U.S. patent application number 14/851966 was filed with the patent office on 2015-12-31 for method and system for measuring the pharmacokinetics of liposomal curcumin and its metabolite tetrahydrocurcumin.
The applicant listed for this patent is SignPath Pharma Inc.. Invention is credited to Lawrence Helson.
Application Number | 20150377894 14/851966 |
Document ID | / |
Family ID | 49756241 |
Filed Date | 2015-12-31 |
United States Patent
Application |
20150377894 |
Kind Code |
A1 |
Helson; Lawrence |
December 31, 2015 |
Method and System for Measuring the Pharmacokinetics of Liposomal
Curcumin and its Metabolite Tetrahydrocurcumin
Abstract
The present invention includes a stabilized curcumin
composition. The composition includes a curcumin composition and a
phosphate composition, wherein the phosphate composition is
non-buffering and is provided in an amount sufficient to stabilize
and/or prevent the degradation of curcumin and/or a curcuminoid in
a biological sample.
Inventors: |
Helson; Lawrence;
(Quakertown, PA) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
SignPath Pharma Inc. |
Quakertown |
PA |
US |
|
|
Family ID: |
49756241 |
Appl. No.: |
14/851966 |
Filed: |
September 11, 2015 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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13918112 |
Jun 14, 2013 |
9170257 |
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14851966 |
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61659660 |
Jun 14, 2012 |
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Current U.S.
Class: |
435/29 ; 424/450;
436/128; 514/679; 568/304 |
Current CPC
Class: |
A61P 29/00 20180101;
G01N 2500/10 20130101; G01N 33/5308 20130101; G01N 33/5091
20130101; G01N 33/64 20130101; A61K 31/12 20130101; A61P 17/02
20180101; A61K 9/127 20130101; A61P 35/00 20180101; G01N 33/50
20130101; G01N 33/84 20130101; C07C 45/86 20130101 |
International
Class: |
G01N 33/64 20060101
G01N033/64; A61K 9/127 20060101 A61K009/127; A61K 31/12 20060101
A61K031/12; C07C 45/86 20060101 C07C045/86 |
Claims
1. A method of determining a curcumin level in a sample comprising
the steps of: providing the sample suspected of comprising a
curcuminoid; adding a strong acid to the sample, wherein the strong
acid composition is non-buffering; and detecting the amount of
curcuminoid in the sample, wherein the non-buffering strong acid
reduces the degradation of the curcuminoid in the sample.
2. The method of claim 1, wherein the sample is an in vitro
sample.
3. The method of claim 1, wherein the sample is an aqueous sample,
a supernatant sample, a tears sample, a sputum sample, a blood
sample or a bile sample.
4. The method of claim 1, wherein the curcuminoid composition
comprises curcumin and analogues and derivatives selected from
curcumin; tetrahydrocurcumin; hexahydrocurcumin and
hexahydrocurcuminol; curcumin glucuronide; and curcumin
sulfate.
5. The method of claim 1, wherein the strong acid is selected from
at least one of a phosphoric acid; an orthophosphoric acid; a
phosphate salt; or a Na-phosphate.
6. The method of claim 1, wherein the curcuminoid composition
further comprises a liposome, a phospholipid or a polymer
composition to form an encapsulated curcuminoid composition.
7. The composition of claim 6, wherein the liposome, the
phospholipid or the polymer composition is selected from the group
consisting of phosphatidylcholine (lecithin), lysolecithin,
lysophosphatidylethanol-amine, phosphatidylserine,
phosphatidylinositol, sphingomyelin, phosphatidylethanolamine
(cephalin), cardiolipin, phosphatidic acid, cerebrosides,
dicetylphosphate, phosphatidylcholine, and
dipalmitoyl-phosphatidylglycerol, stearylamine, dodecylamine,
hexadecyl-amine, acetyl palmitate, glycerol ricinoleate, hexadecyl
sterate, isopropyl myristate, amphoteric acrylic polymers, fatty
acid, fatty acid amides, cholesterol, cholesterol ester,
diacylglycerol, and diacylglycerolsuccinate; or wherein the polymer
composition is selected from the group consisting of polyesters,
polylactides, polyglycolides, polycaprolactones, polyanhydrides,
polyamides, polyurethanes, polyesteramides, polydioxanones,
polyacetals, polyketals, polycarbonates, polyorthocarbonates,
polyorthoesters, polyphosphoesters, polyphosphazenes,
polyhydroxybutyrates, polyhydroxyvalerates, polyalkylene oxalates,
polyalkylene succinates, poly(malic acid), poly(amino acids),
copolymers, terpolymers, and combinations or mixtures thereof.
8. The composition of claim 6, wherein the encapsulated curcuminoid
composition has a size of about 10-900 nm.
9. A kit for detecting curcumin comprising: a first vial for a
biological sample, and a second vial with an amount of a strong
acid that is a non-buffering sufficient to stabilize a curcuminoid
in a biological sample; and a set of instructions for stabilizing a
curcumin sample using the non-buffering phosphate composition,
wherein the non-buffering phosphate composition comprises a
phosphoric acid; an orthophosphoric acid; a phosphate salt; or a
Na-phosphate to stabilize curcumin; tetrahydrocurcumin;
hexahydrocurcumin and hexahydrocurcuminol; curcumin glucuronide;
and curcumin sulfate.
10. A method of stabilizing stabilizing a curcumin composition in
plasma sample or a bile sample against degradation during an
analytical processes comprising the steps of: providing a sample
comprising a curcumin composition, wherein the sample is a bile
sample or a blood sample and the curcumin composition is selected
from curcumin; tetrahydrocurcumin; hexahydrocurcumin and
hexahydrocurcuminol; curcumin glucuronide; and curcumin sulfate;
adding a phosphate composition to the sample, wherein the phosphate
composition is non-buffering and is selected from a phosphoric
acid; an orthophosphoric acid; a phosphate salt; or a Na-phosphate;
and detecting the amount of curcuminoid in the sample, wherein the
non-buffering phosphate composition reduces the degradation of the
curcuminoid in the sample.
11. A stabilized curcumin composition comprising: a curcumin
composition and a phosphate composition, wherein the phosphate
composition is non-buffering and is provided in an amount
sufficient to at least one of reduce the degradation or stabilize
the curcumin in a sample.
12. The stabilized curcumin composition of claim 14, wherein the
curcumin composition selected from Curcumin; tetrahydrocurcumin;
hexahydrocurcumin and hexahydrocurcuminol; curcumin glucuronide;
and curcumin sulfate.
13. The stabilized curcumin composition of claim 14, wherein the
phosphate composition is selected from at least one of a phosphoric
acid, a orthophosphoric acid, a phosphate salt, or a
Na-phosphate.
14. The stabilized curcumin composition of claim 14, further
comprising a liposome to form a liposomal curcumin composition.
15. The stabilized curcumin composition of claim 14, wherein the
stabilized curcumin composition comprises a solution dosage
form.
16. A method of performing a clinical trial to evaluate a candidate
drug comprising a curcumin or curcuminoid believed to be useful in
treating a medical condition, the method comprising: (a) obtaining
a first tissue samples prior to providing the candidate substance
from tissue suspected from a set of patients; (b) administering the
candidate drug to a first subset of the patients, and a placebo to
a second subset of the patients; (c) repeating step (a) after the
administration of the candidate drug or the placebo; and (d)
obtaining a second tissue sample from the first and second set of
patients and stabilizing the curcumin or curcuminoids in the second
tissue samples by adding an effective amount of a non-buffering
phosphate; and (e) determining of there is a statistically
significant difference in the amount of curcumin or curcuminoids in
the second tissue samples between the first and second subset of
patients, wherein a statistically significant reduction indicates
that the candidate drug is useful in treating said disease
state.
17. The stabilized curcumin composition of claim 19, wherein the
curcumin composition selected from at least one of curcumin;
tetrahydrocurcumin; hexahydrocurcumin and hexahydrocurcuminol;
curcumin glucuronide; or curcumin sulfate.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application is a continuation of U.S. patent
application Ser. No. 13/918,112 filed Jun. 14, 2013, which is a
non-provisional application of U.S. Provisional Application Ser.
No. 61/659,660, filed Jun. 14, 2012, the entire contents of which
are incorporated herein by reference.
STATEMENT OF FEDERALLY FUNDED RESEARCH
[0002] None.
INCORPORATION-BY-REFERENCE OF MATERIALS FILED ON COMPACT DISC
[0003] None.
TECHNICAL FIELD OF THE INVENTION
[0004] The present invention relates generally to compositions and
methods for stabilizing curcumin and tetrahydrocurcumin (THC) and
in particular, to compositions and methods for stabilizing curcumin
and THC in plasma and bile against degradation occurring during
analytical processes by lowering the pH with phosphoric acid.
BACKGROUND OF THE INVENTION
[0005] Without limiting the scope of the invention, its background
is described in connection with methods of stabilizing curcumin and
THC in plasma and bile against degradation occurring during
analytical processes. Curcumin is the major yellow pigment of
turmeric, derived from the rhizome of the herb Curcuma longa Linn,
and has traditionally been used as a treatment for inflammation,
skin wounds, and tumors. In addition, preclinical animal models,
curcumin has shown cancer chemo preventive, antineoplastic and
anti-inflammatory properties. Curcumin
[1,7-bis(4-hydroxy-3-methoxyphenyl)-1,6-heptadiene-3,5-dione] has
the structure below:
##STR00001##
[0006] Curcumin acts as a scavenger of hydroxyl radical, superoxide
anion and singlet oxygen and other oxygen species. Curcumin plays a
role in cellular signal induction pathways pertinent to growth,
differentiation and malignant transformations, including inhibiting
protein kinases, c-Jun/AP-1 activation, prostaglandin biosynthesis,
and may play a role in the activation of the transcription factor
NF-.kappa.B. However, it has been thought that the bioavailability
of curcumin in animals remains low with a poor bioavailability
which may be related to its inadequate absorption and fast
metabolism. Indirect evidence suggests that curcumin is metabolized
in the intestinal tract where curcumin undergoes metabolic
O-conjugation to curcumin glucuronide and curcumin sulfate and
bioreduction to THC, hexahydrocurcumin and hexahydrocurcuminol.
