U.S. patent application number 14/521000 was filed with the patent office on 2015-06-04 for pharmaceutical anti-infective composition for inhalation.
The applicant listed for this patent is Philippe Baudier, Arthur DEBOECK, Francis VANDERBIST. Invention is credited to Philippe Baudier, Arthur DEBOECK, Francis VANDERBIST.
Application Number | 20150150895 14/521000 |
Document ID | / |
Family ID | 39232962 |
Filed Date | 2015-06-04 |
United States Patent
Application |
20150150895 |
Kind Code |
A1 |
DEBOECK; Arthur ; et
al. |
June 4, 2015 |
PHARMACEUTICAL ANTI-INFECTIVE COMPOSITION FOR INHALATION
Abstract
An anti-infective composition for inhalation, containing, at
least an effective amount of an antimicrobial aminoglycoside
compound or a salt thereof; and an effective amount of a biofilm
modifier which is a macrolide compound or salt thereof.
Inventors: |
DEBOECK; Arthur; (Gurabo,
PR) ; Baudier; Philippe; (Brussels, BE) ;
VANDERBIST; Francis; (Brussels, BE) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
DEBOECK; Arthur
Baudier; Philippe
VANDERBIST; Francis |
Gurabo
Brussels
Brussels |
PR |
US
BE
BE |
|
|
Family ID: |
39232962 |
Appl. No.: |
14/521000 |
Filed: |
October 22, 2014 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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13137826 |
Sep 15, 2011 |
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14521000 |
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12385518 |
Apr 9, 2009 |
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13137826 |
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PCT/EP2007/060855 |
Oct 11, 2007 |
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12385518 |
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Current U.S.
Class: |
514/29 |
Current CPC
Class: |
A61K 9/0078 20130101;
A61K 47/28 20130101; A61K 31/7048 20130101; A61K 47/183 20130101;
A61K 9/0075 20130101; A61P 31/04 20180101; A61K 31/7036 20130101;
A61K 47/24 20130101; A61K 9/4816 20130101; A61K 31/7032 20130101;
A61K 47/26 20130101; A61K 9/0073 20130101; A61K 31/7052
20130101 |
International
Class: |
A61K 31/7048 20060101
A61K031/7048; A61K 9/00 20060101 A61K009/00; A61K 47/26 20060101
A61K047/26; A61K 31/7036 20060101 A61K031/7036 |
Foreign Application Data
Date |
Code |
Application Number |
Oct 11, 2006 |
EP |
PCT/EP2006/000112 |
Oct 11, 2007 |
EP |
PCT/EP2006/060855 |
Claims
1. An inhalation composition for the treatment of lung disease
caused by biofilm producing bacteria, fungal or viral pathogens,
comprising at least: (a) an antimicrobial aminoglycoside compound
or a salt thereof, and (b) a biofilm modifier which is a macrolide
selected from the group consisting of azythromycin, clarithromycin
and erythromycin; or a pharmaceutical salt thereof or both; said
components a) and b) being in an amount sufficient to disrupt said
biofilm and treat said disease.
2. The composition of claim 1, in a form of a dry powder.
3. The composition according to claim 1, in a form of a suspension,
solution, or a combination thereof.
4. The composition of claim 1, wherein a ratio
aminoglycoside/macrolide is between 0.2 to 5.
5. The composition of claim 4, wherein the ratio is between 0.3 and
3.
6. The composition of claim 4, wherein the ratio is between 0.8 and
2.
7. The composition of claim 1, wherein the total amount of active
ingredients represents more than 20% of the composition.
8. The composition claim 7, wherein the total amount of the active
ingredients is more than 30% of the composition.
9. The composition of claim 8, wherein the total amount of the
active ingredients is more than 40% of the composition.
10. The composition of claim 1, containing at least one or more
pharmaceutically acceptable excipients for inhalation to the
lungs.
