U.S. patent application number 14/580877 was filed with the patent office on 2015-04-23 for compositions and methods for treating vaginal infections and pathogenic vaginal biofilms.
This patent application is currently assigned to Toltec Pharmaceuticals, LLC. The applicant listed for this patent is Toltec Pharmaceuticals, LLC. Invention is credited to Dawn Flynn, Suzanne Gordon.
Application Number | 20150110898 14/580877 |
Document ID | / |
Family ID | 40678965 |
Filed Date | 2015-04-23 |
United States Patent
Application |
20150110898 |
Kind Code |
A1 |
Gordon; Suzanne ; et
al. |
April 23, 2015 |
Compositions and Methods for Treating Vaginal Infections and
Pathogenic Vaginal Biofilms
Abstract
The present invention generally relates to methods and
compositions for treating pathogenic vaginal biofilms. More
specifically, the invention relates to pharmaceutical compositions
comprising a combination of boric acid and diethylaminetetracetic
acid (EDTA) and to methods of using such compositions to treat or
prevent biofilm formation associated with vaginal infections, such
as bacterial vaginosis (BV), vulvovaginal candidiasis (VVC),
trichomoniasis or mixed infections.
Inventors: |
Gordon; Suzanne; (Chicago,
IL) ; Flynn; Dawn; (Elmhurst, IL) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Toltec Pharmaceuticals, LLC |
Chicago |
IL |
US |
|
|
Assignee: |
Toltec Pharmaceuticals, LLC
|
Family ID: |
40678965 |
Appl. No.: |
14/580877 |
Filed: |
December 23, 2014 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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13709740 |
Dec 10, 2012 |
8956663 |
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14580877 |
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12323585 |
Nov 26, 2008 |
8349368 |
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13709740 |
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60991308 |
Nov 30, 2007 |
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Current U.S.
Class: |
424/659 |
Current CPC
Class: |
A61K 33/22 20130101;
A61P 33/02 20180101; A61K 33/22 20130101; A61K 45/06 20130101; A61P
15/02 20180101; A61P 31/02 20180101; A61K 31/4164 20130101; A61K
31/198 20130101; A61P 13/02 20180101; A61K 2300/00 20130101; A61K
31/198 20130101; A61P 31/00 20180101; A61K 2300/00 20130101; A61P
31/04 20180101; A61K 31/4174 20130101 |
Class at
Publication: |
424/659 |
International
Class: |
A61K 33/22 20060101
A61K033/22; A61K 31/4174 20060101 A61K031/4174 |
Claims
1. A pharmaceutical composition for treating and/or preventing a
vaginal infection and/or pathogenic vaginal biofilms consisting of:
a) from about 100 mg to about 1 g of boric acid; b) from about 10
mg to about 500 mg of ethylene-diamine-tetra-acetic acid (EDTA); c)
optionally, one or more of a surfactant, a gelling agent, a buffer,
a preservative, a detergent, an oil, an alcohol, an emulsifier, a
solubilizer, a humectant, a bioadhesive, an additional
therapeutically active substance, and a bioactive agent selected
from the group consisting of bergamot oil, tea tree oil or other
essential oils, and zinc ion; and d) a pharmaceutically acceptable
carrier suitable for vaginal and/or vulvar drug administration, and
wherein said pharmaceutical composition has a pH of about 3.5 to
about 7.5.
2. The pharmaceutical composition of claim 1 wherein the additional
therapeutically active substance is a nitroimidazole.
3. The pharmaceutical composition of claim 2 wherein the
nitroimidazole is selected from the group consisting of
metronidazole and tinidazole.
4. The pharmaceutical composition of claim 1 wherein said
pharmaceutically acceptable carrier is selected from the group
consisting of one of more of an ointment, cream, gel, tablet,
capsule, ovule, suppository, solution, suspension, foam, film,
liposomal composition, a vaginal ring, tampon, sponge, pillow,
puff, and osmotic pump system.
5. An article of manufacture comprising a packaging material and a
pharmaceutical composition of claim 1 within said packaging
material.
Description
FIELD OF THE INVENTION
[0001] The invention generally relates to compositions and methods
for treating vaginal infections and pathogenic vaginal
biofilms.
BACKGROUND OF THE INVENTION
[0002] According to the Centers for Disease Control (CDC),
vaginitis is an extremely common diagnosis among women of
reproductive age, resulting in more than 10 million medical office
visits every year. The vast majority of cases of vaginitis are
related to one of three infections: bacterial vaginosis (BV),
vulvovaginal candidiasis (VVC) (aka "vaginal yeast infection"), and
trichomoniasis. Bacterial vaginosis accounts for approximately
40-45% of all infections, while VVC and trichomoniasis account for
about 20-25% and 15-20%, respectively. In some instances, the
etiology of vaginitis may be mixed. Unfortunately, national
surveillance data on vaginitis are lacking, as none are reportable
diseases; however, prevalence estimates by the National Health and
Nutrition Examination Survey for BV show that nearly one-third
(29%) of women in the general U.S. population between the ages of
14-49 years of age are positive for this infection.
[0003] Although a number of FDA-approved therapies are available
for the management of vaginitis, treatment is often challenging and
further complicated by the increasing rates of treatment-resistant
organisms and recurrent and persistent infections. In addition,
there is a growing body of scientific evidence that identifies
these infections as important risk factors for more serious health
complications, particularly during pregnancy. Estimates of the
direct cost of vaginitis for medical office visits and self
treatment are reportedly more than $1 billion annually; however,
indirect costs related to secondary complications (e.g., morbidity
in pregnancy, pelvic inflammatory disease, and postoperative
infections) and lost work productivity are far greater.
