Note: Descriptions are shown in the official language in which they were submitted.
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Title of the Invention
METHODS AND COMPOSITIONS FOR WOUND MANAGEMENT
The present invention is a continuation-in-part of non-provisional U.S.
Application Serial No. 09/955,657 filed September 18, 2001, and also claims
benefit
of provisional U.S. Application Serial No. 60/435,413 filed December 19, 2002,
both
of which are incorporated herein by reference in their entireties.
Field of the Invention
The present invention relates generally to methods and compositions for
managing wounds, including surface wounds or skin lesions of a human or animal
patient. The present invention further relates to methods useful in treating
wounds
infected by microorganisms. The present invention also relates to methods of
promoting the repair of wounds of mouth and the skin surfaces.
Background
The outermost layers of the skin form a physical barner that protects an
animal from microbial invasion and the establishment of opportunistic
infections.
Injuries to the skin, for instance an abrasion, incision, laceration, a burn
from thermal,
radiant or chemical exposure or a necrotic lesion of the surface tissue,
destroy the
integrity of the cornified layer, epidermis or dermis and will allow microbes
to
penetrate into the underlying tissues. An established infection can become
systemic
and ultimately life-threatening.
A breach in the skin combined with a compromised immune system means
that burn patients are highly susceptible to opportunistic infections. Of
about seventy
thousand burn victims per year serious enough to require hospitalization, as
many as
ten thousand will die, usually from a nosocomial infection. Many burn
survivors will
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suffer permanent disfigurement from the tissue damage that accompanies
microbial
infections. Greater treatment needs and prolonged care of burns patients in
the
hospital also mean that infections represent a significant financial drain.
Long-term
recuperative costs can also be significant.
Ulcers are exposed surface lesions of the skin or a mucoid layer such as the
lining of the mouth, where inflamed and necrotic tissue sloughs off. This
exposed
tissue is also highly susceptible to opportunistic microbial invasion. In this
instance,
the primary infected site is localized and best treated by a topical
application of an
antimicrobial agent, sometimes supplemented with systemic antibiotic
administration.
Although tissue damage is not usually as great as for a burn, infected ulcers
are
discomforting to the patient, disfiguring and also life-threatening if leading
to a
systemic infection.
The repair of damaged skin requires an ongoing process of physiologic
debridement, development of highly vascular granulation tissue that provides
wound
protection, wound closure through contraction and epithelial migration to
provide
long-term surface protection. The interval between wounding and skin repair
depends
on the location of the injury, the degree of damage, the apposition of the
wound
edges, the medical condition of the patient and the type and severity of wound
contamination with foreign debris and microbes.
Trauma and necrosis of the skin decreases vascularization to a wound and slows
the influx of immunologic proteins and white blood cells. The severity of the
damage
and number of invading microbes determine whether or not a - clinical
infection
occurs. While clinicians frequently focus on the type of microbes that may
contaminate a wound, some studies suggest that the number of invading microbes
is
more important than the species. Proliferating microbes cause additional and
accelerated tissue damage through both direct (toxins and cellular damage) and
indirect (edema and accumulation of pus) impairment of vascular supply. These
changes further impair access of immune system components to the wound as well
as
reducing the clearance of necrotic debris and preventing systemically
delivered
antibiotics from reaching contaminated tissues. Collagenase and proteases that
accumulate in association with degenerating inflammatory cells damage
connective
tissue proteins and further inhibit wound healing.
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The wound healing cascade is delayed until the inflammatory and physiologic
debridement phases have killed and removed contaminating microbes and necrotic
tissues. Experiments in rats demonstrated that infection decreased the
bursting
strength of a wound (increased likelihood of dehiscence) as well as increasing
angiogenesis and vessel thrombosis. While some data suggests infected wounds
that
heal by second intention are stronger than ones closed primarily, it is
generally
considered best to resolve and prevent infection.
Macrophages that enter a wound serve a phagocytic function as well as the
critical role of stimulating the maturation of fibroblast that are ultimately
responsible
for skin repair. Epithelial cell proliferation and migration are necessary for
re-
epithelization. This process is enhanced in a moist environment that is well
oxygenated by a viable blood supply.
Initiating treatment in a timely manner is most important to reduce or prevent
microbial colonization and additional pathogen-associated wound damage.
Contaminating bacteria begin to invade adjacent tissue within 6 hours and most
data
suggest that a wound should be considered infected when 105 to 10~ microbes
are
present per gram of tissue.
Surgical preparation of the skin surrounding the wound followed by copious
flushing under pressure of the wound with solutions that will remove dirt and
bacteria
while preserving cellular integrity is critical. The two most commonly used
wound
irrigants are Betadine and NOLVASANTM. Some studies suggest that Betadine
solution will reduce the number of bacteria in a wound while others suggest no
effect
on microbial colonization In vivo data suggests that significant numbers of
bacteria
can survive in all noncytotoxic concentrations of Betadine solution and
Nolvasan.
However, other studies have demonstrated that either of these antimicrobials
facilitated improved wound healing compared to saline-treated controls. In a
human
study, flooding a wound with penicillin significantly reduced the incidence of
wound
infection. Solutions containing ampicillin, neomycin, kanamycin and gentamicin
have also been used for flushing wounds. However" persistent use of
antimicrobials
or antiseptics can result in the proliferation of multi-drug resistant
bacteria within a
hospital, an animal facility or among the animal population cared for by a
veterinary
facility.
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There is still a need, however, not addressed in the prior art, for methods of
treating and infection susceptible wounds of humans and animals that also
promote
wound healing. Such methods should also offer relief from pain and provide
protection from infection. Safe and effective means of topically administering
such
compositions to a wound are also desirable. Suitable composition preferably
would
also have enhanced activity against drug resistant strains of infectious
microbes.
Summary of the Invention
The present invention addresses the need to manage a wound, such as a burn, a
lesion, an incision or a laceration of the skin, or a lesion of the oral
mucosa, of an
animal or human patient to promote wound repair and reduce the likelihood of
infection. Briefly described, the present invention provides methods and
compositions for use in the methods for wound management that comprise
contacting
a wound of a patient with an effective amount of a therapeutic composition
comprising a pharmaceutically acceptable chelating agent, a pharmaceutically
acceptable pH buffering agent, an antimicrobial agent, Vitamin E a surfactant
and a
pharmaceutically acceptable carrier. The antimicrobial agents) has increased
antimicrobial activity because of the synergy with the chelating agent and
maintenance of the treated area at a pH suitable for sustained antibiotic
activity. The
antimicrobial agent can, therefore, be used in effective doses that are less
than would
be required for the same level of antimicrobial activity in the absence of the
chelator.
The compositions of the present invention are, therefore, useful in
counteracting or
preventing an infection or will be more effective against infections caused by
drug-
resistant strains of microbes.
The Vitamin E promotes tissue repair, thereby reducing the likelihood of
opportunistic infection, improving wound healing and reducing pain sensation.
