U.S. patent application number 15/837004 was filed with the patent office on 2018-04-12 for use of alginate compositions in preventing or reducing liver damage caused by a hepatotoxic agent.
This patent application is currently assigned to Hadasit Medical Research Services and Development Ltd.. The applicant listed for this patent is B.G. Negev Technologies & Applications Ltd., at Ben-Gurion University, Hadasit Medical Research Services and Development Ltd.. Invention is credited to Ami BEN-YA'ACOV, Smadar COHEN, Yaron ILAN, Eyal SHTEYER.
Application Number | 20180099006 15/837004 |
Document ID | / |
Family ID | 51020000 |
Filed Date | 2018-04-12 |
United States Patent
Application |
20180099006 |
Kind Code |
A1 |
COHEN; Smadar ; et
al. |
April 12, 2018 |
USE OF ALGINATE COMPOSITIONS IN PREVENTING OR REDUCING LIVER DAMAGE
CAUSED BY A HEPATOTOXIC AGENT
Abstract
Disclosed herein are compositions, methods and uses utilizing
alginate compositions, for treating, preventing and/or reducing
liver damage induced by a hepatotoxic agent, and for treating a
medical condition treatable by a hepatotoxic agent, in which an
alginate composition is administered prior to, concomitant with, or
shortly after exposure to a hepatotoxic agent. Also disclosed are
pharmaceutical compositions comprising a hepatotoxic agent and an
alginate composition and uses thereof for treating medical
conditions treatable by the hepatotoxic agent.
Inventors: |
COHEN; Smadar; (Beer-Sheva,
IL) ; ILAN; Yaron; (Kfar-Tavor, IL) ; SHTEYER;
Eyal; (Mevaseret Zion, IL) ; BEN-YA'ACOV; Ami;
(Jerusalem, IL) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Hadasit Medical Research Services and Development Ltd.
B.G. Negev Technologies & Applications Ltd., at Ben-Gurion
University |
Jerusalem
Beer-Sheva |
|
IL
IL |
|
|
Assignee: |
Hadasit Medical Research Services
and Development Ltd.
Jerusalem
IL
B.G. Negev Technologies & Applications Ltd., at Ben-Gurion
University
Beer-Sheva
IL
|
Family ID: |
51020000 |
Appl. No.: |
15/837004 |
Filed: |
December 11, 2017 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
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14758603 |
Jun 30, 2015 |
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PCT/IL2013/051088 |
Dec 30, 2013 |
|
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15837004 |
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61747325 |
Dec 30, 2012 |
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61747328 |
Dec 30, 2012 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A61K 9/0053 20130101;
A61K 31/167 20130101; A61P 1/16 20180101; A61P 29/00 20180101; A61K
45/06 20130101; A61K 31/734 20130101; A61K 31/734 20130101; A61K
2300/00 20130101 |
International
Class: |
A61K 31/734 20060101
A61K031/734; A61K 45/06 20060101 A61K045/06; A61K 9/00 20060101
A61K009/00; A61K 31/167 20060101 A61K031/167 |
Claims
1. A pharmaceutical composition comprising a therapeutically
effective amount of a hepatotoxic agent and a therapeutically
effective amount of an alginate composition, said alginate
composition comprising a pharmaceutically acceptable carrier,
wherein said alginate composition comprises alginate characterized
by a molecular weight in a range of from 10 to 75 kDa.
2. The composition of claim 1, being a unit dosage form.
3. The composition of claim 1, wherein said carrier is an aqueous
carrier.
4. The composition of claim 1, wherein said alginate composition
comprises alginate at a concentration in a range of from 0.4% to
10% (w/v).
5. The composition of claim 1, wherein said alginate composition
comprises alginate in a form of a sodium salt.
6. The composition of claim 1, being formulated for systemic
administration.
7. The composition of claim 1, being formulated for oral
administration and/or intraperitoneal administration.
8. The composition of claim 1, wherein said hepatotoxic agent is
selected from the group consisting of ethanol, paracetamol,
acarbose, amiodarone, bosentan, bromfenac, dantrolene, diclofenac,
dihydralazine, disulfiram, felbamate, fluoxetine, halothane,
isoniazid, kava, ketoconazole, labetalol, leflunomide,
methotrexate, methyldopa, nefazodone, nicotinic acid, paroxetine,
pemoline, propylthiouracil, pyrazinamide, rifampin, ritonavir,
sertraline, statins, tacrine, tetracycline antibiotics, tolcapone,
troglitazone, trovafloxacin, valproic acid, ximelagatran,
zafirlukast, zileuton, anabolic steroids, azathioprine,
azithromycin, captopril, cimetidine, ciprofloxacin, clopidogrel,
dicloxacillin, erythromycin, estrogens, flucloxacillin, naproxen,
phenobarbital, phenothiazine antipsychotics, phenytoin, sulindac,
terbinafine, tricyclic antidepressants, amoxicillin-clavulanic
acid, carbamazepine, cyclosporine, enalapril, flutamide,
methimazole, nitrofurantoin, sulfonamides, trazodone, trimethoprim,
verapamil, allopurinol, aspirin, betahistine, busulfan,
cephalosporins, chlorpheniramine, clarithromycin, codeine,
corticosteroids, cyclophosphamide, cytarabine, danazol,
dihydrocodeine, fluconazole, hydralazine, indinavir, ma-huang,
mebeverine, metoclopramide, oxycodone, penicillamine,
phenylbutazone, procainamide, quinidine, retinol, reverse
transcriptase inhibitors, sulpiride, tamoxifen and
telithromycin.
9. A method of reducing a liver damage caused by paracetamol, the
method comprising administering to a subject exposed to paracetamol
a therapeutically effective amount of an alginate composition, said
administering being effected prior to, concomitant with, or shortly
after exposure to paracetamol, thereby reducing liver damage,
wherein administration of said alginate composition is effected by
oral administration.
10. The method of claim 9, wherein said administering is effected
up to 50 minutes after said exposure to paracetamol.
11. The method of claim 9, wherein said administering is effected
during a time period ranging from 100 minutes prior to said
exposure to paracetamol to 50 minutes subsequent to said exposure
to paracetamol.
12. The method of claim 9, wherein said alginate composition
comprises an aqueous carrier.
13. The method of claim 12, wherein said alginate composition is
characterized by a solution viscosity in a range of from 3 to 50
mPa*seconds, at a shear rate of 1 second.sup.-1 and at a
concentration of 2% (w/v) alginate in said aqueous carrier.
14. The method of claim 9, wherein said alginate composition
comprises alginate is in a form of a sodium salt.
15. The method of claim 9, wherein said alginate composition and
said paracetamol are co-formulated within the same composition.
16. A method of treating a medical condition treatable by
paracetamol in a subject in need thereof, the method comprising
co-administering to the subject a therapeutically effective amount
of paracetamol and a therapeutically effective amount of an
alginate composition, said co-administering being effected such
that said alginate composition is administered to the subject
during a time period ranging from 100 minutes prior to
administration of paracetamol to 50 minutes subsequent to
administration of paracetamol, thereby treating the medical
condition.
17. The method of claim 16, wherein said medical condition is
selected from the group consisting of fever and pain.
18. The method of claim 16, wherein said alginate composition
comprises alginate characterized by a molecular weight in a range
of from 10 to 75 kDa.
19. The method of claim 16, wherein said alginate composition
comprises alginate is in a form of a sodium salt.
20. The method of claim 16, wherein said alginate composition and
said paracetamol are co-formulated within the same composition.
Description
RELATED APPLICATIONS
[0001] This application is a continuation of U.S. patent
application Ser. No. 14/758,603 filed on Jun. 30, 2015, which is a
National Phase of PCT Patent Application No. PCT/IL2013/051088
having International Filing Date of Dec. 30, 2013, which claims the
benefit of priority under 35 USC .sctn. 119(e) of U.S. Provisional
Patent Application Nos. 61/747,325 filed on Dec. 30, 2012 and
61/747,328 filed on Dec. 30, 2012. The contents of the above
applications are all incorporated by reference as if fully set
forth herein in their entirety.
FIELD AND BACKGROUND OF THE INVENTION
[0002] The present invention, in some embodiments thereof, relates
to a therapy and, more particularly, but not exclusively, to novel
therapeutic methodologies for reducing or preventing liver damage
by utilizing alginate-based compositions.
[0003] Alginate is an anionic polysaccharide derived from brown
algae. Alginate is a linear block co-polymer of (1-4)-linked
.beta.-D-mannuronic acid (M) and .alpha.-L-guluronic acid (G). The
monomers can appear in homopolymeric blocks of consecutive
G-residues (G-blocks), consecutive M-residues (M-blocks), or
alternating M- and G-residues (MG-blocks).
[0004] Sodium alginate is soluble in water, and, in the presence of
divalent cations, such as calcium ions, alginate forms a hydrogel.
In nature, alginate exists in both the soluble form and as a
hydrogel. The hydrogel protects brown algae from stress caused by
the hydrostatic pressure of water and by waves.
[0005] The ratio of mannuronic acid to guluronic acid (M/G) in
alginate differs according to the type of algae, and according to
environmental conditions. The G residues in alginate have a
particularly high affinity to calcium ions. Consequently, the
amount of G and length of G-sequences influences the extent of
alginate crosslinking and the mechanical properties of the formed
hydrogel.
[0006] In addition to diversity in M/G ratio, alginate may vary in
molecular weight. In nature, alginate usually has a molecular
weight in a range of 100-200 kDa. Using different treatment
protocols, such as heat and .gamma.-radiation, the molecular weight
of alginate can be reduced. Alginates with molecular weights of
approximately 50 kDa are commercially available.
[0007] International Patent Application PCT/IL97/00191 (published
as WO 97/44070) describes implantable polysaccharide (e.g.
alginate) sponges for use as a matrix, substrate or scaffold for
the cultivation of mammalian cells in vitro prior to their
implantation to replace damaged or removed tissue.
[0008] International Patent Application PCT/IL2004/000371
(published as WO 2004/098669) describes injectable cross-linked
alginate, which forms a hydrogel in vivo, for use in the repair of
cardiac tissue damage and ablation of cardiac arrhythmias, when
locally applied onto the cardiac tissue.
[0009] International Patent Application PCT/IL2008/001552
(published as WO 2009/069131) describes treatment of hepatic
disorders via administration of cross-linked or non-cross-linked
alginate biomaterial. Local administration to the liver of alginate
in solid, hydrogel or liquid form is described, as well as systemic
administration by injection of alginate in liquid form.
[0010] Landa, N. et al. [Circulation 117:1388-1396 (2008)]
describes calcium-cross-linked alginate in an injectable
low-viscosity solution, which can undergo phase transition into a
hydrogel after injection. Injection of the alginate solution into a
cardiac infarct was reported to prevent adverse cardiac remodeling
and dysfunction.
[0011] Orally administered calcium alginate (403 kDa) has been
reported to reduce liver damage caused by ingestion of CCl.sub.4
for one week in mice [Khotimchenko & Khotimchenko, Mar Drugs
2:108-122 (2004)].
[0012] Tsur-Gang et al. [Biomaterials 30:189-195 (2009)] describes
modification of alginate with the adhesion peptide RGD in order to
cause alginate to better interdigitate with the host. RGD-modified
alginate is also described in International Patent Application
PCT/IL2004/000371 (published as WO 2009/069131).
[0013] Liver disease represents a worldwide health problem in
humans, which can be managed pharmacologically in only a few cases.
Development of new drugs depends primarily on the availability of
suitable animal models. The pathophysiology of liver disease
includes complex phenomena such as interrelationships on humoral
basis, the highly sophisticated morphological organization of the
organ itself, and the integrity of metabolic and immunologic
pathways and their regulation in the individual cell types of the
liver.
[0014] The liver is responsible for the synthesis of serum
proteins; intermediary metabolism of amino acids, lipids, and
carbohydrates; and detoxification of foreign compounds. These
functions are usually seriously hampered in the various animal
models of liver diseases.
[0015] More than 900 drugs have been implicated in causing liver
injury [Friedman et al. (2003), Current Diagnosis & Treatment
in Gastroenterology, New York: Lang Medical Books/McGraw-Hill. pp.
664-679]. Hepatotoxicity is the most common reason for a drug to be
withdrawn from the market, and also accounts for a substantial
number of compound failures during drug development. Drug-induced
liver injury (DILI) is responsible for 5% of all hospital
admissions and 50% of all acute liver failures. Liver function
tests are routinely used to monitor subjects taking any of a
variety of drugs (e.g., methotrexate, carbamazepine).
[0016] Paracetamol intoxication, which may be intentional or
unintentional, is one of the major causes of death from drug
overdose and may lead to acute liver failure, sometimes
irreversibly. Paracetamol-induced liver toxicity is the most
prevalent cause of acute liver failure in the Western world.
Currently, an accepted treatment is N-acetylcysteine
administration, which has several drawbacks, mainly due to its
limited therapeutic window.
[0017] Knowledge of the mechanisms of paracetamol hepatotoxicity
derives to a large extent from studies performed in mice treated
with paracetamol. In mice, covalent binding of APAP metabolites to
liver proteins begins within 15 minutes of the overdose,
concurrently with the beginning of glutathione depletion, and peaks
within 1-2 hours. This is followed by other pathogenetic events
such as disturbance of intracellular calcium homeostasis, oxidative
and nitrosative stress, massive hepatic congestion, and activation
of the innate immunity including natural-killer and natural-killer
cells with T-cell receptors, macrophages and neutrophils. Oncotic
necrosis is the main mode of hepatocyte cell death.
[0018] Alcoholic liver disease is the major cause of liver disease
in Western countries. Chronic consumption of alcohol results in the
secretion of pro-inflammatory cytokines (TNF-.alpha., IL-6 and
IL-8), oxidative stress, lipid peroxidation, and acetaldehyde
toxicity. These factors cause inflammation, apoptosis and
eventually fibrosis of liver cells.
[0019] Additional art includes International Patent Application
Publication WO 95/19743; International Patent Application
Publication WO 98/12228; International Patent Application
Publication WO 2004/082594; German Patent Application Publication
DE 19723155 A1; Ichi et al. [J Nutr Sci Vitaminol 53:53-56 (2007)];
Seifert & Phillips [Biotechnol Prog 13:562-568 (1997)];
Balakrishnan & Jayakrishnan [Biomaterials 26:3941-3951 (2005)];
Maruyama et al. [J Surg Res 58:29-294 (1995)]; and Dvir-Ginzberg et
al. [Tissue Engineering 9:757-766 (2003)].
SUMMARY OF THE INVENTION
[0020] As exemplified herein, systemically administered alginate is
surprisingly effective at treating liver damage associated with
chemical hepatotoxicity, when the alginate is administered prior
to, concomitant with, or shortly after exposure to hepatotoxicity.
Following such administration, liver damage may be reduced,
nullified or even not induced, rendering administration of
hepatotoxic agents without an alginate generally inadvisable. The
high degree of biocompatibility of alginate renders alginate
unusually suitable for being administered prior to, shortly after,
or in combination with exposure to hepatotoxic agents, or for being
co-formulated with a hepatotoxic agent.
[0021] According to an aspect of some embodiments of the invention,
there is provided a method of reducing or preventing a liver damage
caused by a hepatotoxic agent, the method comprising administering
to a subject exposed to the hepatotoxic agent a therapeutically
effective amount of an alginate composition, the administering
being effected prior to, concomitant with, or shortly after
exposure to the hepatotoxic agent, thereby reducing or preventing
liver damage.
[0022] According to an aspect of some embodiments of the invention,
there is provided a method of treating a medical condition
treatable by a hepatotoxic agent in a subject in need thereof, the
method comprising co-administering to the subject a therapeutically
effective amount of the hepatotoxic agent and a therapeutically
effective amount of an alginate composition, the co-administering
being effected such that the alginate composition is administered
to the subject during a time period ranging from 100 minutes prior
to administration of the hepatotoxic agent to 50 minutes subsequent
to administration of the hepatotoxic agent, thereby treating the
medical condition.
[0023] According to an aspect of some embodiments of the invention,
there is provided a method of treating a medical condition selected
from the group consisting of the medical conditions listed in Table
1 (right column) herein in a subject in need thereof, the method
comprising co-administering to the subject a therapeutically
effective amount of a hepatotoxic agent selected from the group
consisting of the hepatotoxic drugs listed in Table 1 (left column)
herein, the aforementioned hepatotoxic drug being respective to the
aforementioned condition, and a therapeutically effective amount of
an alginate composition, the co-administering being effected such
that the alginate composition is administered to the subject during
a time period ranging from 100 minutes prior to administration of
the hepatotoxic agent to 50 minutes subsequent to administration of
the hepatotoxic agent, thereby treating the medical condition.
[0024] According to an aspect of some embodiments of the invention,
there is provided a method of treating a liver damage induced by a
hepatotoxic agent, the method comprising administering to a subject
in need thereof a therapeutically effective amount of an alginate
composition, the administering being effected prior to, concomitant
with, or up to 50 minutes after administration of the hepatotoxic
agent, thereby treating the liver injury.
