one of embodiments
19-22, wherein the liposome further comprises a phospholipid
selected from the group consisting of phosphatidylcholine (PC),
phosphatidylethanolamine (PE), phosphatidylserine (PS),
phosphatidylglycerol (PG), phosphatidylinositol (PI), and any
combination thereof. 24. The composition of any one of embodiments
19-22, wherein the liposome further comprises phosphatidylcholine
(PC) and phosphatidylethanolamine (PE). 25. The composition of any
one of embodiments 19-24, wherein the liposome further comprises
phosphatidylcholine (PC) at a concentration of at least 70 molar
percent. 26. The composition of any one of embodiments 19-25,
wherein the liposome further comprises a PEGylated neutral lipid.
27. The composition of embodiment 26, wherein the PEGylated neutral
lipid is present in the lipid particle at a concentration of at
least 1 molar percent. 28. The composition of any one of
embodiments 19-27, wherein the liposome further comprises
cholesterol. 29. The composition of any one of embodiments 19-28,
wherein the liposome further comprises a fatty acid. 30. The
composition of embodiment 29, wherein the fatty acid comprises a
carbon chain that ranges in length from 14-18 carbon atoms. 31. The
composition of any one of embodiments 1-17, wherein the lipid
particle is a micelle. 32. The composition of any one of
embodiments 1-17, wherein the lipid particle is a solid lipid
nanoparticle. 33. The composition of any one of embodiments 1-17,
wherein the lipid particle is a niosome. 34. The composition of any
one of embodiments 1-33, wherein the endogenous carbohydrate is
selected from the group consisting of a monosaccharide, a
phosphorylated monosaccharide, a disaccharide, a phosphorylated
disaccharide, an oligosaccharide, a phosphorylated oligosaccharide,
a polysaccharide, a phosphorylated polysaccharide, mannose, a
phosphorylated mannose, a mannofuranose, a phosphorylated
mannofuranose, a mannopyranos, a phosphorylated mannopyranos,
mannose-1-phosphate, a nucleotide sugar, a uridine diphosphate, a
guanine diphosphate, a cytosine monophosphate, fucose, GDP-fucose,
a sialic acid, CMP-sialic acid, N-acetylneuraminic acid (Neu5Ac),
and CMP-Neu5Ac. 35. The composition of any one of embodiments 1-33,
wherein the endogenous carbohydrate is mannose-1-phosphate. 36. The
composition of any one of embodiments 1-33, wherein the endogenous
carbohydrate is GDP-fucose. 37. The composition of any one of
embodiments 1-33, wherein the endogenous carbohydrate is CMP-sialic
acid. 38. The composition of any one of embodiments 1-37, wherein
the lipid particle has an average particle size that ranges from
0.05 to 0.5 microns in diameter. 39. The composition of any one of
embodiments 1-38, wherein the lipid particle is capable of
delivering the endogenous carbohydrate to a cell interior. 40. The
composition of any one of embodiments 1-39, wherein the composition
when administered to a subject in need thereof induces at least a
0.05-fold to at least a 3-fold increase in cellular production of
higher-order lipid-linked oligosaccharides in the subject, as
compared to cellular production of higher-order lipid-linked
oligosaccharides in the subject in the absence of administering the
composition to the subject. 41. The composition of embodiment 40,
wherein the higher-order lipid-linked oligosaccharides comprise
Man4GlcNAc2, Man5GlcNAc2, Man6GlcNAc2, Man7GlcNAc2, Man8GlcNAc2,
Man9GlcNAc2, or any combination thereof. 42. A composition
comprising:
[0121] a lipid particle comprising ethylene oxide, an ethylene
oxide oligomer, an ethylene oxide polymer, polyethylene glycol
(PEG), or any combination thereof; and
[0122] a carbohydrate encapsulated in the lipid particle.