Much of this is confirmed through examination and analysis of
curcumin present in samples (e.g., tissue extracts) before and
after treatment. In studies it has been shown that perorally
administered curcumin has poor bioavailability and only low or
non-measurable blood levels were observed. Others have administered
piperine along with curcumin to enhance the bioavailability of
curcumin; however, the level of enhancement was only modest and no
curcumin could be detected after 3 hours even when supplemented
with piperine.
[0007] U.S. Pat. No. 8,153,172, entitled "Composition to Enhance
the Bioavailability of Curcumin," discloses a composition having a
curcuminoid and an essential oil of turmeric. A composition having
a curcuminoid and an essential oil of turmeric, wherein the
essential oil is present in an amount sufficient to cause an
enhancement of bioavailability of curcumin when the composition is
administered to a human as compared to bioavailability of curcumin
obtained upon administration of a composition prepared without
adding essential oil to the curcuminoid. A method to prepare a
composition having a curcuminoid and an essential oil of
turmeric.
[0008] U.S. Pat. No. 7,067,159, entitled "Methods for Treating
Prostate Cancer with Herbal Compositions," discloses methods for
treating prostate cancer, comprising administration of a
composition comprising therapeutically effective amounts of
supercritical extracts of rosemary, turmeric, oregano and ginger;
and therapeutically effective amounts of hydroalcoholic extracts of
holy basil, ginger, turmeric, Scutellaria baicalensis, rosemary,
green tea, huzhang, Chinese goldthread, and barberry.
[0009] U.S. Pat. No. 7,060,733, entitled "Methods for Treating
Pancreatitis with Curcumin Compounds and Inhibitors of Reactive
Oxygen Species," discloses methods of treating, preventing,
modulating, attenuating, or inhibiting a disease or a disorder
associated with inflammation related to NF-.kappa.B activation in a
subject which comprises administering to the subject at least one
curcumin compound. Also disclosed are combination therapies
comprising the administration of at least one curcumin compound and
at least one ROS inhibitor. Pharmaceutical compositions and kits
are also disclosed.
[0010] U.S. Pat. No. 5,679,864, entitled "Process for the Synthesis
of Curcumin-Related Compounds," discloses a process for the
synthesis of curcumin and curcumin-related compounds by reacting
the enol form of a 2,4-diketone with a monocarbocyclic aldehyde in
the presence of an organic amine catalyst. The reactants are
dissolved in a highly polar, aprotic organic solvent. The
curcumin-related product is recovered in crystalline form by
precipitation from the reaction mass and solvent
recrystallization.
SUMMARY OF THE INVENTION
[0011] The present invention provides a method of stabilizing
curcumin and tetrahydrocurcumin in the plasma and bile against
degradation occurring during analytical processes by lowering the
pH with phosphoric acid. One embodiment of the present invention
provides a method of determining a curcumin level in a biological
sample by providing a biological sample comprising a curcuminoid
composition and adding a strong acid, e.g., a phosphate
composition, to the sample, wherein the phosphate composition is
non-buffering and detecting the amount of curcuminoid in the
sample, wherein the non-buffering strong acid reduces the
degradation of the curcuminoid in the sample. The biological sample
may be an in vitro sample and include an aqueous sample, a
supernatant sample, a tears sample, a sputum sample, a blood sample
or a bile sample. The curcuminoid composition may include curcumin
and analogues and derivatives selected from curcumin;
tetrahydrocurcumin; hexahydrocurcumin and hexahydrocurcuminol;
curcumin glucuronide; and curcumin sulfate and the phosphate
composition may include a phosphoric acid; an orthophosphoric acid;
a phosphate salt; or a Na-phosphate. The curcuminoid composition
may include a liposome, a phospholipid or a polymer composition to
form an encapsulated curcuminoid composition and the liposome, the
phospholipid or the polymer composition is selected from the group
consisting of phosphatidylcholine (lecithin), lysolecithin,
lysophosphatidylethanol-amine, phosphatidylserine,
phosphatidylinositol, sphingomyelin, phosphatidylethanolamine
(cephalin), cardiolipin, phosphatidic acid, cerebrosides,
dicetylphosphate, phosphatidylcholine, and
dipalmitoyl-phosphatidylglycerol, stearylamine, dodecylamine,
hexadecyl-amine, acetyl palmitate, glycerol ricinoleate, hexadecyl
sterate, isopropyl myristate, amphoteric acrylic polymers, fatty
acid, fatty acid amides, cholesterol, cholesterol ester,
diacylglycerol, and diacylglycerolsuccinate; or wherein the polymer
composition is selected from the group consisting of polyesters,
polylactides, polyglycolides, polycaprolactones, polyanhydrides,
polyamides, polyurethanes, polyesteramides, polydioxanones,
polyacetals, polyketals, polycarbonates, polyorthocarbonates,
polyorthoesters, polyphosphoesters, polyphosphazenes,
polyhydroxybutyrates, polyhydroxyvalerates, polyalkylene oxalates,
polyalkylene succinates, poly(malic acid), poly(amino acids),
copolymers, terpolymers, and combinations or mixtures thereof and
have a size of about 10-900 nm.
[0012] One embodiment of the present invention provides a
stabilized curcumin composition. The composition includes a
curcumin composition and a phosphate composition, wherein the
phosphate composition is non-buffering, wherein and wherein the
non-buffering strong acid reduces the degradation of the
curcuminoid in the sample. As a result the phosphate composition
does not include an aqueous solution consisting of a mixture of a
weak acid and its conjugate base or a weak base and its conjugate
acid. The curcumin composition may be a curcumin composition, a
modified curcumin composition or a product of a curcumin
degradation, for example, the curcumin composition may be selected
from curcumin; tetrahydrocurcumin; hexahydrocurcuminol; curcumin
glucuronide; curcumin sulfate or other related products. In
addition the curcumin composition may be a mixture of the 2 or more
modified curcumin compositions, a product of a curcumin
degradation, modified curcumin or synthetic curcumin compositions.
The phosphate composition is a phosphate containing composition
that is non-buffering and as a result is not mixture of a weak acid
and its conjugate base or a weak base and its conjugate acid, e.g.,
not a PBS. In one embodiment of the present invention, the
phosphate composition is a phosphoric acid. In other embodiments
the phosphate composition can be an orthophosphoric acid; a
phosphate salt; a Na-phosphate; a K-phosphate; or other counter ion
phosphate. In other embodiments the phosphate composition can be a
mixture of phosphate compositions as long as the final composition
is not a buffer, i.e., not mixture of a weak acid and its conjugate
base or a weak base and its conjugate acid. The stabilized curcumin
can further comprising a liposome to form a liposomal curcumin
composition and can be in any common dosage form known to the
skilled artisan including infusion nanoparticle, tablet, capsule,
liquid and the like.
[0013] One embodiment of the present invention includes a method of
analyzing a curcumin sample by providing a sample comprising a
curcumin composition and adding a phosphate composition to the
sample, wherein the phosphate composition is non-buffering and at
least one of stabilizes or reduced the degradation of the curcumin
in the sample. The method can then include the step of analyzing at
least one property of the sample and in some cases the sample is an
aqueous sample, a blood sample or a bile sample. As a result the
phosphate composition does not include an aqueous solution
consisting of a mixture of a weak acid and its conjugate base or a
weak base and its conjugate acid. The curcumin composition may be a
curcumin composition, a modified curcumin composition or a product
of a curcumin degradation, for example, the curcumin composition
may be selected from curcumin; tetrahydrocurcumin;
hexahydrocurcuminol; curcumin glucuronide; curcumin sulfate or
other related products. In addition the curcumin composition may be
a mixture of the 2 or more modified curcumin compositions, a
product of a curcumin degradation, modified curcumin or synthetic
curcumin compositions. The phosphate composition is a phosphate
containing composition that is non-buffering and as a result is not
mixture of a weak acid and its conjugate base or a weak base and
its conjugate acid, e.g., not a PBS. In one embodiment of the
present invention, the phosphate composition is a phosphoric acid.
In other embodiments the phosphate composition can be an
orthophosphoric acid; a phosphate salt; a Na-phosphate; a
K-phosphate; or other counter ion phosphate. In other embodiments,
the phosphate composition can be a mixture of phosphate
compositions as long as the final composition is not a buffer,
i.e., not mixture of a weak acid and its conjugate base or a weak
base and its conjugate acid.
[0014] One embodiment of the present invention provides a method of
stabilizing a curcumin or tetrahydrocurcumin sample by providing a
sample comprising curcumin composition and adding a phosphate
composition to the sample, wherein the phosphate composition is
non-buffering and at least one of stabilizes or reduced the
degradation of the curcumin in the sample. As a result, the
phosphate composition does not include an aqueous solution
consisting of a mixture of a weak acid and its conjugate base or a
weak base and its conjugate acid. The curcumin composition may be a
curcumin composition, a modified curcumin composition or a product
of a curcumin degradation; for example, the curcumin composition
may be selected from curcumin, tetrahydrocurcumin,
hexahydrocurcuminol, curcumin glucuronide, curcumin sulfate or
other related products. In addition, the curcumin composition may
be a mixture of the 2 or more modified curcumin compositions, a
product of a curcumin degradation, modified curcumin or synthetic
curcumin compositions. The phosphate composition is a phosphate
containing composition that is non-buffering and as a result is not
a mixture of a weak acid and its conjugate base or a weak base and
its conjugate acid, e.g., not a PBS. In one embodiment of the
present invention, the phosphate composition is a phosphoric acid.
In other embodiments the phosphate composition can be an
orthophosphoric acid, a phosphate salt, a Na-phosphate, a
K-phosphate, or other counter ion phosphate. In other embodiments,
the phosphate composition can be a mixture of phosphate
compositions as long as the final composition is not a buffer,
i.e., not a mixture of a weak acid and its conjugate base or a weak
base and its conjugate acid.