11. The composition of claim 10, wherein at least one of said
pharmaceutically acceptable excipients is a carbohydrate or a
mixture of two or more carbohydrates.
12. The composition of claim 11, wherein at least one of said
pharmaceutically acceptable carbohydrate comprises anhydrous
lactose, lactose monohydrate, mannitol, xylitol, dextrose,
saccharose, a cyclodextrin compound or a mixture thereof.
13. The composition of claim 10, wherein at least one of said
pharmaceutically acceptable excipient is a lipidic excipient.
14. The composition of claim 13, wherein at least one of said
lipidic excipients is selected from the group consisting of
cholesterol compounds, phospholipid, ethers of fatty alcohols,
esters of fatty acids, hydrogenated oils, polyoxyethylenated
compounds and esters of glycerol.
15. The composition of claim 14, wherein said composition contains
a mixture of cholesterol or a cholesterol compound and a
phospholipid or a phospholipid.
16. The composition of claim 1, further containing one or more
antioxidant(s) selected from the group consisting of cysteine
and/or esters thereof ascorbic acid and/or salts thereof,
tocopherol esters, propylgallate, butylhydroxyanisole, or
butylhydroxytoluene.
17. The composition of claim 1, wherein the aminoglyscoside is
selected from the group consisting of Tobramycin, Kanamycin,
Streptomycin, Gentamicin, Amikacin, Arbekacin, Bekanamycin,
Astromycin, Dihydrostreptomycin, Framycetin, Neomycin, Netilmicin,
lsepamicin, Micronomicin, Sisomicin or a pharmaceutically
acceptable salt thereof.
18. The composition of claim 1, wherein the aminoglyscoside
compound is Tobramycin or a salt thereof and the macrolide compound
is clarithromycin or a salt thereof, said composition being free of
another antimicrobial agent.
19. The composition of claim 1, wherein said composition is free of
excipients.
20. The composition of claim 2, wherein said dry powder composition
is filled in pharmaceutically acceptable capsules.
21. The composition of claim 20, wherein said pharmaceutically
acceptable capsule comprises, as main polymer, gelatin,
hydroxypropylcellulose or starch.
22. The composition of claim 2, wherein the said dry powder
composition is filled into a multidose dry powder inhaler
device.
23. The composition of claim 1, which is filled into a nebulizer
inhaler.
24. The composition of claim 1, wherein the biofilm produced is a
mature biofilm.
25. The composition of claim 1, whereby each active ingredient (a)
and (b) are delivered in an amount of from 5 to 20 times above the
MIC for the pathogens causing the infection.
Description
FIELD OF THE INVENTION
[0001] The invention relates to pharmaceutical compositions for the
treatment of lung infections caused by biofilm producing bacterial,
fungal or viral pathogens.
[0002] The invention discloses inhaled combinations of at least one
antibiotic agent which is an aminoglycoside or a salt thereof and
at least one biofilm modifyer agent which is a macrolide or a salt
thereof, for the treatment of recurrent lung infections associated
with a biofilm.
[0003] The compositions of the present invention are meant to be
administered locally in the lungs of patients, said lung
administration being performed using a dry powder inhaler system or
a nebulizer.
[0004] The weight ratio of each of said antibiotic and said biofilm
modifyer may be superior or equal to 10% of said dry powder
inhaler. The weight ratio between the aminoglycoside and the
macrolide in the compositions of the invention can be comprised
between 0.2 and 5, and the total amount of both active ingredients
per pharmaceutical composition can be comprised between 1 and 50 mg
for a total weight of dry powder per composition being comprised
between 1 and 100 mg.