43. The composition of embodiment 42, wherein the carbohydrate is
an endogenous carbohydrate. 44. The composition of embodiment 43,
wherein the endogenous carbohydrate is selected from the group
consisting of a monosaccharide, a phosphorylated monosaccharide, a
disaccharide, a phosphorylated disaccharide, an oligosaccharide, a
phosphorylated oligosaccharide, a polysaccharide, a phosphorylated
polysaccharide, mannose, a phosphorylated mannose, a mannofuranose,
a phosphorylated mannofuranose, a mannopyranos, a phosphorylated
mannopyranos, mannose-1-phosphate, a nucleotide sugar, a uridine
diphosphate, a guanine diphosphate, a cytosine monophosphate,
fucose, GDP-fucose, a sialic acid, CMP-sialic acid,
N-acetylneuraminic acid (Neu5Ac), and CMP-Neu5Ac. 45. The
composition of embodiment 42, wherein the carbohydrate is
mannose-1-phosphate. 46. The composition of embodiment 42, wherein
the carbohydrate is GDP-fucose. 47. The composition of embodiment
42, wherein the carbohydrate is CMP-sialic acid. 48. The
composition of any one of embodiments 42-47, wherein the lipid
particle comprises an ethylene oxide oligomer. 49. The composition
of any one of embodiments 42-47, wherein the lipid particle
comprises an ethylene oxide polymer. 50. The composition of any one
of embodiments 42-47, wherein the lipid particle comprises
polyethylene glycol (PEG). 51. The composition of any one of
embodiments 42-50, wherein the lipid particle further comprises
choline, ethanolamine, serine, glycerol, inositol, or any
combination thereof. 52. The composition of any one of embodiments
42-51, wherein the lipid particle further comprises a lipid
selected from the group consisting of a phospholipid, a
glycerolipid, a sphingolipid, and any combination thereof. 53. The
composition of embodiment 52, wherein the lipid comprises a polar
head group. 54. The composition of embodiment 53, wherein the polar
head group is selected from the group consisting of choline,
ethanolamine, serine, glycerol, inositol, and any combination
thereof. 55. The composition of any one of embodiments 42-52,
wherein the lipid particle further comprises a phospholipid. 56.
The composition of embodiment 55, wherein the phospholipid is
selected from the group consisting of phosphatidylcholine (PC),
phosphatidylethanolamine (PE), phosphatidylserine (PS),
phosphatidylglycerol (PG), phosphatidylinositol (PI), and any
combination thereof. 57. The composition of any one of embodiments
42-56, wherein the lipid particle further comprises a sterol. 58.
The composition of embodiment 57, wherein the sterol is
cholesterol. 59. The composition of embodiment 58, wherein the
cholesterol is present at a concentration of at most 15 molar
percent. 60. The composition of any one of embodiments 42-59,
wherein the lipid particle further comprises a fatty acid. 61. The
composition of any one of embodiments 42-60, wherein the lipid
particle further comprises a PEGylated neutral lipid. 62. The
composition of any one of embodiments 42-61, wherein the lipid
particle is a lamellar nanostructure comprising an amphiphilic
lipid. 63. The composition of any one of embodiments 42-62, wherein
the lipid particle is selected from the group consisting of a
liposome, a micelle, a solid lipid nanoparticle, and a niosome. 64.
The composition of any one of embodiments 42-62, wherein the lipid
particle is a liposome. 65. The composition of embodiment 64,
wherein the liposome comprises polyethylene-glycol (PEG). 66. The
composition of embodiment 65, wherein the liposome is a stealth
liposome. 67. The composition of embodiment 66, wherein the stealth
liposome is immunotolerant. 68. The composition of any one of
embodiments 64-67, wherein the liposome further comprises a
phospholipid selected from the group consisting of
phosphatidylcholine (PC), phosphatidylethanolamine (PE),
phosphatidylserine (PS), phosphatidylglycerol (PG),
phosphatidylinositol (PI), and any combination thereof. 69. The
composition of any one of embodiments 64-67, wherein the liposome
further comprises phosphatidylcholine (PC) and
phosphatidylethanolamine (PE). 70. The composition of any one of
embodiments 64-69, wherein the liposome comprises
phosphatidylcholine (PC) at a concentration of at least 70 molar
percent. 71. The composition of any one of embodiments 64-70,
wherein the liposome further comprises a PEGylated neutral lipid.
72. The composition of embodiment 71, wherein the PEGylated neutral
lipid is present in the lipid particle at a concentration of at
least 1 molar percent. 73. The composition of any one of
embodiments 64-72, wherein the liposome further comprises
cholesterol. 74. The composition of any one of embodiments 64-73,
wherein the liposome further comprises a fatty acid. 75. The
composition of embodiment 74, wherein the fatty acid comprises a
carbon chain that ranges in length from 14-18 carbon atoms. 76. The
composition of any one of embodiments 42-62, wherein the lipid
particle is a micelle. 77. The composition of any one of
embodiments 42-62, wherein the lipid particle is a solid lipid
nanoparticle. 78. The composition of any one of embodiments 42-62,
wherein the lipid particle is a niosome. 79. The composition of any
one of embodiments 42-78, wherein the lipid particle has an average
particle size that ranges from 0.05 to 0.5 microns in diameter. 80.