[0004] Accordingly, the public health implications of these
infections are significant, and there is a need for improved
therapeutic approaches.
[0005] Biofilm-related infections were first described in 1978 and
are now believed to be a causative factor in more than 60% of human
infections, particularly in their persistence and recurrence.
Biofilms have been described for a wide range of chronic infections
caused by bacterial and fungal organisms, including skin wounds and
burns, otitis media, periodontal disease, endocarditis, urinary
tract infections and device-related infections (e.g., catheters,
heart valves), but are not recognized as important in the causation
of vaginal infections and, thus, have not been addressed in
practice by those skilled in the art of treating vaginitis.
[0006] Biofilms are highly organized populations of microorganisms
embedded in a protective exopolysaccharide (i.e., carbohydrate)
matrix that adhere to inert and living membrane surfaces (i.e.,
sessile populations) by way of adhesion proteins. In contrast to
their free-floating or "planktonic" counterparts,
biofilm-associated microorganisms are notoriously resistant to
antimicrobial therapy--up to 1000-fold or greater--and are a source
of many recurrent and recalcitrant infections. It is believed that
their persistence is related, in part, to the up regulation of
genes that confer a highly distinct and resistant biofilm phenotype
that perpetuates growth and survival of the biofilm community. This
includes the formation of biofilm matrix material, which restricts
antimicrobial penetration and interferes with normal host defense
mechanisms, and the generation of "persister" organisms that are
essentially intolerant to killing.
[0007] The ability of biofilms to migrate over solid surfaces away
from areas of high stress--a capability known as swarming--and
their slow rate of growth are also believed to contribute to their
pathogenicity and persistence. Evidence further suggests that
established biofilms play a role in the persistence of other
secondary pathogens, such as viruses, by acting as protective
reservoirs that shield these organisms from destruction by the
immune system and conventional antimicrobial therapies.
[0008] Boric acid or boracic acid [B(OH).sub.3] is a weak inorganic
acid with weak antimicrobial properties. In vitro, boric acid is
weakly fungistatic against clinical isolates of C. albicans as well
as non-albicans species, including C. tropicalis, C. glabrata and
C. parapsilosis (Shubair, 1990; Prutting 1998). While boric acid
also displays bacteriostatic activity in vitro against a range of
common bacterial pathogens, including staphylococci and
streptococci, P. aeruginosa, E. coli, and Proteus, Klebsiella and
Enterobacter species (Meers 1990), the antibacterial effects of
boric acid are slow acting and, in contrast to many antibiotics,
appear to be independent of cell growth as dividing and
stationary-phase cells have been shown to be equally affected
(Meers 1990). The weak antimicrobial properties of boric acid
render it surprising that boric acid would be effective in the
treatment of vaginal infections, particularly those that are
resistant, persistent and recurrent in nature.
[0009] Boric acid also displays other biological effects. For
example, boric acid has been shown to play a role in the modulation
of calcium and to stimulate wound healing through action on
extracellular matrix formation and synthesis of growth factors
(Dzondo 2002). Boric acid has also been shown to have
anti-proliferative effects in prostate cancer cell lines and
cytoprotective effects in animal models of gastric injury (Barranco
2006); (Alsaif 2004). There is also evidence to further suggest
that boric acid has antiviral activity, specifically against herpes
simplex virus (Skinner 1979; Rodu 1988). On a molecular level,
boric acid binds cis-diol compounds, including membrane
polysaccharides and carbohydrate moieties of nucleic acids involved
in cell metabolism and signaling (e.g., RNA, NAD, ATP), which may
explain in part the reported effects of boric acid on membrane and
cellular functioning (Kim 2006); whereas the combination of boric
acid and ethylene-diamine-tetra-acetic acid (EDTA) has demonstrated
a unique synergy on corneal membrane permeability in vitro (Kikuchi
2005).
SUMMARY OF THE INVENTION
[0010] The invention generally relates to the surprising discovery
that boric acid may be used to treat and/or prevent vaginal
infections due to its role as a vaginal biofilm disrupter. We
postulate that biofilms play an important role in vaginal
infections, specifically in vaginitis, and more specifically, in
bacterial vaginosis (BV), vulvovaginal candidiasis (VVC) (aka
"vaginal yeast infection"), and trichomoniasis.
[0011] In one embodiment, the invention relates to a pharmaceutical
composition for treating and/or preventing vaginal infections
and/or pathogenic vaginal biofilms comprising boric acid and
ethylene-diamine-tetra-acetic acid (EDTA).
[0012] The amount of boric acid in the pharmaceutical compositions
of the invention is preferably from about 1 mg to about 100 g, more
preferably from about 10 mg to about 10 g, and even more preferably
from about 100 mg to about 1 g.
[0013] The amount of EDTA in the pharmaceutical compositions of the
invention is preferably from about 1 mg to about 1 g, and more
preferably from about 10 mg to about 500 mg.
[0014] In one embodiment, the relative amount of boric acid to EDTA
is in the range of from about 1:1 to about 1000:1 on a weight
basis.
[0015] In another embodiment, the relative amounts of EDTA and
boric acid are in the range of from about 2:1 to 100:1.
[0016] In another embodiment, the pharmaceutical compositions of
this invention further comprise one or more bioactive agents
selected from the group consisting of bergamot oil, tea tree oil or
other essential oils, zinc ion, and gallium.