[0015] One embodiment of the present invention provides a curcumin
diagnostic kit including a non-buffering phosphate composition and
a set of instructions for stabilizing a curcumin sample using the
non-buffering phosphate composition, wherein the amount of a
non-buffering phosphate composition sufficient to stabilize a
curcuminoid in a biological sample, and wherein the non-buffering
phosphate composition comprises a phosphoric acid; an
orthophosphoric acid; a phosphate salt; or a Na-phosphate to
stabilize Curcumin; tetrahydrocurcumin; hexahydrocurcumin and
hexahydrocurcuminol; curcumin glucuronide; and curcumin sulfate. As
a result the phosphate composition does not include an aqueous
solution consisting of a mixture of a weak acid and its conjugate
base or a weak base and its conjugate acid.
[0016] Another embodiment of the present invention provides a
method of stabilizing a curcumin composition in plasma sample or a
bile sample against degradation during an analytical processes by
providing a sample comprising a curcumin composition, wherein the
sample is a bile sample or a blood sample and the curcumin
composition is selected from Curcumin; tetrahydrocurcumin;
hexahydrocurcumin and hexahydrocurcuminol; curcumin glucuronide;
and curcumin sulfate and adding a phosphate composition to the
sample, wherein the amount of a non-buffering phosphate composition
is sufficient to stabilize a curcuminoid in a biological sample. As
a result the phosphate composition does not include an aqueous
solution consisting of a mixture of a weak acid and its conjugate
base or a weak base and its conjugate acid. The curcumin
composition may be a curcumin composition, a modified curcumin
composition or a product of a curcumin degradation, for example,
the curcumin composition may be selected from curcumin;
tetrahydrocurcumin; hexahydrocurcuminol; curcumin glucuronide;
curcumin sulfate or other related products. In addition the
curcumin composition may be a mixture of the 2 or more modified
curcumin compositions, a product of a curcumin degradation,
modified curcumin or synthetic curcumin compositions. The phosphate
composition is a phosphate containing composition that is
non-buffering and as a result is not mixture of a weak acid and its
conjugate base or a weak base and its conjugate acid, e.g., not a
PBS. In one embodiment of the present invention, the phosphate
composition is a phosphoric acid. In other embodiments the
phosphate composition can be an orthophosphoric acid; a phosphate
salt; a Na-phosphate; a K-phosphate; or other counter ion
phosphate. In other embodiments the phosphate composition can be a
mixture of phosphate compositions as long as the final composition
is not a buffer, i.e., not mixture of a weak acid and its conjugate
base or a weak base and its conjugate acid. The stabilized curcumin
can further comprising a liposome to form a liposomal curcumin
composition and can be in any common dosage form known to the
skilled artisan including infusion nanoparticle, tablet, capsule,
liquid and the like.
[0017] Another embodiment of the present invention provides a
method of performing a clinical trial to evaluate a candidate drug
comprising a curcumin or curcuminoid believed to be useful in
treating a medical condition, the method comprising: (a) obtaining
a first tissue samples prior to providing the candidate substance
from tissue suspected from a set of patients; (b) administering the
candidate drug to a first subset of the patients, and a placebo to
a second subset of the patients; (c) repeating step (a) after the
administration of the candidate drug or the placebo; and (d)
obtaining a second tissue sample from the first and second set of
patients and stabilizing the curcumin or curcuminoids in the second
tissue samples by adding an effective amount of a non-buffering
phosphate; and (e) determining of there is a statistically
significant difference in the amount of curcumin or curcuminoids in
the second tissue samples between the first and second subset of
patients, wherein a statistically significant reduction indicates
that the candidate drug is useful in treating said disease
state.
BRIEF DESCRIPTION OF THE DRAWINGS
[0018] For a more complete understanding of the features and
advantages of the present invention, reference is now made to the
detailed description of the invention along with the accompanying
figures and in which:
[0019] FIGS. 1A-1D are graphs of the plasma levels of curcumin and
THC as a function of time after infusion.
[0020] FIGS. 2A-2D are graphs of the plasma, bile and urine
curcumin levels as a function of time after infusion.
DETAILED DESCRIPTION OF THE INVENTION
[0021] While the making and using of various embodiments of the
present invention are discussed in detail below, it should be
appreciated that the present invention provides many applicable
inventive concepts that can be embodied in a wide variety of
specific contexts. The specific embodiments discussed herein are
merely illustrative of specific ways to make and use the invention
and do not delimit the scope of the invention.
[0022] To facilitate the understanding of this invention, a number
of terms are defined below. Terms defined herein have meanings as
commonly understood by a person of ordinary skill in the areas
relevant to the present invention. Terms such as "a", "an" and
"the" are not intended to refer to only a singular entity, but
include the general class of which a specific example may be used
for illustration. The terminology herein is used to describe
specific embodiments of the invention, but their usage does not
delimit the invention, except as outlined in the claims.
[0023] The term "liposome" refers to a capsule wherein the wall or
membrane thereof is formed of lipids, especially phospholipid, with
the optional addition therewith of a sterol, especially
cholesterol.
[0024] As used herein, the term "in vivo" refers to being inside
the body. The term "in vitro" used as used in the present
application is to be understood as indicating an operation carried
out in a non-living system.
[0025] The terms "effective amount" or "therapeutically effective
amount" described herein means the amount of the subject compound
that will elicit the biological or medical response of a tissue,
system, animal or human that is being sought by the researcher,
veterinarian, medical doctor or other clinician.
[0026] The term "pharmaceutically acceptable" as used herein to
describe a carrier, diluent or excipient must be compatible with
the other ingredients of the formulation and not deleterious to the
recipient thereof.
[0027] The term "curcumin" as used herein to describe (i) curcumin
derivatives or combinations thereof dissolved or dispersed in an
aqueous or a non-aqueous solvent with one or more optional related
co-factors, proteins, antibodies, pain medications, and other
pharmaceutically active agents dissolved, dispersed or suspended in
the solvent, (ii) a suitable aqueous or non-aqueous dispersion
medium, wherein the one or more spherical liposomes are dispersed
in the dispersion medium, and (iii) one or more optional
excipients, diluents, extended or controlled release agents,
lubricants, preservatives or any combinations thereof.
[0028] The present invention provides the stabilization of curcumin
and/or THC in plasma; however, the stabilization is more
complicated than the acidification of plasma with H.sub.3PO.sub.4.
For example, in Phase 1, human plasma samples were stabilized with
the addition of Na-phosphate, however, the bench stability of both
curcumin and THC was minimal and no curcumin and/or THC were
detected after the samples sat on the bench at room temperature for
a couple of hours. The addition of Na-phosphate stabilized the
plasma, although not as efficient as with H.sub.3PO.sub.4. As a
result, one embodiment of the present invention provides the
stability of curcumin and THC in plasma by the addition of
Na-phosphate. Another embodiment of the present invention provides
the stability of curcumin and THC in plasma by the addition of
O-Phosphoric acid. Another embodiment of the present invention
provides the stability of curcumin and THC in plasma by the
addition of Na-phosphate and O-Phosphoric acid. O-Phosphoric acid
can be more easily incorporated in the plasma or body fluids. The
phosphate molecule is essential for stabilizing the curcumin and
THC, both the sodium phosphate and phosphoric acid can be used. In
contrast, a phosphate buffer does not stabilize curcumin and THC in
plasma. The phosphate buffer is not specific to which salt it is
made from and can be made with either sodium or potassium phosphate
(monobasic or dibasic) whereas sodium phosphate monobasic is
specific as is orthophosphoric acid. The main purpose of adding the
phosphate/phosphoric acid is to stabilize the curcumin and THC in
the plasma. As we have seen the presence of the phosphoric acid in
the plasma shows a higher concentration for the analytes. This
stabilisation process may as well be achieved by using other
phosphate salts known to the skilled artisan. The addition of
H.sub.3PO.sub.4 leads to higher curcumin concentrations because of
the shift in pH confirmation. For example, the pH of EDTA plasma is
7.95, which is a little higher than the pH of plasma without EDTA.
After the addition of 50 .mu.l % H.sub.3PO.sub.4 to 950 ml plasma
the pH was 4.4. As curcumin is stable at pH values below 5.5 the
addition of H.sub.3PO.sub.4 stabilizes curcumin in the plasma
samples. Other embodiments may use other acidification agents (e.g.
ACD, acidic citrate, etc.,) and may also use anticoagulants. In
addition, the urine tends to be more acidic with a pH of between 5
and 7; however, no stabilization was observed with
H.sub.3PO.sub.4.
[0029] The present invention provides a method of stabilizing
curcumin and THC in the plasma and bile against degradation
occurring during analytical processes by lowering the pH with
phosphoric acid. In one study of 4 dogs, 2 males and 2 females were
infused with 10 mg/kg liposomal curcumin (LIPOCURC.TM.) over 2
hours, and another 4 dogs, 2 males and 2 females were infused with
10 mg/kg liposomal curcumin (LIPOCURC.TM.) over 8 hours. Plasma
levels of curcumin and THC were obtained at necropsy 15 minutes
following the infusion. THC levels were 6.3-9.6 fold higher than
curcumin at both infusion rates suggesting a combination of a high
rate of enzymatic curcumin metabolism and a comparatively slower
rate of blood THC clearance. Compared to the 8 hour infusion, the 2
hour infusion levels of both curcumin and its metabolite THC were
significantly higher. The plasma half lives of both compounds
following the 2 hour infusion ranged from 0.4-0.7 hours, and was a
consequence of both hepatic and renal clearance. However at higher
plasma concentrations renal excretion predominates particularly
with THC. Enhanced clearance rates were noted during the 8 hour
infusions which prevented achieving a steady state. These
observations suggest that for hematopoietic malignancies including
leukemia, lymphoma, and bone marrow metastases, the 2 hour infusion
may be advantageous based upon higher concentration profiles, and
the unstimulated clearance rates.