The
wound management methods of the present invention may further include
delivering
an effective amount of the therapeutic composition by contacting the wound
with a
medical dressing having an effective amount of the composition deposited
thereon.
The present invention further provides methods suitable for delivering the
therapeutic composition, comprising an antimicrobial agent, a chelating agent
and a
buffer and a surfactant to an oral wound.
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Additional objects, features, and advantages of the invention will become
more apparent upon review of the detailed description set forth below when
taken in
conjunction with the accompanying drawing figures, which are briefly described
as
follows.
Brief Descriution of the Figures
Figure 1. IsoboloGram, illustrating the combined effect of EDTA and neomycin
(in
50 mM Tris) on Staphylococcus aureus.
Figure 2. IsoboloGram, illustrating the combined effect of EDTA and neomycin
(in
50 mM Tris) on Pseudomonas aeruginosa.
Figure 3. IsoboloGram, illustrating the combined effect of EDTA and neomycin
(in
50 mM Tris) on Enterococcus faecalis.
Figure 4. Growth of Staphylococcus aureus on Mueller Hinton agar when treated
alone or with combinations of EDTA, water, and neomycin.
Figure 5. Growth of Pseudornonas aeruginosa on Mueller Hinton agar when
treated
alone or with combinations of EDTA, water, and neomycin.
Figure 6. Growth of Enterococcus faecalis on M Enterococcus agar when treated
alone or with combinations of EDTA, water, and neomycin.
Detailed Description of the Invention
A full and enabling disclosure of the present invention, including the best
mode known to the inventor of carrying out the invention is set forth more
particularly
in the remainder of the specification, including reference to the Examples.
This
description is made for the purpose of illustrating the general principles of
the
invention and should not be taken in the limiting sense.
The present invention addresses the need to manage a wound, such as a burn, a
lesion, an incision or a laceration of the skin, or a lesion of the oral
mucosa, of an
animal or human patient to promote wound repair and reduce the likelihood of
infection. The present invention provides methods and compositions for wound
management, the methods comprising contacting a wound of a patient with an
effective amount of a therapeutic composition comprising a pharmaceutically
acceptable chelating agent, a pharmaceutically acceptable pH buffering agent,
an
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antimicrobial agent, Vitamin E, a surfactant and a pharmaceutically acceptable
carrier. The antimicrobial agents) has increased antimicrobial activity
because of the
synergy with the chelating agent and maintenance of the treated area at a pH
suitable
for sustained antibiotic activity. The antimicrobial agent can, therefore, be
used in
effective doses that are less than would be required for the same level of
antimicrobial
activity in the absence of the chelator. The compositions of the present
invention are,
therefore, useful in counteracting or preventing an infection or will be more
effective
against infections caused by drug-resistant strains of microbes.
The Vitamin E promotes tissue repair, thereby reducing the likelihood of
opportunistic infection, improving wound healing and reducing pain sensation.
The
wound management methods of the present invention may further include
delivering
an effective amount of the therapeutic composition by contacting the wound
with a
medical dressing having an effective amount of the composition deposited
thereon.
The present invention further provides methods for delivery of a therapeutic
composition, comprising an antimicrobial agent, a chelating agent and a buffer
and a
surfactant for repairing a wound and inhibiting a microbial colonization of
the oral
mucosa, .
De araitions
The term "therapeutically effective" amount of a composition of the present
invention is an amount that results in wound repair and/or a reduction in pain
sensation at the site of a treated wound and the inhibition or prevention of
microbial
invasion of the treated wound. A skilled artisan or scientist using routine
protocols,
such as those disclosed in the Examples below or in the literature, may
readily
confirm the utility of the compositions described herein.
The term "wound" as used herein refers to a lesion or open wound that can
expose underlying epidermal, dermal, muscular or adipoidal tissue to the air.
Wounds
include, but are not limited to, a puncture wound, an incision, a laceration,
a
penetrating wound, a perforating wound, a tunnel wound and the like. Wounds
also
include open wounds that have been sutured or otherwise mechanically closed
but
have not healed or repaired the break in the skin or oral mucosal layer or of
the
surface layers of the eye including the conjunctiva and cornea..
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The terms "lesion" and "surface lesion" as used herein refer to a
circumscribed
area of pathologically altered tissue, an injury or wound. Primary lesions are
the
immediate result of the pathological condition and include, but are not
limited to, cuts,
abrasions, vesicles, blebs, bullae chancres, pustules, tubercles or any other
such
condition of the skin or a surface of the mouth, nose, anus or any other
orifice of the
body of a human or animal, or to the surface layers of the eye including the
conjunctiva and cornea., or secondary lesions that later develop from a
primary lesion
and includes, but is not limited to, fissures and ulcers and other wounds.
The term "wound management" refers to therapeutic methods that induce
and/or promote repair of a wound including, but not limited to, arresting
tissue
damage such as necrotization, promoting tissue growth and repair, reduction or
elimination of an established microbial infection of the wound and prevention
of new
or additional microbial infection or colonization. The term may further
include
reducing or eliminating the sensation of pain attributable to a wound.
The terms "wound healing" and "wound repair" refer to a process involving
tissue growth that partially or totally closes a wound, repairs a breach in
the dermis or
epidermis and partially or totally restores the barrier properties of the skin
or the
repair of the surface layers of the eye including the conjunctiva and cornea.
The term "microbial infection" as used herein refers to any pathological
presence of at least one bacterial species on or in an injury or lesion to the
skin of a
human or animal. It is further understood that a "microbial infection" may
include
any systemic infection that is amenable to inhibition by application of the
antimicrohial compositions of the present invention.
The term "burn" as used herein refers to tissue injury of the skin caused by
thermal, chemical, or radiation exposure or abrasive friction. A burn may be a
"first-
degree burn" with superficial damage to the outer cornified layer, a "second-
degree
burn" with damage extends down into the epidermal layer of cells but is not of
sufficient extent that regeneration of the skin is prevented, or a "third-
degree burn"
where the injury extends below the dermis to the underlying tissue and wherein
repair
of the skin is not possible without grafting.
The term "ulcer" as used herein refers to an open sore or lesion of the skin
or a
mucous membrane that involves the sloughing off of inflamed and necrotized
tissue
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and includes, but is not limited to, callous ulcers, chronic leg ulcers,
decubitus,
denture ulcers of the oral mucosa, traumatic ulcers of the mouth, infections
stomatitis
of the mouth and any type of secondary lesion that is a breach of the
cornified and the
epidermal layer of the skin or the mucosal surface of the mouth.
The term "antimicrobial agent" as used herein refers to the compounds and
combinations thereof, including bacteristatic or bactericidal compositions or
agents,
that may be administered to an animal or human and which inhibit the
proliferation of
a microbial infection.
The term "pharmaceutically acceptable" as used herein refers to a compound
or combination of compounds that will not impair the physiology of the
recipient
human or animal to the extent that the viability of the recipient is
compromised.