[0025] According to an aspect of some embodiments of the invention,
there is provided a use of an alginate composition in the
manufacture of a medicament for reducing or preventing a liver
damage caused by a hepatotoxic agent, the medicament being for
administration prior to, concomitant with, or shortly after
exposure to the hepatotoxic agent.
[0026] According to an aspect of some embodiments of the invention,
there is provided a use of an alginate composition in the
manufacture of a medicament for treating a medical condition
treatable by a hepatotoxic agent, the treating comprising
co-administering a therapeutically effective amount of the
hepatotoxic agent and a therapeutically effective amount of the
alginate composition, the co-administering being effected such that
the alginate composition is administered to a subject during a time
period ranging from 100 minutes prior to administration of the
hepatotoxic agent to 50 minutes subsequent to administration of the
hepatotoxic agent.
[0027] According to an aspect of some embodiments of the invention,
there is provided a use of an alginate composition in the
manufacture of a medicament for treating a medical condition
selected from the group consisting of the medical conditions listed
in Table 1 (right column) herein, the treating comprising
co-administering a therapeutically effective amount of a
hepatotoxic agent selected from the group consisting of the
hepatotoxic drugs listed in Table 1 (left column) herein, the
aforementioned hepatotoxic drug being respective to the
aforementioned condition, and a therapeutically effective amount of
the alginate composition, the co-administering being effected such
that the alginate composition is administered to a subject during a
time period ranging from 100 minutes prior to administration of the
hepatotoxic agent to 50 minutes subsequent to administration of the
hepatotoxic agent.
[0028] According to an aspect of some embodiments of the invention,
there is provided a use of an alginate composition in the
manufacture of a medicament for treating a liver damage induced by
a hepatotoxic agent, the treating being effected prior to,
concomitant with, or up to 50 minutes after administration of the
hepatotoxic agent.
[0029] According to an aspect of some embodiments of the invention,
there is provided an alginate composition, for use in reducing or
preventing a liver damage caused by a hepatotoxic agent, the
alginate composition being for administration prior to, concomitant
with, shortly after exposure to the hepatotoxic agent.
[0030] According to an aspect of some embodiments of the invention,
there is provided an alginate composition, for use in treating a
medical condition treatable by a hepatotoxic agent, the treating
comprising co-administering a therapeutically effective amount of
the hepatotoxic agent and a therapeutically effective amount of the
alginate composition, the co-administering being effected such that
the alginate composition is administered to a subject during a time
period ranging from 100 minutes prior to administration of the
hepatotoxic agent to 50 minutes subsequent to administration of the
hepatotoxic agent.
[0031] According to an aspect of some embodiments of the invention,
there is provided an alginate composition, for use in treating a
medical condition selected from the group consisting of the medical
conditions listed in Table 1 (right column) herein, the treating
comprising co-administering a therapeutically effective amount of a
hepatotoxic agent selected from the group consisting of the
hepatotoxic drugs listed in Table 1 (left column), the
aforementioned hepatotoxic drug being respective to the
aforementioned condition, and a therapeutically effective amount of
the alginate composition, the co-administering being effected such
that the alginate composition is administered to a subject during a
time period ranging from 100 minutes prior to administration of the
hepatotoxic agent to 50 minutes subsequent to administration of the
hepatotoxic agent.
[0032] According to an aspect of some embodiments of the invention,
there is provided an alginate composition, for use in treating a
liver damage induced by a hepatotoxic agent, the treating being
effected prior to, concomitant with, or up to 50 minutes after
administration of the hepatotoxic agent.
[0033] According to an aspect of some embodiments of the invention,
there is provided a pharmaceutical composition comprising a
therapeutically effective amount of a hepatotoxic agent and a
therapeutically effective amount of an alginate composition, the
alginate composition comprising a pharmaceutically acceptable
carrier.
[0034] According to some embodiments of the invention,
administration is effected up to 50 minutes after exposure to the
hepatotoxic agent.
[0035] According to some embodiments of the invention,
administration is effected during a time period ranging from 100
minutes prior to exposure to the hepatotoxic agent to 50 minutes
subsequent to exposure to the hepatotoxic agent.
[0036] According to some embodiments of the invention, the treating
is effected prior to, concomitant with, or up to 50 minutes after
administration of the hepatotoxic agent.
[0037] According to some embodiments of the invention, the treating
is effected during a time period ranging from 100 minutes prior to
administration of the hepatotoxic agent to 50 minutes subsequent to
administration of the hepatotoxic agent.
[0038] According to some embodiments of the invention, the alginate
composition comprises a pharmaceutically acceptable carrier.
[0039] According to some embodiments of the invention, the carrier
is an aqueous carrier.
[0040] According to some embodiments of the invention, the alginate
composition is characterized by a solution viscosity in a range of
from 3 to 50 mPa*seconds, at a shear rate of 1 second.sup.-1 and at
a concentration of 2% (w/v) alginate in the aqueous carrier.
[0041] According to some embodiments of the invention, the solution
viscosity is in a range of from 3 to 20 mPa*seconds.
[0042] According to some embodiments of the invention, the alginate
composition comprises alginate at a concentration in a range of
from 0.4% to 10% (w/v).
[0043] According to some embodiments of the invention, the alginate
composition comprises alginate characterized by a molecular weight
in a range of from 10 to 75 kDa.
[0044] According to some embodiments of the invention, the
molecular weight is in a range of from 30 to 50 kDa.
[0045] According to some embodiments of the invention, the alginate
composition comprises alginate in a form of a sodium salt.
[0046] According to some embodiments of the invention,
administration of the alginate composition is effected by systemic
administration.
[0047] According to some embodiments of the invention,
administration of the alginate composition is effected by oral
administration.
[0048] According to some embodiments of the invention,
administration of the alginate composition is effected by
intraperitoneal administration.
[0049] According to some embodiments of the invention,
administration of the alginate composition comprises
co-administration of the alginate composition and the hepatotoxic
agent.
[0050] According to some embodiments of the invention, the alginate
composition and the hepatotoxic agent are co-formulated within the
same composition.
[0051] According to some embodiments of the invention, the
hepatotoxic agent is selected from the group consisting of ethanol,
paracetamol, acarbose, amiodarone, bosentan, bromfenac, dantrolene,
diclofenac, dihydralazine, disulfiram, felbamate, fluoxetine,
halothane, isoniazid, kava, ketoconazole, labetalol, leflunomide,
methotrexate, methyldopa, nefazodone, nicotinic acid, paroxetine,
pemoline, propylthiouracil, pyrazinamide, rifampin, ritonavir,
sertraline, statins, tacrine, tetracycline antibiotics, tolcapone,
troglitazone, trovafloxacin, valproic acid, ximelagatran,
zafirlukast, zileuton, anabolic steroids, azathioprine,
azithromycin, captopril, cimetidine, ciprofloxacin, clopidogrel,
dicloxacillin, erythromycin, estrogens, flucloxacillin, naproxen,
phenobarbital, phenothiazine antipsychotics, phenytoin, sulindac,
terbinafine, tricyclic antidepressants, amoxicillin-clavulanic
acid, carbamazepine, cyclosporine, enalapril, flutamide,
methimazole, nitrofurantoin, sulfonamides, trazodone, trimethoprim,
verapamil, allopurinol, aspirin, betahistine, busulfan,
cephalosporins, chlorpheniramine, clarithromycin, codeine,
corticosteroids, cyclophosphamide, cytarabine, danazol,
dihydrocodeine, fluconazole, hydralazine, indinavir, ma-huang,
mebeverine, metoclopramide, oxycodone, penicillamine,
phenylbutazone, procainamide, quinidine, retinol, reverse
transcriptase inhibitors, sulpiride, tamoxifen and
telithromycin.
[0052] According to some embodiments of the invention, the
hepatotoxic agent is paracetamol.
[0053] According to some embodiments of the invention, the
hepatotoxic agent is ethanol.
[0054] According to some embodiments of the invention, the
pharmaceutical composition is for treating a medical condition
treatable by the hepatotoxic agent.
[0055] According to some embodiments of the invention, the alginate
composition in the pharmaceutical composition is for reducing or
preventing a liver damage caused by the hepatotoxic agent.
[0056] According to some embodiments of the invention, the
composition is a unit dosage form.
[0057] According to some embodiments of the invention, the
composition is formulated for systemic administration.
[0058] According to some embodiments of the invention, the
composition is formulated for oral administration.
[0059] According to some embodiments of the invention, the
composition is formulated for intraperitoneal administration.
[0060] Unless otherwise defined, all technical and/or scientific
terms used herein have the same meaning as commonly understood by
one of ordinary skill in the art to which the invention pertains.
Although methods and materials similar or equivalent to those
described herein can be used in the practice or testing of
embodiments of the invention, exemplary methods and/or materials
are described below. In case of conflict, the patent specification,
including definitions, will control. In addition, the materials,
methods, and examples are illustrative only and are not intended to
be necessarily limiting.
BRIEF DESCRIPTION OF THE SEVERAL VIEWS OF THE DRAWINGS
[0061] The patent or application file contains at least one drawing
executed in color. Copies of this patent or patent application
publication with color drawing(s) will be provided by the Office
upon request and payment of the necessary fee.
[0062] Some embodiments of the invention are herein described, by
way of example only, with reference to the accompanying drawings.
With specific reference now to the drawings in detail, it is
stressed that the particulars shown are by way of example and for
purposes of illustrative discussion of embodiments of the
invention. In this regard, the description taken with the drawings
makes apparent to those skilled in the art how embodiments of the
invention may be practiced.
[0063] In the drawings:
[0064] FIG. 1 is a bar graph showing serum ALT activity in mice 24
hours after administration of 4 mg paracetamol (APAP), with or
without per os (PO) administration of 200 .mu.l of a 2% solution of
VLVG alginate, 30 minutes prior to administration of paracetamol
(p<0.0005 between the two groups);
[0065] FIGS. 2A and 2B present images (FIG. 2A; magnified .times.40
and .times.100) showing staining with IgG of necrotic cells in
liver tissue from mice after administration of 4 mg paracetamol
(APAP), with or without per os (PO) administration of a 2% solution
of VLVG alginate, 30 minutes prior to administration of
paracetamol, as well as a bar graph (FIG. 2B) showing the amount of
necrosis (scale bars are 500 .mu.m for .times.40 magnification and
200 .mu.m for .times.100 magnification, *p<0.007);
[0066] FIG. 3 is a bar graph showing serum ALT activity in mice 24
hours after administration of 4 mg paracetamol (APAP), with or
without per os (PO) administration of a 50, 100 or 200 .mu.l of a
2% solution of VLVG alginate, 30 minutes prior to administration of
paracetamol (*p<0.02, **p<0.000001 relative to APAP
alone);
[0067] FIG. 4 presents images (magnified .times.100) showing
staining of nitrotyrosine in liver tissue from mice after
administration of 4 mg paracetamol (APAP), with or without per os
(PO) administration of 50 .mu.l or 200 .mu.l of a 2% solution of
VLVG alginate, 30 minutes prior to administration of
paracetamol;
[0068] FIG. 5 presents images (magnified .times.100) showing
staining of Ki-67 in liver tissue from mice after administration of
4 mg paracetamol (APAP), with or without per os (PO) administration
of 50 .mu.l or 200 .mu.l of a 2% solution of VLVG alginate, 30
minutes prior to administration of paracetamol;
[0069] FIG. 6 is a bar graph showing serum paracetamol (APAP)
levels in mice 1, 4 and 24 hours after administration of 4 mg
paracetamol, with per os (PO) administration of 100 .mu.l or 200
.mu.l of a 2% solution of VLVG alginate, 30 minutes prior to
administration of paracetamol;
[0070] FIG. 7 is a bar graph showing serum ALT activity in mice 24
hours after administration of 4 mg paracetamol (APAP), with or
without per os (PO) administration of 2% solutions of VLVG
alginate, LVG54 alginate or LVG150 alginate, or i.p. administration
of a 2% solution of VLVG alginate, 30 minutes prior to
administration of paracetamol (*p<0.05, **p<0.01,
***p<0.0002 relative to APAP alone; & p<0.04 relative to
2% VLVG i.p. treatment);
[0071] FIG. 8 is a bar graph showing serum ALT activity in mice 24
hours after administration of 8 mg paracetamol (APAP), with or
without per os (PO) administration of a 2% solution of VLVG
alginate, 30 minutes prior to administration of paracetamol
(*p=0.005 between the two groups);
[0072] FIG. 9 is a bar graph showing serum ALT activity in mice 24
hours after administration of vehicle or 8 mg paracetamol (APAP),
with or without per os (PO) administration of a 2% solution of VLVG
alginate, 30 minutes prior to administration of paracetamol or
vehicle (*p>0.004 relative to APAP alone);
[0073] FIG. 10 is a bar graph showing serum ALT activity in mice 24
hours after administration of 4 mg paracetamol (APAP), with or
without per os (PO) administration of a 2% solution of VLVG
alginate, 30 or 60 minutes after administration of paracetamol
(p<0.002 for difference between 30 and 60 minutes);
[0074] FIG. 11 is a bar graph showing serum ALT activity in mice 24
hours after administration of 4 mg paracetamol (APAP), with or
without per os (PO) administration of a 2% solution of VLVG
alginate, 30 minutes after administration of paracetamol or
concurrently (mix) with paracetamol (*p<0.01, **p<0.002
relative to APAP without VLVG);
[0075] FIG. 12 presents images of immunohistochemical staining of
biotin-labeled VLVG alginate in liver parenchyma (at magnifications
of .times.40 and .times.100, scale bar represents 500 .mu.m for
magnification of .times.40 and 200 .mu.m for magnification of
.times.100); and
[0076] FIGS. 13A and 13B present images showing presence of stained
biotin in liver and pancreas (FIG. 13B) and absence of stained
biotin in colon and spleen (FIG. 13A), following administration of
a 2% solution of biotin-labeled VLVG alginate (no staining is
visible after administration of saline as control, scale bars
represent 200 .mu.m).
DESCRIPTION OF SPECIFIC EMBODIMENTS OF THE INVENTION
[0077] The present invention, in some embodiments thereof, relates
to therapy and, more particularly, but not exclusively, to novel
therapeutic methodologies for reducing or preventing liver damage
caused by hepatotoxic agents by utilizing alginate-based
compositions.
[0078] Before explaining at least one embodiment of the invention
in detail, it is to be understood that the invention is not
necessarily limited in its application to the details set forth in
the following description or exemplified by the Examples. The
invention is capable of other embodiments or of being practiced or
carried out in various ways.
[0079] The present inventors have investigated the effects of
alginate on liver damage. Through serendipity, the inventors
uncovered that systemically administered alginate is particularly
effective at treating liver damage associated with chemical
hepatotoxicity, when the alginate is administered prior to,
concomitant with, or shortly after (e.g., less than 60 minutes
after) exposure to hepatotoxicity. The inventors have further
envisioned that although such a time frame is normally not
particularly useful for treating liver diseases or disorders, the
high degree of biocompatibility of alginate (which is widely used
as a food additive) and the abovementioned time frame during which
alginate is particularly effective makes alginate unusually
suitable for being administered prior to, shortly after, or in
combination with exposure to hepatotoxic agents. In such a usage,
liver damage may be reduced, nullified or even not induced,
rendering administration of hepatotoxic agents without an alginate
generally inadvisable.
[0080] Referring now to the drawings, FIGS. 1-5 and 7-9 show that
oral administration of a solution of alginate reduces liver damage
caused by paracetamol, when the alginate is administered prior to
the paracetamol. FIGS. 3-5 show that a dose of 100 or 200 .mu.l of
a 2% solution of alginate in mice results in almost complete
elimination of signs of hepatotoxicity caused by 4 mg paracetamol,
whereas 50 .mu.l is only partially effective. FIG. 6 shows that the
reduction in liver damage is not due to any alteration of
paracetamol absorption. FIG. 7 shows that very low viscosity
alginate is the most potent alginate (compared to alginate
characterized by higher viscosity), that oral administration and
intraperitoneal administration of alginate are both effective, and
that oral administration is particularly effective. FIGS. 8 and 9
show that the alginate protects against sub-lethal doses of
paracetamol. FIG. 10 shows that the alginate exhibits a superior
protection against liver damage when administered 30 minutes after
paracetamol, (compared to alginate administered 60 minutes after
paracetamol). FIG. 11 shows that the alginate protects against
liver damage when administered concomitantly with, or 30 minutes
after paracetamol. FIGS. 12-13B show that systemically administered
alginate localizes in the liver, further indicating that alginate
can act on the liver following systemic administration.
[0081] Hence, according to one aspect of embodiments of the
invention, there is provided a method of protecting a subject from
liver damage caused by a hepatotoxic agent, the method comprising
administering to a subject exposed to the hepatotoxic agent a
therapeutically effective amount of an alginate composition.
[0082] Herein, the phrase "protecting a subject from liver damage"
refers to treating damage that has already been caused by reducing
or eliminating the damage, and/or to preventing or reducing future
liver damage.
[0083] It is to be understood that the protection may be due to a
local effect (e.g., a direct effect of alginate in contact with
liver tissue) and/or due to a systemic effect, such as a systemic
anti-inflammatory effect (e.g., by reducing levels of
pro-inflammatory cytokines).