BACKGROUND OF THE INVENTION
[0005] Antibiotic resistance and persistent infections refractory
to per os or injected treatments are a major problem in
bacteriological transmissions, resistance to eradication and
ultimately pathogenesis. While the consequences of bacterial
resistance and bacterial recalcitrance are the same, there are two
different mechanisms that explain the two processes. [0006]
Antibiotic/Antimicrobial Resistance. In the case of antibiotic or
antimicrobial resistance, biofilms provide the unique opportunity
for bacterial to reside in close proximity with one another for
long periods of time. This prolonged juxtaposition of bacterial
allows gene transfer between and among bacteria, allowing the genes
of resistance to be transferred to same or different strains of
bacteria to neighboring cells that are not resistant. Consequently,
a virulent cell can transfer its virulence genes to a non-virulent
cell, making it resistant to antibiotics. [0007]
Antibiotic/Antimicrobial Recalcitrance. In the case of antibiotic
or antimicrobial recalcitrance, there are two possible
explanations, both of which involve the biofilm and both of which
may be operative simultaneously. While gene transfer may occur, it
is not a factor in recalcitrance.
[0008] Biofilms are matrix-enclosed accumulations of microorganisms
such as bacteria (with their associated bacteriophages), fungi,
protozoa and viruses that may be associated with these elements.
While biofilms are rarely composed of a single cell type, there are
common circumstances where a particular cellular type
predominates.
[0009] Biofilms are the most important primitive structure in
nature. In a medical sense, biofilms are important because the
majority of infections that occur in animals are biofilm-based.
Infections from planktonic bacteria, for example, are only a minor
cause of infectious disease.
[0010] In summary, the biofilm formation consists of planktonic
cells adsorbing onto a surface, experience phenotypic
transformations and form colonies. Once the colonizing cells become
established, they secrete exopolysaccharides that serve as the
backbone for the growing biofilm. While the core or backbone of the
biofilm is derived from the cells themselves, other components
e.g., lipids, proteins etc, over time, become part of the biofilm.
Thus a biofilm is heterogeneous in its total composition,
homogenous with respect to its backbone and heterogeneous with
respect to its depth, creating diffusion gradients for materials
and molecules that attempt to penetrate the biofilm structure.
[0011] The first of the explanatory mechanisms of resistance
offered by biofilm is simply a physical phenomenon: the biofilm
structures present a barrier to the penetration of antibiotics and
antimicrobial agents and a protective shroud to physical agents
such as ultraviolet radiation.
[0012] Another biofilm resistance mechanism is based on biochemical
or metabolic principles. Just as the deep-seated bacterial are
protected from chemical and physical agents by the "barrier" effect
of the biofilm, the biofilm also acts as a barrier to nutrients
that are necessary for normal metabolic activity. Further, the
nutrient-limited bacteria are in a reduced state of metabolic
activity, which make them less susceptible to chemical and physical
agents because the maximal effects of these killing agents are
achieved only when the bacteria are in a metabolically active
state. In addition, biofilms are linked to other virulence factors
of pathogens (like efflux pumps or alginate secretion).
[0013] In particular, biofilms constitute a growing problem for the
treatment of respiratory diseases associated with infection like
cystic fibrosis, diffuse panbronchiolitis, exacerbation of chronic
obstructive pulmonary diseases, pneumonia, etc. . . . . The
treatments of those diseases with antibiotics become consequently
more and more difficult due to the resistance offered by said
biofilm.
[0014] Biofilms are associated with various bacteria, fungi and
viruses among which Pseudomonas aeruginosa and Staphylococcus
aureus cause the most dramatical consequences in the above cited
respiratory diseases.
[0015] Whichever the mechanistic explanations for either resistance
or recalcitrance, the removal or disruption of the biofilm is a
mandatory requirement for the successful treatment of the
infection.
[0016] Aminoglycoside antibiotics are very active antimicrobial
agents but their use has been limited because of their high
frequence of serious and irreversible adverse events associated
with their use. The most common and important toxicity are
nephrotoxicity and ototoxicity. Aminoglycosides are usually
administered by intra-venous injection because they are very poorly
absorbed by the oral route. Nevertheless, a nebulized formulation
of Tobramycin (TOBI.RTM.) to treat lung infections due to P.
aeruginosa in Cystic Fibrosis patients is available.