The composition of any one of embodiments 42-79, wherein the lipid
particle is capable of delivering the carbohydrate to a cell
interior. 81. The composition of any one of embodiments 42-80,
wherein the composition when administered to a subject induces at
least a 0.05-fold to at least a 3-fold increase in cellular
production of higher-order lipid-linked oligosaccharides in the
subject, as compared to cellular production of higher-order
lipid-linked oligosaccharides in the subject in the absence of the
composition. 82. The composition of embodiment 81, wherein the
higher-order lipid-linked oligosaccharides comprise Man4GlcNAc2,
Man5GlcNAc2, Man6GlcNAc2, Man7GlcNAc2, Man8GlcNAc2, Man9GlcNAc2, or
any combination thereof. 83. A composition comprising:
[0123] a lipid particle; and
[0124] a carbohydrate encapsulated in the lipid particle,
[0125] wherein the lipid particle comprises choline, ethanolamine,
glycerol, inositol, or any combination thereof.
84. The composition of embodiment 83, wherein the lipid particle
further comprises a molecule that is capable of minimizing
degradation of the lipid particle and/or enhancing retention of the
lipid particle when administered to a subject, and/or makes the
lipid particle immunotolerant when administered to a subject. 85.
The composition of embodiment 83 or 84, wherein the carbohydrate is
an endogenous carbohydrate. 86. The composition of embodiment 85,
wherein the endogenous carbohydrate is selected from the group
consisting of a monosaccharide, a phosphorylated monosaccharide, a
disaccharide, a phosphorylated disaccharide, an oligosaccharide, a
phosphorylated oligosaccharide, a polysaccharide, a phosphorylated
polysaccharide, mannose, a phosphorylated mannose, a mannofuranose,
a phosphorylated mannofuranose, a mannopyranos, a phosphorylated
mannopyranos, mannose-1-phosphate, a nucleotide sugar, a uridine
diphosphate, a guanine diphosphate, a cytosine monophosphate,
fucose, GDP-fucose, a sialic acid, CMP-sialic acid,
N-acetylneuraminic acid (Neu5Ac), and CMP-Neu5Ac. 87. The
composition of embodiment 83 or 84, wherein the carbohydrate is
mannose-1-phosphate. 88. The composition of embodiment 83 or 84,
wherein the carbohydrate is GDP-fucose. 89. The composition of
embodiment 83 or 84, wherein the carbohydrate is CMP-sialic acid.
90. The composition of any one of embodiments 83-89, wherein the
lipid particle comprises ethylene oxide, an ethylene oxide
oligomer, an ethylene oxide polymer, polyethylene glycol (PEG), or
any combination thereof. 91. The composition of embodiment 90,
wherein the lipid particle comprises an ethylene oxide oligomer.
92. The composition of embodiment 90, wherein the lipid particle
comprises an ethylene oxide polymer. 93. The composition of
embodiment 90, wherein the lipid particle comprises polyethylene
glycol (PEG). 94. The composition of any one of embodiments 83-93,
wherein the lipid particle further comprises a lipid selected from
the group consisting of a phospholipid, a glycerolipid, a
sphingolipid, and any combination thereof; optionally wherein the
lipid has a polar head group. 95. The composition of embodiment 94,
wherein the polar head group is selected from the group consisting
of choline, ethanolamine, glycerol, inositol, and any combination
thereof. 96. The composition of any one of embodiments 83-93,
wherein the lipid particle further comprises a phospholipid. 97.
The composition of embodiment 96, wherein the phospholipid is
selected from the group consisting of phosphatidylcholine (PC),
phosphatidylethanolamine (PE), phosphatidylglycerol (PG),
phosphatidylinositol (PI), and any combination thereof. 98. The
composition of any one of embodiments 83-97, wherein the lipid
particle further comprises a sterol. 99. The composition of
embodiment 98, wherein the sterol is cholesterol. 100. The
composition of embodiment 99, wherein the cholesterol is present at
a concentration of at most 15 molar percent. 101. The composition
of any one of embodiments 83-100, wherein the lipid particle
further comprises a fatty acid. 102. The composition of any one of
embodiments 83-101, wherein the lipid particle further comprises a
PEGylated neutral lipid. 103. The composition of any one of
embodiments 83-102, wherein the lipid particle is a lamellar
nanostructure comprising an amphiphilic lipid. 104. The composition
of any one of embodiments 83-103, wherein the lipid particle is
selected from the group consisting of a liposome, a micelle, a
solid lipid nanoparticle, and a niosome. 105. The composition of
any one of embodiments 83-103, wherein the lipid particle is a
liposome. 106. The composition of embodiment 105, wherein the
liposome comprises polyethylene-glycol (PEG). 107. The composition
of embodiment 105 or 106, wherein the liposome is a stealth
liposome. 108. The composition of embodiment 107, wherein the
stealth liposome is immunotolerant. 109. The composition of any one
of embodiments 105-108, wherein the liposome further comprises a
phospholipid selected from the group consisting of
phosphatidylcholine (PC), phosphatidylethanolamine (PE),
phosphatidylglycerol (PG), phosphatidylinositol (PI), and any
combination thereof. 110. The composition of any one of embodiments
105-108, wherein the liposome further comprises phosphatidylcholine
(PC) and phosphatidylethanolamine (PE). 111. The composition of any
one of embodiments 105-110, wherein the liposome comprises
phosphatidylcholine (PC) at a concentration of at least 70 molar
percent. 112. The composition of any one of embodiments 105-111,
wherein the liposome further comprises a PEGylated neutral lipid.