[0084] As used herein, the terms "treat", "treating" and
"treatment" encompass abrogating, substantially inhibiting, slowing
or reversing the progression of a condition, substantially
ameliorating clinical or aesthetical symptoms of a condition or
substantially preventing or reduce the appearance of clinical or
aesthetical symptoms of a condition.
[0085] Herein, the "subject" encompasses any subject who is exposed
or is about to be exposed to a hepatotoxic agent, as further
detailed hereinbelow. The subject may be already diagnosed as being
previously exposed to the hepatotoxic agent and/or diagnosed with
liver injury; subjects having a medical condition which is to be
treated with a hepatotoxic agent and/or has already begun treatment
with a hepatotoxic agent; and any subject at risk for liver injury
associated with a hepatotoxic agent (e.g., an alcoholic).
[0086] By "exposed" it is meant that the subject has consumed a
hepatotoxic agent (e.g., by administration of the hepatotoxic
agent, or by eating, drinking or inhaling the hepatotoxic agent).
If a hepatotoxic agent is administered, then "exposure" includes
any time between administration and the presence of the agent or of
metabolites thereof in the liver.
[0087] The subject may be any human or non-human animal. In some
embodiments, the subject is a mammal. In exemplary embodiments, the
subject is a human.
[0088] As used herein, the phrase "alginate composition"
encompasses an alginate per se (e.g., as a solid), or an alginate
within a carrier (e.g., a liquid carrier). The carrier may be a
pure substance (e.g., a solvent) or may comprise additional
ingredients (e.g., a solution consisting of a solvent and
solutes).
[0089] In exemplary embodiments, an alginate composition as
described herein contains an aqueous carrier, such that the
composition comprises an aqueous solution of the alginate. The
aqueous carrier may be, for example, water or an aqueous solution,
such as a sodium chloride solution.
[0090] In some embodiments, the carrier is a pharmaceutically
acceptable carrier.
[0091] As used herein, the phrase "therapeutically effective
amount" generally describes an amount of the compound being
administered which will relieve to some extent one or more of the
symptoms of the condition being treated. The relief may be of an
existing symptom and/or of a future symptom (e.g., a symptom of a
condition which is to be prevented or reduced).
[0092] In the context of protecting a subject from liver damage, a
therapeutically effective amount is sufficient to result in a
reduction in liver damage by at least 25%, and preferably at least
50%, as compared to liver damage in individuals who are in a
similar condition but are not administered alginate. Liver damage
may be quantified according to serum levels of any biomolecule
which is used in the art as a marker for liver damage (e.g., as
exemplified herein), where liver damage is represented by the
difference between measured serum levels of the biomolecule and the
normal range for serum levels of the biomolecule (as recognized in
the art). For example, when liver damage is indicated by serum
levels of alanine transaminase (ALT) above the upper limit of
normal (ULN), a reduction by 50% of the difference between serum
levels of ALT and the ULN (in comparison with individuals not
administered alginate) indicates a reduction of 50% in liver
damage. In addition, to ALT, aspartate transaminase (AST), albumin,
and alkaline phosphatase (ALP) are examples of markers used in the
art for determining liver damage.
[0093] In the context of using a hepatotoxic agent (e.g., a
hepatotoxic drug) to treat a medical condition treatable by the
hepatotoxic agent, as described herein, a therapeutically effective
amount is an amount which will relieve to some extent one or more
of the symptoms of the condition treatable by the hepatotoxic
agent, and which is an amount sufficient to cause liver damage in
at least some subjects.
[0094] According to some embodiments, administration of the
alginate composition is effected prior to, concomitant with, or
shortly after exposure to the hepatotoxic agent.
[0095] In any aspect of embodiments of the invention described
herein, a treatment regimen according to any of the embodiments
described herein (e.g., with respect to a time and/or route of
administration, and/or a condition being treated) may be used in
association with an alginate according to any one of the
embodiments described herein regarding an alginate, and/or with a
hepatotoxic agent according to any of the embodiments described
herein regarding a hepatotoxic agent, unless otherwise
indicated.
[0096] Herein, "shortly after exposure" means up to 24 hours after
exposure (i.e., not later than 24 hours after exposure). In some of
any one of the embodiments described herein, administration is
effected up to 12 hours after exposure. In some embodiments,
administration is effected up to 6 hours after exposure. In some
embodiments, administration is effected up to 4 hours after
exposure. In some embodiments, administration is effected up to 3
hours after exposure. In some embodiments, administration is
effected up to 2 hours after exposure.
[0097] In preferred embodiments from among any one of the
embodiments described herein, administration is effected less than
1 hour after exposure. In some embodiments, administration is
effected up to 50 minutes after exposure. In some embodiments,
administration is effected up to 40 minutes after exposure. In some
embodiments, administration is effected up to 30 minutes after
exposure. In some embodiments, administration is effected up to 20
minutes after exposure. In some embodiments, administration is
effected up to 10 minutes after exposure.
[0098] As used herein, the terms "concomitant" and "concomitantly"
refer to an event (e.g., administration of an alginate composition)
being performed as closely in time as is practically possible to
another event (e.g., exposure to a hepatotoxic agent). In some
embodiments, "concomitant" and "concomitantly" refer to events
separated by no more than 1 hour. In some embodiments,
"concomitant" and "concomitantly" refer to events separated by no
more than 45 minutes. In some embodiments, "concomitant" and
"concomitantly" refer to events separated by no more than 30
minutes. In some embodiments, "concomitant" and "concomitantly"
refer to events separated by no more than 20 minutes. In some
embodiments, "concomitant" and "concomitantly" refer to events
separated by no more than 10 minutes.
[0099] In some of any one of the embodiments described herein, the
alginate composition is administered no more than 24 hours prior to
exposure to the hepatotoxic agent. In some embodiments, the
alginate composition is administered no more than 12 hours prior to
exposure to the hepatotoxic agent. In some embodiments, the
alginate composition is administered no more than 6 hours prior to
exposure to the hepatotoxic agent. In some embodiments, the
alginate composition is administered no more than 3 hours prior to
exposure to the hepatotoxic agent.
[0100] Exposure to the hepatotoxic agent may be effected, for
example, by co-administration of the hepatotoxic agent (e.g., a
hepatotoxic drug) and the alginate composition, as described
herein.
[0101] In some of any one of the embodiments described herein,
co-administration is such that the alginate composition is
administered during a time period ranging from 100 minutes prior to
exposure to the hepatotoxic agent to 50 minutes subsequent to
exposure to the hepatotoxic agent. In some embodiments, the
alginate composition is administered no more than 75 minutes prior
to exposure to the hepatotoxic agent. In some embodiments, the
alginate composition is administered no more than 50 minutes prior
to exposure to the hepatotoxic agent. In some embodiments, the
alginate composition is administered no more than 40 minutes prior
to exposure to the hepatotoxic agent. In some embodiments, the
alginate composition is administered no more than 30 minutes prior
to exposure to the hepatotoxic agent. In some embodiments, the
alginate composition is administered no more than 20 minutes prior
to exposure to the hepatotoxic agent. In some embodiments, the
alginate composition is administered no more than 10 minutes prior
to exposure to the hepatotoxic agent.
[0102] In some of any one of the embodiments described herein, the
alginate composition is administered no more than 40 minutes
subsequent to exposure to the hepatotoxic agent. In some
embodiments, the alginate composition is administered no more than
30 minutes subsequent to exposure to the hepatotoxic agent. In some
embodiments, the alginate composition is administered no more than
20 minutes subsequent to exposure to the hepatotoxic agent. In some
embodiments, the alginate composition is administered no more than
10 minutes subsequent to exposure to the hepatotoxic agent.
[0103] In some of any one of the embodiments described herein, the
alginate composition is administered during a time period ranging
from 75 minutes prior to exposure to the hepatotoxic agent to 50
minutes subsequent to exposure to the hepatotoxic agent. In some
embodiments, the alginate composition is administered during a time
period ranging from 50 minutes prior to exposure to the hepatotoxic
agent to 40 minutes subsequent to exposure to the hepatotoxic
agent. In some embodiments, the alginate composition is
administered during a time period ranging from 40 minutes prior to
exposure to the hepatotoxic agent to 40 minutes subsequent to
exposure to the hepatotoxic agent. In some embodiments, the
alginate composition is administered during a time period ranging
from 30 minutes prior to exposure to the hepatotoxic agent to 30
minutes subsequent to exposure to the hepatotoxic agent. In some
embodiments, the alginate composition is administered during a time
period ranging from 20 minutes prior to exposure to the hepatotoxic
agent to 20 minutes subsequent to exposure to the hepatotoxic
agent. In some embodiments, the alginate composition is
administered during a time period ranging from 10 minutes prior to
exposure to the hepatotoxic agent to 10 minutes subsequent to
exposure to the hepatotoxic agent.
[0104] In some of any one of the embodiments described herein
wherein the alginate composition is administered subsequent to
exposure to the hepatotoxic agent, the alginate composition is
administered during a time period ranging from 10 to 50 minutes
subsequent to exposure to the hepatotoxic agent. In some
embodiments, the alginate composition is administered during a time
period ranging from 20 to 40 minutes subsequent to exposure to the
hepatotoxic agent. In exemplary embodiments, the alginate
composition is administered about 30 minutes subsequent to exposure
to the hepatotoxic agent. In some embodiments, the alginate
composition is administered during a time period ranging from 100
minutes prior to exposure to the hepatotoxic agent to 50 minutes
subsequent to exposure to the hepatotoxic agent. In some
embodiments, the alginate composition is administered during a time
period ranging from 100 minutes prior to exposure to the
hepatotoxic agent to 50 minutes subsequent to exposure to the
hepatotoxic agent.
[0105] Without being bound by any particular theory, it is believed
that the alginate composition is particularly effective when the
time during which alginate is present in the liver of the subject
overlaps considerably with the time during which the liver is
exposed to dangerous levels of the hepatotoxic agent, or a
hepatotoxic metabolite of the agent. For example, paracetamol
hepatotoxicity is mediated by accumulation of hepatotoxic
metabolites (e.g., N-acetyl-p-benzoquinone imine) in the liver
following glutathione depletion, and such accumulation begins
somewhat later (e.g., 15 minutes later) than the actual ingestion
of the paracetamol.
[0106] In some of any of the embodiments described herein,
protecting the subject from the liver damage comprises reducing or
preventing a liver damage caused by the hepatotoxic agent.
[0107] In some of any one of the embodiments described herein, the
method reduces by at least 25%, and preferably by at least 50%, the
amount liver damage caused by the hepatotoxic agent subsequently to
the time at which the alginate composition is administered. Liver
damage caused by the hepatotoxic agent subsequently to
administration of the alginate composition is determined according
to changes in levels of a marker (e.g., as described herein) for
liver damage after the time of alginate administration.
[0108] In some of any one of the embodiments described herein, the
method is for treating a medical condition treatable by the
hepatotoxic agent (e.g., a hepatotoxic drug) in a subject in need
thereof. In such embodiments, the method comprises co-administering
to the subject a therapeutically effective amount of the
hepatotoxic agent (so as to treat the medical condition) and the
alginate composition (so as to reduce or prevent liver damage by
the therapeutically effective amount of the hepatotoxic agent). The
therapeutically effective amount of the hepatotoxic agent is
sufficient to be capable of causing liver damage, as described
herein.
[0109] In some of any one of the embodiments described herein, the
method is for treating a liver damage induced by a hepatotoxic
agent. In some embodiments, the liver damage induced by a
hepatotoxic agent is a drug-induced liver injury (DILI), as this
term is defined in the art, the hepatotoxic agent being a drug.
[0110] In some of any one of the embodiments described herein, the
liver damage (e.g., DILI) is characterized by relatively
predictable reactions, for example, hepatotoxicity is dose-related,
has a high incidence, occurs with a short latency (within a few
days), results from direct toxicity of the hepatotoxic agent (e.g.,
a hepatotoxic drug) or its metabolite and/or is reproducible in
animal models. Paracetamol is an exemplary agent which induces DILI
characterized by a predictable reaction.
[0111] In some of any one of the embodiments described herein, the
liver damage (e.g., DILI) is characterized by idiosyncratic
reactions, for example, occurs with variable latency (at least 1
week), has a low incidence, hepatotoxicity may not be dose-related,
levels of ALT are more than 3 times the upper limit of normal (ULN)
and/or alkaline phosphatase (ALP) levels are more than twice the
ULN. Most hepatotoxic drugs are associated with DILI characterized
by idiosyncratic reactions.
[0112] In some of any one of the embodiments described herein, the
liver damage (e.g., DILI) is mediated by an immune reaction.
[0113] In some of any one of the embodiments described herein, the
liver damage (e.g., DILI) is not mediated by an immune
reaction.
[0114] The hepatotoxic agent described herein throughout may be any
therapeutically active agent known in the art of medicine which can
cause liver damage (examples of which are described in detail
below), as well as any other substance which is hepatotoxic when
consumed. Examples of such substances include, without limitation,
alcohol (e.g., ethanol) and beverages which contain alcohol (e.g.,
beer, wine, liquors), as well as chemicals found in industry and in
household products (e.g., methanol, carbon tetrachloride, vinyl
chloride and other volatiles, and arsenic).
[0115] Exposure to alcohol by self-administration (e.g., drinking
alcoholic beverages) is very common, and is a leading cause of
liver damage. In some embodiments, the alginate composition is
included in an alcoholic beverage (e.g., beer, wine, liquor) or in
any other alcoholic composition which may be drunk (e.g., rubbing
alcohol), so as to prevent or reduce liver damage caused by the
alcohol.
[0116] In some of any one of the embodiments described herein, the
hepatotoxic agent is a therapeutically active agent (e.g., a
conventional drug), namely, an agent administered in order to treat
a medical condition. Examples of hepatotoxic drugs and medical
conditions treatable by the drugs are presented in Table 1.