[0017] In another embodiment, the pharmaceutical compositions of
this invention further comprise at least one additive selected from
the group consisting of gelling agents, buffers, preservatives,
surfactants, detergents, oils, alcohols, emulsifiers, solubilizers,
humectants, and bioadhesives.
[0018] In one embodiment, the pharmaceutical compositions of this
invention further comprise a pharmaceutically acceptable carrier
suitable for vaginal and/or vulvar drug administration.
[0019] In another embodiment, the invention relates to a method of
treating and/or preventing vaginal infections and/or pathogenic
vaginal biofilms comprising administering to a patient in need
thereof a therapeutically effective amount of boric acid in the
absence of any therapeutic amount of acetic acid.
[0020] In another embodiment, the invention relates to a method of
treating and/or preventing a vaginal infection and/or pathogenic
vaginal biofilms comprising administering to a patient in need
thereof a therapeutically effective amount of a pharmaceutical
composition comprising boric acid and EDTA.
[0021] In one embodiment, the methods of this invention comprise
applying the pharmaceutical compositions to a vagina and/or a vulva
from once a day to three times a day.
[0022] In another embodiment, the methods of this invention
comprise applying the pharmaceutical compositions to a vagina
and/or a vulva intermittently, such as twice weekly, as
prophylaxis.
[0023] In another embodiment, the pharmaceutical compositions of
the invention are applied topically to a vagina and/or a vulva of a
patient.
[0024] The pharmaceutical compositions may be applied in the form
of a suppository, an ointment, cream, solid (e.g., tablet, capsule,
ovule, suppository), solution, suspension, gel, foam, film or
liposomal composition. The pharmaceutical compositions may also be
contained within a vaginal ring, tampon, suppository, sponge,
pillow, puff, or osmotic pump system.
[0025] The pharmaceutical compositions of the invention can be
co-administered with other pharmaceutically active compounds, as
for example, with metronidazole.
[0026] The dosage forms of the pharmaceutical compositions may also
be formulated in a sustained-release form, employing various
polymers, fibers, resins, waxes, oils, or other pharmaceutical
excipients used by those skilled in the art of medicinal chemistry
to produce a prolonged release of the active constituents of the
pharmaceutical compositions.
[0027] In one embodiment, the vaginal infection is a bacterial
infection.
[0028] In another embodiment, the vaginal infection is vulvovaginal
candidiasis.
[0029] In yet another embodiment, the vaginal infection is
trichomoniasis.
[0030] In yet another embodiment, the vaginal infection is a viral
infection.
[0031] If not properly treated, vaginal infections may result in
urinary tract infections.
Thus, in another embodiment, the invention relates to the treatment
and prevention of urinary tract infections.
[0032] In yet another embodiment, the vaginal infection is caused
by two or more pathogens selected from the group consisting of
bacteria, fungi, parasites, and viruses and is referred to as a
mixed infection.
[0033] In another embodiment, the invention relates to an article
of manufacture comprising packaging material and a pharmaceutical
composition of the invention within the packaging material. The
pharmaceutical composition is present in an amount sufficient to
treat a vaginal infection or pathogenic vaginal biofilms in a
patient, preferably in an amount equivalent to at least one unit
dose. The packaging material comprises a label that indicates that
the pharmaceutical composition can be used for treating vaginal
infections and associated pathogenic vaginal biofilms. Preferably
the label includes other printed indicia such as a listing of
ingredients, the manufacturer's name and address, and the like.
Preferably the packaging material also includes a printed insert
including detailed information on the composition, its method of
administration for treatment of vaginal infections and pathogenic
biofilms, side effects, contraindications, and the like indicia,
which may be required by governmental agencies responsible for
regulation of pharmaceutical products.
BRIEF DESCRIPTION OF THE DRAWINGS
[0034] FIG. 1 is a diagram that illustrates a symbiotic
relationship of BV pathogens;
[0035] FIG. 2 is a chart that illustrates the effect of boric acid
and EDTA on C. albicans biofilm in terms of log reduction in
biofilm load vs. untreated biofilm controls.
[0036] FIG. 3 is a chart that illustrates the effect of boric acid
and EDTA on G. vaginalis biofilm in terms of log reduction in
biofilm load vs. untreated biofilm controls.
[0037] FIG. 4 is a Kaplan-Meier chart that illustrates the cure
probability of women with refractory BV that were treated with
boric acid.
DETAILED DESCRIPTION OF THE INVENTION
Definitions
[0038] A "biofilm" as used herein refers to a surface-attached,
highly structured, single- or multi-species population of
microorganisms enmeshed in a self-produced exopolysaccharide matrix
that facilitates adherence, colonization and growth on a surface,
such as the vaginal mucosa, and displays a distinct biofilm
phenotype.
[0039] When referring to a compound as an active agent, applicants
intend the term "compound" or "active agent" to encompass not only
the specified molecular entity but also its pharmaceutically
acceptable, pharmacologically active salts.
[0040] The terms "treating" and "treatment" as used herein refer to
causing a reduction in severity and/or frequency of symptoms,
elimination of symptoms and/or underlying cause, prevention of the
occurrence of symptoms and/or their underlying cause, and/or
improvement . Thus, "treating" a patient with said compositions of
the invention includes prevention of a particular disorder in a
susceptible individual, as well as management of a clinically
symptomatic individual to inhibit or cause regression of a disorder
or disease. Treatment can include prophylaxis, therapy, or cure.