113. The composition of embodiment 112, wherein the PEGylated
neutral lipid is present in the lipid particle at a concentration
of at least 1 molar percent. 114. The composition of any one of
embodiments 105-113, wherein the liposome further comprises
cholesterol. 115. The composition of any one of embodiments
105-114, wherein the liposome further comprises a fatty acid. 116.
The composition of embodiment 115, wherein the fatty acid comprises
a carbon chain that ranges in length from 14-18 carbon atoms. 117.
The composition of any one of embodiments 83-103, wherein the lipid
particle is a micelle. 118. The composition of any one of
embodiments 83-103, wherein the lipid particle is a solid lipid
nanoparticle. 119. The composition of any one of embodiments
83-103, wherein the lipid particle is a niosome. 120. The
composition of any one of embodiments 83-119, wherein the lipid
particle has an average particle size that ranges from 0.05 to 0.5
microns in diameter. 121. The composition of any one of embodiments
83-120, wherein the lipid particle is capable of delivering the
carbohydrate to a cell interior. 122. The composition of any one of
embodiments 83-121, wherein the composition when administered to a
subject induces at least a 0.05-fold to at least a 3-fold increase
in cellular production of higher-order lipid-linked
oligosaccharides in the subject, as compared to cellular production
of higher-order lipid-linked oligosaccharides in the subject in the
absence of the composition. 123. The composition of embodiment 122,
wherein the higher-order lipid-linked oligosaccharides comprise
Man4GlcNAc2, Man5GlcNAc2, Man6GlcNAc2, Man7GlcNAc2, Man8GlcNAc2,
Man9GlcNAc2, or any combination thereof. 124. A composition
comprising:
[0126] a liposome; and
[0127] mannose-1-phosphate encapsulated in the liposome,
wherein the liposome comprises cholesterol and
phosphatidylethanolamine (PE) attached to polyethylene glycol
(PEG). 125. The composition of embodiment 124, wherein the liposome
is a stealth liposome. 126. The composition of embodiment 125,
wherein the stealth liposome is immunotolerant. 127. The
composition of any one of embodiments 124-126, wherein the liposome
further comprises a phospholipid selected from the group consisting
of phosphatidylcholine (PC), phosphatidylserine (PS),
phosphatidylglycerol (PG), phosphatidylinositol (PI), and any
combination thereof. 128. The composition of any one of embodiments
124-126, wherein the liposome further comprises phosphatidylcholine
(PC). 129. The composition of embodiment 127 or 128, wherein the
liposome comprises phosphatidylcholine (PC) at a concentration of
at least 70 molar percent. 130. The composition of any one of
embodiments 124-129, wherein the liposome further comprises a
PEGylated neutral lipid. 131. The composition of embodiment 130,
wherein the PEGylated neutral lipid is present in the lipid
particle at a concentration of at least 1 molar percent. 132. The
composition of any one of embodiments 124-131, wherein the
cholesterol is present at a concentration of at most 15 molar
percent. 133. The composition of any one of embodiments 124-132,
wherein the liposome further comprises a fatty acid. 134. The
composition of embodiment 133, wherein the fatty acid comprises a
carbon chain that ranges in length from 14-18 carbon atoms. 135.