[0030] The parenteral administration of liposomal curcumin
(LIPOCURC.TM.) with therapeutic intent poses several questions
relating to deciding an optimal rate of administration for patients
with neoplastic diseases. Options ranging from bolus intravenous
injections to constant infusions are impacted by enzymatic
metabolism, pH dependent degradation, renal and hepato-biliary
excretion mechanisms. During pre-clinical toxicological evaluation
in dogs, dose dependent hemolysis was noted following brief
infusions of 20 mg/kg and greater curcumin content. Ten mg/kg doses
infused over 2 hours were nontoxic. This same 2 hour infusion
schedule was used in an ascending dose Phase 1 trial in normal
human subjects where the highest intravenous dose administered (5
mg/kg) was without adverse reaction. To avoid toxicity from a
too-high C.sub.max we used a two hour infusion, however in view of
the unknown metabolic and elimination factors in dogs we compared 2
hour and 4 fold longer infusions (8 hours) to determine any
advantages.
[0031] Plasma concentration data arising from the infusion of
liposomal curcumin (LIPOCURC.TM.) in 8 dogs (4 females and 4 males)
of the Beagle breed were used. The results and analysis for the
study are presented for intravenous infusion dosing of a total dose
of 10 mg/kg infused over a period of either 2 or 8 hours. Plasma
levels of curcumin and its metabolite, THC were measured at timed
intervals post-dosing. All animals were euthanized and subject to
necropsy 15 minutes post-infusion and samples of tissues, plasma,
bile, and urine taken to determine, the tissue distribution and
pharmacokinetics of curcumin and THC following two different rates
of infusion and two different analyte preservation/stabilization
methods, e.g., with and/or without phosphoric acid
(H.sub.3PO.sub.4) and the plasma pharmacokinetics, urine and bile
levels of curcumin and THC reviewed. A summary of the treatment
groups is presented in Table 1 below.
TABLE-US-00001 TABLE 1 Summary of Treatment Groups Concentration of
Infusion Duration of Number of Beagle Dose Curcumin Rate Infusion
Dogs On Study.sup.a Groups (mg/kg) (mg/mL) mL/kg/hr (hr) M F Part
A, Liposomal 10 0.5 10 2 2 2 Curcumin Part B, Liposomal 10 0.125 10
8 2 2 Curcumin
Liposomal curcumin (LIPOCURC.TM.) was administered to 8 Beagle dogs
by intravenous infusion over two hours (Part A) or eight hours
(Part B). For the 2 hour infusion, blood samples were taken at
predose and 0.25, 0.5, 1.5 and at 2 hours during infusion and at 15
minutes post-infusion. For the 8 hour infusion, blood samples were
taken at predose and 0.25, 0.5, 1.5, 4, 4, 6 and at 8 hours during
infusion and at 15 minutes post-infusion.
[0032] For all groups, plasma curcumin and THC were determined
using a method developed by the Bioanalytical Department at
Nucro-Technics [1]. Bioanalysis was performed on two sets of
samples, one set that was treated with phosphoric acid and one set
that was not treated with phosphoric acid. Phosphoric acid was used
to treat one set of samples based on preliminary studies indicating
that phosphate increased the stability of curcumin and THC in the
tissue matrix. Values that were below the limit of quantification
were assigned a value of 0.
[0033] As there were no consistent differences between the plasma
levels of curcumin in male dogs or female dogs, the average plasma
concentrations from male and female dogs were used to perform the
PK analysis. Plasma concentration vs. time profiles were analyzed
using (unless otherwise stated) the data from 4 dogs. Plasma
profiles for the test articles are presented as the mean data.+-.SE
of 4 dogs. Average plasma concentrations were used to perform the
PK analysis. Plasma concentration vs. time profiles were analyzed
and the PK parameters estimated using WinNonlin Version 5.2.1
employing the intravenous infusion model with first order
elimination. Unless stated otherwise, the plasma concentration-time
profiles for the test articles are presented as the mean data.+-.SE
of 4 dogs.
[0034] FIGS. 1A-1D are graphs of the plasma levels of curcumin as a
function of time after infusion. FIG. 1A is a graph of the plasma
level of curcumin following a 2 hour infusion of 5 mg/kg/hr of
curcumin. FIG. 1B is a graph of the plasma level of curcumin
following an 8 hour infusion of 1.25 mg/kg curcumin. FIG. 1C is a
graph of the plasma level of THC following a 2 hour infusion of 5
mg/kg/hr curcumin. FIG. 1D is a graph of the plasma level of THC
following an 8 hour infusion of 1.25 mg/kg/hr curcumin. Values are
presented as the mean.+-.standard error of 4 dogs.
[0035] The plasma levels and AUC of curcumin and THC following
either 2 hours (high rate) or 8 hours (low rate) infusion were
clearly higher in the presence of phosphoric acid (Tables 2 and 3),
suggesting that phosphoric acid increased the stability of curcumin
and THC in plasma samples.
[0036] Table 2 below is a table of the AUC of plasma concentration
vs. time for curcumin and THC upon bioanalysis in the presence and
absence of phosphoric acid. Phosphoric acid was added to the plasma
samples in the form of phosphoric acid; C.sub.max represents the
observed value and AUC is the area under the curve to 15 minutes
post-infusion calculated using the linear trapezoidal rule.
TABLE-US-00002 AUC (ng/mL*hr) C.sub.max (ng/mL) Infusion Time
Curcumin THC Curcumin THC 2 hr 65 1318 46 891 2 hr + phosphate 394
3797 320 2983 8 hr 52 411 15 77 8 hr + Phosphate 187 1171 66
293
[0037] Table 3 below is a table of the plasma concentration vs.
time for curcumin and THC upon bioanalysis in the presence and
absence of phosphoric acid.
TABLE-US-00003 Infusion Rate [Plasma], ng/mL [Plasma + PO.sub.4],
ng/mL and Time Curcumin THC Curcumin THC 5 mg/kg/hr Pre-Dose 0 .+-.
0 0 .+-. 0 0 .+-. 0 0 .+-. 0 15 min 20 .+-. 2 483 .+-. 50 8 .+-. 3
284 .+-. 89 30 min 25 .+-. 5 566 .+-. 77 77 .+-. 39 1116 .+-. 318
90 min.sup.2 36 .+-. 3 891 .+-. 238 319 .+-. 91 2352 .+-. 441 2 hr
46 .+-. 23 454 .+-. 79 257 .+-. 46 2983 .+-. 852 1.25 mg/kg/hr
Pre-Dose 0 .+-. 0 0 .+-. 0 0 .+-. 0 0 .+-. 0 15 min 0 .+-. 0 72
.+-. 15 13 .+-. 8 59 .+-. 24 30 min 5 .+-. 2 63 .+-. 15 32 .+-. 12
121 .+-. 28 90 min 9 .+-. 1 64 .+-. 14 65 .+-. 16 293 .+-. 73 2 hr
3 .+-. 1 68 .+-. 11 38 .+-. 14 226 .+-. 64 4 hr 12 .+-. 1 77 .+-. 8
30 .+-. 28 193 .+-. 127 6 hr 0 .+-. 0 0 .+-. 0 0 .+-. 0 64 .+-. 10
8 hr 15 .+-. 4 67 .+-. 29 6 .+-. 2 62 .+-. 26
Values are presented as the mean.+-.SE of 4 values. This was also
the case for bile, but less so, while for urine the impact of the
addition of phosphoric acid was variable.
[0038] FIGS. 2A-2D are graphs of the plasma, bile, and urine
curcumin post-infusion levels as a function of time after infusion
plasma, bile, and urine levels. FIG. 2A is a graph of curcumin
levels following a 2 hour infusion of 5 mg/kg/hr curcumin. FIG. 2B
is a graph of curcumin levels following an 8 hour infusion of 1.25
mg/kg curcumin. FIG. 2C is a graph of THC levels following a 2 hour
infusion of 5 mg/kg/hr curcumin. FIG. 2D is a graph of THC levels
following an 8 hour infusion of 1.25 mg/kg/hr curcumin. Table 3
shows THC in the absence of phosphoric acid, the value is presented
as the mean.+-.SE of three determinations, otherwise all values are
presented as the mean.+-.standard error of 4 dogs.
[0039] Equivocal data for the bioanalysis of curcumin in the plasma
of rats has been observed in the literature following oral
administration of high doses [2]. Detection methods rather than
plasma stability were speculated as the reason for the discrepancy,
however, it appears that plasma/tissue stability would also be an
issue in the bioanalysis of curcumin. One embodiment of the present
invention provides the quantification of curcumin and THC
stabilized by phosphoric acid in plasma, bile, and urine
samples.
[0040] Upon a 2 hour infusion of curcumin at 5 mg/kg/hr (total dose
10 mg/kg), the plasma levels of curcumin rose to attain a maximum
concentration of 320 ng/mL by 1.5 hours and then began to
stabilize/fall during the infusion. Upon cessation of the infusion,
there was a rapid drop in plasma concentrations of curcumin from
257 ng/mL to 65 ng/mL in 15 minutes. THC had a similar
concentration-time profile. For the 8 hour infusion of curcumin at
a rate of 1.25 mg/kg/hr (total dose 10 mg/kg), peak plasma
concentrations of 187 ng/mL were also reached by 1.5 hours and then
began to fall during the infusion period and thus, steady-state
levels were not achieved; a similar concentration-time profile was
also observed for THC. The ratio of THC to curcumin based on AUC
was 9.6 for the 2 hour infusion and 6.3 for the 8 hour infusion.
The drop in plasma levels of both curcumin and its metabolite, THC,
upon the 8 hour infusion suggests that infusion of curcumin may
activate or enhance its own elimination.
[0041] Computer assisted pharmacokinetic analysis of the plasma
concentration data was only shown for the 2 hour infusion. The
estimated PK parameters for curcumin and THC are shown in Table 4,
while the C.sub.max observed and calculated AUC are shown in Table
2.