[0017] TOBI.RTM. presents the advantage to allow to treat lung
infections locally with a lower systemic exposure than the
intravenous formulation and is thus responsible of less
side-effects. However, due to low respiratory fraction compositions
administered through nebulisation (5 to 8% of the nominal dose),
the nominal dose to administer (300 mg of Tobramycin b.i.d.) is
still too high and can be responsible of a significant frequency
and/or severity of side-effects.
[0018] Although, aminoglycoside antibiotics are very effective
against several planktonic bacteria, they are much less effective
if not ineffective against the same bacterial which have formed a
biofilm. This phenomenon of resistance of biofilms is also true for
other antibiotics and represents a major public health concern. In
some specific lung diseases like cystic fibrosis and diffuse
panbronchiolitis, it is of major importance to dispose of
compositions which are able to destroy, disorganize, inhibit the
biofilm and/or able to prevent its formation.
[0019] Oral dosage formulations of macrolides antibiotics are
widely used to treat, among others, respiratory infections like
acute exacerbations of chronic bronchitis, sinusitis,
rhinopharyngitis, . . . .
[0020] While macrolides antibiotics are mostly available as oral
dosage forms containing several hundreds of milligrams of the
antibiotic, no inhaled form has been available up to now, because
amounts of hundreds of mg are impossible to administer ambulatorily
by inhalation through systems like dry powder inhalers or metered
dose inhalers.
[0021] In summary, oral and intravenous antibiotic compositions
used to treat bacterial infections are efficient not and safe
against bacterial biofilms responsible for lung infections or
surinfections.
[0022] Consequently, there is an urgent need for efficient and safe
antibiotic compositions, administered directly in the lung and able
to treat lung infections due to biofilms. The present invention
discloses a dry powder composition for inhalation allowing to
obtain a) a high pulmonary amount of aminoglycoside antibiotic and
b) high dose of a biofilm modifyer which is selected from the group
of macrolides and derivatives, which is efficacious against said
biofilm when administered directly into the lungs.
STATE OF THE ART
[0023] Several inventors have already described attempts to act on
bacterial biofilms.
[0024] JP 726 7868 relates to a biofilm-removing agent containing a
macrolide antibiotic at a low concentration, capable of removing
biofilm formed by periodontal pathogens, making a drug effectively
penetrate to and act on an affected part and also suppressing the
formation of the biofilm and useful for self care. This is a
removing agent of biofilm periodontal pathogens containing a
macrolide antibiotic having a 14-membered ring, preferably
belonging to an erythromycin, a clarithromycin, a
triacetyloleandomycin or a roxithromycin. For administration, the
antibiotic may be contained in e.g. a slow-releasing ointment or
film or in a solvent such as a mouth-washing preparation. The dose
of the macrolide antibiotic is preferably 0.5-10 mg/day, especially
preferably 1-5 mg/day.
[0025] WO 200 602 9893 provides the use of a compound selected from
the group comprising an anthraquinone and a naphtoquinone,
stereoisomeric forms, racemic mixtures, metabolites, esters or
salts thereof, or mixtures thereof, and/or of at least one plant
extract or active fraction thereof comprising said compound for
preventing and/or inhibiting biofilm formation. The present
invention further relates to compositions for preventing and/or
inhibiting the formation of a biofilm, oral health products and a
method for preventing and/or inhibiting biofilm formation.
[0026] US 2005/0049181 A1 describes a synergistic antimicrobial
composition for inhibiting biofilm formation includes an
iron-sequestering glycoprotein, a cationic polypeptide and a
chelating agent, or an iron-sequestering glycoprotein and a
chelating agent, or an iron-sequestering glycoprotein and a
cationic polypeptide. Additionally, surfactants and quaternary
ammonium compounds may also be advantageously combined with
iron-sequestering glycoproteins in an antimicrobial composition.