For example, treatment of a biofilm encompasses prevention of
formation of a biofilm in a patient susceptible to development of a
biofilm as well as treatment of a patient with a biofilm by
inhibiting, controlling or causing regression of the underlying
disease.
[0041] The term "vagina" as used herein is intended to be inclusive
of the vaginal region generally, including also the vulva,
vestibule, and the cervix.
[0042] The term "vaginal infection" includes any infectious
disorder, including biofilm-associated infections, of any vaginal
area, including the vulva, vestibule, and the cervix. The term also
encompasses plural vaginal infections.
[0043] The term "disrupting" is intended to encompass inflicting
any and all damage to biofilms.
[0044] The term "therapeutically effective amount" of the
pharmaceutical compositions of the invention refers to a sufficient
amount of the composition to treat disorders, at a reasonable
benefit/risk ratio applicable to any medical treatment. It will be
understood, however, that the total daily usage of the compositions
of the present invention will be decided by the attending physician
within the scope of sound medical judgment. The specific effective
dose level for any particular patient will depend upon a variety of
factors including the disorder being treated and the severity of
the disorder; activity of the specific compound employed; the
specific composition employed; the age, body weight, general
health, sex and diet of the patient; the time of administration,
route of administration, and rate of excretion of the specific
compound employed; the duration of the treatment; drugs used in
combination or coincidental with the specific compound employed;
and like factors well known in the medical arts. For example, it is
well within the skill of the art to start doses of the composition
at levels lower than required to achieve the desired therapeutic
effect and to gradually increase the dosage until the desired
effect is achieved.
[0045] As used in this specification and the appended claims, the
singular forms "a," "an" and "the" include plural referents unless
the context clearly dictates otherwise. Thus, for example,
reference to "a bioactive agent" includes a mixture of two or more
bioactive agents, reference to "a pharmaceutically acceptable
excipient" includes mixtures of such excipients, and the like.
[0046] The invention relates to a pharmaceutical composition for
treating and/or preventing a vaginal infection and/or pathogenic
vaginal biofilms comprising boric acid and EDTA.
[0047] The amount of boric acid in the pharmaceutical compositions
of the invention is preferably from about 1 mg to about 100 g, more
preferably from about 10 mg to about 10 g, and even more preferably
from about 100 mg to about 1 g.
[0048] The amount of EDTA in the pharmaceutical compositions of the
invention is preferably from about 1 mg to about 1 g, and more
preferably from about 10 mg to about 500 mg.
[0049] In one embodiment, the relative amount of boric acid to EDTA
is in the range of from about 1:1 to about 1000:1 on a weight
basis.
[0050] In another embodiment, the relative amounts of EDTA to boric
acid are in the range of from about 2:1 to 100:1.
[0051] The pharmaceutical compositions of this invention may
further comprise one or more bioactive agents selected from the
group consisting of bergamot oil, tea tree oil or other essential
oils, zinc ion, and gallium. These bioactive agents may enhance
and/or complement the biofilm-disrupting effects of boric acid and
EDTA.
[0052] For example, gallium may be added to the pharmaceutical
composition to enhance the biofilm disruption effects of boric acid
and EDTA by targeting the iron metabolism of pathogenic
microorganisms and interfering with its signaling role in the
formation of biofilms.
[0053] Zinc ion or a salt thereof may be added to enhance the
biofilm disrupting effect of boric acid and EDTA.
[0054] Surfactants may be added to the pharmaceutical compositions.
The surfactants may provide for better surface contact of the
compositions with the vaginal mucosa by further reducing surface
tension and promoting dispersal of the active substances, or may
possess anti-biofilm properties that may complement those of said
boric acid and EDTA compositions. Surfactants that may be added
include but are not limited to lauryl sulfate and teepol, and the
like.
[0055] The pharmaceutical compositions of this invention may
further comprise other additives such as gelling agents, buffers,
preservatives, detergents, oils, alcohols, emulsifiers,
solubilizers, humectants, and bioadhesives.
[0056] The pharmaceutical compositions of this invention further
comprise a pharmaceutically acceptable carrier suitable for vaginal
and/or vulvar drug administration.
[0057] The compositions of the present invention may include a
physiologically tolerable preservative.
[0058] Suitable physiologically tolerable preservatives include
bacteriostats, preservatives, inhibitors, and the like, such as
methyl, ethyl, propyl, and butyl esters of parahydroxybenzoic acid
(paraben); propyl gallate; sorbic acid and its sodium and potassium
salts; propionic acid and its calcium and sodium salts;
6-acetoxy-2,4-dimethyl-m-dioxane; 2-bromo-2-nitropropane-1,3 -diol;
salicylanilides such as dibromosalicylanilide and
tribromosalicylamilide, the cis isomer of
1-(3-chloroallyl-3,5,7-triaza-1-azanidadamantane chloride;
hexachlorophene; sodium benzoate; phenolic compounds such as butyl
hydroxyanisol, butyl hydroxytoluene, chloro- and bromo-cresols, and
chloro- and bromo-oxylenols; quaternary ammonium compounds such as
benzalkonium chloride; aromatic alcohols such as 2-phenylethyl
alcohol and benzyl alcohol; chlorobutanol; quinoline derivatives
such as iodochlorohydroxyquinoline; and the like. Preferably, the
preservative is included in an amount in the range of about 0.05 to
about 0.2 weight percent, on a total composition weight basis.