The composition of any one of embodiments 124-134, wherein the
liposome has an average particle size that ranges from 0.05 to 0.5
microns in diameter. 136. The composition of any one of embodiments
124-135, wherein the 1 liposome is capable of delivering the
mannose-1-phosphate to a cell interior. 137. The composition of any
one of embodiments 124-136, wherein the composition when
administered to a subject induces at least a 0.05-fold to at least
a 3-fold increase in cellular production of higher-order
lipid-linked oligosaccharides in the subject, as compared to
cellular production of higher-order lipid-linked oligosaccharides
in the subject in the absence of the composition. 138. The
composition of embodiment 137, wherein the higher-order
lipid-linked oligosaccharides comprise Man4GlcNAc2, Man5GlcNAc2,
Man6GlcNAc2, Man7GlcNAc2, Man8GlcNAc2, Man9GlcNAc2, or any
combination thereof. 139. Use of the composition of any one of
embodiments 1-138 in the manufacture of a medicament for delivering
a carbohydrate to a subject. 140. Use of the composition of any one
of embodiments 1-138 in the manufacture of a medicament for
delivering a carbohydrate to a cell interior. 141. Use of the
composition of any one of embodiments 1-138 in the manufacture of a
medicament for treating a congenital disorder of glycosylation
(CDG). 142. The composition of any one of embodiments 1-138 for use
in delivering a carbohydrate to a subject in need thereof. 143. The
composition of any one of embodiments 1-138 for use in delivering a
carbohydrate to a cell interior of a subject in need thereof. 144.
The composition of any one of embodiments 1-138 for use in treating
a congenital disorder of glycosylation (CDG) to a subject in need
thereof. 145. A pharmaceutical composition comprising the
composition of any one of embodiments 1-138, and a pharmaceutically
acceptable carrier. 146. A kit comprising the composition of any
one of embodiments 1-138. 147. The kit of embodiment 146, wherein
the kit further comprises a package insert comprising instructions
for using the composition to treat a congenital disorder of
glycosylation (CDG) in a subject in need of such treatment. 148.
The kit of embodiment 146, wherein the kit further comprises a
package insert comprising instructions for using the composition to
deliver a carbohydrate to a subject in need of such treatment. 149.
The kit of embodiment 146, wherein the kit further comprises a
package insert comprising instructions for using the composition to
deliver a carbohydrate to a cell interior. 150. A method for
delivering a carbohydrate to a subject in need thereof, comprising
administering to the subject the composition of any one of
embodiments 1-138. 151. The method of embodiment 150, wherein the
subject has a congenital disorder of glycosylation (CDG). 152. The
method of embodiment 151, wherein the congenital disorder of
glycosylation (CDG) is selected from the group consisting of a CDG
type I disorder, a CDG-Ia disorder, a CDG type II disorder, a
CDG-IIc disorder, and a CDG-IIf disorder. 153. The method of
embodiment 151, wherein the congenital disorder of glycosylation
(CDG) is a CDG-Ia disorder. 154. A method for delivering a
carbohydrate to a cell interior of a subject in need thereof,
comprising administering to the subject the composition of any one
of embodiments 1-138, wherein the administered composition
traverses the cell plasma membrane to deliver the carbohydrate to
the cell interior. 155. The method of embodiment 154, wherein the
subject has a congenital disorder of glycosylation (CDG). 156. The
method of embodiment 155, wherein the congenital disorder of
glycosylation (CDG) is selected from the group consisting of a CDG
type I disorder, a CDG-Ia disorder, a CDG type II disorder, a
CDG-IIc disorder, and a CDG-IIf disorder. 157. The method of
embodiment 155, wherein the congenital disorder of glycosylation
(CDG) is a CDG-Ia disorder. 158. A method for treating a congenital
disorder of glycosylation (CDG) in a subject in need thereof,
comprising administering to the subject the composition of any one
of embodiments 1-138. 159. The method of embodiment 158, wherein
the congenital disorder of glycosylation (CDG) is selected from the
group consisting of a CDG type I disorder, a CDG-Ia disorder, a CDG
type II disorder, a CDG-IIc disorder, and a CDG-IIf disorder. 160.
The method of embodiment 158, wherein the congenital disorder of
glycosylation (CDG) is a CDG-Ia disorder. 161. The method of any
one of embodiments 150-160, wherein administration of the
composition induces at least a 0.05-fold to at least a 2-fold
increase in cellular production of higher-order lipid-linked
oligosaccharides in the subject, as compared to cellular production
of higher-order lipid-linked oligosaccharides in the subject in the
absence of the composition. 162. The method of embodiment 161,
wherein the higher-order lipid-linked oligosaccharides comprise
Man4GlcNAc2, Man5GlcNAc2, Man6GlcNAc2, Man7GlcNAc2, Man8GlcNAc2,
Man9GlcNAc2, or any combination thereof. 163. The method of any one
of embodiments 150-162, wherein administration of the composition
minimizes degradation of the lipid particle and/or enhances
retention of the lipid particle when administered to the subject,
and/or makes the lipid particle immunotolerant when administered to
the subject. 164. The method of any one of embodiments 150-163,
wherein the composition is administered orally, topically,
dermally, nasally, intravenously, intramuscularly,
intraperitoneally, intracerobrospinally, intracranially,
intraspinally, subcutaneously, intra-articularly, intrasynovialy,
or intrathecaly. 165. Pharmaceutical nanocarriers of dolichol
pyrophosphate linked oligosaccharide to endoplasmic reticulum are
capable of treating all CDG type I disorders. 166. Pharmaceutical
nanocarriers of mannose-1 phosphate to cell interior are capable of
treating CDG-Ia disorder. 167. Pharmaceutical nanocarriers of
GDP-fucose to Golgi are capable of treating CDG-IIc disorder. 168.