Preferably, the administered compound or combination of compounds will elicit,
at
most, a temporary detrimental effect on the health of the recipient human or
animal.
The term "chelating agent" as used herein refers to any organic or inorganic
compound that will bind to a metal ion having a valence greater than one.
The term "pH buffering agent" as used herein refers to any pharmaceutically
acceptable organic or inorganic compound or combination of compounds that will
maintain the pH of an antibiotic-containing solution within 0.5 pH units of a
selected
pH value.
The term "carrier" as used herein refers to any pharmaceutically acceptable
solvent of antibiotics, chelating agents and pH buffering agents that will
allow a
therapeutic composition to be administered directly to a wound of the skin or
to the
oral mucosa. The carrier will also allow a composition to be applied to a
medical
dressing for application to such a wound. A "carrier" as used herein,
therefore, refers
to such solvent as, but not limited to, water, saline, physiological saline,
ointments,
creams, oil-water emulsions, gels, or any other solvent or combination of
solvents and
compounds known to one of skill in the art that is pharmaceutically and
physiologically acceptable to the recipient human or animal. The term
"carrier" is
understood not to include surfactants such as detergents, non-ionic
surfactants such as
lecithin, and the like.
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The term "surfactant" as used herein refers to any detergent or other
pharmaceutically acceptable non-ionic compound that lowers surface tension of
an
aqueoussolution.
One aspect of the present invention provides methods for wound management
wherein a wound of a human or animal patient is contacted with an effective
amount a
therapeutic composition comprising a chelating agent, a buffer, an
antimicrobial
agent, Vitamin E, a surfactant and a carrier. More than one antimicrobial
agent may
be used to inhibit the proliferation of a single invasive organism, or a mixed
population of invasive organisms. The antimicrobial agents) should be selected
after
determining the composition and antibiotic resistance spectrum of the invading
microbial population.
Administration of the compositions of the present invention to a wound results
in accelerated wound repair with reduced or no sepsis, as described in
Examples S-14
below. Even with wounds that penetrated the dermal layer, there is reduced
pain
sensation, more extensive and quicker tissue growth and less overall
discomfort to the
patient. An additional benefit is that the tissue repair restrict
opportunistic infections
that would otherwise prolong the period of wound healing, increase the extent
of the
wound or even develop to threaten the life of the infected patient.
Before applying the therapeutic composition to the patient, the wound can be
debrided to clean the wound of necrotic or infected tissue. Debridation may be
mechanical by cutting or pulling away damaged tissue from the wound or, if
readily
inaccessible, other methods including, but not limited to, the application of
sterile
maggots may be used. Optionally, the wound may be prewashed before the
application of the therapeutic composition using a composition comprising a
chelating
agent having a concentration from about 1 mM to about 250 mM, a buffering
agent
having a concentration of about 10 mM to about 250 mM and a detergent having a
concentration from about 1 to about 30% v/v as given, for instance, in Example
15
below. In one embodiment, the composition comprises about 8 mM EDTA and 20
mM Tris with between about 2% and about 30% v/v of cocamidopropyl betaine
The therapeutic compositions used in the methods of wound management
herein described may be applied to a wound by any number of methods including
as a
lavage where the wound is washed or irrigated. In one embodiment, for example,
the
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compositions are absorbed onto the surface of the fibers of a wound dressing
before or
during the treatment, ensuring that while the wound is ventilated it is still
subject to
contact with the therapeutic compositions for a prolonged period.
In various embodiments of the methods of the present invention, the treated
wound is in an oral mucosal surface of the patient. In this instance, the
therapeutic
composition can be applied as a mouth wash or rinse or in combination with a
dressing that may be secured over the wound. In other embodiments, the
therapeutic
compositions of the invention are ophthalmic compositions suitable for
administering
to the surface of an eye for the repair or healing of a wound to the
conjunctiva or
corneal surface. The therapeutic compositions of the present invention may
also be
used as a bath for the total or partial immersion of a human or animal for the
treatment of multiple skin lesions such as for managing or burnt foot, or
hand, or large
wound, of a human or animal.
The pharmaceutically acceptable chelating agent of the therapeutic
compositions of the present methods may be selected from
ethylenediamenetetracetic
acid (EDTA), triethylene tetramine dihydrochloride (TRIEN), ethylene glycol-
bis
(beta-aminoethyl ether)-N, N, N', N'-tetracetic acid (EGTA), diethylenetriamin-
pentaacetic acid (DPTA), triethylenetetramine hexaacetic acid (TTG),
deferoxamine,
Dimercaprol, edetate calcium disodium, zinc citrate, penicilamine succimer and
Editronate or any other pharmaceutically acceptable chelating agent, salt or
combination thereof, known to one of ordinary skill in the art, and which will
chelate
divalent metal ions such as, but not only, Caz+, Mg2+, Mn2+, Fez+, and Zn2+.
The
chelating agent, when delivered to a wound of a human or animal patient will
have a
concentration between from about 1 mM to about 250 mM, more preferably from
about 1 mM to about 100 mM, most preferably from about 1 mM to about 50 mM. In
a preferred embodiment the chelating agent is EDTA having a concentration of
about
8 mM.
The therapeutic compositions of the present invention also include a
pharmaceutically acceptable pH buffering agent that preferably will maintain
the pH
of the antimicrobial composition, when delivered to the skin injury or skin
lesion, to
between about pH 7.0 and about pH 9Ø A pH buffering agent may be selected
from,
but is not limited to, Tris (hydroxymethyl) aminomethane (tromethaprim; TRIZMA
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base), or salts thereof, phosphates or any other buffering agent such as, for
example,
phosphate-buffered saline that is biologically acceptable. In a preferred
embodiment,
the pH of the antimicrobial composition in solution is about 8Ø The
buffering agent,
when delivered to a wound, has an effective dose of between about 5 mM and
about
250 mM, more preferably between about 5 mM and about 100 mM, most preferably
between about 10 mM and about 100 mM. In a preferred embodiment the buffer
agent has a concentration of about 20 mM.
The compositions of the present invention may also comprise at least one
antimicrobial agent. The infections that may be treated by the methods and
compositions of the present invention may be any opportunistic infection of a
wound
by a bacterium, or a multiple infection of more than one species of bacteria.
Microbial species that may cause infections inhibited by the methods of the
present
invention include fungi and bacterial species that may cause infections of a
burn,
lesion, oral mucosal lesion or other wound of a human or animal including, but
are
not limited to, Aerobacter aerongenes, Aeromonas spp., Bacillus spp.,
Bordetella spp,
Camp3~lobacter spp., Chlarnydia spp., Corynebacterium spp., Desulfovibrio
spp.,
Escherichia coli, enteropathogenic E. coli, Enterotoxin producing E coli,
Helicobacter pylori, Klebsiella pneumoniae, Legionella pneumophiia, Leptospira
spp., Mycobacterium tuberculosis, M. bovis, Neisseria gonorrhoeae, N.
rneningitidis,
Nocardia spp., Proteus mirabilis, P vulgaris, Pseudomonas aerugiraosa,
Rhodococcus
equi, Salmonella enteridis, S. typhimurium, S. typhosa, Shigella sonnei, S
dysenterae,
Staphylococcus aureus, Staph. epidermidis, Streptococcus anginosus, S. mutans,
Vibrio cholerae, Yersinia pesos, Y. pseudotuberculosis, Actinomycetes spp.,
and
Streptonzyces spp.