[0059] Pharmaceutically acceptable excipients that can be included
in the pharmaceutical compositions of the present invention
include, for example, physiologically tolerable surfactants,
solvents, emollients, colorants, fragrances, and the like, which
are well known in the art. The compositions preferably have a pH
value in the range of about 3.5 to about 7, more preferably in the
range of about 4 to 4.5.
[0060] The invention also relates to a method of treating and/or
preventing vaginal infections and/or pathogenic vaginal biofilms
comprising administering to a patient in need thereof a
therapeutically effective amount of boric acid.
[0061] The invention also relates to a method of treating and/or
preventing vaginal infections and/or pathogenic vaginal biofilms
comprising administering to a patient in need thereof a
therapeutically effective amount of boric acid and EDTA.
[0062] Without being bound to any specific theory, it is believed
that boric acid may act by binding preferentially with cis-diol
carbohydrate components of the pathogenic biofilm matrix and
stimulating the synthesis of protease enzymes, thereby disrupting
the structural integrity of the biofilm and its adherence to the
vaginal mucosa. Boric acid may also offer an additional advantage
in BV biofilms because of its ability to chemically "trap" ammonia
by forming ammonium borate (Thiex NJ et al. 2002). The relevance of
this relates to the symbiotic relationship described for G.
vaginalis, the primary bacterial constituent of BV biofilms, and P.
bivia--a secondary BV pathogen (Pybus 1997). In vitro research
suggests that G. vaginalis thrives off the ammonia by-products of
P. bivia metabolism while P. bivia thrives off the amino acid
by-products of G. vaginalis metabolism. FIG. 1 shows this symbiotic
relationship in the form of a diagram. Accordingly, vaginal
administration of boric acid may interrupt this symbiosis by
trapping ammonia and depleting the vaginal microenvironment of
essential BV nutrients required for bacterial growth and biofilm
formation and survival.
[0063] The invention also relates to a method of treating and/or
preventing a vaginal infection comprising administering to a
patient a therapeutically effective amount of a pharmaceutical
composition comprising boric acid and EDTA.
[0064] The amount of boric acid in the pharmaceutical compositions
of the invention is preferably from about 1 mg to about 100 g, more
preferably from about 10 mg to about 10 g, and even more preferably
from about 100 mg to about 1 g.
[0065] The amount of EDTA in the pharmaceutical compositions of the
invention is preferably from about 1 mg to about 1 g, and more
preferably from about 10 mg to about 500 mg.
[0066] In one embodiment, the relative amount of boric acid to EDTA
is in the range of from about 1:1 to about 1000:1 on a weight
basis.
[0067] In another embodiment, the relative amount of EDTA to boric
acid is in the range of from about 2:1 to 100:1.
[0068] In one embodiment, the methods of this invention comprise
applying the pharmaceutical compositions to a vagina and/or a vulva
from once a day to three times a day.
[0069] In another embodiment, the methods of this invention
comprise applying the pharmaceutical compositions to a vagina
and/or a vulva intermittently, such as twice weekly, as
prophylaxis.
[0070] The pharmaceutical compositions of the invention are applied
topically to a vagina and/or a vulva of a patient.
[0071] The pharmaceutical compositions may be applied in the form
of a suppository, an ointment, cream, solid (e.g., tablet, capsule,
ovule, and suppository), solution, suspension, gel, foam, film, or
liposomal composition. Ointments and creams, may, for example, be
formulated with an aqueous or oily base with the addition of
suitable thickening and/or gelling agents. One technique for
applying the compositions of the claimed invention is to employ a
single use packet (such as a small envelope-like structure, or the
like) containing an intended single unit dose. The packet is
initially sealed, but is opened at the time of use by tearing,
cutting, or the like at a desired or planned location in the packet
after which the contents are directly administrable as labeled.
[0072] The pharmaceutical compositions may also be contained within
a vaginal ring, tampon, suppository, sponge, pillow, puff, or
osmotic pump system.
[0073] The pharmaceutical compositions of the invention can be
co-administered with other pharmaceutical compositions, for
example, with metronidazole.
[0074] The dosage forms of the pharmaceutical compositions may also
be formulated in a sustained-release form, employing various
polymers, fibers, resins, waxes, oils, or other pharmaceutical
excipients used by those skilled in the art of medicinal chemistry
to produce a prolonged release of the active constituents of the
pharmaceutical compositions.
[0075] In one embodiment, the vaginal infection is a bacterial
infection.
[0076] In another embodiment, the vaginal infection is vulvovaginal
candidiasis.
[0077] In yet another embodiment, the vaginal infection is
trichomoniasis.
[0078] In yet another embodiment, the vaginal infection is a viral
infection.
[0079] If not properly treated, vaginal infections may result in
urinary tract infections.
Thus, in another embodiment, the invention relates to the treatment
and prevention of urinary tract infections.
[0080] In yet another embodiment, the vaginal infection is caused
by two or more pathogens selected from the group consisting of
bacteria, fungi, parasites, and viruses.
[0081] The methods and pharmaceutical compositions can be used in
combination with other antimicrobial therapies, including
antibacterial, antifungal, antiparasitic or antiviral agents, to
potentiate their antimicrobial efficacy and/or reduce their dosage
requirements when employed for the treatment of vaginal infections
associated with biofilm formation.
[0082] In addition, the methods and the pharmaceutical compositions
of the invention may also be used as a carrier material for other
medicines, such as antibiotics, antifungals, antiparasitics and
anti-inflammatories, thereby further broadening the compositions'
medical efficacy.