Pharmaceutical nanocarriers of CMP-sialic acid to Golgi are capable
of treating CDG-IIf disorder.
EXAMPLES
[0128] The invention will be more fully understood by reference to
the following Examples. They should not, however, be construed as
limiting the scope of the invention. All citations throughout the
disclosure are hereby expressly incorporated by reference.
Abbreviations
[0129] "PC" corresponds to phosphatidylcholine.
[0130] "PE" corresponds to phosphatidylethanolamine.
[0131] "PEG" corresponds to
1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[amino(polyethylene
glycol)-2000] or just Poly-ethylene-glycol.
[0132] "M1P" corresponds to mannose-1-phosphate.
[0133] "PBS" corresponds to phosphate buffered saline solution.
[0134] "MWCO" corresponds to molecular weight cut-off membrane.
Materials and Methods
Liposomes
[0135] Materials for liposome preparation (PC, PE, and PE-PEG) were
purchased from Avanti Polar Lipids.
Cell Lines
[0136] CDG-Ia human dermal fibroblasts and wild-type human dermal
fibroblasts were purchased from Coriell Institute.
Culture Media
[0137] Dulbecco's min. Eagle's media was purchased from
Mediatech.
[0138] Alpha-minimal essential medium was purchased from
Mediatech.
[0139] Glutamine and penicillin-streptomycin were purchased from
Omega Scientific.
[0140] FBS was purchased from Thermo.
Example 1: Preparation of Mannose-1-Phosphate Liposomes
[0141] The following Example demonstrates the preparation of
liposomes containing mannose-1-phosphate (M1P-liposomes). The
structure of mannose-1-phosphate (M1P) is depicted in FIG. 1A.
While a specific isomer is depicted in FIG. 1A, it should be
understood that the carbohydrates used in the compositions and
methods provided herein may be any isomeric form, provided that
such carbohydrates are endogenous carbohydrates for a given
subject.
Liposome Preparation Method
[0142] Liposomes were prepared by thin-film hydration from
phosphatidylcholine (PC), cholesterol, and phosphatidylethanolamine
(PE) conjugated to poly-ethylene-glycol (PEG). The structure of PEG
is depicted in FIG. 1B. In FIG. 1B, in one variation, n may be in
the range of 800-5000. In other variations, the value of n may be
2000. In yet other variations, PEG may be modified either with
meleimide or NHS-ester depending on what the PEG will be conjugated
to. The structure of phosphatidylethanolamine attached to PEG
(PEG-PE) is depicted in FIG. 1C. The structure of
phosphatidylcholine (PC) is depicted in FIG. 1D.
[0143] The bulk solvent from the lipid film of PC and cholesterol
was removed by rotary evaporation for 30 minutes, followed by
freeze drying for 4 hours to remove trace amounts of solvent.
Following this, the liposomes were rehydrated first with a solution
of mannose-1-phosphate (M1P) in 1.times. phosphate buffered saline
pH 7.4 (PBS) at a M1P concentration of 10 mg/mL. Then, PBS was
further added so as to achieve a lipid concentration of 20 mg/mL.
Liposomes were then heated to 39.degree. C. and sized or extruded
through 200 nm pore size and then 100 nm pore size polycarbonate
membranes using a mini handheld extruder. Liposomes were then
dialyzed against PBS using a 2 kDa molecular weight cut-off
membrane (MWCO).
[0144] Following this step,
1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[amino(polyethylene
glycol)-2000] (PEG) was added by post-insertion. Briefly, a thin
film of PEG was obtained by drying off solvent using a stream of
N.sub.2 gas and freeze drying as mentioned previously. This film
was then rehydrated using the prepared M1P-loaded liposomal
solution. After vortexing for 5 minutes, the solution was incubated
at 37.degree. C. overnight to allow for post-insertion of PEG into
the core of the nanoparticle. The structure of the
mannose-1-phosphate-loaded liposomes is depicted in FIG. 2.
TABLE-US-00001 TABLE 1 Formulation Molar percentage (%) Weight (mg)
PC 70 16.7 Cholesterol 30 3.64 DSPE-PEG.sub.2000 2 1.75 M1P 5
1.0
[0145] In Table 1, the final volume of the liposome formulation was
1 mL.