[0042] Table 4 below illustrates the estimated PK parameters of
curcumin and THC. For a 2 hour intravenous infusion at a dose rate
of 2 mg/kg/hr; total dose 10 mg/kg. The estimated PK parameters
were determined by fitting the data to a first-order elimination
continuous intravenous infusion model.
TABLE-US-00004 Parameter Units Curcumin THC AUC ng*hr/mL 485 5185
C.sub.max ng/mL 233 2429 t.sub.1/2(e).sup.1 hr 0.4 0.5 Ke.sup.1
hr.sup.-1 1.6 1.4 MRT.sup.1 hr 0.6 0.7 CL L/hr/kg 20.6 Vss L/kg
12.7
[0043] The rapid decrease in plasma concentration of curcumin is
consistent with short t.sub.1/2(e) and MRT values of 0.4 and 0.6
hours respectively as a result of a high clearance of 20.6 L/kg/hr
from a volume of distribution of 12.7 L/kg. The fitted C.sub.max
and AUC values of 233 ng/mL and 485 ng*hr/mL are close to the
observed C.sub.max of 320 ng/mL and calculated AUC of 394 ng*hr/mL.
THC had estimated t.sub.1/2(e) and MRT values close to those of
curcumin with the estimated values being 0.5 and 0.7 hours,
respectively with C.sub.max and AUC values of 2429 ng/mL and 5185
ng*hr/mL, compared to the observed values of 2983 ng/mL and 3797
ng*hr/mL. The observed C.sub.max values for curcumin at infusion
dose rates of 1.25 and 5.0 mg/kg/hr were close to being
dose-proportional to the dosing rate, with dosing rate normalized
C.sub.max values (C.sub.max/Dosing rate in mg/kg/hr) of 64 and 53
ng/mL observed for the 2 and 8 hour infusions. The AUDs and
infusion dose rate normalized AUDs up to 2 hours for the high and
the low infusion rates were 354 and 82 ng*hr/mL and 59 and 66
ng*hr/mL, respectively, also consistent with
dose-proportionality.
[0044] Measurement of the levels of curcumin and THC in the plasma,
urine, and bile provide additional information concerning the
disposition of curcumin (FIG. 2A-2D; Table 5 below). For bile, the
levels of curcumin and THC were somewhat higher in female dogs
compared to the male dogs. At both the high and low infusion rate
of 1.25 mg/kg/hr, curcumin was found at higher concentrations in
the urine and bile compared to plasma. At the low infusion rate,
the urine and bile to plasma concentration ratios were 10 and 32,
respectfully while at the higher infusion rate, the values observed
were 44 and 16, respectfully.
[0045] Table 5 illustrates plasma, urine, and bile levels of
curcumin and THC 15 minutes, 2 hours and 8 hours post infusion.
TABLE-US-00005 Mattrix 2 hour 8 hour 2 hour 8 hour [Curcumin],
ng/mL [Curcumin + H.sub.3PO.sub.4], ng/mL Plasma 0 .+-. 0 0 .+-. 0
65 .+-. 28 14 .+-. 141 Urine 3657 .+-. 932 369 .+-. 247 2842 .+-.
170 148 .+-. 87 Bile 590 .+-. 224 292 .+-. 83 1028 .+-. 539 449
.+-. 96 [THC], ng/mL) [THC + H.sub.3PO.sub.4], (ng/mL) Plasma 38
.+-. 4 20 .+-. 42 1167 .+-. 379 142 .+-. 122 Urine 6417 .+-. 1450
2451 .+-. 84 3587 .+-. 1083 621 .+-. 206 Bile 187 .+-. 74 84 .+-.
12 391 .+-. 197 168 .+-. 53
Unless indicated otherwise, values are the mean.+-.SE of 4
determinations. Three values were 0 and one value was 58 ng/mL.
Mean.+-.SE of 3 determinations.
[0046] The liver and the kidney can eliminate curcumin from the
plasma and at higher plasma concentrations the kidney can excrete
more curcumin while biliary excretion is approaching saturation.
This is consistent with studies in rats where tissue disposition
studies of intravenously administered curcumin demonstrated the
highest exposure in the liver and kidney [3]. Modulation of renal
transporters may play an important role in the enhancement of the
elimination of curcumin previously mentioned. For THC the urine to
plasma concentration ratios were higher than the bile to plasma
concentration ratios, both at the low and high infusion rates, with
values of 3.1 and 4.4 compared to 0.3 and 1.2, respectively. This
is consistent with metabolism of curcumin to THC by the hepatic and
extra-hepatic tissues, accumulation of THC in the plasma and
excretion via the urine.
[0047] These data demonstrate drug stability, dose, and schedule of
administration represent important and malleable components of
curcumin clinical therapeutics. Tissue phenotype, metabolism,
excretion routes, transport mechanisms and distribution are
important but less subject to modification. Of these parameters
curcumin degradation prior to and during analytic procedures is
critically important and contributes to the variences and validity
of plasma levels reported in animal studies of oral and parenteral
curcumin administration. The high susceptibility to ambient light
and pH of curcumin was resolved by the addition of phosphoric acid
to stabilize curcumin prior to analytical processing.
[0048] Another factor contributing to misinformation regarding
curcumin blood levels in animal models is the effect of metabolic
activity. Curcumin can be released as free curcumin from any of the
delivery vehicles, and distributes mainly to circulating and tissue
lipids because of low aqueous solubility or is metabolized to a
number of secondary compounds via conjugation with glucuronides or
sulfates, or reduced to dihydrocurcumin, THC and octahydrocurcumin.
Although the specific and collective biological activity of these
metabolites in animal models has not been published. The
predominant reduced metabolite is THC and has a similar biological
activity to curcumin and can be converted by NADH-dependent
dihydrocurcumin by intestinal E. Coli. THC can also be converted
from curcumin via a specific enzyme reductase, which has a
molecular mass of 82 KDa and consists of two identical subunits
with a restricted substrate spectrum, preferentially acting on
curcumin. Its mechanism of action on curcumin is rendered in two
steps (i.e., two enzyme reactions). The first is a NADPH-dependent
reduction to an intermediate dihydrocurcumin and the second is
NADPH-dependent curcumin/dihydrocurcumin reductase to THC. The
enzyme is part of the medium chain dehydrogenase-reductase
superfamily, and its presence raises intriguing issues of enzyme
origins and distribution. It is found in the blood of mice
following intraperitoneal administration of curcumin, and it is
assumed that the enzyme is also present in human blood and tissues:
particularly the liver in humans. It is also found in a particular
strain of human origin intestinal E. coli: K-12 substr. MG1655
version 15.1. While there are no published studies reporting on
levels of this reducing enzyme in animal models, the significant
presence of THC in the plasma of the dogs strongly suggests the
presence of the enzyme in tissues and blood.
[0049] The addition of phosphoric acid to plasma and bile samples
in dogs prevented the degradation of curcumin and THC, which raises
issues of validity of published data on curcumin distribution and
excretion. Infusion of liposomal curcumin (LIPOCURC.TM.) in dogs at
two different infusion rates resulted in higher plasma levels of
curcumin and THC with a 2 hour infusion compared to an 8 hour
infusion. The C.sub.max and AUC.sub.2 hr normalized to the infusion
dose rate were proportional. The plasma levels of THC were higher
than curcumin with the ratio of plasma THC to curcumin ranging from
6.3-9.6. These data emphasize the putative presence of a curcumin
reducing enzyme in blood or tissues.
[0050] Analysis of the 2 hour curcumin infusion data provided
estimates of the plasma t.sub.1/2(e) and the mean residence times
(MRT) which were short, ranging from 0.4-0.7 hours. The short
plasma t.sub.1/2(e) and MRT are likely a consequence of the
clearance of curcumin by both hepatic and renal routes. Clearances
of curcumin and THC over 8 hours infusion are augmented, preventing
attainment of a steady-state. The mechanism may potentially be
through modulation of renal transporters. The present invention
provides a 2 hour infusion of curcumin, THC or curcumin and THC
would be preferable for liquid malignancies while the 8 hour
infusion of curcumin, THC or curcumin and THC for solid tumors in
the absence of tumor cell/tissue data.
[0051] In addition the present invention may be administered
intravenously a therapeutically effective amount of a
pharmaceutical composition curcumin, curcumin analogues, curcumin
derivatives or combinations thereof dissolved or dispersed in a
suitable aqueous or non-aqueous medium, wherein the curcumin is
enclosed in one or more spherical liposomes or is conjugated to one
or more biodegradable polymers. In another aspect the liposomes
comprise a lipid or a phospholipid wall, wherein the lipids or the
phospholipids are selected from the group consisting of
phosphatidylcholine (lecithin), lysolecithin,
lysophosphatidylethanol-amine, phosphatidylserine,
phosphatidylinositol, sphingomyelin, phosphatidylethanolamine
(cephalin), cardiolipin, phosphatidic acid, cerebrosides,
dicetylphosphate, phosphatidylcholine, and
dipalmitoyl-phosphatidylglycerol, stearylamine, dodecylamine,
hexadecyl-amine, acetyl palmitate, glycerol ricinoleate, hexadecyl
sterate, isopropyl myristate, amphoteric acrylic polymers, fatty
acid, fatty acid amides, cholesterol, cholesterol ester,
diacylglycerol, and diacylglycerolsuccinate. In a specific aspect
the one or more liposomes have a size of about 100 nm. In another
aspect the therapeutically effective amount comprises 50 nM/kg of
body weight of the subject. In yet another aspect the
pharmaceutical composition is optionally administered along with
related co-factors, proteins, antibodies, pain medications, and
other pharmaceutically active agents. In another aspect of the
method disclosed hereinabove the one or more pharmaceutically
active agents are selected from the group consisting of L-dopa,
Carbidopa, benserazide, Tolcapone, dopamine agonists bromocriptine,
pergolide, pramipexole, ropinirole, piribedil, cabergoline,
apomorphine, lisuride, MAO inhibitors, selegiline, and
rasagiline.