TABLE-US-00001 TABLE 1 Hepatotoxic drug Medical condition(s)
treatable by drug Paracetamol Fever; pain (acetaminophen, APAP)
Acarbose Diabetes Amiodarone Cardiac arrhythmia Bosentan
Hypertension Bromfenac Inflammation; pain Dantrolene Malignant
hyperthermia; neuroleptic malignant syndrome; muscle spasticity;
Ecstasy intoxication; serotonin syndrome; 2,4-dinitrophenol
poisoning Diclofenac Pain; inflammation; dysmenorrhea Dihydralazine
Hypertension Disulfiram Alcohol dependence; cocaine dependence;
scabies; protozoal infections Felbamate Epilepsy Fluoxetine
Depression; obsessive-compulsive disorder; eating disorders; panic
disorder; body dysmorphic disorder; premenstrual dysphoric
disorder; trichotillomania; cataplexy; alcohol dependence Halothane
Surgery Isoniazid Bacterial infections Kava Anxiety disorder
Ketoconazole Fungal infections; alopecia Labetalol Hypertension
Leflunomide Rheumatoid arthritis; psoriatic arthritis Methotrexate
Cancer; rheumatoid arthritis; psoriasis; psoriatic arthritis;
lupus; inflammatory bowel disease; pregnancy; ectopic pregnancy
Methyldopa Hypertension; pre-eclampsia Nefazodone Depression;
migraine Nicotinic acid Pellagra; atherosclerosis Paroxetine
Depression; obsessive-compulsive disorder; post-traumatic stress
disorder; panic disorder; anxiety disorder; premenstrual dysphoric
disorder; premature ejaculation Pemoline Narcolepsy;
attention-deficit hyperactivity disorder Propylthiouracil
Hyperthyroidism Pyrazinamide Bacterial infections Rifampin
Bacterial infections; cholestatic pruritis; vaccinia virus
infection Ritonavir Retroviral infections Sertraline Depression;
obsessive-compulsive disorder; post-traumatic stress disorder;
panic disorder; body dysmorphic disorder; anxiety disorder; eating
disorders; premenstrual dysphoric disorder; syncope Statins
Cardiovascular disease Tacrine Alzheimer's disease Tetracycline
Bacterial infections; malaria; antibiotics balantidiasis Tolcapone
Parkinson's disease Troglitazone Diabetes Trovafloxacin Bacterial
infections Valproic acid Epilepsy; bipolar disorder; depression;
migraine; schizophrenia; colorectal polyps; basal cell carcinoma;
acne; Alzheimer's disease Ximelagatran Deep venous thrombosis;
venous thromboembolism; atrial fibrillation Zafirlukast Asthma
Zileuton Asthma Anabolic Hypoplastic anemia; growth failure;
steroids cancer; AIDS; delayed puberty; bone loss; gender identity
disorder Azathioprine Graft-versus-host reaction; rheumatoid
arthritis; pemphigus; lupus; Behcet's disease; autoimmune
hepatitis; atopic dermatitis; myasthenia gravis; neuromyelitis
optica; restrictive lung disease; inflammatory bowel disease;
multiple sclerosis Azithromycin Bacterial infections;
toxoplasmosis; malaria Captopril Hypertension; congestive heart
failure; diabetic nephropathy Cimetidine Heartburn; peptic ulcer;
herpes zoster; calcific tendinitis; interstitial cystitis; cancer
Ciprofloxacin Bacterial infections Clopidogrel Atherosclerosis;
coronary artery disease; peripheral vascular disease;
cerebrovascular disease; coronary stent implantation Dicloxacillin
Bacterial infections Erythromycin Bacterial infections Estrogens
Pregnancy Flucloxacillin Bacterial infections Naproxen Fever; pain;
inflammation; dysmenorrhea Phenobarbital Epilepsy; benzodiazepine
dependence; Gilbert's syndrome; cyclic vomiting syndrome
Phenothiazine Schizophrenia; acute psychosis; bipolar
antipsychotics disorder; hallucination; delusion disorder;
congestive heart failure; porphyria; tetanus; amoebic
meningoencephalitis; insomnia; pruritus; migraine; opioid addiction
Phenytoin Epilepsy Sulindac Pain; inflammation; colorectal polyps;
preterm labor; Alzheimer's disease Terbinafine Fungal infections
Tricyclic Depression; anxiety disorder; obsessive- antidepressants
compulsive disorder; panic disorder; post-traumatic stress
disorder; body dysmorphic disorder; personality disorder;
attention-deficit hyperactivity disorder; eating disorders; bipolar
disorder; pain; neuralgia; fibromyalgia; migraine; smoking
addiction; Tourette syndrome; trichotillomania; irritable bowel
syndrome; interstitial cystitis; nocturnal enuresis; narcolepsy;
insomnia; pathological crying and/or laughing; chronic hiccups;
ciguatera poisoning; schizophrenia; biliary dyskinesia Amoxicillin-
Bacterial infections clavulanic acid Carbamazepine Epilepsy;
bipolar disorder; neuropathic pain; attention-deficit hyperactivity
disorder; schizophrenia; phantom limb syndrome; complex regional
pain syndrome; paroxysmal extreme pain disorder; neuromyotonia;
intermittent explosive disorder; personality disorder; myotonia
congenita; post-traumatic stress disorder Cyclosporine
Graft-versus-host reaction; psoriasis; atopic dermatitis; pyoderma
gangrenosum; autoimmune urticaria; rheumatoid arthritis; dry eye
Enalapril Hypertension; chronic heart failure Flutamide Prostate
cancer Methimazole Hyperthyroidism Nitrofurantoin Bacterial
infections Sulfonamides Bacterial infections; retroviral
infections; diabetes; heart failure; liver cirrhosis; hypertension;
glaucoma; epilepsy; altitude sickness; cystinuria; dural ectasia;
periodic paralysis; osteoarthritis; rheumatoid arthritis; pain;
colorectal polyps; burns; gout; hyperuricemia; cardiac arrhythmia;
inflammatory bowel disease; migraine Trazodone Depression; bipolar
disorder; anxiety disorder; insomnia; fibromyalgia; panic disorder;
diabetic neuropathy; eating disorders; obsessive-compulsive
disorder; alcohol dependence; schizophrenia; complex regional pain
syndrome Trimethoprim Bacterial infections Verapamil Hypertension;
angina pectoris; cardiac arrhythmia; cluster headache; migraine;
malaria Allopurinol Hyperuricemia; gout; tumor lysis syndrome;
ischemic reperfusion injury; uric acid nephrolithiasis; protozoal
infections; epilepsy; hypertension Aspirin Pain; migraine; fever;
cardiovascular disease; percutaneous coronary intervention; cancer;
rheumatic fever; Kawasaki disease Betahistine Meniere's disease;
balance disorder Busulfan Cancer Cephalosporins Bacterial
infections Chlorpheniramine Allergy Clarithromycin Bacterial
infections Codeine Pain; cough; diarrhea; irritable bowel syndrome
Corticosteroids Adrenal insufficiency; congenital adrenal
hyperplasia; pain; inflammation; arthritis; temporal arteritis;
dermatitis; allergy; asthma; hepatitis; lupus; inflammatory bowel
disease; sarcoidosis; Addison's disease; brain tumor
Cyclophosphamide Cancer; lupus; rheumatoid arthritis; Wegener's
granulomatosis; multiple sclerosis Cytarabine Cancer; herpesvirus
infection Danazol Endometriosis; menorrhagia; fibrocystic breast
disease; immune thrombocytopenic purpura; mastodynia; hereditary
angioedema Dihydrocodeine Pain; cough; dyspnea; irritable bowel
syndrome; opioid addiction Fluconazole Fungal infections
Hydralazine Hypertension Indinavir Retroviral infections Ma-huang
Excess weight Mebeverine Irritable bowel syndrome Metoclopramide
Nausea; vomiting; gastroparesis; migraine Oxycodone Pain; diarrhea;
irritable bowel syndrome Penicillamine Rheumatoid arthritis;
Wilson's disease; cystinuria; scleroderma; arsenic poisoning
Phenylbutazone Fever; pain Procainamide Cardiac arrhythmia
Quinidine Cardiac arrhythmia; malaria Retinol Acne; acute
promyelocytic leukemia; vitamin A deficiency Reverse Retroviral
infections transcriptase inhibitors Sulpiride Schizophrenia;
depression Tamoxifen Cancer; McCune-Albright syndrome; anovulatory
disorder; retroperitoneal fibrosis; gynecomastia; bipolar disorder;
Riedel's thyroiditis Telithromycin Bacterial infections
[0117] Thus, in some of any one of the embodiments described
herein, the method is for treating any of the medical conditions
listed in Table 1 (right column), and is effected by
co-administering to the subject a therapeutically effective amount
of a hepatotoxic agent that is respective to (e.g., suitable for
treating) the medical condition (as listed in Table 1 (left
column)) and a therapeutically effective amount of an alginate
composition.
[0118] Examples of hepatotoxic anabolic steroids include, without
limitation, 4-androstenedione, androstenone, boldenone,
fluoxymesterone, methandienone, methandrostenolone,
methyltestosterone, nandrolone decanoate, nortestosterone,
oxandrolone, oxymetholone, testosterone and trenbolone.
[0119] Examples of hepatotoxic cephalosporins include, without
limitation, cefacetrile, cefadroxil, cephalexin, cefaloglycin,
cefalonium, cefaloridine, cefalotin, cefapirin, cefatrizine,
cefazaflur, cefazedone, cefazolin, cefradine, cefroxadine,
ceftezole, cefaclor, cefonicid, cefprozil, cefuroxime, cefuzonam,
cefmetazole, cefotetan, cefoxitin, cefcapene, cefdaloxime,
cefdinir, cefditoren, cefetamet, cefixime, cefmenoxime, cefodizime,
cefotaxime, cefovecin, cefpimizole, cefpodoxime, cefteram,
ceftibuten, ceftiofur, ceftiolene, ceftizoxime, ceftriaxome,
cefoperazone, ceftazidime, cefclidine, cefepime, cefluprenam,
cefoselis, cefozopran, cefpirome, cefquinome, ceftobiprole and
ceftaroline.
[0120] Examples of hepatotoxic corticosteroids include, without
limitation, aclometasone dipropionate, amcinonide, betamethasone,
betamethasone dipropionate, betamethasone sodium phosphate,
betamethasone valerate, budesonide, clobetasone-17-butyrate,
clobetasol-17-propionate, cortisone acetate, desonide,
dexamethasone, dexamethasone sodium phosphate, fludrocortisone,
fluocinonide, fluocinonide acetonide, fluocortolone, fluocortolone
caproate, fluocortolone pivalate, fluprednidene acetate,
halcinonide, hydrocortisone, hydrocortisone-17-aceponate,
hydrocortisone acetate, hydrocortisone-17-buteprate,
hydrocortisone-17-butyrate, hydrocortisone-17-valerate,
methylprednisolone, mometasone, prednicarbate, prednisolone,
prednisone, tixocortol pivalate, triamcinolone acetonide, and
triamcinolone alcohol.
[0121] Examples of hepatotoxic estrogens include, without
limitation, oral contraceptive ingredients such as ethinyl
estradiol, estradiol valerate and mestranol. The estrogen may be
formulated alone or in a form of a combined oral contraceptive, as
is common in the art.
[0122] Examples of hepatotoxic phenothiazine antipsychotics
include, without limitation, promethazine, chlorpromazine,
promazine, triflupromazine, methotrimeprazine, mesoridazine,
thioridazine, fluphenazine, perphenazine, prochlorperazine and
trifluoperazine. In some embodiments, the phenothiazine
antipsychotic is chlorpromazine. Conditions treatable by
chlorpromazine include, without limitation, schizophrenia; acute
psychosis; bipolar disorder; porphyria; tetanus; amoebic
meningoencephalitis; insomnia; pruritus; migraine; and opioid
addiction.
[0123] Examples of hepatotoxic reverse transcriptase inhibitors
include, without limitation, nevirapine and nucleoside analog
reverse transcriptase inhibitors (NRTIs) such as abacavir,
apricitabine, didanosine, emtricitabine, entecavir, lamivudine,
nevirapine, stavudine, zalcitabine and zidovudine.
[0124] Examples of hepatotoxic statins include, without limitation,
atorvastatin, cerivastatin, fluvastatin, lovastatin, mevastatin,
pitavastatin, rosuvastatin and simvastatin.
[0125] Examples of hepatotoxic sulfonamides include, without
limitation, anti-bacterial drugs such as sulfamethoxazole,
sulfisomidine, sulfacetamide, sulfadoxine and dichlorphenamide;
anti-diabetic agents such as carbutamide, acetohexamide,
chlorpropamide, tolbutamide, tolazamide, glipizide, gliclazide,
glibenclamide, glibornuride, gliquidone, glisoxepide,
glyclopyramide and glimepiride; diuretics such as acetazolamide,
ethoxzolamide, sultiame and zonisamide; protease inhibitors such as
darunavir, amprenavir, fosamprenavir and tipranavir; mafenide;
celecoxib; probenecid; sotalol; sulfasalazine; and sumatriptan.
[0126] Examples of hepatotoxic tetracycline antibiotics include,
without limitation, tetracycline, chlortetracycline,
oxytetracycline, demeclocycline, doxycycline, lymecycline,
meclocycline, methacycline, minocycline, PTK 0796, rolitetracycline
and tigecycline.
[0127] Examples of hepatotoxic tricyclic antidepressants include,
without limitation, amitriptyline, amitriptylinoxide, butriptyline,
clomipramine, demexiptiline, desipramine, dibenzepin, dimetacrine,
dosulepin, imipramine, imipraminoxide, lofepramine, melitracen,
metapramine, nitroxezapine, nortriptyline, noxiptiline, pipofezine,
propizepine, protriptyline, quinupramine, amineptine, iprindole,
opipramol, tianeptine and trimipramine. In some embodiments, the
tricyclic antidepressant is amitriptyline. Conditions treatable by
amitriptyline include, without limitation, depression; anxiety
disorder; attention-deficit hyperactivity disorder; migraine;
eating disorders; bipolar disorder; neuralgia; insomnia; nocturnal
enuresis; pain; and biliary dyskinesia.
[0128] Eating disorders treatable by the hepatotoxic drugs
described herein (see, Table 1) include, without limitation,
bulimia nervosa, anorexia nervosa, night eating syndrome, obesity,
and binge eating disorder.
[0129] Examples of anxiety disorder treatable by hepatotoxic drugs
described herein (see, Table 1) include, without limitation,
generalized anxiety disorder and social anxiety disorder.
[0130] Borderline personality disorder is a non-limiting example of
a personality disorder treatable by hepatotoxic drugs described
herein.
[0131] In some of any one of the embodiments described herein, the
liver damage is characterized by hepatocellular injury and/or
hepatitis. Examples of hepatotoxic agents which can cause such
liver damage include, without limitation, ethanol, paracetamol,
acarbose, amiodarone, bosentan, bromfenac, dantrolene, diclofenac,
dihydralazine, disulfiram, felbamate, fluoxetine, halothane,
isoniazid, kava, ketoconazole, labetalol, leflunomide,
methotrexate, methyldopa, nefazodone, nicotinic acid, paroxetine,
pemoline, propylthiouracil, pyrazinamide, rifampin, ritonavir,
sertraline, statins, tacrine, tetracycline antibiotics, tolcapone,
troglitazone, trovafloxacin, valproic acid, ximelagatran,
zafirlukast, and zileuton. Paracetamol is an exemplary hepatotoxic
agent.
[0132] In some of any one of the embodiments described herein, the
liver damage is characterized by cholestasis. Examples of
hepatotoxic agents which can cause such liver damage include,
without limitation, anabolic steroids, azathioprine, azithromycin,
captopril, chlorpromazine, cimetidine, ciprofloxacin, clopidogrel,
dicloxacillin, erythromycin, estrogens, flucloxacillin, naproxen,
phenobarbital, phenothiazine antipsychotics, phenytoin, sulindac,
terbinafine, and tricyclic antidepressants.
[0133] In some of any one of the embodiments described herein, the
liver damage is characterized by a combination of hepatitis and
cholestasis. Examples of hepatotoxic agents which can cause such
liver damage include, without limitation, amitriptyline,
amoxicillin-clavulinic acid, carbamazepine, cyclosporine,
enalapril, flutamide, methimazole, nitrofurantoin, sulfonamides,
trazodone, trimethoprim, and verapamil.
[0134] In some of any one of the embodiments described herein, the
liver damage is characterized by fibrosis and/or cirrhosis.
Methotrexate is an exemplary hepatotoxic agent which can cause such
liver damage.
[0135] In some of any one of the embodiments described herein, the
liver damage is characterized by granulomas. Examples of
hepatotoxic agents which can cause such liver damage include,
without limitation, allopurinol, amoxicillin-clavulanic acid,
carbamazepine, hydralazine, methyldopa, penicillamine,
phenylbutazone, phenytoin, procainamide, quinidine and
sulfonamides.
[0136] In some of any one of the embodiments described herein, the
liver damage is characterized by microvesicular steatosis. Examples
of hepatotoxic agents which can cause such liver damage include,
without limitation, nucleoside analog reverse transcriptase
inhibitors (e.g., such as described herein) and valproate.
[0137] In some of any one of the embodiments described herein, the
liver damage is characterized by neoplasms. Examples of hepatotoxic
agents which can cause such liver damage include, without
limitation, anabolic steroids and estrogens.
[0138] In some of any one of the embodiments described herein, the
liver damage is characterized by non-alcoholic steatohepatitis.
Examples of hepatotoxic agents which can cause such liver damage
include, without limitation, amiodarone and tamoxifen.
[0139] In some of any one of the embodiments described herein, the
liver damage is characterized by vascular lesions. Examples of
hepatotoxic agents which can cause such liver damage include,
without limitation, anabolic steroids, estrogens, azathioprine,
retinol, methotrexate, busulfan and cyclophosphamide.
[0140] According to another aspect of embodiments of the invention,
there is provided a use of an alginate composition in the
manufacture of a medicament for protecting a subject from liver
damage caused by a hepatotoxic agent (e.g., an agent described
herein). Treatment utilizing the medicament is effected by
administering the medicament prior to, concomitant with, or shortly
after (e.g., up to 50 minutes after) exposure to the hepatotoxic
agent (e.g., in accordance with a method described herein).
[0141] In some of any one of the embodiments described herein, the
medicament is for reducing or preventing a liver damage caused by
the hepatotoxic agent, as described herein.
[0142] In some of any one of the embodiments described herein, the
medicament is for treating a medical condition treatable by a
hepatotoxic agent, as described herein. Such treating comprises
co-administering, as described herein, a therapeutically effective
amount of the hepatotoxic agent and a therapeutically effective
amount of alginate composition.
[0143] In some of any one of the embodiments described herein, the
medicament is for treating a medical condition such as the medical
conditions listed in Table 1 (right column). Such treating
comprises co-administering a therapeutically effective amount of a
hepatotoxic agent respective to (or suitable for treating) the
medical condition (e.g., a hepatotoxic drug as listed in Table 1
(left column)), as described herein, and a therapeutically
effective amount of an alginate composition.
[0144] In some of any one of the embodiments described herein, the
medicament is for treating a liver damage induced by a hepatotoxic
agent (e.g., a drug-induced liver injury), as described herein.
[0145] According to another aspect of embodiments of the invention,
there is provided an alginate composition for use in protecting a
subject from liver damage caused by a hepatotoxic agent (e.g., an
agent described herein). Treatment utilizing the alginate
composition effected by administering the alginate composition
prior to, concomitant with, or shortly after (e.g., up to 50
minutes after) exposure to the hepatotoxic agent (e.g., in
accordance with a method described herein).
[0146] In some of any one of the embodiments described herein, the
alginate composition is for reducing or preventing a liver damage
caused by the hepatotoxic agent, as described herein.
[0147] In some of any one of the embodiments described herein, the
alginate composition is for treating a medical condition treatable
by a hepatotoxic agent, as described herein. Such treating
comprises co-administering, as described herein, a therapeutically
effective amount of the hepatotoxic agent, as described herein and
a therapeutically effective amount of the alginate composition.
[0148] In some of any one of the embodiments described herein, the
alginate composition is for treating a medical condition such as
the medical conditions listed in Table 1 (right column). Such
treating comprises co-administering a therapeutically effective
amount of a hepatotoxic agent respective to (or suitable for
treating) the medical condition as listed in Table 1 (left column),
as described herein, and a therapeutically effective amount of the
alginate composition.