The action of the antimicrobial agent can be either bacteriostatic wherein the
antibiotic arrests the proliferation of, but does not necessarily kill, the
microorganism
or the activity of the antibiotic can be bacteriocidal and kill the organism
or a
combination of activities. Antibiotics suitable for use in the wound
management
methods of the present invention include, but are not limited to, ~3-lactams
(penicillins
and cephalosporins), vancomycins, bacitracins, macrolides (erythromycins),
lincosamides (clindomycin), chloramphenicols, tetracyclines, aminoglycosides
(gentamicins), amphotericns, cefazolins, clindamycins, mupirocins,
sulfonamides and
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trimethoprim, rifampicins, metronidazoles, quinolones, novobiocins, polymixins
and
Gramicidins and the like and any salts or variants thereof. It also understood
that it is
within the scope of the present invention that the tetracyclines include, but
are not
limited to, immunocycline, chlortetracycline, oxytetracycline, demeclocycline,
methacycline, doxycycline and minocycline and the like. It is also further
understood
that it is within the scope of the present invention that aminoglycoside
antibiotics
include, but are not limited to, gentamicin, amikacin and neomycin and the
like.
Techniques to identify the infecting microorganism and to determine the
concentration of the antibiotic that will inhibit or kill fifty percent
(MICSO) of the
organisms will be considered well known to one of ordinary skill in the art
and will
not require undue experimentation. The techniques to determine the antibiotic
sensitivity of a bacterial isolate, and the methods of determining the
synergistic effect
of adding a chelating agent to a solution of an antibiotic are described in
Manual of
Methods for General Microbiology, Eds: Gerhardt et al., American Society of
Microbiology, 1981, and incorporated herein in its entirety by reference.
Before the application to a wound of a composition that includes an
antibiotic,
it is be useful to identify the species and the antibiotic sensitivity
spectrum of the
invasive microbe(s). Routine tests well known to one of ordinary skill in the
art,
including determining the MIC and FIC of antibiotics in the absence and/or
presence
of a chelating agent may be used and the amount of the antimicrobial
composition
may be adjusted accordingly so as to inhibit growth of the microorganism. The
concentrations and amounts of the antimicrobial agent and chelating agent may
be
adjusted to levels that are physiologically accepted by the exposed tissue of
the injury
or lesion and effective against the microbial infection of the skin injury or
skin lesion.
In one embodiment of the present invention, the concentration of the
antibiotic is in
the range of about 0.04 mg/ml to about 25 mg/ml and the concentration of the
chelating agent in the carrier is in the range of about 0.1 mM to about 100.0
mM.
In various embodiments, the antibiotic is a penicillin, an aminoglycoside, a
vancomycin, a chloramphenicol, an erythromycin, a tetracycline, gentamicin,
nalidixic acids, or a streptomycin. In another embodiment the antibiotic is
tetracycline. In a preferred embodiment of the present invention, the
antibiotic is
neomycin. In another embodiment of the present invention, the antibiotic is
amikacin.
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In yet another embodiment, the antibiotic is gentamicin. However, a
combination of
antibiotics may be used depending upon the antibiotic resistance profiles of
the
microbial population of the wound.
The therapeutic compositions for use in the methods of wound management
also comprise a surfactant that can useful in cleaning a wound or contributing
to
bactericidal activity of the administered compositions. Suitable surfactants
include,
but are not limited to, phospholipids such as lecithin, including soy lecithin
and
detergents. Preferably, the surfactant selected for application to a wound or
skin
surface is mild and not lead to extensive irritation or promote further tissue
damage to
the patient.
Suitable nonionic surfactants which can be used are, for example: fatty
alcohol
ethoxylates (alkylpolyethylene glycols); alkylphenol polyethylene glycols;
alkyl
mercaptan polyethylene glycols; fatty amine ethoxylates
(alkylaminopolyethylene
glycols); fatty acid ethoxylates (acylpolyethylene glycols); polypropylene
glycol
ethoxylates (Pluronic); fatty acid alkylolamides (fatty acid amide
polyethylene
glycols); alkyl polyglycosides, N-alkyl-, N-alkoxypolyhydroxy fatty acid
amide, in
particular N-methyl-fatty acid glucamide, sucrose esters; sorbitol esters,and
esters of
sorbitol polyglycol ethers. A preferred surfactant is polypropylene glycol
ethoxylates
with a preferred concentration of between about 5% wt % and about 25% wt %. A
most preferred surfactant is Pluronic F-127 (Poloxamer 407). In other
embodiments
of the composition, the surfactant comprises lecithin with or without the
addition of
Pluronic F-127, the Pluronic F-127 being between about 2 and about 20 wt % for
increasing the viscosity or gelling of the compositions.
The therapeutic compositions further include Vitamin E that will promote
tissue growth and repair and reduce the pain experienced at the site of the
skin injury.
By promoting tissue repair, not only is discomfort to the patient reduced, but
there
may be less scar tissue formation and hence less permanent disfiguring of the
patient.
Furthermore, faster healing of a skin injury or lesion is useful for reducing
the
likelihood of a nosocomial infection and the problems associated therewith.
The
Vitamin E, when delivered to a wound of a patient by the methods of the
present
invention, has a concentration of between about 20 IU/ml and 500 IU/ml,
preferably
between about 50 IU/ml and 500 IU/ml, more preferably between about 100 IU/ml
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and about 500 IU/ml. In one embodiment of the therapeutic composition, the
Vitamin
E has a concentration of about 100 IU/ml. In another embodiment, the
concentration
is between about 325 IU/ml and about 360 ILJ/ml..
The therapeutic compositions for use in the methods of the invention
preferably include a pharmaceutically acceptable Garner that provides the
medium in
which are dissolved or suspended the constituents of the compositions.
Suitable
carriers include any aqueous medium, oil, emulsion, ointment and the like that
will
allow the therapeutic compositions to be delivered to the target wound without
increasing damage to the tissues of the wound.
It is also contemplated that the therapeutic compositions of the invention can
be prepared as precursor solutions, or as sterile powders or concentrates that
are
useful for the extemporaneous preparation of the administered compositions.
Optionally, the compositions may further include a preservative to extend the
shelf
life of the composition. A particularly useful preservative is scorbic acid,
preferably
as the sodium or potassium salt. A preferred amount of the preservative is
between
about 0.1 wt % to about 5 wt %. A more preferred amount is about 0.2 wt %.