[0052] In one aspect of the composition disclosed hereinabove the
one or more spherical liposome or the polymer conjugate may be
dispersed in a dispersion medium, wherein the dispersion medium is
an aqueous or non-aqueous dispersion medium. In related aspects the
lipid or the phospholipid is selected from the group consisting of
phosphatidylcholine (lecithin), lysolecithin,
lysophosphatidylethanol-amine, phosphatidylserine,
phosphatidylinositol, sphingomyelin, phosphatidylethanolamine
(cephalin), cardiolipin, phosphatidic acid, cerebrosides,
dicetylphosphate, phosphatidylcholine, and
dipalmitoyl-phosphatidylglycerol, stearylamine, dodecylamine,
hexadecyl-amine, acetyl palmitate, glycerol ricinoleate, hexadecyl
sterate, isopropyl myristate, amphoteric acrylic polymers, fatty
acid, fatty acid amides, cholesterol, cholesterol ester,
diacylglycerol, and diacylglycerolsuccinate and the one or more
biodegradable polymers are selected from the group consisting of
polyesters, polylactides, polyglycolides, polycaprolactones,
polyanhydrides, polyamides, polyurethanes, polyesteramides,
polydioxanones, polyacetals, polyketals, polycarbonates,
polyorthocarbonates, polyorthoesters, polyphosphoesters,
polyphosphazenes, polyhydroxybutyrates, polyhydroxyvalerates,
polyalkylene oxalates, polyalkylene succinates, poly(malic acid),
poly(amino acids), copolymers, terpolymers, and combinations or
mixtures thereof.
[0053] In another aspect the composition is administered
intravenously, sub-cutaneously, intra-muscularly, or
intra-peritoneally. In a specific aspect the one or more liposomes
have a size of about 100 nm. In yet another aspect the composition
is administered intravenously.
[0054] In another aspect the present invention may include lipid or
the phospholipid selected from the group consisting of
phosphatidylcholine (lecithin), lysolecithin,
lysophosphatidylethanol-amine, phosphatidylserine,
phosphatidylinositol, sphingomyelin, phosphatidylethanolamine
(cephalin), cardiolipin, phosphatidic acid, cerebrosides,
dicetylphosphate, phosphatidylcholine, and
dipalmitoyl-phosphatidylglycerol, stearylamine, dodecylamine,
hexadecyl-amine, acetyl palmitate, glycerol ricinoleate, hexadecyl
sterate, isopropyl myristate, amphoteric acrylic polymers, fatty
acid, fatty acid amides, cholesterol, cholesterol ester,
diacylglycerol, and diacylglycerolsuccinate. In yet another aspect
the composition is administered intravenously, sub-cutaneously,
intra-muscularly or intra-peritoneally. In another aspect the one
or more liposomes have a size of about 100 nm. In a specific aspect
the composition is administered intravenously.
[0055] In specific aspects of the method described hereinabove the
one or more liposomes have a size of about 100 nm and the
therapeutically effective amount comprises 50 nM/kg of body weight
of the subject. In a related aspect the pharmaceutical composition
is optionally administered along with related co-factors, proteins,
antibodies, pain medications, and other pharmaceutically active
agents, wherein the pharmaceutically active agents comprise
serotonin reuptake inhibitors sertraline and paroxetine.
[0056] Tissue concentration data arising from the infusion of
liposomal curcumin in 8 (4 female and 4 male) Beagle dogs were used
to assemble this report. The results and analysis are presented for
12 tissue samples (brain cortex, hippocampus, striatum, brain stem,
heart, lungs, muscle, liver, kidney, pancreas, intestinal wall and
urinary bladder) following the termination of intravenous infusion
at a total dose of 10 mg/kg infused over a period of either 2 or 8
hours. Tissue levels of curcumin and its metabolite,
tetrahydrocurcumin (THC) were measured in animals that were killed
and subject to necropsy 15 minutes post-infusion to determine the
tissue distribution and pharmacokinetics of curcumin and THC
following two different rates of infusion and two different analyte
preservation/stabilization methods (with and without
H.sub.3PO.sub.4).
[0057] The test article will be administered to 8 Beagle dogs by
intravenous infusion over two hours (Part A) or eight hours (Part
B) as shown in Table 6.
TABLE-US-00006 TABLE 6 Summary of Treatment Groups Concentration of
Infusion Duration of Number of Beagle Dose Curcumin Rate Infusion
Dogs On Study.sup.a Groups (mg/kg) (mg/mL) mL/kg/hr (hr) M F 1.
Part A, Liposomal 10 0.5 10 2 2 2 Curcumin 2. Part B, Liposomal 10
0.125 10 8 2 2 Curcumin
[0058] Fifteen minutes following either the 2 hour or 8 hour
infusion, blood, urine and bile samples were taken, prior to the
dogs being necropsied and organs removed for the isolation of
tissues. Multiple samples of tissue weighing approximately 1 gram
were removed and snap frozen in the presence or absence of
phosphoric acid (H.sub.3PO.sub.4). For all tissue samples, the
levels of curcumin and THC were determined using a method developed
by the Bioanalytical Department at Nucro-Technics. Phosphoric acid
was used to treat one set of samples based on preliminary studies
indicating that phosphate increased the stability of curcumin and
THC in the tissue matrix. Values that were below the limit of
quantification were assigned a value of 0. As there were no
consistent differences between the tissue levels of curcumin in
males and female dogs, the average plasma concentrations from male
and female dogs was used to assess the tissue distribution results.
Tissue distribution data was analyzed using, unless stated, the
data from 4 dogs and are presented as the mean.+-.standard error
(S.E.).
[0059] The distribution of curcumin and THC in tissues is
illustrated in Tables 7-11. In general, curcumin and THC were
widely distributed amongst the 12 tissues assessed. While in plasma
the addition of phosphoric acid had a clear stabilizing effect on
both the levels of curcumin and THC, the effects in tissues was
less clear and to some extent tissue dependent and more evident for
THC. Thus, despite the high degree of variability for some tissues,
for brain tissue, phosphoric acid had a clear stabilizing effects,
again more prominent for THC, while in other tissues, the
stabilizing effect of phosphoric acid was minor or absent (i.e.
heart and kidney). These differences may arise as a consequence of
differing metabolic capabilities for each tissue.
TABLE-US-00007 TABLE 7 Tissue Distribution of Curcumin in the
Presence and Absence of H.sub.3PO.sub.4 following 2 hour infusions.
Levels (ng/g).sup.1 Tissue No H.sub.3PO.sub.4 S.E. Plus
H.sub.3PO.sub.4 S.E. Cortex, Brain 0.52 0.05 0.74 0.13 Hippocampus
0.09 0.00 0.09 0.09 Striatum 0.33 0.10 0.48 0.07 Brain Stem 0.30
0.04 0.45 0.06 Heart 0.49 0.08 0.48 0.09 Lungs 86.82 24.99 22.86
2.14 Muscle 1.23 0.32 0.19 0.02 Liver 4.28 1.90 1.82 0.45 Kidney
1.03 0.17 0.89 0.15 Pancreas 2.02 0.71 0.92 0.35 Intestinal Wall
2.97 0.98 1.14 0.26 Urinary Bladder 0.60 0.07 0.69 0.12
.sup.1Phosphate was added to the tissue samples in the form of
phosphoric acid
TABLE-US-00008 TABLE 8 Tissue Distribution of THC in the Presence
and Absence of H.sub.3PO.sub.4 following 2 hour infusions. Levels
(ng/g).sup.1 Tissue No H.sub.3PO.sub.4 S.E. Plus H.sub.3PO.sub.4
S.E. Cortex, Brain 0.68 0.05 3.08 0.30 Hippocampus 0.75 0.09 6.46
1.82 Striatum 6.22 3.10 11.12 1.42 Brain Stem 2.34 0.34 10.62 1.30
Heart 2.51 0.68 0.69 0.42 Lungs 24.99 5.11 2.14 2.67 Muscle 5.26
1.33 4.19 1.03 Liver 1.90 0.67 0.45 0.81 Kidney 3.06 0.63 4.25 0.61
Pancreas 2.02 0.71 0.92 0.35 Intestinal Wall 0.73 0.40 2.12 0.89
Urinary Bladder 0.84 0.20 0.87 0.34 .sup.1Phosphate was added to
the tissue samples in the form of phosphoric acid.
TABLE-US-00009 TABLE 9 Tissue Distribution of Curcumin in the
Presence and Absence of H.sub.3PO.sub.4 following 8 hour infusions.
Levels (ng/g).sup.1 Tissue No H.sub.3PO.sub.4 S.E. Plus
H.sub.3PO.sub.4 S.E. Cortex, Brain 0.72 0.18 0.81 0.15 Hippocampus
0.00 0.00 0.01 0.01 Striatum 0.15 0.02 0.49 0.08 Brain Stem 0.41
0.10 0.58 0.04 Heart 0.67 0.15 0.75 0.17 Lungs 317.93 101.28 250.75
56.42 Muscle 3.25 1.31 0.79 0.24 Liver 39.38 13.70 28.38 10.30
Kidney 2.71 0.65 2.77 1.04 Pancreas 1.88 0.62 2.84 0.76 Intestinal
Wall 1.79 0.53 0.84 0.17 Urinary Bladder 3.24 1.37 2.26 0.51
.sup.1Phosphate was added to the tissue samples in the form of
phosphoric acid.