[0149] In some of any one of the embodiments described herein, the
alginate composition is for treating a liver damage induced by a
hepatotoxic agent (e.g., drug-induced liver injury), as described
herein.
[0150] The administration of the alginate composition (e.g., in a
form of a medicament described herein) may be during any time
period for administration described herein. For example, in some of
any one of the embodiments described herein, co-administration is
effected by administering the alginate composition/medicament
during a time period ranging from 100 minutes prior to exposure to
the hepatotoxic agent to 50 minutes subsequent to exposure to the
hepatotoxic agent (e.g., as described herein).
[0151] In some of any one of the embodiments described herein of
any of the aspects of embodiments of the invention described
herein, administration of the alginate composition comprises
co-administration of a therapeutically effective amount of the
alginate composition and a therapeutically effective amount of the
hepatotoxic agent.
[0152] The co-administration of a therapeutically effective amount
of a hepatotoxic agent with an alginate composition is superior to
current methodologies and regimens for administration of
therapeutically effective amounts of hepatotoxic agents, in view of
the hepatoprotection provided by the alginate. The
co-administration described herein may therefore be effected
without required monitoring of liver function, or with less
frequent monitoring, in a subject being co-administered the
hepatotoxic agent and alginate composition, in contrast to current
methodologies (e.g., in which a subject is administered the
hepatotoxic agent without an alginate composition). Hence, in some
embodiments, a treatment of a medical condition as described herein
is devoid of monitoring liver function or is effected while
monitoring liver function less frequent (compared to subjects
receiving the hepatotoxic drug without an alginate composition as
described herein). For example, for a subject receiving a
hepatotoxic drug for treating a chronic medical condition, liver
function is monitored once every 6 month, or even a year, of
treatment.
[0153] In some of any one of the embodiments described herein, the
alginate composition and a therapeutically effective amount of the
hepatotoxic agent are formulated separately (e.g., as two separate
pharmaceutical compositions). The alginate composition may
optionally be formulated as described herein (e.g., a medicament
described herein), and the hepatotoxic agent may optionally be
formulated in a standard form (e.g., a commercially available
composition). In some such embodiments, the compositions comprising
the alginate composition and the hepatotoxic agent are administered
concomitantly.
[0154] In some of any one of the embodiments described herein, the
alginate composition and hepatotoxic agent are administered via
different routes of administration. For example, in some
embodiments, the alginate composition is administered by oral
administration, and the hepatotoxic agent is administered by
intraperitoneal administration (e.g., wherein intraperitoneal
administration is a standard route of administration for the agent)
or by buccal administration, transdermal administration,
transmucosal administration, inhalation, rectal administration,
and/or intravenous injection (e.g., wherein such a route of
administration is a standard route of administration for the
agent). In some embodiments, the alginate composition is
administered by intraperitoneal administration, and the hepatotoxic
agent is administered by oral administration (e.g., wherein oral
administration is a standard route of administration for the
agent).
[0155] In some of any one of the embodiments described herein, the
alginate composition and hepatotoxic agent are co-formulated within
the same composition (i.e., as one composition). Pharmaceutical
compositions comprising both alginate composition and hepatotoxic
agent are described herein.
[0156] In some of any one of the embodiments described herein, the
alginate composition and hepatotoxic agent, either co-formulated or
formulated separately, are administered via the same route of
administration. For example, in some embodiments, the alginate
composition and hepatotoxic agent are administered by oral
administration (e.g., wherein oral administration is a standard
route of administration for the hepatotoxic agent), and in some
embodiments, the alginate composition and hepatotoxic agent are
administered by intraperitoneal administration (e.g., wherein
intraperitoneal administration is a standard route of
administration for the hepatotoxic agent).
[0157] In any aspect of embodiments of the invention described
herein, an alginate according to any one of the embodiments
described herein may be used in association with a hepatotoxic
agent according to any of the embodiments described herein
regarding a hepatotoxic agent, unless otherwise indicated.
[0158] In some of any one of the embodiments described herein, the
alginate comprises an alginate which may be in a form of alginic
acid (i.e., a protonated form) and/or in a form of an alginate salt
(i.e., a non-protonated form). In some embodiments, the alginate
includes both protonated and non-protonated carboxylic groups.
[0159] Examples of suitable alginate salts include, without
limitation, a sodium salt, a potassium salt, a calcium salt, and a
magnesium salt. In some embodiments, the alginate salt comprises at
least one univalent cation, such as sodium and/or potassium. In
exemplary embodiments, the alginate is in a form of a sodium
salt.
[0160] Herein, a "sodium salt" of alginate refers to alginate which
comprises at least 0.2 sodium ions per carboxylate group (of the
alginate) bound to the alginate (e.g., by ionic bonds). In some of
any one of the embodiments described herein, the alginate comprises
at least 0.3 sodium ions per carboxylate group. In some
embodiments, the alginate comprises at least 0.4 sodium ions per
carboxylate group. In some embodiments, the alginate comprises at
least 0.5 sodium ions per carboxylate group. In some embodiments,
the alginate comprises at least 0.6 sodium ions per carboxylate
group. In some embodiments, the alginate comprises at least 0.7
sodium ions per carboxylate group. In some embodiments, the
alginate comprises at least 0.8 sodium ions per carboxylate group.
In some embodiments, the alginate comprises at least 0.9 sodium
ions per carboxylate group. In some embodiments, the alginate
comprises at least 0.95 sodium ions per carboxylate group. In some
embodiments, the alginate comprises at least 0.99 sodium ions per
carboxylate group.
[0161] In some of any one of the embodiments described herein, a
sodium salt is prepared by mixing an alginate in a solution
containing sodium (e.g., a sodium chloride solution). In some
embodiments, mixing is performed by homogenization (e.g., as
exemplified herein). In exemplary embodiments, the sodium solution
contains about 0.15 M sodium ions.
[0162] In some of any one of the embodiments described herein, the
alginate comprises at least 0.2 univalent cations per carboxylate
group (of the alginate) bound to the alginate (e.g., by ionic
bonds). In some embodiments, the alginate comprises at least 0.3
univalent cations per carboxylate group. In some embodiments, the
alginate comprises at least 0.4 univalent cations per carboxylate
group. In some embodiments, the alginate comprises at least 0.5
univalent cations per carboxylate group. In some embodiments, the
alginate comprises at least 0.6 univalent cations per carboxylate
group. In some embodiments, the alginate comprises at least 0.7
univalent cations per carboxylate group. In some embodiments, the
alginate comprises at least 0.8 univalent cations per carboxylate
group. In some embodiments, the alginate comprises at least 0.9
univalent cations per carboxylate group. In some embodiments, the
alginate comprises at least 0.95 univalent cations per carboxylate
group. In some embodiments, the alginate comprises at least 0.99
univalent cations per carboxylate group.
[0163] In some of any one of the embodiments described herein, the
alginate composition comprises alginate that is characterized by a
molecular weight of up to 300 kDa. In some embodiments, the
alginate is characterized by a molecular weight in a range of from
3 to 300 kDa. In some embodiments, the alginate is characterized by
a molecular weight in a range of from 5 to 200 kDa. In some
embodiments, the alginate is characterized by a molecular weight in
a range of from 8 to 160 kDa. In some embodiments, the alginate is
characterized by a molecular weight in a range of from 10 to 75
kDa. In some embodiments, the alginate is characterized by a
molecular weight in a range of from 20 to 60 kDa. In some
embodiments, the alginate is characterized by a molecular weight in
a range of from 30 to 50 kDa.
[0164] In some of any one of the embodiments described herein, the
alginate composition consists of an alginate as described herein
and an aqueous carrier described herein, the alginate composition
being characterized by a solution viscosity in a range of from 3 to
50 mPa*seconds, at a shear rate of 1 second.sup.-1 and at a
concentration of 2% weight/volume (20 grams per liter) in the
aqueous carrier. In some embodiment, the solution viscosity is in a
range of from 3 to 20 mPa*seconds. In some embodiment, the solution
viscosity is in a range of from 5 to 20 mPa*seconds. In some
embodiment, the solution viscosity is in a range of from 10 to 20
mPa*seconds. In some embodiment, the solution viscosity is in a
range of from 5 to 50 mPa*seconds. In some embodiment, the solution
viscosity is in a range of from 10 to 50 mPa*seconds. In exemplary
embodiments, the solution viscosity is about 15.5 mPa*seconds.
[0165] It is to be understood that the phrase "at a concentration
of 2%" does not indicate that the concentration of alginate in the
alginate composition is necessarily 2%. Rather, the phrase means
that a solution viscosity is determined for a composition
consisting of the alginate (at a concentration of 2%) and the
aqueous carrier, that is the composition for which solution
viscosity is determined may differ from the alginate composition
described herein only in the relative amount of the aqueous
carrier.
[0166] Techniques for determining a molecular weight and/or a
solution viscosity of alginate are exemplified herein. Additional
techniques will be known by the skilled person.
[0167] The abovementioned ranges of alginate molecular weight and
solution viscosity correspond to a relatively small molecular size
of the alginate, including alginate known in the art as VLVG (very
low viscosity high G alginate). As shown in the Examples herein,
VLVG alginate exhibits more efficacy than do alginates
characterized by larger molecular size.
[0168] Without being bound by any particular theory, it is believed
that such relatively small molecular size facilitates diffusion of
the alginate and transport in vivo, and that this enhances the
therapeutic efficacy of the alginate composition. It is further
believed that such relatively small molecular sizes are so small as
to significantly alter the properties of alginate in the alginate
composition which are believed to be play a role in the therapeutic
efficacy of the alginate composition, for example, by inhibiting
the ability of alginate to form a hydrogel.
[0169] In some of any one of the embodiments described herein, the
alginate composition is a composition described in co-filed
International Patent Application having Attorney's Docket No.
58819, which claims priority from U.S. Provisional Patent
Application No. 61/747,328, the contents of which are incorporated
herein by reference in their entirety. Such a composition comprises
alginate, a source of sodium ions (e.g., sodium salt) and a carrier
(e.g., as described herein). As described therein, homogenization
of alginate with the sodium ions (e.g., in saline) alters the
physical properties of alginate composition considerably, in such a
manner as to significantly enhance dissolution and diffusion of the
alginate.
[0170] In some of any one of the embodiments described herein, the
alginate-containing composition (as described in the abovementioned
International Patent Application) is characterized by at least one
of the following properties: [0171] (i) A zeta potential weaker
than -25 mV (i.e., closer to 0 mV), at a concentration of 0.5%
(weight per volume) alginate in the abovementioned carrier; [0172]
(ii) A diffusion coefficient of at least 10.sup.-8
cm.sup.2/millisecond, at a concentration of 0.5% (weight per
volume) alginate in the abovementioned carrier; [0173] (iii) A
solution viscosity in a range of from 3 to 20 mPa*seconds, at a
shear rate of 1 second.sup.-1 and at a concentration of 2%
weight/volume (20 grams per liter) in the carrier; [0174] (iv) A
small angle X-ray scattering (SAXS) pattern characterized by an
absence of a peak in scattering intensity in the interval
0.012<q<0.7 .ANG..sup.-1; and [0175] (v) An absence of
structures observable by transmission electron microscopy (e.g.,
cryogenic transmission electron microscopy) which are more than 5
nm in width.
[0176] Techniques for determining a diffusion coefficient, zeta
potential, SAXS pattern, transmission electron microscopy image
and/or a solution viscosity of alginate are exemplified therein.
Additional techniques will be known by the skilled person. Values
of diffusion coefficients, zeta potentials and solution viscosities
refer to values at a temperature of 25.degree. C.
[0177] In order to determine a presence or absence of structures by
transmission electron microscopy, specimens are preferably
vitrified (e.g., by rapid plunging into liquid ethane pre-cooled
with liquid nitrogen) in a controlled-environment vitrification
system, and examined using low-dose imaging, as exemplified herein,
so as to prevent microstructural changes by ice crystallization
and/or radiation damage. In addition, microscopy is preferably
performed without heavy metal staining, so as to avoid structural
changes due to interactions between alginate and heavy metal.
[0178] In some of any one of the embodiments described herein, the
alginate in the composition is characterized by at least two of the
aforementioned properties. In some embodiments, the alginate in the
composition is characterized by at least 3 of the aforementioned
properties. In some embodiments, the alginate in the composition is
characterized by at least 4 of the aforementioned properties. In
some embodiments, the alginate in the composition is characterized
by all of the aforementioned properties.
[0179] In some of any one of the embodiments described herein, the
composition is characterized by a diffusion coefficient of at least
10.sup.-8 cm.sup.2/millisecond at a concentration of 0.5%, as
described herein. In some embodiments, the diffusion coefficient is
at least 2.times.10.sup.-8 cm.sup.2/millisecond. In some
embodiments, the diffusion coefficient is at least
3.times.10.sup.-8 cm.sup.2/millisecond. In some embodiments, the
diffusion coefficient is at least 4.times.10.sup.-8
cm.sup.2/millisecond. In some embodiments, the diffusion
coefficient is at least 5.times.10.sup.-8 cm.sup.2/millisecond. In
some embodiments, the diffusion coefficient is at least
6.times.10.sup.-8 cm.sup.2/millisecond. In some embodiments, the
diffusion coefficient is at least 7.times.10.sup.-8
cm.sup.2/millisecond. In some embodiments, the diffusion
coefficient is at least 8.times.10.sup.-8 cm.sup.2/millisecond. In
some embodiments, the diffusion coefficient is at least
9.times.10.sup.-8 cm.sup.2/millisecond. In some embodiments, the
diffusion coefficient is at least 10.sup.-7 cm.sup.2/millisecond.
In some embodiments, the diffusion coefficient is at least
2.times.10.sup.-7 cm.sup.2/millisecond. In some embodiments, the
diffusion coefficient is at least 3.times.10.sup.-7
cm.sup.2/millisecond.
[0180] It is to be understood that the phrase "at a concentration
of 0.5%", recited herein with respect to diffusion coefficients and
zeta potentials, does not indicate that the concentration of
alginate in the composition described herein is necessarily 0.5%.
Similarly, the phrase "at a concentration of 2%", recited herein
with respect to solution viscosities, does not indicate that the
concentration of alginate in the composition described herein is
necessarily 2%. Rather, the phrases mean that a diffusion
coefficient, zeta potential and solution viscosity are determined
for a composition consisting of the alginate (at a concentration of
2% or 0.5%) and the carrier (and a source of sodium ions), that is,
the composition for which diffusion coefficient, zeta potential or
solution viscosity is determined may differ from the composition of
embodiments of the invention in the amount of the carrier relative
to the alginate. Thus, for example, a diffusion coefficient and/or
zeta potential may be characterized by measuring a composition as
described herein after the composition has been diluted with
carrier (with a source of sodium ions) to result in an alginate
concentration of 0.5%, as exemplified herein.
[0181] Diffusion coefficients may be determined using dynamic light
scattering measurement, by recording the real time fluctuations in
the intensity of the scattered light. Equations for calculating a
diffusion coefficient based on measurement of the intensity time
correlation function as a function of the decay time, are described
in the Examples section herein.
[0182] In some of any one of the embodiments described herein, the
alginate composition is characterized by at least one of a zeta
potential described herein and a diffusion coefficient described
herein.
[0183] In some of any one of the embodiments described herein, the
alginate composition is characterized by a zeta potential described
herein.
[0184] In some of any one of the embodiments described herein, the
composition is characterized by both a diffusion coefficient as
described herein, and a zeta potential as described herein.
[0185] Herein and in the art, the phrase "zeta potential" refers to
electric potential difference between a dispersion medium (e.g., a
liquid medium of the composition described herein) and a stationary
layer of fluid attached to a dispersed particle (e.g., alginate).
As alginate is negatively charged (due to the presence of
carboxylate groups), the zeta potential of alginate in the carrier
is negative. Thus, a "weaker" zeta potential refers to a value
closer to 0 mV (i.e., less negative).
[0186] In some of any one of the embodiments described herein, the
zeta potential is weaker than -23 mV, under the abovementioned
measurement conditions. In some embodiments, the zeta potential is
weaker than -21 mV. In some embodiments, the zeta potential is
weaker than -20 mV. In some embodiments, the zeta potential is
weaker than -19 mV.
[0187] Without being bound by any particular theory, it is believed
that a weak zeta potential (e.g., weaker than -25 mV) indicates a
high degree of masking by sodium ions of the negative charge of
alginate carboxylate groups, which in turn weakens intermolecular
forces between alginate molecules.
[0188] In some of any one of the embodiments described herein, the
zeta potential is at least -10 mV (i.e., -10 mV or a more negative
value). In some embodiments, the zeta potential is at least -12.5
mV. In some embodiments, the zeta potential is at least -15 mV. In
some embodiments, the zeta potential is in a range of from about
-17.8 mV to about -21.2 mV. In some embodiments, the zeta potential
is about -17.8 mV.
[0189] Without being bound by any particular theory, it is believed
that a very weak zeta potential (e.g., weaker than -10 mV)
indicates a high degree of instability, as there is little
electrostatic forces to repel alginate molecules from one
another.