Medical dressings suitable for use in the methods of the present invention for
contacting a wound with the therapeutic compositions can be any material that
is
biologically acceptable and suitable for placing over any wound such as a
burn, or a
surface lesion of the skin or the oral mucosa or teeth of the mouth. In
exemplary
embodiments, the support may be a woven or non-woven fabric of synthetic or
non-
synthetic fibers, or any combination thereof. The dressing may also comprise a
support, such as a polymer foam, a natural or man-made sponge, a gel or a
membrane
that may absorb or have disposed thereon, a therapeutic composition. A gel
suitable
for use as a support for the antimicrobial composition of the present
invention is
KYTM (sodium carboxymethylcellulose 7H 4F (Hercules, Inc., Wilmington, DE))..
A film, a natural or synthetic polymer, or a rigid or malleable material that
is
known to one of ordinary skill in the art as being acceptable for insertion in
the mouth
of a human or animal, and which will place an antimicrobial composition
according to
the present invention in contact with a tooth or a lesion of the oral mucosa.
In one
such embodiment of the medical dressing of the present invention, the support
is a
gauze. The gauze may be absorbent and can be wetted with an antimicrobial
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composition of the present invention before applying the gauze to an infected
wound
or other site.
The present invention also contemplates that the gauze may be impregnated
with the therapeutic composition and then dried. This allows the impregnated
dressing to be stored for later use, or to avoid excessively dampening an
injured area.
In yet another embodiment of the present invention, a therapeutic composition
is
absorbed on the surface of the support material of the medical dressing. The
composition may be applied to the surface by wetting the surface with a
solution of
the composition and drying the support to deposit the composition thereon. A
concentration of the composition that is effective for promoting wound repair
and/or
against the proliferation of a microorganism may be attained when the dressing
is
wetted by the patient's body.
Another aspect of the present invention is therapeutic compositions
comprising about 2 to about 50 wt % of a pharmaceutically acceptable chelating
agent, about 2 to about 50 wt % of a pharmaceutically acceptable buffering
agent, and
a surfactant. In one embodiment, the surfactant is a detergent such as, but
not limited
to, a mild detergent that will avoid irritation to the surface of the skin or
other tissue to
which the therapeutic composition is applied. For instance, one suitable
detergent is
cocamidopropyl betaine, but it is contemplated that any detergent known to
those of
ordinary skill and which will clean a wound or skin surface without triggering
an
inflammatory reaction or otherwise further extends the extent of injury of the
recipient
patient can be used. In one embodiment, the detergent is about 3 to about 33
wt % of
a therapeutic composition. In another embodiment the surfactant is lecithin.
In various embodiments of the invention, the composition further comprises
from about 1 to about 25 wt % of an antimicrobial agent. The embodiments may
also
include from about 2 to about 50 wt % of Vitamin E and/or from about 2 to
about 98
wt % of a pharmaceutically acceptable carrier. Optionally, the therapeutic
compositions include a preservative that will increase the shelf life of the
compositions. A typical preservative is scorbic acid, or the salts thereof.
In the compositions of the present invention, the chelating agent is selected
from the group consisting of ethylenediamenetetracetic acid (EDTA),
triethylene
tetramine dihydrochloride (TRIEN), ethylene glycol-bis (beta-aminoethyl ether)-
N,
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N, N', N'-tetracetic acid (EGTA), diethylenetriamin-pentaacetic acid (DPTA),
triethylenetetramine hexaacetic acid (TTG), deferoxamine, Dimercaprol, edetate
calcium disodium, zinc citrate, penicilamine succimer and Editronate.
Preferably, the
chelating agent is ethylenediamenetetracetic acid (EDTA).
The various embodiments of the compositions can further comprise 1 to 20 wt
of an anti-inflammatory agent such as, but not limited to dexamethasone.
The antimicrobial agents) that may be included in the various embodiments
of the compositions include, but are not limited to, a (3-lactam, an
aminoglycoside, a
vancomycin, a bacitracin, a macrolide, an erythromycin, a lincosamide, a
chloramphenicol, a tetracycline, a gentamicin, an amphotericin, a cefazolin, a
clindamycin, a mupirocin, a nalidixic acid, a sulfonamide and trimethoprim, a
streptomycin, a rifampicin, a metronidazole, a quinolone, a novobiocin, a
polymixin
and a gramicidin. More preferably, the antibiotic s selected from the group
consisting
of a ~-lactam, an aminoglycoside, a vancomycin, a chloramphenicol, an
erythromycin, a tetracycline, gentamicin, nalidixic acid and a streptomycin.
In one
embodiment, the antimicrobial agent is oxytetracycline. In another embodiment,
the
antimicrobial agent is amikacin. In yet another embodiment, the antimicrobial
agent
~s neomycin.
Compositions of the present invention may also include a carrier, as described
above, for dissolving or suspending the components of the therapeutic
composition.
In various embodiments of the therapeutic compositions, the pharmaceutically
acceptable pH buffering agent can be Tris (hydroxymethyl) aminomethane (TRIZMA
Base) which, when dissolved in a carrier will have a .concentration of between
about
5 mM and about 250 mM, preferably between about 5 mM and about 100mM, more
preferably between about 10 mM and about 100 mM. In a most preferred
embodiment, theconcentration of the buffering agent is about 20 mM.
In various embodiments of the therapeutic compositions, the Vitamin E, when
dissolved in a carrier has a concentration of between about 20 IU/ml and 500
IU/ml,
preferably between about 50 IU/ml and 500 ILT/ml, more preferably yet between
about
100 IU/ml and about 500 IU/ml. In one embodiment the concentration of the
Vitamin
E is between about 325 IU/ml and about 360 IU/ml. Optionally, the Vitamin E
may
be prepared by dissolving in hydrous lanolin as a carrier in a volume ratio
from about
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10:1 to about 1:1, preferably having a volume ratio of about 35:15.
Preferably, in all
of embodiments, the pharmaceutically acceptable carrier is non-allergenic.
Another aspect of the invention is kits that comprise therapeutic compositions
as described above, or the components to prepare the compositions, and
packaging
that includes instructions on how to prepare and use the compositions to
manage a
wound and promote healing thereof. One embodiment of the invention, therefore,
comprises a vessel containing a pharmaceutically acceptable chelating agent, a
pharmaceutically acceptable buffering agent suitable for maintaining the pH of
the
site of a treated wound, a pharmaceutically acceptable antimicrobial agent,
Vitamin E,
a pharmaceutically acceptable carrier, a surfactant and packaging material.
The
packaging material comprises instructions directing the use of the kit for
preparing the
therapeutic composition of the present invention and delivering the
composition to a
wound or to the mouth of a human or animal to accelerate healing of a wound.
The
kit may further comprise separate vessels containing Vitamin E and/or
surfactant,
instructions for adding the Vitamin E to the therapeutic composition and for
administering the therapeutic compositions to the animal or human patient.
Even though the invention has been described with a certain degree of
particularity, it is evident that many alternatives, modifications, and
variations will be
apparent to those skilled in the art in light of the present disclosure.