Example 2: Measurement of M1P Content in Liposomes
[0146] CDG-Ia disorder is a gycosylation disorder characterized by
the deficiency or abnormalities associated with the enzyme
phosphomannomutase (PMM). This disorder is diagnosed by testing the
serum glycoproteins of hepatic origin (transferrin is the
clinically tested protein), which are commonly underglycosylated.
Typically transferrin is assessed using isoelectric focusing, which
would underscore a dramatic decrease in PMM activity (PMM activity
in CDG-Ia patients is approximately 0.29 nM/min/mg, as compared to
normal PMM activity of approximately 2.97 nM/min/mg). PMM enzyme is
responsible for catalyzing the conversion of mannose-6-phosphate to
mannose-1-phosphate, which is in turn a precursor for the ultimate
synthesis of G.sub.3M.sub.9Gn.sub.2-P-P-Dolichol, which in turn is
a precursor to lipid-linked-oligosaccharide (LLO). This is
represented by the following schematic representation of the
metabolic glycosylation pathway is:
glucose.fwdarw.glucose-6-phosphate.fwdarw.fructose-6-phosphate.fwdarw.man-
nose.fwdarw.mannose-6-phosphate.fwdarw.mannose-1-phosphate.fwdarw.GDP-mann-
ose.fwdarw.peptide N-glycosylation.
[0147] Typical LLO oligosaccharides include mannose subunits (M)
linked to an N-acetylglucosamine disaccharide (Gn.sub.2). The
decrease in PMM activity thus hampers the full maturation of LLO
oligosaccharides, which typically range in size from
M.sub.5Gn.sub.2 to M.sub.9Gn.sub.2. The resulting immature LLO
oligosaccharides are not fully functional LLO and are typically
truncated. Immature LLO oligosaccharides typically have sizes of
M.sub.2Gn.sub.2 and M.sub.3Gn.sub.2.
[0148] Analysis of synthesized oligosaccharide chains is typically
performed by incubating fibroblasts with radiolabeled mannose in
order to verify any divergence from established patterns of
oligosaccharide maturation. This is typically accomplished by
labeling newly-synthesized oligosaccharides within the human
fibroblasts with [2-.sup.3H]mannose. Following incubation with
labeled mannose, cell lysates are digested to further analyze the
labeled sugars by chromatography. However, a steady supply of
mannose is necessary to ensure a continuous production of the
saccharide chain. (S. Catherine Hubbard and Phillips W. Robbins.
Journal of Biological Chemistry (1980) 255: 11782-11793). A problem
with the approach of systematically supplying mannose within the
body to treat disorders, such as CDG-Ia, is the inability of
mannose to traverse the plasma membrane. Hence various mannose
derivatives have been evaluated for their ability to accumulate
within the cytosol of cells. However, this approach is also
problematic, due to the toxicity and instability of such
derivatives.
[0149] The following Example demonstrates the capacity of
M1P-liposomes to drive the biosynthesis and full maturation of
various LLO oligosaccharides. This approach bypasses the need to
either permeabilize cells or to use a mannose derivative.
Lipid Sample Preparation
[0150] Lipids were precipitated with 10% Triton X-100 using a ratio
of 1:1 liposome:detergent, respectively. Lipid precipitate was
removed by centrifugation. Isolated M1P was treated with HCl at a
final concentration of 0.1 N; 1-phospho linkage was hydrolyzed for
15 min at 100.degree. C. Samples were dried down in a speed vacuum
in preparation for labeling with a fluorophore for detection and
quantiation as described by Alwael et al. (Anal. Methods, 2011, 3,
62). Concentration of M1P in liposomes was 1.0 mg per milliliter of
formulation equivalent to 5% of total lipid concentration.
Therapeutic/Trials
[0151] Cellular internalization into the cytosol and subsequent
integration of the M1P-liposomes (as prepared in Example 1), was
assessed in vitro using primary human dermal fibroblasts derived
from patients affected with CDG-Ia disorder (CDG-Ia fibroblasts)
and wild-type human dermal fibroblasts. For evaluation of the
ability of the M1P-liposomes to correct protein glycosylation
defects, M1P-liposomes were delivered to CDG Type I primary
fibroblasts derived from patients affected with CDG-Ia disorder.
The ability to drive glycosylation forward was evaluated by
comparing glycoprotein N-glycan structure before and after
treatment. Such comparison required N-glycan analysis using normal
phase chromatography.