[0083] The total daily dose of the compositions of this invention
administered to a human or lower animal may range from about 100 mg
to about 15 g/day. More preferable doses can be in the range of
from about 500 mg to about 5 g/day. If desired, the effective daily
dose can be divided into multiple doses for purposes of
administration; consequently, single dose compositions may contain
such amounts or submultiples thereof to make up the daily dose.
[0084] The invention also relates to an article of manufacture
comprising packaging material and a pharmaceutical composition of
the invention within the packaging material. The pharmaceutical
composition is present in an amount sufficient to treat a vaginal
infection and/or disrupt a pathogenic vaginal biofilm in a patient,
preferably in an amount equivalent to at least one unit dose. The
packaging material comprises a label that indicates that the
pharmaceutical composition can be used for treating vaginal
infections and/or disrupting a pathogenic vaginal biofilm.
Preferably, the label includes other printed indicia such as a
listing of ingredients, the manufacturer's name and address, and
the like. Preferably, the packaging material also includes a
printed insert including detailed information on the composition,
its method of administration for treatment of pathogenic vaginal
biofilms and vaginal infections, side effects, contraindications,
and the like indicia, which may be required by governmental
agencies responsible for regulation of pharmaceutical products.
[0085] As the Examples show, provided compositions and methods
demonstrate unexpectedly advantageous properties of the methods and
pharmaceutical compositions according to the invention. The
Examples are presented for illustrative purposes only and are not
meant to limit the invention unless otherwise stated in the
claims.
EXAMPLE 1
Disruption of C. albicans and G. vaginalis Biofilms by Boric Acid
and EDTA
A) Experimental Methods:
[0086] The biofilm disrupting effects of boric acid and EDTA at
various test concentrations, alone and in combination, against C.
albicans and G. vaginalis biofilms were studied in vitro using CDC
biofilm reactors (Biosurface Technologies Corporation). The
experiments were carried out at the Center for Biofilm Engineering
in Bozeman, Montana. The CDC biofilm reactor method was selected
based on its reproducibility and ability to provide consistent
biofilm samples and growth conditions necessary for the evaluation
of antimicrobial agents. As a prelude to the biofilm experiments,
the planktonic (i.e., free-floating vs. biofilm-associated) MIC
values for boric acid and EDTA against both test organisms were
established utilizing standard MIC methodologies. In contrast to C.
albicans, the planktonic MIC values for boric acid and EDTA against
G. vaginalis had not previously been reported.
[0087] The CDC reactor consists of eight polycarbonate coupon
holders suspended from a ported lid. Each holder accommodates 3
separate coupons. The lid with coupon holders and coupons, 24 in
total, is mounted in a 1 liter glass vessel with a side-arm
discharge port. Biofilms are formed by circulating liquid growth
media through the vessel while mixing and shear force is generated
by a magnetic stir bar/vane rotated by a magnetic stir plate.
[0088] In preparation of the biofilm experiments relating to the
invention, overnight cultures of C. albicans and G. vaginalis were
prepared by adding 1 mL frozen stock to 9 mL growth media (either
G. vaginalis ATCC #14018 in 50% BHI broth+1% starch, or C. albicans
ATCC #96113 in 50% SD broth +1% sucrose) and incubated for 24 hours
at 37.degree. C. C. albicans was grown under atmospheric conditions
while G. vaginalis was grown with the addition of 5% CO.sub.2.
[0089] Once the overnight cultures were ready, 24 polycarbonate
coupons were placed in the CDC biofilm reactor coupon holders and
secured using a screwdriver. The influent/effluent tubing ends were
then foiled and clamped, and the entire vessel autoclaved for 20
minutes. The appropriate medium was added directly into the reactor
up to the effluent sidearm (approximately 350 mL). The reactor was
then inoculated with 4 mL of overnight culture. To facilitate
optimal biofilm growth, C. albicans reactors were incubated under
atmospheric conditions at 37.degree. C., with continuous stirring
at the lowest setting for 24 hours. G. vaginalis reactors were
placed in a 5% CO.sub.2 incubator with continuous stirring at the
lowest setting for 120 hours. This phase of biofilm formation is
referred to as "batch mode." After the batch mode was completed,
growth medium flow was initiated at 1 mL/min for an additional 24
hours via a peristaltic pump.
[0090] Next, the 24 biofilm-covered coupons were aseptically
removed from the CDC reactor and placed in a 24-well plate, with
each well containing 2 mL of test solution. The test solutions were
run in triplicate and included varied concentrations of boric acid
(BA) and EDTA or a combination thereof. A growth control and an
active control--based on current treatment standards for the
respective infections--were also included. Fluconazole at a
concentration of 8 .mu.g/mL was selected as the treatment control
for C. albicans. This represents the higher end of the range of
drug levels achieved in biologic fluids and tissues (4.12-8.08
.mu.g/mL), including vaginal secretions and tissues, following oral
dosing of fluconazole at a maximum daily dose of 400 mg (more than
double the current FDA-approved dose for vaginal candidiasis). The
active control for G. vaginalis was metronidazole at a
concentration of 192 .mu.g/mL. This concentration was based in part
on the planktonic MIC value of metronidazole against G. vaginalis
that was found in this study (128 .mu.g/mL), and represents a
conservative level that falls on the higher end of MIC values
reported by other investigators. Note also that this level is more
than 8 times higher than peak systemic levels of metronidazole
achieved with the approved 750-mg BV regimen (Flagyl ER.RTM.).