Accordingly, it is
intended that all such alternatives, modifications, and variations that fall
within the
spirit and the scope of the invention be embraced by the defined claims.
The following examples are presented to describe preferred embodiments and
utilities of the present invention, but should not be construed as limiting
thereof.
Examine 1: Determination of syner~istic actions and fractional inhibitory
concentration (FIC) index
The antibacterial action of combinations of EDTA-Tris and neomycin was
measured by a two-dimensional microtiter checkerboard technique described in
Gilman et al., The Pharmacological Basis of Therapeutics, eds Goodman and
Gilman, 1085-1086 (Macvnillan Publishing Co., New York, 1985),. Sabath, L. D,
Arztimicrob. Agents and Chem. 210-217. (1967) and Sparks et al., Vet. Res.
Comm..
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18, 241-249 (1994), incorporated herein by reference in their entireties.
Each well of a round-bottomed 96-well microtiter plate was inoculated with
0.05 ml of 2-fold dilutions of neomycin and EDTA in 50 mM Tris. Then 0.05 ml
of
an 18-hour old culture of a test organism, containing 106 colony-forming units
(CFU)/ml, was added to each well. Controls for the culture and media were
included
in each plate. Plates were covered and incubated at 37° Celsius for 18-
24 hours.
Results were plotted as isobolograms for the determination of antagonistic,
neutral or additive, or synergistic effects. To generate isobolograms, FICs of
the two
test solutions were plotted individually on the x-axis and y-axis to determine
the
effect of combining the two test solutions on bacterial growth. A line that
curves
away from the zero point and the coordinates indicates antagonism. A straight
line
indicates neutral or additive effects. Lines that curves toward the zero point
and the
coordinates are indicative of synergism if there is at least a 4-fold decrease
in the MIC
of each compound, when used in combination, as compared with the MIC of each
test
compound alone.
A numerical score or fractional inhibitory concentration (FIC) index was
determined. The FIC index is equal to the sum of the values of FIC for the
individual
drugs:
MIC of Drug A with Drug B MIC of Drug B with Drug A
FIC = +
MIC of Drug A MIC of Drug B
An FIC index greater than 1.0 indicates an antagonistic interaction, an FIC
index of 1.0 indicates addition, and an FIC index of less than or equal to 0.5
indicates
synergism between the two test agents.
Example 2: Inhibition of the growth of microorganisms infecting burns
The organisms of this study were isolated from human burn patients. They
included strains of methicillin resistant Staphylococcus aureus, and
vancomycin
resistant strains of Pseudomonas aeruginosa and Enterococcus faecalis. The
bacterial
isolates were propagated in or on Brain Heart Infusion broth (BHI), Mueller-
Hinton
Broth (MHB), blood agar (BA), Mueller-Hinton agar (MHA), enterococcus agar
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(EA), or 2X nutrient agar (2xNA).
The EDTA-Tris treatment solutions were prepared from a stock solution
containing 0.5 mols/1 sodium EDTA and 1.0 mols/1 Tris-HC1, pH 8Ø The
treatment
solutions contained SmM sodium EDTA and 50 mM Tris-HC 1 with or without of
neomycin sulfate 1 mg / ml.
Antibiotic resistance profiles were determined by the disc diffusion method on
MHA (5). Antibiotics tested included ampicillin (AM-10), chloramphenicol (C-
30),
ciprofloxacin (CIP-5), kanamycin (K-30), gentamicin (GM-10), nalidixic acid
(NA-
30), neomycin (N-30), streptomycin (S-10), sulfisoxazole (G-25), tetracycline
(Te-
30), and vancomycin (Va-30).
Minimal Inhibitory Concentrations (MICs) and Minimal Bactericidal
Concentrations (MBCs) for EDTA-Tris and neomycin were determined by the broth-
dilution microtiter method in MHB or BHI according to the method of Blair et
al.,
Manual of Clinical Microbiology. p.307 (pub: Am. Soc. Microbiol. Williams and
Wilkins, Baltimore 1970), incorporated herein by reference in its entirety.
Example 3: In vitro effect of EDTA-Tris and neomycin on Enterococcus faecalis,
Pseudomouas aeru.~inosa, and Staphylococcus aureus
2xNA plates were swabbed with 200 ml of an overnight culture containing
about 10' colony-forming-units of a test organism. The plates were sampled
with
multipoint contactors as described in Wooley et al., Am. J. Vet. Res. 35, 807-
810
( 1974). Each multipoint contactor consisted of an array of 27 mm sewing
needles
mounted to an aluminum plate measuring 1 mm x 50 mm x 50 mm. The needles were
set 3.5 mm apart. The multipoint contactors were sterilized by autoclaving. To
collect
samples, a multipoint contactor was touched to an overnight bacterial culture
grown
on 2xNA as described above. Replicate plates were then inoculated by lightly
pressing the needles bearing the test bacteria onto either MHA plates, BA
plates or
EA plates for Ps. aeruginosa, Staph. aureus and Ent. faecalis respectively.
The agar
plates were incubated at 37°C and colonies were counted at 24 hours and
48 hours.
Each strain of microorganism was tested on a control agar plate (plate 1), and
on plates wherein the inoculated bacteria were covered with a sterile surgical
gauze
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saturated with 7 ml of: 5 mM EDTA-Tris (plate 2); 5 mM EDTA-Tris and 1 mg/ml
neomycin (plate 3); 1 mg/ml neomycin (plate 4); sterile water (plate 5).
Samples
were taken at 0 mins, and at 30 mins, 1 hour, 2 hours, 4 hours, 6 hours, 8
hours, and
24 hours of incubation.
Example 4: The antibiotic resistance profiles, MIC and MBC values for test
strains of Staph. aureus, Ps. aeru.~inosa, and Ent. faecalis
The antibiotic resistance profiles and MIC values for test strains of Staph.
aureus, Ps. aeruginosa, and Ent. faecalis are shown on Table 1.
Table 1. Antibiotic resistance profiles of Staphylococcus aureus, Pseudomonas
aeruginosa, and Enterococcus faecalis.
Antimicrobial
Agents"
Am C Cip Gm K NA N S G Te Va
Staphylococcus R'~ I R S R R R S S S S
aureus
Pseudomonas aeruginosaR R I I R R R R R R R
Enterococcus faecalisS R R R R R R R R R R
" Am= ampicillin; C= chloramphenicol; Cip= ciprofloxacin; K= kanamycin;
Gm= gentamicin; NA= nalidixic acid; N= neomycin; S= streptomycin; G=
sulfisoxazole; Te= tetracycline; Va= vancomycin; B R= resistant; I=
intermediate; S=
sensitive.
Fractional inhibitory concentrations (FICs) and isoboloGrams for the EDTA-
Tris-neomycin combination to determine a synergistic, additive, or
antagonistic
reaction, as described in Example I, were determined for Staph. aureus, Ps.
aeruginosa, and Ent. faecalis. MIC and MBC values for concentrations of
neomycin,
ampicillin, chloramphenicol, amikacin and oxytetracycline and EDTA
administered
individually, and the FIC values for Staph. aureus, Ps. aeruginosa, and Ent.
faecalis
are shown in Table 2 (Columns 2 and 3). MIC values for mixtures of the above
antibiotics and EDTA in the presence of each other are shown in Table 2
(Columns 4
and 5 respectively).