TABLE-US-00010 TABLE 10 Tissue Distribution of THC in the Presence
and Absence of H.sub.3PO.sub.4 following 8 hour infusions. Levels
(ng/g).sup.1 Tissue No H.sub.3PO.sub.4 S.E. Plus H.sub.3PO.sub.4
S.E. Cortex, Brain 0.06 0.04 0.49 0.12 Hippocampus 0.01 0.01 1.13
0.35 Striatum 1.12 0.11 3.14 0.26 Brain Stem 0.83 0.08 3.02 0.37
Heart 0.51 0.08 0.03 0.03 Lungs 10.81 2.50 6.36 2.13 Muscle 0.38
0.25 0.93 0.20 Liver 2.63 0.62 2.25 0.57 Kidney 1.32 0.18 2.04 0.32
Pancreas 0.34 0.19 1.34 0.52 Intestinal Wall 0.31 0.31 0.21 0.12
Urinary Bladder 1.37 0.42 1.11 0.41 .sup.1Phosphate was added to
the tissue samples in the form of phosphoric acid
TABLE-US-00011 TABLE 11 H.sub.3PO.sub.4 stabilized Tissue Partition
Coefficients (Kp) for Curcumin and THC following 2 hour and 8 hour
infusions. Kp [tissue]/[plasma].sup.1 Tissue Curcumin, 2 hr THC, 2
hr Curcumin, 8 hr THC, 8 hr Cortex, Brain 0.0134 0.0006 0.0544
0.0047 Hippocampus 0.0016 0.0014 0.0007 0.0108 Striatum 0.0087
0.0039 0.0329 0.0300 Brain Stem 0.0081 0.0037 0.0389 0.0289 Heart
0.0087 0.0000 0.0503 0.0003 Lungs 0.4126 0.0078 16.8289 0.0609
Muscle 0.0034 0.0011 0.0530 0.0089 Liver 0.0329 0.0028 1.9047
0.0215 Kidney 0.0161 0.0025 0.1859 0.0195 Pancreas 0.0166 0.0017
0.1906 0.0128 Intestinal Wall 0.0206 0.0003 0.0564 0.0020 Urinary
Bladder 0.0125 0.0014 0.1517 0.0106
[0060] .sup.1The plasma concentrations used to calculate the tissue
partition coefficients were an average of the plasma concentration
measured at the end of the infusion period and 15 minutes post
infusion and were for 2 and 8 hours curcumin concentrations, 55.4
and 14.9 ng/mL, respectively and for 2 and 8 hours THC
concentrations, 810.9 and 104.5 ng/mL, respectively.
[0061] For the purpose of consistency with the discussion of the
plasma, bile and urine PK of curcumin and THC, the tissue
distribution results will be discussed for tissue levels determined
in the presence of phosphoric acid. Curcumin and THC were
distributed in all of the tissues investigated to different
extents. Following the 2 hours infusion, the tissue distribution
was high for curcumin in the lung (22.86 ng/g) compared to other
tissues (13-254-fold). The next highest tissue was the liver (1.82
ng/g), with distribution in other tissues ranging from 0.09-1.14
ng/g. The high distribution of curcumin into the lung may be due
related to fact that it is a very lipophilic compound. A similar
pattern was observed for THC following the 2 hour infusion, with
comparable tissue levels of THC to curcumin observed.
[0062] Upon 8 hours of infusion, albeit at a lower infusion
concentration, the extent of curcumin and THC changed. While the
lung and liver again had the highest and second highest levels of
curcumin, there were clearly increased concentrations of curcumin
and THC in the liver and lungs with 2 hours versus 8 hours levels
of 22.86 vs 250.75 ng/g and 1.82 vs 28.38 ng/mL, respectively. The
highest level in the lung observed, 250.75 ng/g of curcumin
translates into a tissue concentration of 0.68 .mu.M accepting that
1 gram tissue is equivalent to 1 mL of volume. Curcumin levels
ranged from 0.01-2.84 ng/g in other tissues. The levels of THC in
the pancreas, kidney and urinary bladder were also increased
following 8 hours of infusion, while other tissues were comparable
to those observed with the 2 hour infusion. The levels of THC were
also increased following 8 hours of infusion compared to 2 hours
infusion. The increased tissue incorporation of curcumin in the
lung and liver with 8 hours of infusion is consistent with the
previously reported inability to achieve steady-state plasma levels
of curcumin during 8 hours of infusion, further supporting an
enhancement of tissue uptake during the course of infusion. A
comparison of the tissue partition coefficients (Kp) further
support this point and sheds additional light on the impact of
short versus longer infusions of curcumin on tissue distribution in
dogs (Tables 10-11). Firstly, both following 2 and 8 hours of
infusion, the majority of the Kp values for curcumin and THC are
below one, suggesting a poor tissue distribution of curcuminoids
into tissues and consistent with the low oral bioavailability of
curcumin. Low Kp values have also been observed in rodent studies
and ranged from 0.06-0.25 in the rat. Exceptions to this are the
liver and lung with >1 values of 1.9 and 16.8 respectively with
8 hours of infusion. Secondly, the Kp values are higher for
curcumin than for THC, which to some extent makes sense with the
lower lipophilicity of THC. Thirdly, the Kp values are higher
amongst all tissues for both curcumin and THC following the 8 hour
infusion compared to the 2 hour infusion. This latter point highly
supports and enhancement of the tissue distribution of curcuminoids
with longer infusion. In the literature, curcumin has been reported
to inhibit the transporter mediated efflux of drugs from cells. At
the mechanistic level, this may indeed explain the increased uptake
of curcumin into tissues with a longer infusion and inability to
attain steady-state plasma levels. Essentially, as infusion
proceeds, curcumin levels build-up in tissues and begins to
progressively inhibit efflux, resulting in greater tissue
sequestration over time, the extent of which in any one tissue
being dependent on the balance between uptake and efflux
transporter activity. The higher levels of THC in tissues at 8
hours may be a consequence of the metabolism of the higher tissues
levels of curcumin. Thus, in addition to the conclusions reached
from analysis of the plasma levels of curcumin, the rapid clearance
of curcumin from the circulation in addition to the impact of the
liver and kidney, may also involve a number of tissues and be
dependent on their balance of transporter mediated uptake and
efflux. Curcumin and THC were distributed amongst all of the
tissues investigated with very high levels compared to other
tissues observed in the lung. The liver had the second highest
levels. With 8 hour infusion, the tissue levels of curcumin in the
lung and liver increased substantially compared to 2 hour infusion,
with the pancreas, kidney and urinary bladder also displaying
higher tissue levels. Tissue partition coefficients for curcumin
and THC were higher for the 8 hour infusion compared to the 2 hour
infusion, suggesting that prolonged infusion of curcumin may
facilitate tissue distribution via a transporter-dependent
mechanism.
TABLE-US-00012 TABLE 12 Effect of duration of a single dose: 10
mg/kg: 2 hours vs 8 hours intravenous curcumin infusion on the
ratio of tissue distribution of curcumin: THC in dogs.
Concentration of THC A Concentration of curcumin 2 h infusion >
8 h B C infusion 8 h infusion > 2 h infusion 8 h infusion = 2 h
infusion lung lung intestinal wall intestinal wall muscle heart
heart spleen bladder muscle liver brainstem bladder kidney cortex
spleen pancreas hippocampus liver striatum kidney pancreas
brainstem cortex striatum hippocampus
[0063] As seen in Table 12 above: In column A the THC
concentrations are higher in all 13 organs tested following a 2
hour infusion of liposomal curcumin compared to an 8 hour liposomal
curcumin infusion. In column B the curcumin concentrations of
following intravenous infusions of liposomal curcumin appear to be
both tissue specific and time dependent. The longer infusion (8
hour) distributes preferably to 6 tissues. In column C the curcumin
concentrations are not significantly different in the 2 hour and 8
hour infusions in 7 other tissues. Intertissue variance. Variance
following infusions may be due to several causes: vascular supply,
penetration, local tissue clearance/excretion, enzymatic reduction
to THC from curcumin.
TABLE-US-00013 TABLE 13 Average H.sub.3PO.sub.4 stabilized tissue
concentrations (ng/gm) of curcumin and THC of 4 dogs 2 male and 2
female following 2 hour and 4 dogs following 8 hour infusions of 10
mg/kg liposomal curcumin. Two hour infusion Eight hour infusion
Tissue Curcumin THC Curcumin THC Lung 22.86 9.37 250.75 6.36 Liver
1.81 4.58 28.38 2.24 Spleen 0.075 1.60 22.90 0.42 Pancreas 0.85
0.91 2.84 1.34 Kidney 0.89 4.25 2.76 2.03 Bladder 0.66 2.25 0.87
1.11 Heart 0.47 0.68 0.74 0.03 Intestinal wall 1.14 2.11 1.11 0.09
Muscle 0.18 4.19 0.68 2.42 Brainstem 0.45 10.6 0.57 3.02 Cortex
0.73 3.08 0.80 0.49 Striatum 0.48 11.11 0.49 3.13 Hippocampus 0.09
6.46 0.01 1.12
[0064] Interpretation. The distribution of intravenous liposomal
curcumin to various body tissues is not homogeneous, and it appears
that the lung, liver and spleen either collect or retain
significantly more curcumin than the remaining tissues. These data
show the 8 hour infusion leads to significantly higher levels of
curcumin in the lung, liver, spleen, pancreas, kidney, and muscle
hypothetically due to low enzymatic reduction to THC or decreased
clearance. In other tissues: muscle, bladder, heart, and intestinal
wall there is no significant difference. Levels of THC are
significantly reduced in all tissues receiving the 8 hour infusion.
These data reflect the net result of the tissue dependent presence
of reductive enzymes, the delivery of curcumin to the tissues
leading to lesser amounts of THC and the pharmacokinetic profile of
THC. The brain tissues are remarkably clear for supporting the
presence of THC over curcumin, in this case prolonged infusion
leads to greater clearance and lesser concentrations. The infusion
duration does affect curcumin and THC metabolism, and may have to
be taken into consideration when treating different tissue
pathologies. For example, cerebral disorders may be better treated
with brief infusions to achieve higher levels of THC, assuming THC
is equally or better effective against brain based disorders than
curcumin.