[0190] Zeta potentials may be determined by analyzing
experimentally measured electrophoretic mobility distributions
using a standard theoretical model. Devices for performing the
measurements and analysis include, for example, a Zeta Plus.TM.
zeta potential analyzer (Brookhaven Instruments Corp., NY).
[0191] In some of any one of the embodiments described herein, the
composition comprising alginate as described herein is
characterized by a solution viscosity in a range of from 5 to 20
mPa*seconds, under the measurement conditions described herein. In
some embodiment, the solution viscosity is in a range of from 10 to
20 mPa*seconds. In some embodiments, the solution viscosity is
about 15.5 mPa*seconds.
[0192] Solution viscosity can be determined using commercially
available stress-control rheometers (e.g., AR 2000 stress-control
rheometer, TA Instruments). In exemplary embodiments, the rheometer
is operated in the coneplate mode with a cone angle of 1.degree.
and a 60 mm diameter, as exemplified herein.
[0193] The phrase "absence of a peak in scattering intensity in the
interval 0.012<q<0.7 .ANG..sup.-1" means that for the
aforementioned range of values for the variable q, where q is
defined as:
q = 4 .pi. .lamda. sin .theta. ##EQU00001##
where 2.theta. is the scattering angle, and .lamda. is the
radiation wavelength (e.g., about 1.542 .ANG.), there is no value
of q for which the scattering intensity obtained by small angle
X-ray scattering is greater than for slightly lower and higher
values of q. Typically, in such a situation, the scattering
intensity is simply correlated to the values of q, such that the
highest scattering intensities are obtained for the highest values
of q measured, as exemplified herein.
[0194] Small angle X-ray scattering measurements (e.g., utilizing
Cu K.alpha. radiation) may be performed at ambient temperature
(e.g., 25.degree. C.) using commercially available devices (e.g.,
SAXSLAB GANESHA 300-XL system). The 2D SAXS images are azimuthally
averaged to produce one-dimensional profiles of intensity, I vs. q,
using commercially available data analysis programs, as exemplified
herein. The scattering spectra of the capillary and control
composition (i.e., composition lacking alginate) are collected and
subtracted from the corresponding composition data, to produce the
above-described scattering intensity as a function of q.
[0195] In some of any one of the embodiments described herein, the
alginate composition is characterized by a diffusion coefficient
described herein and by a solution viscosity described herein.
[0196] In some of any one of the embodiments described herein, the
alginate composition is characterized by a zeta potential described
herein and by a solution viscosity described herein.
[0197] In some of any one of the embodiments described herein, the
composition is characterized by a diffusion coefficient as
described herein, a zeta potential as described herein, and by a
solution viscosity described herein.
[0198] In some embodiments, the alginate composition is
characterized by a diffusion coefficient described herein and by a
scattering pattern described herein.
[0199] In some embodiments, the alginate composition is
characterized by a zeta potential described herein and by a
scattering pattern described herein.
[0200] In some embodiments, the composition is characterized by a
diffusion coefficient as described herein, a zeta potential as
described herein, and by a scattering pattern described herein.
[0201] In some embodiments, the alginate composition is
characterized by a diffusion coefficient described herein and by an
absence of observable structures more than 5 nm in width, as
described herein.
[0202] In some embodiments, the alginate composition is
characterized by a zeta potential described herein and by an
absence of observable structures more than 5 nm in width, as
described herein.
[0203] In some embodiments, the composition is characterized by a
diffusion coefficient as described herein, a zeta potential as
described herein, and by an absence of observable structures more
than 5 nm in width, as described herein.
[0204] The alginate may be characterized by any ratio of mannuronic
acid residues (M) to guluronic acid residues (G).
[0205] In some of any one of the embodiments described herein,
mannuronic acid represents at least 5% of the residues. In some
embodiments, mannuronic acid represents at least 10% of the
residues. In some embodiments, mannuronic acid represents at least
20% of the residues. In some embodiments, mannuronic acid
represents at least 30% of the residues. In some embodiments,
mannuronic acid represents at least 45% of the residues. In some
embodiments, mannuronic acid represents at least 50% of the
residues. In some embodiments, mannuronic acid represents at least
60% of the residues. In some embodiments, mannuronic acid
represents at least 70% of the residues. In some embodiments,
mannuronic acid represents at least 80% of the residues. In some
embodiments, mannuronic acid represents at least 90% of the
residues. In some embodiments, mannuronic acid represents at least
95% of the residues.
[0206] In some of any one of the embodiments described herein,
mannuronic acid represents from 25% to 65% of the residues, and
guluronic acid represents from 35% to 75% of the residues.
[0207] In some of any one of the embodiments described herein, the
alginate is cross-linked or partially cross-linked, for example, by
cations (e.g., divalent or trivalent cations) such as calcium,
copper, aluminum, magnesium, strontium, barium, tin, zinc and/or
chromium ions, and/or by organic cations and/or cationic polymers.
In some embodiments, the alginate is not cross-linked by
molybdenum, vanadium, tungsten or chromium ions.
[0208] In some embodiments, the alginate is non-cross-linked (e.g.,
substantially free of cations other than univalent cations).
[0209] In some of any one of the embodiments described herein, the
alginate is not silylated alginate.
[0210] In some of any one of the embodiments described herein, the
alginate described herein is derived from brown algae. Suitable
brown algae sources include, without limitation, Laminaria
hyperborea and Macrocystis pyrifera.
[0211] Types of alginate suitable for use in the context of
embodiments of the present invention are also described in
International Patent Application WO 2009/069131, the contents of
which are incorporated herein by reference.
[0212] According to any of the aspects described herein, the
alginate composition and/or the hepatotoxic agent described herein
can be administered either as alginate and/or hepatotoxic agent per
se (e.g., alginate composition described herein consists of
alginate per se and is administered as such), or as alginate and/or
hepatotoxic agent as a part of a composition which further
comprises a pharmaceutically acceptable carrier (e.g., the alginate
composition described herein consists of alginate and a
pharmaceutically acceptable carrier).
[0213] Hence, according to still another aspect of the present
invention, there is provided a pharmaceutical composition, which
comprises a therapeutically effective amount of a hepatotoxic agent
and a therapeutically effective amount of an alginate composition,
the alginate composition comprising a carrier which is a
pharmaceutically acceptable carrier.
[0214] As used herein a "pharmaceutical composition" refers to a
preparation of one or more active agents (e.g., alginate and/or
hepatotoxic agent) as described herein, with other chemical
components such as pharmaceutically acceptable and suitable
carriers and excipients. The purpose of a pharmaceutical
composition is to facilitate administration of a compound to an
organism.
[0215] Herein, the term "pharmaceutically acceptable carrier"
refers to a carrier or a diluent that does not cause significant
irritation to an organism and does not abrogate the biological
activity and properties of the administered compound. Examples,
without limitations, of pharmaceutically acceptable carriers are:
propylene glycol, saline, emulsions and mixtures of organic
solvents with water, as well as solid (e.g., powdered) and gaseous
carriers.
[0216] Herein the term "excipient" refers to an inert substance
added to a pharmaceutical composition to further facilitate
administration of a compound. Examples, without limitation, of
excipients include calcium carbonate, calcium phosphate, various
sugars and types of starch, cellulose derivatives, gelatin,
vegetable oils and polyethylene glycols.
[0217] It is to be understood that an alginate composition may
consist of an alginate and a pharmaceutically acceptable carrier
(as described herein), wherein the carrier is a solution containing
a hepatotoxic agent (as described herein), and that such an
alginate composition is, in itself, a pharmaceutical composition
such as described herein. However, for convenience and simplicity,
such a pharmaceutical composition is generally described herein as
a combination of a hepatotoxic agent and an alginate composition,
that is, the hepatotoxic agent is not considered as being a
component of the alginate composition.
[0218] It is to be further understood that a combination of a
hepatotoxic agent and an alginate composition comprising alginate
and a carrier, as described herein, does not indicate that the
carrier is necessarily contacted with the alginate (to form an
alginate composition) prior to being contacted with the hepatotoxic
agent. Rather, the components of the alginate composition (carrier
and alginate) and the hepatotoxic agent may be combined in any
order.
[0219] In some embodiments wherein the alginate composition and/or
pharmaceutical composition described herein comprises a liquid
carrier (e.g., an aqueous carrier), the composition comprises
alginate at a concentration in a range of from 0.4% to 10% (w/v).
In some embodiments, the concentration is in a range of from 1% to
4% (w/v). In exemplary embodiments, the concentration is about 2%
(w/v).
[0220] Techniques for formulation and administration of drugs may
be found in "Remington's Pharmaceutical Sciences" Mack Publishing
Co., Easton, Pa., latest edition, which is incorporated herein by
reference.
[0221] Pharmaceutical compositions of embodiments of the present
invention may be manufactured by processes well known in the art,
e.g., by means of conventional mixing, dissolving, granulating,
dragee-making, levigating, emulsifying, encapsulating, entrapping
or lyophilizing processes.
[0222] Pharmaceutical compositions for use in accordance with
embodiments of the present invention thus may be formulated in
conventional manner using one or more pharmaceutically acceptable
carriers comprising excipients and auxiliaries, which facilitate
processing of the alginate and/or hepatotoxic agent into
preparations which, can be used pharmaceutically. Proper
formulation is dependent upon the route of administration
chosen.
[0223] In some embodiments, the alginate composition and/or
hepatotoxic agent described herein are formulated for systemic
administration.
[0224] In some embodiments, the alginate composition and/or
hepatotoxic agent described herein are formulated for
intraperitoneal administration (e.g., intraperitoneal
injection).
[0225] For injection, the alginate and/or hepatotoxic agent
described herein may be formulated in aqueous solutions, preferably
in physiologically compatible buffers such as Hank's solution,
Ringer's solution, or physiological saline buffer with or without
organic solvents such as propylene glycol, polyethylene glycol. For
transmucosal administration, penetrants are used in the
formulation. Such penetrants are generally known in the art.
[0226] As exemplified herein, oral administration is a particularly
effective and convenient route for administering an alginate
composition.
[0227] For oral administration, the alginate composition and/or
hepatotoxic agent described herein can be formulated readily by
combining the alginate and/or hepatotoxic agent with
pharmaceutically acceptable carriers well known in the art. Such
carriers enable the alginate and/or hepatotoxic agent described
herein to be formulated as tablets, pills, dragees, capsules,
liquids, gels, syrups, slurries, suspensions, and the like, for
oral ingestion by a patient. Pharmacological preparations for oral
use can be made using a solid excipient, optionally grinding the
resulting mixture, and processing the mixture of granules, after
adding suitable auxiliaries if desired, to obtain tablets or dragee
cores. Suitable excipients are, in particular, fillers such as
sugars, including lactose, sucrose, mannitol, or sorbitol;
cellulose preparations such as, for example, maize starch, wheat
starch, rice starch, potato starch, gelatin, gum tragacanth, methyl
cellulose, hydroxypropylmethyl-cellulose, sodium
carboxymethylcellulose; and/or physiologically acceptable polymers
such as polyvinylpyrrolidone (PVP). If desired, disintegrating
agents may be added, such as cross-linked polyvinyl pyrrolidone or
agar. It is to be appreciated that alginate may also be an
effective disintegrating agent.
[0228] Dragee cores are provided with suitable coatings. For this
purpose, concentrated sugar solutions may be used which may
optionally contain gum arabic, talc, polyvinyl pyrrolidone,
carbopol gel, polyethylene glycol, titanium dioxide, lacquer
solutions and suitable organic solvents or solvent mixtures.
Dyestuffs or pigments may be added to the tablets or dragee
coatings for identification or to characterize different
combinations of alginate and/or hepatotoxic agent doses.
[0229] Pharmaceutical compositions, which can be used orally,
include push-fit capsules made of gelatin as well as soft, sealed
capsules made of gelatin and a plasticizer, such as glycerol or
sorbitol. The push-fit capsules may contain the active ingredients
in admixture with filler such as lactose, binders such as starches,
lubricants such as talc or magnesium stearate and, optionally,
stabilizers. In soft capsules, the alginate and/or hepatotoxic
agent described herein may be dissolved or suspended in suitable
liquids, such as fatty oils, liquid paraffin, or liquid
polyethylene glycols. In addition, stabilizers may be added. All
formulations for oral administration should be in dosages suitable
for the chosen route of administration.
[0230] For buccal administration, the compositions may take the
form of tablets or lozenges formulated in conventional manner.
[0231] For administration by inhalation, the alginate composition
and/or hepatotoxic agent described herein may be conveniently
delivered in the form of an aerosol spray presentation (which
typically includes powdered, liquified and/or gaseous carriers)
from a pressurized pack or a nebulizer, with the use of a suitable
propellant, e.g., dichlorodifluoromethane, trichlorofluoromethane,
dichloro-tetrafluoroethane or carbon dioxide. In the case of a
pressurized aerosol, the dosage unit may be determined by providing
a valve to deliver a metered amount. Capsules and cartridges of,
e.g., gelatin for use in an inhaler or insufflator may be
formulated containing a powder mix of the alginate composition
and/or hepatotoxic agent described herein and a suitable powder
base such as, but not limited to, lactose or starch.
[0232] The alginate composition and/or hepatotoxic agent described
herein may be formulated for parenteral administration, e.g., by
bolus injection or continuous infusion. Formulations for injection
may be presented in unit dosage form, e.g., in ampoules or in
multidose containers with optionally, an added preservative. The
compositions may be suspensions, solutions or emulsions in oily or
aqueous vehicles, and may contain formulatory agents such as
suspending, stabilizing and/or dispersing agents.
[0233] The phrase "unit dosage form", as used herein, describes
physically discrete units, each unit containing a predetermined
quantity of active ingredients (e.g., an alginate and/or
hepatotoxic agent as described herein) calculated to produce the
desired effect, in association with at least one pharmaceutically
acceptable carrier, diluent, excipient, or combination thereof.
[0234] Pharmaceutical compositions for parenteral administration
include aqueous solutions of the alginate and/or hepatotoxic agent
described herein in water-soluble form. Additionally, suspensions
of the alginate and/or hepatotoxic agent described herein may be
prepared as appropriate oily injection suspensions and emulsions
(e.g., water-in-oil, oil-in-water or water-in-oil in oil
emulsions). Suitable lipophilic solvents or vehicles include fatty
oils such as sesame oil, or synthetic fatty acids esters such as
ethyl oleate, triglycerides or liposomes. Aqueous injection
suspensions may contain substances, which increase the viscosity of
the suspension, such as sodium carboxymethyl cellulose, sorbitol or
dextran. Optionally, the suspension may also contain suitable
stabilizers or agents, which increase the solubility of the
alginate and/or hepatotoxic agent described herein to allow for the
preparation of highly concentrated solutions.
[0235] Alternatively, the alginate composition and/or hepatotoxic
agent described herein may be in powder form for constitution with
a suitable vehicle, e.g., sterile, pyrogen-free water or saline,
before use.
[0236] The alginate composition and/or hepatotoxic agent described
herein may also be formulated in rectal compositions such as
suppositories or retention enemas, using, e.g., conventional
suppository bases such as cocoa butter or other glycerides.
[0237] The pharmaceutical compositions herein described may also
comprise suitable solid of gel phase carriers or excipients.
Examples of such carriers or excipients include, but are not
limited to, calcium carbonate, calcium phosphate, various sugars,
starches, cellulose derivatives, gelatin and polymers such as
polyethylene glycols.
[0238] Pharmaceutical compositions suitable for use in context of
the present invention include compositions wherein the active
ingredients are contained in an amount effective to achieve the
intended purpose. More specifically, a therapeutically effective
amount of alginate composition and/or hepatotoxic agent described
herein is an amount effective to prevent, alleviate or ameliorate
symptoms of liver damage and/or disease treatable by hepatotoxic
agent, and/or prolong the survival of the subject being
treated.
[0239] Determination of a therapeutically effective amount of
alginate composition and/or hepatotoxic agent described herein is
well within the capability of those skilled in the art, especially
in light of the detailed disclosure provided herein, as well as
knowledge in the art regarding dosage of hepatotoxic agents.
[0240] For any alginate composition and/or hepatotoxic agent
described herein, the therapeutically effective amount or dose can
be estimated initially from activity assays in animals (e.g., as
exemplified herein). For example, a dose can be formulated in
animal models to achieve a circulating concentration range that
includes the IC.sub.50 as determined by activity assays (e.g., the
concentration of the alginate, which achieves a half-maximal
reduction of liver damage, as quantified according to assays known
in the art (e.g., assays exemplified herein). Such information can
be used to more accurately determine useful doses in humans.
[0241] A therapeutically effective amount for the alginate in a
composition may range from about 8 mg/kg body weight to about 3,200
mg/kg body weight. In some embodiments, the therapeutically
effective amount for the alginate is from about 16 mg/kg body
weight to about 3,200 mg/kg body weight. In some embodiments, the
therapeutically effective amount for the alginate is more than 40
mg/kg body weight. In some embodiments, the therapeutically
effective amount for the alginate is at least about 80 mg/kg body
weight. As is demonstrated in the Examples section that follows, an
amount of an alginate of 100 mg/kg or higher was shown to exhibit a
more potent protective effect in mice than 50 mg/kg.