Table 2. Minimal Inhibitory Concentration (MIC) data concerning the amounts
(mM)
of EDTA in 50 mM Tris and antibiotics (mg/ml) when reacting alone and in
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combination against Pseudomonas aeruginosa, Staphylococcus aureus, and
Enterococcus faecalis.
MIC
Individually
Co-administered
Administered
Neomycin EDTA Neomycin FIC
+ EDTA
Ps. aeruginosa1.0 1.25 0.063 0.156 0.19
Staph. aureus3.13 1.0 1.56 0.25 0.75
Erat. faecalis3.13 15.63 1.17 1.88 0.5
Ampicillin EDTA Ampicillin FIC
+ EDTA
Ps. aeruginosa0.49 1.25 0.123 0.156 0.38
Staph. aureus0.24 1.0 0.0075 0.25 0.28
Ent. faecalis0.001 15.63 0.00025 7.82 0.75
ChloramphenicolEDTA Chloramphenicol FIC
+ EDTA
Ps. aeruginosa12.5 1.25 1.56 0.313 0.37
Staph. aureus0.39 1.0 0.39 1.0 2.0
Ent. faecalis0.4 15.63 0.2 3.9 0.75
Amikacin EDTA Amikacin FIC
+ EDTA
Ps. aeruginosa0.001 1.25 0.001 1.25 2.0
Staph. aureus0.12 1.0 0.03 0.5 0.75
Ent. faecalis2.0 15.63 1.0 7.8 1.0
OxytetracyclineEDTA Oxytetracycline FIC
+ EDTA
Ps. aeruginosa0.003 1.25 0.00075 0.313 0.5
Staph. aureus0.0001 1.0 0.00005 0.5 1.0
Ent. faecalis0.05 15.63 0.025 3.91 0.75
* Synergistic reaction (FIC = # 0.5)
Additive reaction (FIC = > .05 to # 1.0)
Antagonistic reaction (FIC = > 1.0)
The MBC values for EDTA and neomycin were decreased by at least 75% for
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bacterial killing (MBC) in those situations in which synergistic potentation
occurred
(Ps. aeruginosa and Ent. faecalis) as shown in Table 3. A decrease of about
50% was
observed with Staph. aureus.
Table 3. Minimal Bactericidal Concentrations (MBC), of Staphylococcus aureus,
Pseudomonas aeruginosa, and Enterococcus faecalis reacted with EDTA (mM) and
neomycin (mg/ml) in 50 mM Tris.
Bacterial Species IndividuallyCo-administered
Administered
Staph. aureus EDTA (mM) 7.81 3.9
Neom cin (mg/ml)3.13 1.56
Ps. aeruginosaEDTA (mM) 250 20.0
Neomycin (mg/ml)5.0 0.04
En. faecalis EDTA (mM) 250 62.5
~ ~
Neomycin (mg/ml)25 0 6 25
Specifically in the case of Staph. aureus, the MBC values for EDTA and
neomycin when combined were decreased by 50% as compared to the bactericidal
effect of each when individually administered.
With Ps. aeruginosa, the MBC values for EDTA and neomycin when in
combination were decreased 99.2% compared to when EDTA or neomycin were
individually administered. In the case of Ertt. faecalis, MBC values of EDTA
and
neomycin were both reduced 75% compared to when EDTA and neomycin were
administered individually.
Synergistic effects were observed when various concentrations of EDTA-Tris
and neomycin were reacted with Ps. aeruginosa and Ent. faecalis, while an
additive
effect was observed with Staph. aureus as shown in Figs. 1 - 3.
Example 5: Inhibition of growth of Ps aeru~inosa and Staph aureus
In the in vitro gauze-point-contactor study, the potentation effect was seen
for
EDTA-Tris-neomycin reactions with Ps. aeruginosa and Staph. aureus. These
reactions are illustrated in Figs. 4 and 5. When the same combinations of EDTA-
Tris
and neomycin were reacted with Ent. faecalis, no antibacterial activity was
noted at
these concentrations as shown in Fig. 6.
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Example 6: Treatment of a skin burn of a dog and antimicrobial
protection of draft donor sites by Vitamin E with EDTA-Tris and antibiotic
A mixed-breed, 35 lb spayed female canine, 1-2 years old, had been doused
with gasoline, set on fire and burned over 30% of body. The dog was given
initial
emergency treatment for 5 days and the burned area cultured for microbial
infection,
identifying: (3-hemolytic E coli, Klebsiella oxytoca, Proteus,sp., and
Enterococcus sp.
The dog was administered cefazolin systemically and the burned area cleared of
tissue
debris and wetted with a solution of EDTA-Tris and neomycin daily. The burn
area
was tree of the four bacteria after 3 days of systemic and topical EDTA-Tris-
neomycin therapy. After approximately 10 days, neomycin was replaced with
amikacin. The dog received an autologous skin graft approximately three weeks
after
the burn incident and the donor site treated with EDTA-Tris-amikacin and 100
IU of
Vitamin E. The dog was discharged from veterinarian hospital care two weeks
later.
Example 7: Treatment of microbially infected skin and oral lesions
A composition comprising EDTA, Tris and neomycin in KYTM gel Garner was
applied to skin ulcers of a turtle, a snake and a frog. Infection was reduced
until
eliminated, and the treated animals fully healed of their injuries and
infections.
A 13 year old domestic short hair cat had developed proliferative gingivitis.
The mouth was swabbed with a cotton-tipped swab twice daily for a week with a
solution containing 5 mM EDTA, 50 mM Tris, and 2 mg/ml neomycin. After the
first
week, the mouth and gums were swabbed twice weekly for a further month.
Following clearance of the infection from the animal's mouth, there was no
recurrence for at least one year. A similar human oral lesion also responded
to this
treatment. Likewise, mouthwashes also containing EDTA, Tris and neomycin, as
above, were used to treat and heal infections stomatitis of the oral cavities
of iguanas
and snakes.
Example 8: Vitamin E does not reduce the antibacterial efficacy of EDTA Tris
and antibiotics
The addition of Vitamin E to solutions of EDTA-Tris enhanced the
antibacterial effect of the solution. Solutions of Vitamin E alone decreased
the
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numbers of Ps. ueruginosu significantly, as shown in Table 4.
Table 4: Effect of Vitamin E on the Efficacy of EDTA-Tris and Antibiotics when
Tested against Pseudomonas aeruginosa.