TABLE-US-00014 TABLE 14 H.sub.3PO.sub.4 stabilized vs
Non-stabilized tissue analysis following a 2 hour infusion of
Liposomal curcumin. Curcumin THC +H.sub.3PO.sub.4 -H.sub.3PO.sub.4
+H.sub.3PO.sub.4 -H.sub.3PO.sub.4 Lung 22.86 86.80 9.37 17.73
Spleen 0.07 0.48 1.60 1.35 Liver 1.81 4.28 4.58 2.41 Pancreas 0.85
2.81 0.91 2.01 Brainstem 0.45 0.32 10.60 2.33 Cortex 0.73 0.52 3.08
0.67 Striatum 0.48 0.32 11.11 6.21 Hippocampus 0.09 0.00 6.46
0.75
TABLE-US-00015 TABLE 15 H.sub.3PO.sub.4 stabilized vs
non-stabilized tissue analysis following an 2 hour infusion of
Liposomal curcumin. Curcumin THC +H.sub.3PO.sub.4 -H.sub.3PO.sub.4
+H.sub.3PO.sub.4 -H.sub.3PO.sub.4 Lung 250.75 317.90 6.36 10.81
Spleen 22.90 28.63 0.42 0.32 Liver 28.38 39.38 2.24 2.63 Pancreas
2.84 1.87 1.34 0.33 Brainstem 0.57 0.41 3.02 0.83 Cortex 0.80 0.72
0.49 0.12 Striatum 0.49 0.14 3.13 1.12 Hippocampus 0.01 0.00 1.12
0.01
[0065] Following the 2 hour infusion, with regard to curcumin,
higher levels were achieved in the absence of phosphoric acid
addition in the following tissues: lung, spleen, liver, pancreas,
while higher levels in all brain tissues examined were observed
with the addition of phosphoric acid. With regard to THC, all
brain, spleen and liver levels were higher with the addition of
phosphoric acid while lung and pancreas tissues were lower. The
patterns in the 8 hour infusions (Table 15) were as follows: higher
levels of curcumin were achieved in the absence of phosphoric acid
in the following tissues: lung, spleen, liver, while the addition
of phosphoric acid induced higher levels in the pancreas. There was
no significant on the impact of phosphoric acid on curcumin levels
in all brain tissues. Regarding THC levels, the addition of
phosphoric acid increased THC levels in all brain, and pancreatic
tissues. The absence of phosphoric acid addition was associated
with higher THC levels in lung tissue, but had no incremental
impact in other tissues.
[0066] Extrapolating to humans, and based upon the variances in THC
formation, and specific tissue levels of curcumin and THC following
8 hour and 2 hour infusions of liposomal curcumin including the
presence or absence of added phosphoric acid for stabilization,
designing administration schedules may best be adapted for specific
tissue pathologies in order to achieve optimum therapeutic results.
In a 60 kg adult, 370 mg/M2 is equivalent to 10 mg/kg/dose.
Converting 10 mg/kg dose in dogs to humans: .times.0.5=5.0
mg/kg/dose. Clinical applications: decision suggestions for either
2 hour or 8 hour or longer infusions of liposomal curcumin.
[0067] Lung disorders. Curcumin concentrations in the lung are
higher in the 8 hour infusion, than in the 2 hour infusion, and
levels are further elevated when analyzed in the presence of
phosphoric acid. Curcumin may have therapeutic value in treating
scleroderma, as it has already been shown to protect rats from lung
fibrosis induced by a variety of agents. THC concentrations in the
lung are higher after the 2 hour infusion than in the 8 hour
infusion, and levels are lower when analyzed in the presence of
phosphoric acid. Tetrahydrocurcumin has high anti-oxidant activity
potency in three bioassay models, i.e. the linoleic acid
auto-oxidation model, rabbit erythrocyte membrane ghost system, and
rat liver microsome system implying that hydrogenation of
curcuminoids increases anti-oxidant ability.
[0068] Liver disorders: Curcumin concentrations in the liver are
higher in the 8 hour infusion than in the 2 hour infusion, and
further elevated in the presence of phosphoric acid. THC
concentrations in the liver are higher after the 2 hour infusion
than the 8 hour infusion, and are increased in the presence of
phosphoric acid. In the 8 hour infusion there was no advantage to
adding phosphoric acid. Spleen disorders: Curcumin concentrations
in the spleen are higher in the 8 hour infusion than in the 2 hour
infusion, and further elevated in the presence of phosphoric acid.
Curcumin increases sub G1 cell populations with strong
apoptosis-inducing activity. THC concentrations in the spleen are
higher after the 2 hour infusion. Treatment with THC induced
autophagic cell death in human HL-60 promyelocytic leukemia cells
by increasing autophage marker acidic vascular organelle formation.
Flow cytometry also confirmed that THC treatment did not increase
sub-G1 cell population. Western blot analysis showed that THC
significantly down-regulated phosphatidylinositol 3-kinase/protein
kinase B and mitogen-activated protein kinase signalings including
decreasing the phosphorylation of mammalian target of rapamycin,
glycogen synthase kinase 3I.sup.2 and p70 ribosomal protein S6
kinase. Conclusion: these data demonstrated the anticancer efficacy
of THC by inducing autophagy, and provide prevention of human
leukemia. Myelofibrosis (MF) a significant disease burden: 85% of
myelofibrosis patients present with splenomegaly and 60% to 80% of
MF patients report spleen-related symptoms. In MF, splenomegaly of
any degree is clinically relevant, and since the majority of
patients with MF experience debilitating symptoms, appropriate
treatment should be considered. Muscle disorders: Curcumin
concentrations in muscle tissue are higher in the 8 hour infusion
than in the 2 hour infusion. Pancreatic disorders: Curcumin
concentrations in pancreatic disorders are higher in the 8 hour
infusion than in the 2 hour infusion. THC concentrations are higher
after the 2 hour infusion than after an 8 hour infusion. Kidney
disorders: Curcumin concentrations in renal disorders are higher in
the 8 hour infusion than in the 2 hour infusion. THC concentrations
in the kidney are higher after a 2 hour infusion, than after an 8
hour infusion. Neural disorders: Curcumin in the Brainstem, Cortex,
Striatum, and Hippocampus: either 2 hour or 8 hour infusions
produce similar concentrations which are unchanged by phosphoric
acid addition. Curcumin was effective in reducing amyloid plaque
burden, insoluble beta-amyloid peptide.
[0069] In the Parkinson's disease model, depletion of dopamine (DA)
and DOPAC (3, 4-dihydroxy phenyl acetic acid)) occurs with
increased monoamine oxidase (MAO-B) activity. Administration of
curcumin (80 mg/kg i.p.) and tetrahydrocurcumin (60 mg/kg i.p.)
significantly reversed the MPTP-induced depletion of DA and DOPAC.
The MAO-B activity was also significantly inhibited by these
compounds. Both curcumin and THC exert neuroprotection against MPTP
induced neurotoxicity. THC compared with curcumin gavage leads to
dramatically higher drug plasma levels, however resulting brain
levels of parent compounds were similar. Levels in the Brainstem,
Cortex, Striatum and Hippocampus are increased in the 2 hour
infusion and further increased by phosphoric acid in both 2 hour
and 8 hour infusions.
[0070] It is contemplated that any embodiment discussed in this
specification can be implemented with respect to any method, kit,
reagent, or composition of the invention, and vice versa.
Furthermore, compositions of the invention can be used to achieve
methods of the invention.
[0071] It will be understood that particular embodiments described
herein are shown by way of illustration and not as limitations of
the invention. The principal features of this invention can be
employed in various embodiments without departing from the scope of
the invention. Those skilled in the art will recognize, or be able
to ascertain using no more than routine experimentation, numerous
equivalents to the specific procedures described herein. Such
equivalents are considered to be within the scope of this invention
and are covered by the claims.
[0072] All publications and patent applications mentioned in the
specification are indicative of the level of skill of those skilled
in the art to which this invention pertains. All publications and
patent applications are herein incorporated by reference to the
same extent as if each individual publication or patent application
was specifically and individually indicated to be incorporated by
reference.
[0073] The use of the word "a" or "an" when used in conjunction
with the term "comprising" in the claims and/or the specification
may mean "one," but it is also consistent with the meaning of "one
or more," "at least one," and "one or more than one." The use of
the term "or" in the claims is used to mean "and/or" unless
explicitly indicated to refer to alternatives only or the
alternatives are mutually exclusive, although the disclosure
supports a definition that refers to only alternatives and
"and/or." Throughout this application, the term "about" is used to
indicate that a value includes the inherent variation of error for
the device, the method being employed to determine the value, or
the variation that exists among the study subjects.
[0074] As used in this specification and claim(s), the words
"comprising" (and any form of comprising, such as "comprise" and
"comprises"), "having" (and any form of having, such as "have" and
"has"), "including" (and any form of including, such as "includes"
and "include") or "containing" (and any form of containing, such as
"contains" and "contain") are inclusive or open-ended and do not
exclude additional, unrecited elements or method steps.
[0075] The term "or combinations thereof" as used herein refers to
all permutations and combinations of the listed items preceding the
term. For example, "A, B, C, or combinations thereof" is intended
to include at least one of: A, B, C, AB, AC, BC, or ABC, and if
order is important in a particular context, also BA, CA, CB, CBA,
BCA, ACB, BAC, or CAB. Continuing with this example, expressly
included are combinations that contain repeats of one or more item
or term, such as BB, AAA, AB, BBC, AAABCCCC, CBBAAA, CABABB, and so
forth. The skilled artisan will understand that typically there is
no limit on the number of items or terms in any combination, unless
otherwise apparent from the context.
[0076] All of the compositions and/or methods disclosed and claimed
herein can be made and executed without undue experimentation in
light of the present disclosure. While the compositions and methods
of this invention have been described in terms of preferred
embodiments, it will be apparent to those of skill in the art that
variations may be applied to the compositions and/or methods and in
the steps or in the sequence of steps of the method described
herein without departing from the concept, spirit and scope of the
invention. All such similar substitutes and modifications apparent
to those skilled in the art are deemed to be within the spirit,
scope and concept of the invention as defined by the appended
claims.
* * * * *