[0152] By utilizing endocytotic pathways, M1P-liposomes can enter
the cell interior and release their cargo into the cytosol. Due to
the ability to deliver a non-modified mannose-1-phosphate into the
cytosol, analysis was performed on the assembly of LLO
oligosaccharides in wild-type fibroblasts and fibroblasts derived
from a CDG-Ia patient (CDG-Ia fibroblasts). LLO structures were
isolated from the wild-type fibroblasts (having 100% PMM activity)
as well as from the CDG-Ia fibroblasts (having 10% PMM activity)
and the amounts of the LLO structures were compared using
chromatographic methods.
[0153] CDG-Ia dermal fibroblasts make a series of truncated
lipid-linked oligosaccharide (LLO) species ranging in size from
M.sub.2Gn.sub.2 to M.sub.5Gn.sub.2 with M.sub.2Gn.sub.2 and
M.sub.3Gn.sub.2 being predominant. The M1P liposomes from Example 1
were tested in order to evaluate their capacity to normalize the
distribution of various LLO oligosaccharides. CDG-Ia fibroblasts
cultured in low glucose DMEM media were incubated with 300 .mu.M of
bare M1P (M1P) and 300 .mu.M of the M1P-liposomes (M1PL). After
treatment, cells were harvested and LLO oligosaccharides were
recovered, hydrolyzed, modified with fluorophore, and analyzed by
normal phase chromatography as described by Gao et al. (Methods in
Enzymology, 2006, 415, 3-20) and Bones et al. (Anal. Chem, 2011, 83
(13), 5344-52). The peak area of each LLO was normalized to protein
content prior to calculation of percentage abundance.
[0154] The results are depicted in Table 2. The distribution of
various LLO oligosaccharides from wild-type fibroblasts ranged from
M.sub.2Gn.sub.2 to M.sub.7Gn.sub.2. Treatment of CDG-Ia fibroblasts
with M1P-liposomes inhibited the synthesis of truncated LLO
oligosaccharides M.sub.2 Gn.sub.2 and M.sub.3Gn.sub.2, and shifted
production towards larger LLO oligosaccharide species causing an
increase in expression of M.sub.5Gn.sub.2-M.sub.7Gn.sub.2
structures (Table 2). The amount of M.sub.5Gn.sub.2 LLO produced
after treatment of CDG-Ia fibroblasts with M1P-liposomes was three
times greater than the amount in untreated CDG-Ia fibroblasts, and
twice the amount in CDG-Ia fibroblasts treated with bare M1P (Table
2). Additionally, higher order LLO oligosaccharides (e.g., M.sub.6
Gn.sub.2 and M.sub.7Gn.sub.2) were observed in CDG-Ia fibroblasts
after treatment with M1P-liposomes (Table 2). Such higher order LLO
oligosaccharides were not observed in the CDG-Ia fibroblasts prior
to treatment with M1P-liposomes. Moreover, the amount of
M.sub.6Gn.sub.2 and M.sub.7Gn.sub.2LLO oligosaccharides in CDG-Ia
fibroblasts after treatment with M1P-liposomes was comparable to
the amount in wild-type fibroblasts (Table 2).
[0155] Complete normalization was not observed in all cases, as the
analytical approach that was utilized detected both newly
synthesized LLO oligosaccharides and LLO oligosaccharides
previously produced in the cell.
TABLE-US-00002 TABLE 2 Genotype Treatment M.sub.2Gn.sub.2
M.sub.3Gn.sub.2 M.sub.4Gn.sub.2 M.sub.5Gn.sub.2 M.sub.6Gn.sub.2
M.sub.7Gn.sub.2 Normal None 18 10 36 32 4.5 0.4 CDG-Ia None 31 38
24 6.2 CDG-Ia 300 .mu.M M1PL 18 28 31 18 3.9 0.5 CDG-Ia 300 .mu.M
M1P 40 24 27 8.9
[0156] Table 2 depicts normal phase chromatography analyses of LLO
oligosaccharide distribution (expressed as percentage, %) in
wild-type fibroblasts and CDG-Ia fibroblasts. The LLO
oligosaccharides contain two or more mannose (Man) subunits and an
N-acetylglucosamine disaccharide (GlcNAc2). In Table 2,
"M.sub.2Gn.sub.2" corresponds to Man2GlcNAc2, "M.sub.3Gn.sub.2"
corresponds to Man3GlcNAc2, "M.sub.4Gn.sub.2" corresponds to
Man4GlcNAc2, "M.sub.5Gn.sub.2" corresponds to Man5GlcNAc2,
"M.sub.6Gnc.sub.2" corresponds to Man6GlcNAc2, and
"M.sub.7Gn.sub.2" corresponds to Man7GlcNAc2.
* * * * *