[0242] Toxicity and therapeutic efficacy of the alginate
composition and/or hepatotoxicity described herein can be
determined based on prior knowledge in the art regarding such
agents and/or by standard pharmaceutical procedures in experimental
animals, e.g., by determining the EC.sub.50, the IC.sub.50 and the
LD.sub.50 (lethal dose causing death in 50% of the tested animals).
The data obtained from these activity assays and animal studies can
be used in formulating a range of dosage for use in human.
[0243] The dosage may vary depending upon the dosage form employed
and the route of administration utilized. The exact formulation,
route of administration and dosage can be chosen by the individual
physician in view of the patient's condition. (See e.g., Fingl et
al., 1975, in "The Pharmacological Basis of Therapeutics", Ch. 1 p.
1).
[0244] Dosage amount and interval may be adjusted individually to
provide plasma levels of the active moiety which are sufficient to
maintain the desired effects, termed the minimal effective
concentration (MEC). The MEC will vary for each agent, but can be
estimated from in vitro data. Dosages necessary to achieve the MEC
will depend on individual characteristics and route of
administration. HPLC assays or bioassays can be used to determine
plasma concentrations.
[0245] Dosage intervals can also be determined using the MEC value.
Preparations should be administered using a regimen, which
maintains plasma levels above the MEC for 10-90% of the time,
preferable between 30-90% and most preferably 50-90%.
[0246] Depending on the severity and responsiveness of the
condition to be treated, dosing can also be a single administration
of a slow release or delayed release formulation of a composition
described hereinabove, with course of treatment lasting from
several hours to several weeks or until cure is effected or
diminution of the disease state is achieved.
[0247] In some embodiments of any of the aspects described herein,
administration is effected using a composition formulated as a slow
release or delayed release composition, designed to achieve maximal
overlap between exposure of the liver to the effects of the
alginate and exposure of the liver to the hepatotoxic agent. In
order to maximize such overlap, either the alginate composition,
the hepatotoxic agent, or both, may be formulated for slow release
and/or delayed release, depending on the relative pharmacokinetics
of the alginate and hepatotoxic agent.
[0248] For example, in some embodiments, a hepatotoxic drug is
formulated for slow release and/or delayed release (e.g., a
standard commercially available formulation of the drug), and an
alginate composition for co-administration with the drug is
formulated for a slow release and/or delayed release with similar
release kinetics. Alternatively, a hepatotoxic drug is formulated
for delayed release (e.g., a standard commercially available
formulation of the drug), and an alginate composition for
co-administration with the drug is co-administered prior to,
concomitant with or shortly after the hepatotoxic agent is released
(which is the exposure of the subject to the hepatotoxic
agent).
[0249] Techniques for preparing slow release formulations are known
in the art, including suitable coatings for solid compositions
which release the coated compositions according to a pre-determined
time profile, and suitable capsules which release liquid
compositions after a pre-determined period of time.
[0250] The amount of a composition to be administered will, of
course, be dependent on the subject being treated, the severity of
the affliction, the manner of administration, the judgment of the
prescribing physician, etc.
[0251] Compositions of the present invention may, if desired, be
presented in a pack or dispenser device, such as an FDA (the U.S.
Food and Drug Administration) approved kit, which may contain one
or more unit dosage forms containing the alginate composition
and/or hepatotoxic agent described herein. The pack may, for
example, comprise metal or plastic foil, such as, but not limited
to a blister pack or a pressurized container (for inhalation). The
pack or dispenser device may be accompanied by instructions for
administration. The pack or dispenser may also be accompanied by a
notice associated with the container in a form prescribed by a
governmental agency regulating the manufacture, use or sale of
pharmaceuticals, which notice is reflective of approval by the
agency of the form of the compositions for human or veterinary
administration. Such notice, for example, may be of labeling
approved by the U.S. Food and Drug Administration for prescription
drugs or of an approved product insert. Compositions comprising an
alginate composition and/or hepatotoxic agent described herein
formulated in a compatible pharmaceutical carrier may also be
prepared, placed in an appropriate container, and labeled for
treatment of an indicated condition or diagnosis, as is detailed
hereinabove (e.g., treating liver damage, reducing or preventing a
liver damage caused by at least one identified hepatotoxic agent,
treating a condition treatable by the hepatotoxic agent).
[0252] Thus, according to some embodiments of any aspect described
herein, the alginate composition and/or hepatotoxic agent described
herein, for example, in a form of a composition described herein,
are packaged in a packaging material and identified in print, in or
on the packaging material, for use in the treatment of a condition
described herein (e.g., alginate composition identified for
protecting against liver damage, for treating liver damage and/or
for reducing or preventing a liver damage caused by at least one
identified hepatotoxic agent; composition comprising alginate and
hepatotoxic agent identified for treating a condition treatable by
the hepatotoxic agent in the composition).
[0253] The co-formulations described herein of a hepatotoxic agent
with an alginate composition are considerably superior to current
formulations of hepatotoxic agents, in view of the hepatoprotection
provided by the alginate. The co-formulations described herein may
therefore be used without required warnings against the risk of
liver damage (or with less stringent warnings) and/or without
required monitoring of liver function (or with less frequent
monitoring) in a subject being administered the co-formulation, in
contrast to current formulations of hepatotoxic agents.
[0254] It is expected that during the life of a patent maturing
from this application many relevant hepatotoxic agents will be
developed and/or newly identified and the scope of the term
"hepatotoxic agent" is intended to include all such new agents a
priori.
[0255] As used herein the term "about" refers to .+-.10%.
[0256] The terms "comprises", "comprising", "includes",
"including", "having" and their conjugates mean "including but not
limited to".
[0257] The term "consisting of" means "including and limited
to".
[0258] The term "consisting essentially of" means that the
composition, method or structure may include additional
ingredients, steps and/or parts, but only if the additional
ingredients, steps and/or parts do not materially alter the basic
and novel characteristics of the claimed composition, method or
structure.
[0259] As used herein, the singular form "a", "an" and "the"
include plural references unless the context clearly dictates
otherwise. For example, the term "a compound" or "at least one
compound" may include a plurality of compounds, including mixtures
thereof.
[0260] Throughout this application, various embodiments of this
invention may be presented in a range format. It should be
understood that the description in range format is merely for
convenience and brevity and should not be construed as an
inflexible limitation on the scope of the invention. Accordingly,
the description of a range should be considered to have
specifically disclosed all the possible subranges as well as
individual numerical values within that range. For example,
description of a range such as from 1 to 6 should be considered to
have specifically disclosed subranges such as from 1 to 3, from 1
to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6 etc., as
well as individual numbers within that range, for example, 1, 2, 3,
4, 5, and 6. This applies regardless of the breadth of the
range.
[0261] Whenever a numerical range is indicated herein, it is meant
to include any cited numeral (fractional or integral) within the
indicated range. The phrases "ranging/ranges between" a first
indicate number and a second indicate number and "ranging/ranges
from" a first indicate number "to" a second indicate number are
used herein interchangeably and are meant to include the first and
second indicated numbers and all the fractional and integral
numerals therebetween.
[0262] As used herein the term "method" refers to manners, means,
techniques and procedures for accomplishing a given task including,
but not limited to, those manners, means, techniques and procedures
either known to, or readily developed from known manners, means,
techniques and procedures by practitioners of the chemical,
pharmacological, biological, biochemical and medical arts.
[0263] It is appreciated that certain features of the invention,
which are, for clarity, described in the context of separate
embodiments, may also be provided in combination in a single
embodiment. Conversely, various features of the invention, which
are, for brevity, described in the context of a single embodiment,
may also be provided separately or in any suitable subcombination
or as suitable in any other described embodiment of the invention.
Certain features described in the context of various embodiments
are not to be considered essential features of those embodiments,
unless the embodiment is inoperative without those elements.
[0264] Various embodiments and aspects of the present invention as
delineated hereinabove and as claimed in the claims section below
find experimental support in the following examples.
EXAMPLES
[0265] Reference is now made to the following examples, which
together with the above descriptions illustrate some embodiments of
the invention in a non limiting fashion.
Materials and Methods
[0266] Materials:
[0267] Alginates (VLVG and LVG) were obtained from NovaMatrix. VLVG
refers to Very Low Viscosity (high) G alginate, as designated by
the manufacturer. LVG refers to Low Viscosity (high) G alginate, as
designated by the manufacturer. LVG alginate with different
molecular weight are referred to herein as LVG54 and LVG150,
wherein the numbers are those reported by the manufacturer for
solution viscosity of the product for 1% (w/v) solutions.
[0268] Hyaluronan (sodium salt; .about.400 kDa; from Streptococcus
equinus) was obtained from Sigma.
[0269] Concanavalin A (Con A) was obtained from MP Biomedicals.
[0270] Paracetamol syrup (Tiptipot.RTM. syrup) and the vehicle
(paracetamol-free) for the syrup were obtained from CTS
(Israel).
[0271] Preparation of Polysaccharide Solutions:
[0272] Polysaccharides (e.g., alginate) were dissolved in saline
(0.15 M NaCl in water), and mixed intensively with a homogenizer
(28,000 rotations per minute, for 3 minutes). All of the alginate
samples included alginate at a concentration of 2% (w/v). All
samples were sterilized by filtration through a 0.2 .mu.m nylon
membrane. The solutions were maintained at a temperature of
4.degree. C. until use, and appeared clear by eye.
[0273] Animals:
[0274] Male C57BL/6 (B6) mice (11-12 weeks old) were purchased from
Harlan Laboratories (Jerusalem, Israel). All mice were maintained
in specific pathogen-free conditions. Mice were maintained in the
Animal Core of the Hadassah-Hebrew University Medical School. All
mice were administered standard laboratory chow and water ad
libitum and kept in a 12-hour light/dark cycle.
[0275] Rheological Characterization of the Polysaccharide
Solutions:
[0276] The viscosity of the polysaccharide solutions was tested on
an AR 2000 stress-control rheometer (TA Instruments), operated in
the coneplate mode with a cone angle of 1.degree. and a 60 mm
diameter. The apparent viscosities (mPa*seconds) of the solutions
were tested at a shear rate of 1 seconds.sup.-1. The measuring
device was equipped with a temperature control unit (Peltier plate,
.sup..+-.0.05.degree. C.) operated at 25.degree. C.
[0277] Determination of Polysaccharide Molecular Weight:
[0278] Polysaccharide molecular weights were determined by gel
permeation chromatography-multiangle laser light scattering
(GPC-MALLS). Samples were separated on a chromatographic system
comprising a Waters 606 pump followed by two PSS Suprema gel
permeation columns connected in a series (column description:
dimensions 300.times.8 mm.sup.2, particle size 10 mm, porosity of
3000 and 10,000 angstrom). The flow rate was 0.5 ml/minute. The
columns were kept at a constant temperature of 25.degree. C. inside
a Techlab K-4 controlled oven. The chromatographic system was
attached to a Dawn DSP (Wyatt Technology) multiangle laser light
scattering (MALLS) photometer equipped with a He/Ne laser working
at 632.8 nm, a K5 refraction cell and 18 detectors at angles of
14-163.degree..
[0279] Concentration was monitored by a calibrated interferometric
refractometer Optilab DSP (Wyatt Technology). Data processing and
molar mass calculation were performed with Wyatt ASTRA software
version 4.7. Each sample was injected three times to ensure
reproducibility. The dn/dc of the alginate, measured with the
Optilab DSP, controlled by Wyatt dn/dc software, was found to be
0.155 ml/gram (aqueous buffer).
[0280] Aqueous buffer solutions were prepared from ultrapure water
(0.055 .mu.s/cm, USF Seral Purelab RO75, followed by USF Seral
Purelab UV) supplemented with 0.1 M NaNO.sub.3, 0.02% (w/v) NaN3
and 10 mM imidazole. The buffer was titrated with NaNO.sub.3 to a
pH of 7.0 and filtered through a 0.1 .mu.m VacuCap.RTM. 60 filter
(Gelman Sciences).
[0281] Alanine Aminotransferase (ALT) Assays:
[0282] Serum activities of alanine aminotransferase (ALT) was
carried out after 1:10 dilutions, using a Reflovet.RTM. Plus
clinical chemistry analyzer (Roche Diagnostics).
[0283] IL-6 Assays:
[0284] Serum levels of IL-6 were determined by "sandwich" ELISA,
using a Quantikine.RTM. assay kit (R&D Systems), according to
the manufacturer's instructions. Sera from mice were frozen until
ELISA analysis.
[0285] Paracetamol Intoxication Mouse Model:
[0286] Male C57BL/6 (B6) mice (11-12 weeks old) were orally
administered Tiptipot.RTM. paracetamol (acetaminophen,
N-acetyl-p-aminophenol, APAP) syrup, after an overnight fast. The
paracetamol dosages were 160 mg/kg (approximately 4 mg) or 320
mg/kg (approximately 8 mg) in other examples. The syrup was always
diluted with saline to yield a total volume of 350 .mu.l per mouse.
Paracetamol was administered in the morning and food was put back
to the cages 2 hours later. Mice were sacrificed 24 hours after
paracetamol administration.
[0287] In the preventive model, alginate solutions (50, 100 or 200
.mu.l per mouse) were administered per os or i.p. prior to
administration of 160 mg/kg (approximately 4 mg) or 320 mg/kg
(approximately 8 mg) paracetamol. In the therapeutic model,
alginate solutions were administered with 160 mg/kg (approximately
4 mg) paracetamol (by being mixed with the paracetamol) or shortly
thereafter.
[0288] Control mice were administered only with paracetamol syrup
diluted with saline, as described hereinabove. In some experiments
control mice were orally administered with the vehicle for the
Tiptipot.RTM. paracetamol syrup, which was identical to the
paracetamol syrup except for being paracetamol-free.
[0289] In some experiments paracetamol levels in the blood were
determined 30 or 60 min after paracetamol administration, by taking
20 .mu.l of blood from the tail vein of all mice. Paracetamol
levels were measured by using a clinical kit based on fluorescence
polarization immunoassay (FPIA, AxSYM acetaminophen assay (Abbott),
obtained from Ilex Medical Ltd., Israel). Serum activities of ALT
and cytokines were determined as described herein.
[0290] After mice were sacrificed, a portion of each excised liver
was fixed in 10% formalin and was then embedded in paraffin,
sectioned (specimens of 5 .mu.m) and stained with hematoxylin &
eosin (H&E) or with IgG staining for detection of necrosis,
nitrotyrosine and/or Ki-67. Briefly, necrosis staining was as
follows: sections were first de-paraffinized and then were
incubated with rabbit polyclonal IgG. Sections were then incubated
with secondary antibody using the MACH 3 Rabbit HRP Polymer
Detection and then dehydrated, cleared and mounted in synthetic
resin. For color detection, DAB (3,3'-diaminobenzidine) was
applied, followed by counterstaining. Quantification of necrosis
was carried out by the Ariol SL-50 system (Applied Imaging) on
microscope slides. Nitrotyrosine and Ki-67 immunostaining staining
was performed similarly, using a rabbit polyclonal
anti-nitrotyrosine antibody (Abcam) or an anti-Ki-67 antibody
(DAKO).
[0291] Weight changes were monitored by weighing all the mice on
the fasting day, the following morning, just before paracetamol
administration (Day 1) and on the following day before sacrifice
(Day 2).
Example 1
Effect of Alginate on Paracetamol Hepatotoxicity (Preventive
Model)
[0292] Aqueous solutions of 2% alginate (w/v) were prepared by
homogenization in saline, as described in the Materials and Methods
section, from the following types of alginate:
[0293] VLVG (solution viscosity 15.5 mPa*seconds, molecular weight
30-50 kDa);
[0294] LVG54 (solution viscosity 269 mPa*seconds, molecular weight
100 kDa); and
[0295] LVG150 (solution viscosity 879 mPa*seconds, molecular weight
156 kDa).
[0296] Solution viscosity and molecular weight were characterized
as described hereinabove.
[0297] The effect of alginate on paracetamol hepatotoxicity was
investigated in a mouse model, as described in the Materials and
Methods section.
[0298] To this end, 200 .mu.l of a 2% solution of VLVG alginate was
administered per os 30 minutes prior to intoxication with 4 mg
paracetamol. Paracetamol intoxication was evaluated by measuring
levels of ALT and IL-6, by measuring the weight loss typical of
paracetamol intoxication, and by examining necrosis by IgG
staining, according to procedures described in the Materials and
Methods section.
[0299] As shown in FIG. 1, the orally administered VLVG alginate
dramatically reduced serum ALT levels in mice 24 hours after
paracetamol administration, from 6,785.+-.3,230 units per liter to
109.+-.54 units per liter.
[0300] In addition, the orally administered VLVG alginate reduced
serum IL-6 levels from 119.6.+-.66.6 pg/ml to 45.6.+-.33.3 pg/ml
(p=0.05).
[0301] In addition, as shown in Table 2 below, the orally
administered VLVG alginate reversed the weight loss caused by
paracetamol intoxication between Day 1 (the day of intoxication)
and Day 2 (the following day).
[0302] In addition, as shown in FIGS. 2A and 2B, the orally
administered VLVG alginate reduced the degree of necrosis in the
liver of paracetamol intoxicated mice.