Groups Log ~o Colony-Forming-Units/ml
PBS 8.20
EDTA-Tris 3.30
EDTA-Tris + Neomycin No growth
EDTA-Tris + Amikacin No growth
Vitamin E 6.04
Vit E + EDTA-Tris No growth
Vit E + EDTA-Tris + NeomycinNo growth
Vit E + EDTA-Tris + AmikacinNo growth
Vit E + Neomycin 4.40
Vit E. + Amikacin No growth
Example 9:. Self inflicted full thickness flap wound of labial mucosa
The patient had a self inflicted full thickness flap wound of labial mucosa.
Based on previous similar wounds untreated, it was anticipated that severe
swelling
and pain would persist for 7 to 10 days. The wound was rinsed with tap water
and
immediately contacted with. 5 mM EDTA, 50 mM Tris and 1.0 mg/ml Neomycin in
an emulsion containing 100 IU Vitamin E/ml The emulsion was reapplied 3 to 4
times per day. There was minimal pain or swelling associated with the wound.
Example 10: Third degree gasoline burn in dog
A dog suffering fro third degree burns received an application of 5 mM
EDTA, SOmM Tris and 1.0 mg/ml Neomycin in an emulsion containing 100 IU
Vitamin E/ml applied either to bandages or as a spray 3 to 4 times a day.
Continuous
observation of the animal indicated that the applied composition reduced pain,
as
evidenced by the animal's level of relaxation, which typically would otherwise
show
severe stress due to degree of injury and pain associated.
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Example 11: Animal inflicted bite wound
An animal inflicted bite wound of the human knuckle resulting in a 10 mm x 7
mm full thickness flap wound of the skin penetrating to the subdennal tissues.
It was
expected that there would be severe pain and debilitation accompanied by
limited use
of the affected finger for at least a week. The wound was washed immediately
with
tap water and coated with emulsion containing 35 mls of hydrous lanolin, 15
mls of
333 IU/ml Vitamin E, and 0.73 gms of EDTA, 0.6 gms of Tris dissolved in 2 mls
of
distilled water and 0.1 mg/ml of ampicillin. The wound was washed and recoated
with emulsion 3 to 4 times per day. A band-aid was used to protect the wound
from
additional trauma. Minimal to no pain and accelerated healing was observed
compared to similar but untreated wounds.
Example 12: Manaeement of a burn wound
A first to second degree burn wound on the inner surface of the lower ann of
approximately 1 week duration was treated. While the burn was healing, it
remained
crusty and pruritic. Application of 35 mls of hydrous lanolin, 15 mls of 333
IU/ml
Vitamin E, and 0.73 gms of EDTA, 0.6 gms of Tris dissolved in 2 mls of
distilled
water and 0.1 mg/ml of ampicillin resulted in cessation of pruritis within 15
minutes.
Repeat application when the wound begin to itch resulted in similar cessation
of
itching
Example 13: Incision wound
A composition containing 35 mls of hydrous lanolin, 15 mls of 333 IU/ml
Vitamin E, and 0.73 gms of EDTA, 0.6 gms of Tris dissolved in 2 mls of
distilled
water and 0.1 mg/ml of ampicillin was applied once to a small painful cut on a
finger.
After a single application, the pain was gone along with redness. The lesion
healed
quickly thereafter. The medication was applied to a 1 day old very painful toe
lesion
that typically are sore and red for several days. After 1 application, the
pain was gone
and the lesion rapidly healed.
Example 14: Open abcess wound of cat
A cat suffering from an abscess, severe necrotizing dermatitis, fascitis and
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superficial myositis faced either amputation or euthanasia The wound was
debrided,
flushed with sterile saline and 3 gms EDTA, 2.4 gms Tris and 100 mg of
ampicillin
dissolved in a liter of distilled water. Initially, the wound was dressed with
a wet
bandage soaked in above solution. 'The wound was coated with an emulsion of 35
mls
of hydrous lanolin, 1 S mls of 333 IU/ml Vitamin E, and 0.73 gms of EDTA, 0.6
gms
of Tris dissolved in 2 mls of distilled water and 0.1 mg/ml of ampicillin once
per day
(if a bandage was applied) or 3 to 4 times a day (no bandage was applied). The
cat
showed no discomfort even when the wound was left open, and a sufficient
granulation bed was formed to allow surgical closure in two stages.
Example 15: Formulations of Wound Prewash solutions
Percent C'.ncamitlnnrnnvl hPtaina
2.5 5 10 v 15 20 25
0.5 M EDTA 16 1 16 ul 16 ul 16 ul 16 ul 16 ul
1.0 M Tris 20 pl 20 pl 20 pl 20 p.l 20 pl 20 pl
Deionized 939 914 864 814 764 714 pl
water ~l pl pl ~1 pl
Cocamidopropyl25 ~1 50 pl 100 150 200 250 pl
betaine ~1 pl ~1
Total volume = 1.0 ml of prewash.
Final concentrations of EDTA and Tris are 8 mM EDTA and 20 mM Tris.
Example 16: Stock formulations having increased Vitamin E compared to
formulations with hydrous lanolin
(~z) Oleaginous portion:
Vitamin E, mixed tocopherols (333 - 352 ICT/g) SO gms
Lecithin, Soya 50 gms
Sorbic acid 0.2 gm
Lecithin and sorbic acid were dispersed in Vitamin E oil; lecithin was
dissolved over a 12 - 24 hour period to form an oily solution to which could
be added
a lipid soluble antibiotic or other agent (s).
(b) Agueous portion:
Pluronic F-127 (Poloxamer 407) 30 gms
Potassium sorbate 0.2 gm
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Purified water, cold 100 ml
The Pluronic F-127 and potassium sorbate were place in a volumetric flask;
add sufficient purified water to make 100 ml of solution; immediately
refrigerate to
keep pourable, since at room temperature a gel is formed. To the aqueous part
(b),
water soluble antibiotic, or other agent (s) may be added to produce the
desired effect.
Tris-EDTA may be added to the water at a concentration such that the final
formulation administered to the patient was about 8 mM EDTA and 20 mM Tris.
Depending on the desired consistency of the fomulation (emulsion), various
ratios of oleaginous and aqueous parts were prepared. For example, mixing 5 ml
of
each produced a soft creamy emulsion, with a low coefficient of spreading
containing
176 IU/ml of Vitamin E, and 0.2% sorbic acid/potassium sorbate as a
mold/yeast/fungus inhibitor (potassium sorbate is the potassium salt of sorbic
acid;
sorbic acid is used in oleaginous solutions, the salt in aqueous solutions).
Mixing 8 ml of the oleaginous portion (a) with 2 ml of the aqueous portion (b)
also produced an emulsion with a greater amount of Vitamin E (282 IU/ml). A
lower
concentration of Vitamin E could also be prepared by using the ratio of 8 ml
aqueous
part to 2 ml oleaginous part. The effective range of Vitamin E that may be
delivered
by this system is approximately 70 - 282 IU/ml.
Lecithin as the surfactant in the above discussion was 50%; the effective
concentration range for lecithin in the above formulations was 20-50 wt %; it
could
also be formulated to the lower to a range of about 2-50 wt %.
27
ATLANTA 382820v1