[0091] Each test solution was dissolved in the appropriate medium
and filter sterilized before use. The plates were then incubated
for 24 hours under appropriate growth conditions. After the 24-hour
treatment period, all coupons were rinsed by removing the solution
in the well and adding 2 mL of sterile phosphate buffered saline
(PBS). The procedure was repeated for a total of two rinses. The
coupons were then aseptically transferred to a sterile 15 mL
conical vial containing 10 mL of sterile PBS. The tubes were
vortexed for 30 seconds, sonicated for 2 minutes and then vortexed
for an additional 30 seconds. The cells suspension was serially
diluted and plated on agar (SD agar for C. albicans and BHI+1%
starch for G. vaginalis). The plates were incubated for 24-48 hours
and the colonies counted. Results were reported as number of colony
forming units (cfu) per cm.sup.2 and mean log reductions in biofilm
load relative to untreated biofilm controls.
B) Biofilm Disruption:
[0092] In these experiments, boric acid and EDTA displayed profound
effects on established biofilms, well beyond that observed for
active controls and consistently above the 3-log threshold required
for effective killing (i.e., bactericidal vs. bacteriostatic
activity). In fact, neither of the active controls--both
FDA-approved antimicrobials--had an appreciable impact on the
respective pathogenic biofilms.
[0093] For C. albicans, a synergistic effect was observed for the
combination of boric acid and EDTA as shown in FIG. 2.
[0094] FIG. 2 is a chart that illustrates the effect of boric acid
and EDTA on C. albicans biofilm. As FIG. 2 demonstrates, the
greatest log reduction in biofilm load of 3.67 cfu/cm.sup.2 was
achieved when boric acid and EDTA were combined at their planktonic
MIC values (1.56 mg/mL and 0.78 mg/mL, respectively) compared with
higher concentrations of the individual agents alone. This
represents a 99.8% reduction in biofilm load compared with
untreated biofilm controls and shows that boric acid, particularly
when combined with EDTA, confers fungicidal activity against C.
albicans biofilms. These findings are remarkable and completely
unexpected based on boric acid's weak fungistatic activity against
planktonic cells and surprisingly in line with the magnitude of log
kill reported for FDA-approved antimicrobials against susceptible
planktonic pathogens--generally in the range of about 2-4 log
reductions.
[0095] Even more impressive results were demonstrated against G.
vaginalis biofilm, with a clear synergistic effect achieved with
combinations of boric acid and EDTA. FIG. 3 is a data chart that
illustrates this effect as log reduction in biofilm load. Boric
acid at a concentration of 40 mg/mL combined with EDTA at a
concentration of 10 mg/mL resulted in complete eradication of the
G. vaginalis biofilm, whereas the combination of these agents at
their low planktonic MIC values--3.125 mg/mL and 0.19 mg/mL
respectively--led to a 99.7% decrease in biofilm load vs. untreated
biofilm controls. In stark contrast, the metronidazole active
control had little impact on the G. vaginalis biofilm, and the
difference compared with the boric acid and EDTA combinations was
quite dramatic. Again, these findings are unprecedented and
unpredictable based on the weak antimicrobial properties of boric
acid and the planktonic MIC values of these agents against G.
vaginalis that were established for this study.
[0096] The unexpected synergy achieved with the planktonic MIC
values of the inventive agents against both Candida and Gardnerella
biofilms is remarkable when considering that bactericidal agents
(such as beta lactam antibiotics) require concentrations up to
1000-times higher (or more) than their planktonic MICs to
effectively inhibit susceptible organisms in a biofilm mode of
growth. These findings support the proposed mechanism of action of
boric acid as a biofilm disrupter.
[0097] The results are quite remarkable in light of the fact that
biofilm disruption would not have been predicted based on the
relatively weak antimicrobial activity of these agents and the
planktonic MIC values that were established for the first time in
this study against G. vaginalis.
[0098] The therapeutic relevance of these data are three-fold: 1)
superior clinical efficacy against biofilm-associated vaginal
infections compared with available therapies; 2) improved patient
safety with extremely low concentrations of actives and
dramatically reduced local and systemic exposure risks; and 3) low
potential for the development of secondary infections (particularly
vulvovaginal candidiasis), that often complicate vaginitis
treatment.
Comparative Example
[0099] For comparative purposes, reference is made to a study by
Swidsinski, et al, An adherent Gardnerella vaginalis biofilm
persists on the vaginal epithelium after standard therapy with oral
metronidazole. Am J Obstet Gynecol 2008; 198(1): 97.e1-6, which
evaluated BV biofilm characteristics on vaginal biopsies following
standard oral metronidazole therapy. In this study, 18 patients
with BV were treated for 7 days with oral metronidazole, given as
500 mg twice daily and evaluated using standard clinical and
microbiologic methods to confirm diagnosis and assess clinical
response. Each patient was required to return for a single
follow-up visit where they underwent vaginal biopsy, which allowed
for the direct visualization of vaginal biofilms using fluorescent
in situ hybridization techniques. The women were randomly assigned
to return for a follow up visit at one of 6 possible time points at
either Day 3 of treatment or at 7, 14, 21, 28 or 35 days after
completion of therapy.