TABLE-US-00002 TABLE 2 Effect of 4 mg paracetamol and VLVG alginate
on body weight Change in weight from Weight ratio Day 1 to Day 2
Group (Day 2 vs. Day 1) (mg) 4 mg paracetamol alone 99% -0.15 4 mg
paracetamol + VLVG 105% 1.17 alginate (2% w/v, per os) p <
0.0001 for change in weight
[0303] In order to assess the effect of alginate dosage on liver
damage caused by paracetamol, the above experiment was repeated so
as to compare the effects of 50, 100 and 200 .mu.l of a 2% solution
of VLVG alginate, which correspond to doses of 1 mg, 2 mg and 4 mg
of VLVG alginate, respectively.
[0304] As shown in FIG. 3, 100 .mu.l (2 mg alginate) and 200 .mu.l
(4 mg alginate) of the orally administered VLVG alginate solution
dramatically reduced serum ALT levels in mice 24 hours after
paracetamol administration, from 15133.+-.1283 units per liter to
149.+-.28 units per liter and 62.+-.15 (200 .mu.l) units per liter,
respectively, whereas 50 .mu.l (1 mg alginate) of the orally
administered VLVG alginate only moderately reduced serum ALT
levels, from 15133.+-.1283 units per liter to 10813.+-.2253 units
per liter.
[0305] The effects of different dosages of alginate on liver damage
caused by paracetamol were also assessed by staining for
nitrotyrosine (a marker of oxidative stress associated with
peroxynitrite formation due to increased nitric oxide production
following liver injury) and Ki-67 (a marker of cell proliferation),
according to procedures described in the Materials and Methods
section. Paracetamol toxicity develops only after the onset of
oxidative stress and mitochondrial dysfunction, and preventing
these phenomena protects against paracetamol toxicity.
[0306] As shown in FIG. 4, treatment with 4 mg paracetamol resulted
in extensive centrilobular nitrotyrosine staining, which was
limited by 50 .mu.l of 2% VLVG solution, and which was abolished by
200 .mu.l of 2% VLVG solution.
[0307] As shown in FIG. 5, treatment with 4 mg paracetamol resulted
in widespread cell proliferation in the liver, which was limited to
certain areas by 50 .mu.l of 2% VLVG solution, and which was
abolished by 200 .mu.l of 2% VLVG solution.
[0308] These results indicate that VLVG reduces oxidative stress
and cell proliferation associated with paracetamol hepatotoxicity,
and confirm that doses above 50 .mu.l of 2% VLVG solution (1 mg
alginate) can effectively prevent development of adverse effects
associated with paracetamol hepatotoxicity.
[0309] Paracetamol is absorbed rapidly into the blood stream,
reaching peak serum values in about 1-3 hours. In order to
ascertain whether the protective effect is mediated by changes in
paracetamol absorption, paracetamol levels in the blood were
determined after administration of paracetamol.
[0310] As shown in FIG. 6, neither 100 .mu.l nor 200 .mu.l of the
orally administered VLVG alginate solution exhibited any
statistically significant effect on paracetamol levels in the blood
1, 4 or 24 hours after administration of paracetamol.
[0311] Similarly, when paracetamol levels in the blood were
determined 30 minutes after administration of paracetamol, the
paracetamol levels with and without administration of 200 .mu.l of
VLVG alginate solution were essentially identical: 67.72.+-.40.55
.mu.g/ml with VLVG alginate administration and 65.07.+-.43.17
.mu.g/ml without VLVG alginate.
[0312] These results indicate that the alginate does not affect
paracetamol absorption, and that the protective effects of alginate
are not mediated by changes in paracetamol absorption.
[0313] In order to further assess the effect of alginate on liver
damage caused by paracetamol, the above experiment was repeated so
as to compare the effect of orally administered VLVG alginate (200
.mu.l of a 2% solution) with that of intraperitoneally administered
VLVG alginate and with orally administered LVG54 and LVG150
alginate (200 .mu.l of a 2% solution). Liver damage was evaluated
by measuring ALT levels and body weight. Each treatment group
included 4 mice.
[0314] As shown in FIG. 7, intraperitoneally administered VLVG
alginate, orally administered VLVG alginate, LVG54 alginate and
LVG150 alginate each reduced serum ALT levels in mice 24 hours
after paracetamol administration. Furthermore, VLVG alginate was
more effective than LVG54 and LVG150 alginate at reducing ALT
levels, and orally administered VLVG alginate was more effective
than intraperitoneally administered VLVG alginate. ALT levels were
13,250.+-.2,415 units/liter 24 hours after paracetamol
administration alone, and were reduced to 1,987.+-.1,316
units/liter following i.p. administration of VLVG alginate, to
202.+-.159 units/liter following oral administration of VLVG
alginate, to 3,525.+-.3,938 units/liter following oral
administration of LVG54 alginate, and to 6,504.+-.4,277 units/liter
following oral administration of LVG150 alginate.
[0315] In addition, as shown in Table 3 below, intraperitoneally
administered VLVG alginate, orally administered VLVG alginate,
LVG54 alginate and LVG150 alginate each reversed the weight loss
caused by paracetamol intoxication between Day 1 (the day of
intoxication) and Day 2 (the following day).
TABLE-US-00003 TABLE 3 Effect of 4 mg paracetamol and alginate on
body weight Change in weight from Weight ratio Day 1 to Day 2 Group
(Day 2 vs. Day 1) (mg) 4 mg paracetamol alone 96% -1.05 4 mg
paracetamol + VLVG 102% 0.5 alginate (2% w/v, i.p.) 4 mg
paracetamol + VLVG 101% 2.1 alginate (2% w/v, per os) 4 mg
paracetamol + LVG54 108% 2.08 alginate (2% w/v, per os) 4 mg
paracetamol + LVG150 103% 0.8 alginate (2% w/v, per os)
[0316] The above results indicate that alginate can be administered
by various routes to prevent liver damage caused by drug
hepatotoxicity, but that oral administration of alginate is
particularly effective for preventing liver damage caused by drug
hepatotoxicity. The results further indicate that VLVG alginate is
more effective against liver damage than alginates characterized by
different molecular weights and viscosities.
[0317] In order to assess the ability of alginate to protect
against liver damage caused by higher doses of paracetamol, the
above experiments were repeated using doses of 8 mg paracetamol (a
sub-lethal dose) instead of 4 mg. Paracetamol intoxication was
evaluated by measuring levels of ALT and IL-6, and by measuring
body weight. This experiment was performed twice, once with
administration of a vehicle as a control (as described in the
Materials and Methods section), and once without.
[0318] In one experiment, as shown in FIG. 8, VLVG alginate reduced
serum ALT levels in mice 24 hours after administration of 8 mg
paracetamol, from 21,743.+-.5,790 units/liter to 10,903.+-.4,798
units per liter.
[0319] In addition, the orally administered VLVG alginate reduced
serum IL-6 levels from 91.65.+-.28.9 pg/ml to 43.17.+-.15.42 pg/ml
(p<0.005).
[0320] In addition, as shown in Table 4 below, the orally
administered VLVG alginate reversed the weight loss caused by
paracetamol intoxication between Day 1 (the day of intoxication)
and Day 2 (the following day).
TABLE-US-00004 TABLE 4 Effect of 8 mg paracetamol and VLVG alginate
on body weight Change in weight from Weight ratio Day 1 to Day 2
Group (Day 2 vs. Day 1) (mg) 8 mg paracetamol alone 99% -0.17 8 mg
paracetamol + VLVG 110% 2.42 alginate (2% w/v, per os)
[0321] In the second experiment, as shown in FIG. 9, VLVG alginate
reduced serum ALT levels in mice 24 hours after administration of 8
mg paracetamol, from 25,520.+-.3,068 units/liter to 11,845.+-.7,025
units per liter, whereas only 28.+-.1 units/liter ALT was detected
in serum when the vehicle was administered instead of
paracetamol.
[0322] In addition, the orally administered VLVG alginate reduced
serum IL-6 levels from 247.0.+-.234 pg/ml to 84.4.+-.25 pg/ml,
whereas only 18.1.+-.1.5 pg/ml IL-6 was detected in serum when the
vehicle was administered instead of paracetamol.
[0323] In addition, as shown in Table 5 below, the orally
administered VLVG alginate reversed the weight loss caused by
paracetamol intoxication between Day 1 (the day of intoxication)
and Day 2 (the following day).
TABLE-US-00005 TABLE 5 Effect of 8 mg paracetamol and VLVG alginate
on body weight Change in weight from Weight ratio Day 1 to Day 2
Group (Day 2 vs. Day 1) (mg) Syrup vehicle 106% 1.6 8 mg
paracetamol alone 99% -0.3 8 mg paracetamol + VLVG 102% 0.4
alginate (2% w/v, per os)
[0324] In addition, paracetamol levels in the blood were determined
30 minutes after administration of 8 mg paracetamol (or vehicle).
The paracetamol levels with and without administration of VLVG
alginate were essentially identical: 169.9.+-.100.7 .mu.g/ml with
VLVG alginate administration and 153.4.+-.99.8 .mu.g/ml without
VLVG alginate. In contrast, only 5.9.+-.3.7 .mu.g/ml paracetamol
was detected when the vehicle was administered, which corresponds
to the background signal for the assay kit.
[0325] These results indicate that alginate protects against
relatively high doses of paracetamol, and that the protective
effects of alginate are not mediated by changes in paracetamol
absorption.
Example 2
Effect of Alginate on Paracetamol Hepatotoxicity (Therapeutic
Model)
[0326] In view of the results presented in Example 1, which show a
strong protective effect of alginate when administered prior to
administration of paracetamol (a preventive model), the effect of
alginate on paracetamol hepatotoxicity was investigated in a
therapeutic model, in which alginate was administered concurrently
with, or subsequent to, administration of paracetamol, as described
in the Materials and Methods section.
[0327] To this end, a 2% solution of VLVG alginate was prepared as
described hereinabove, and administered per os at a volume of 200
.mu.l 30 or 60 minutes after intoxication with 4 mg paracetamol.
Paracetamol intoxication was evaluated by measuring levels of ALT
and IL-6, and by measuring body weight, as described
hereinabove.
[0328] As shown in FIG. 10, VLVG alginate administered 30 minutes
after paracetamol administration dramatically reduced serum ALT
levels in mice 24 hours later, whereas VLVG alginate administered
60 minutes after paracetamol administration did not reduce serum
ALT levels. Serum ALT levels were 1,025.+-.1,310 units/liter when
VLVG alginate was administered 30 minutes after paracetamol,
10,020.+-.5,031 units/liter when VLVG alginate was administered 60
minutes after paracetamol, and 6,393.+-.7,304 units/liter when
paracetamol was administered without alginate.
[0329] In addition, VLVG alginate administered 30 minutes after
paracetamol reduced serum IL-6 levels from 248.8.+-.98.4 pg/ml to
63.4.+-.42.5 pg/ml. In contrast, when VLVG alginate was
administered 60 minutes after paracetamol, serum IL-6 levels were
131.1.+-.52.4 pg/ml.
[0330] In addition, as shown in Table 6 below, VLVG alginate
administered 30 minutes after paracetamol reversed the weight loss
caused by paracetamol intoxication, whereas VLVG alginate
administered 60 minutes after paracetamol did not.
TABLE-US-00006 TABLE 6 Effect of administration time of VLVG
alginate on body weight Change in weight from Weight ratio Day 1 to
Day 2 Group (Day 2 vs. Day 1) (mg) 4 mg paracetamol alone 94% -1.5
4 mg paracetamol + VLVG 104% 1.03 alginate after 30 minutes 4 mg
paracetamol + VLVG 98% -0.43 alginate after 60 minutes
[0331] The above experiment was repeated using VLVG alginate mixed
with the paracetamol and co-administered.
[0332] As shown in FIG. 11, VLVG alginate dramatically reduced
serum ALT levels in mice 24 hours later, whether administered 30
minutes after paracetamol or when administered mixed with
paracetamol, although the protective effect of the VLVG alginate
was somewhat stronger when administered after paracetamol. Serum
ALT levels were 124.+-.93 units/liter when VLVG alginate was
administered 30 minutes after paracetamol, 902.+-.1,520 units/liter
when VLVG alginate was mixed with paracetamol, and 6,734.+-.3,783
units/liter when paracetamol was administered without alginate.
[0333] In addition, VLVG alginate reduced serum IL-6 levels in mice
24 hours later, whether administered 30 minutes after paracetamol
or when administered mixed with paracetamol. VLVG alginate
administered 30 minutes after paracetamol reduced serum IL-6 levels
from 164.1.+-.85.3 pg/ml to 58.9.+-.31.8 pg/ml, while VLVG alginate
mixed with paracetamol similarly reduced serum IL-6 levels to
58.9.+-.38.7 pg/ml.
[0334] In addition, as shown in Table 7 below, VLVG alginate
reversed the weight loss caused by paracetamol intoxication,
whether administered 30 minutes after paracetamol or when
administered mixed with paracetamol.
TABLE-US-00007 TABLE 7 Effect of administration time of VLVG
alginate on body weight Change in weight from Weight ratio Day 1 to
Day 2 Group (Day 2 vs. Day 1) (mg) 4 mg paracetamol alone 98% -0.52
4 mg paracetamol + VLVG 102% 0.52 alginate after 30 minutes 4 mg
paracetamol mixed 102% 0.56 with VLVG alginate
[0335] These results indicate that alginate exhibits a protective
effect against drug hepatotoxicity, when administered prior to or
concurrently with a hepatotoxic drug, or when administered less
than 60 minutes after administration of the drug.
Example 3
Biodistribution of Alginate
[0336] The biodistribution of alginate was determined using
immunohistochemical staining for biotin-labeled alginate.
[0337] VLVG alginate was labeled with biotin via carbodiimide
chemistry, using procedures as described in Freeman et al.
[Biomaterials 29:3260-3268 (2008)]. No more than 3% of uronic acid
residues were modified, so the biotin-labeled alginate was quite
similar to the non-labeled alginate. A solution of 2% biotinylated
VLVG alginate was administered to mice intraperitoneally (200 .mu.l
per mouse), and the mice were sacrificed after 48 hours. The liver,
spleen, colon or other tissue were then harvested and fixed in 10%
formalin and were then embedded in paraffin and sectioned
(specimens of 5 .mu.m). For biotin detection, slides were first
de-paraffinized, followed by addition of target retrieval solution
and then blocking by peroxidase blocker (DAKO). After incubations
and rinsing, Streptavidin-peroxidase (DAKO) was applied. For color
detection, DAB (3,3'-diaminobenzidine) and chromogen substrate were
applied. The procedure was ended by counterstaining and cover slip
mounting.
[0338] As shown in FIG. 12, intraperitoneally administered VLVG
alginate appeared in the liver parenchyma, as determined by
immunohistochemical staining.
[0339] Biotinylated VLVG alginate was then again administered
intraperitoneally to mice, and the presence of VLVG alginate in
various tissues was examined.
[0340] As shown in FIGS. 13A and 13B, intraperitoneally
administered VLVG alginate appeared in the liver and pancreas of
mice (FIG. 13B), but not in the spleen or colon of mice (FIG.
13A).
[0341] These results indicate that therapeutically effective
amounts of alginate can infiltrate organs such as liver and
pancreas, following systemic administration.
Example 4
Treatment by Co-Administration of Alginate and a Hepatotoxic
Drug
[0342] A subject afflicted by a medical condition (e.g., a
headache, an autoimmune disease, epilepsy) treatable by a
hepatotoxic drug (e.g., paracetamol for treating a headache,
methotrexate for treating an autoimmune disease, carbamazepine for
treating epilepsy) is co-administered the hepatotoxic drug in
combination with alginate. Co-administration is effected by
administering the alginate during a time period ranging from 100
minutes prior to administration of the hepatotoxic drug to 50
minutes after administration of the hepatotoxic drug. The
hepatotoxic drug is administered according to an accepted regimen
for treating the medical condition, and the alginate is
administered in a dose sufficient to prevent or reduce liver damage
caused by the administered dosage of hepatotoxic drug.
[0343] The alginate is administered separately from the hepatotoxic
drug (e.g., in the form of tablets, capsules or a syrup), or the
alginate is co-formulated with the hepatotoxic drug in a form
suitable for administration of the drug (e.g., in the form of
tablets, capsules or a syrup).
[0344] The subject is optionally monitored for signs of liver
damage according to a standard technique (e.g., by serum liver
enzyme assay), and the dose of alginate is optionally increased if
signs of liver damage appear.
[0345] Although the invention has been described in conjunction
with specific embodiments thereof, it is evident that many
alternatives, modifications and variations will be apparent to
those skilled in the art. Accordingly, it is intended to embrace
all such alternatives, modifications and variations that fall
within the spirit and broad scope of the appended claims.
[0346] All publications, patents and patent applications mentioned
in this specification are herein incorporated in their entirety by
reference into the specification, to the same extent as if each
individual publication, patent or patent application was
specifically and individually indicated to be incorporated herein
by reference. In addition, citation or identification of any
reference in this application shall not be construed as an
admission that such reference is available as prior art to the
present invention. To the extent that section headings are used,
they should not be construed as necessarily limiting.
* * * * *