[0100] Results demonstrated resolution of BV based on standard
clinical and microbiologic methods at each time point. Vaginal
biopsy results, however, revealed a different picture, showing a
persistent accumulation of core pathogenic bacteria, particularly
G. vaginalis and Atopobium, in an adherent biofilm that became more
pronounced over time. Notably, all of the G. vaginalis isolates
that were obtained from these women showed good susceptibility to
metronidazole with conventional planktonic MIC testing. This may
well explain the low cure rates reported for standard BV therapies
and further links these outcomes to the inability of these
interventions to effectively disrupt the underlying pathogenic
biofilm. The results obtained in this Comparative Example stand in
stark contrast to the results demonstrated in the Example, which
showed complete eradication of G. vaginalis biofilms with a
combination of boric acid and EDTA and validated the minimal impact
of metronidazole.
EXAMPLE 2
Treatment of Patients with Highly Refractory, Recurrent BV by the
Use of Boric Acid
[0101] A retrospective case review was conducted of the use of
boric acid in a series of patients with recurrent BV. Boric acid
was employed as an antimicrobial agent as part of a 6-month
sequential treatment protocol. To qualify for protocol treatment,
all of the women had previously failed an induction and long-term
maintenance regimen of metronidazole vaginal gel and thus had
highly resistant infections.
[0102] With the exception of boric acid, all of the other
antimicrobials used were FDA approved for BV. The 6-month protocol
was as follows:
[0103] 1. Induction: oral metronidazole or tinidazole twice daily
for 7-10 days
[0104] 2. Boric acid treatment: 600-mg gelatin capsule inserted
vaginally daily for 21 days
[0105] 3. Maintenance: metronidazole vaginal gel twice weekly for 5
months
[0106] Results at the end of 6 months showed that 77% (24/31) of
patients were clinically cured--i.e., asymptomatic and negative for
Amsel's clinical criteria for BV. Sixty-seven percent remained
cured at 9 months (i.e., 3 months after maintenance). The median
duration of remission was 8.7 months. Notably, none of the
treatment failures occurred during boric acid therapy and none of
the patients experienced a secondary yeast infection or
discontinued because of an adverse event or intolerance to any
component of the regimen.
[0107] To further validate these findings, a prospectively designed
clinical study of the use of boric acid in women with highly
refractory BV was undertaken. Similar to the retrospective case
series, boric acid was employed as an antimicrobial agent as part
of a sequential treatment protocol; however, the women enrolled in
this study differed from those in the case series in that their
infections were even more resistant to treatment--all of the women
had failed at least three previous suppressive regimens within 12
months prior to enrollment. Likewise, all patients had florid
symptoms of infection, with malodorous discharge and presented with
all four clinical diagnostic criteria (Amsel's), including
.gtoreq.20% clue cells on vaginal wet mount examination.
[0108] A total of 58 patients were treated "open label" with
standard oral doses of nitroimidazole therapy (metronidazole or
tinidazole) twice daily for 7 days followed by boric acid as a
600-mg gelatin capsule inserted vaginally daily for 21 days.
Patients determined to be clinically cured were then placed on a
maintenance regimen of metronidazole vaginal gel twice weekly for
up to 5 additional months.
[0109] Impressive results were demonstrated early on with a low
rate of clinical failure of 8% following nitroimidazole/boric acid
induction. Similarly, breakthrough infections while on
metronidazole gel maintenance were exceptionally low, at just 12%.
As shown in FIG. 4, the cumulative proportion of cure was
surprisingly high in these refractory cases after 12, 16 and 28
weeks--87%, 78% and 65%, respectively. Likewise, the median
duration of remission was 9 months, which was similar to that noted
in the initial case series (8.7 months).
[0110] These findings are most impressive when juxtaposed against
the short-term cure rates of 30% to 70% reported for women with
primary, uncomplicated BV treated with standard therapies, and the
high likelihood of recurrence--up to 80% within 9 months.
Comparative Example
[0111] For comparative purposes, reference is made to Sobel JD et
al, Suppressive therapy with 0.75% metronidazole vaginal gel to
prevent recurrent bacterial vaginosis, Am J Obstet Gynecol 2006;
194: 1283-1290 which reported results of a more rigorous controlled
clinical study of metronidazole vaginal gel in women with recurrent
BV, using a similar treatment protocol to that described for the
above Example but without boric acid.
[0112] The study involved a 10-day metronidazole vaginal gel
induction phase followed by a 4-month twice-weekly maintenance
regimen and a 3-month follow up period without treatment. At the
end of the 4-month maintenance phase, 74.5% (38/51) of patients
were clinically cured. However, the probability of remaining cured
was just 34% 3 months after maintenance, and the median duration of
remission was between 4 and 5 months (vs. 8.7 to 9 months in the
Example). The rate of break through infections occurring during the
maintenance phase was 25%. This is in stark contrast to the rate
reported in the Example, which was less than half that, at 12%.
Moreover, 59.3% of the women in the Comparative Example required
antifungal therapy for VVC at some point during the study whereas
none of the women in the case series had secondary yeast
infections..sup.1 .sup.1 Note that some patients in the prospective
portion of the Example were placed on prophylactic antifungal
therapy, which precluded an accurate assessment of this
complication.
[0113] The differences between the Example and the Comparative
Example are remarkable, particularly considering that many of these
patients were recruited as failures from the more rigorous
controlled maintenance study of metronidazole vaginal gel. The
relative contribution of boric acid and its mechanism of action in
the Example remain unknown but lend clinical support to the in
vitro discovery that boric acid has therapeutically relevant
biofilm disrupting properties, which are enhanced with the addition
of EDTA. The lack of complicating VVC noted in the Example likely
relates to boric acid's antifungal activity and represents another
important advantage of boric acid over currently approved BV
therapies.
* * * * *