Methods of using a synergistic composition for inhibiting medical
device biofilm formation are also disclosed.
[0027] U.S. Pat. No. 5,718,899 relates to compositions containing a
high concentration of the full repertoire of immunoglobulins,
including IgA, IgM and IgG, are used to combat infections from
microorganisms and viruses at a wound, surgical, or burn site, or
normal tissue at time of risk of infection. The compositions can
contain elevated antibody titers for several specific pathogens
including S. aureus, CNS, Enterococci, S. epidermidis, P.
aeruginosa, E. coli, and Enterobacter spp, etc. The compositions
are applied directly to a wound or burn site as an ointment, creme,
fluid, spray, or the like, prior to vital or bacterial attachment
or biofilm formation such that adhesion of the pathogens is
inhibited and the pathogens closest to the wound or burn site will
be pre-opsonized for phagocytic killing prior to toxin release. The
immunoglobulins in the composition can be immobilize on a
biocompatible material such as collagen, fibrin, hyaluronan,
biodegradable polymers, and fragments thereof, which will be placed
in-situ at the wound, surgical or burn site. In addition, the
immunoglobulins in the composition may be coated on the body
contacting surface of an implantable device such as a catheter,
contact lens, or total joint. These inventive compositions have
particular application in preventing infections.
[0028] US 2004/0109852 A1 relates to methods for preventing or
removing biofilm on a surface, comprising contacting the surface
with an effective amount of a composition comprising one or more
acylases and a carrier to degrade a lactose produced by one or more
microorganisms, wherein the degradation of the lactose prevents or
removes the biofilm.
[0029] U.S. Pat. No. 6,830,745 B1 describes a two component
composition comprises an anchor enzyme complex to degrade biofilm
structures and a second anchor enzyme component having the
capability to act directly upon the bacteria for a bactericidal
effect.
[0030] US 2002/0022005 A1 relates to a composition for degrading
biofilm structure associated with cystic fibrosis .sup.2 and the
debris associated therewith comprises an enzyme selected for its
ability to dismantle the biofilm structure, and an anchor molecule
coupled to an enzyme to form an enzyme-anchor complex. The anchor
molecule is selected for its ability to attach to a surface on or
proximal the biofilm structure. The attachment to the surface
permits prolonged retention time of the enzyme-anchor complex where
the biofilm structure and associated debris are present.
[0031] WO 02/03998 describes formulation containing between 50 and
750 mg of macrolide for delivery by aerosolization to treat
infection where the bacteria is suceptible to said macrolide. This
patent application also describes methods for treatment of
pulmonary infections by a formulation (liquid solution, suspension
or dry powder) delivered as an aerosol having mass median
aerodynamic diameter predominantly inferior to 1 to 5 .mu.m.
[0032] WO2004/075874 relates to a method of treatment and
prevention of acute or chronic Pseudomonas aeruginosa airway
infections through delivery to the lung endobronchial space,
including alveoli, through delivery of an inhalable formulation and
consisting in the inhalation of a macrolide antibiotic alone or in
combination with another antibiotic.
[0033] The problem of resistance of bacterial biofilm to classical
antiinfective agents in respiratory infectious diseases remains
complete. There is still a need to dispose of a safe and efficient
system to destroy, disorganize or prevent the formation of such
biofilms in such diseases and/or to restore the activity of
antibiotic/antiinfective drugs. It is also not only desirable to
degrade biofilms within a biologic system but it is also necessary
to kill the bacterial cells that are released as the biofilm is
undergoing degradation.
OBJECT OF THE INVENTION
[0034] The present invention is defined in appended independent
claim 1. Preferred embodiments are defined in the dependent claims.
[0035] It is an object of the invention to provide a composition
for inhalation to the lungs which contains a combination of at
least an antimicrobial agent consisting in an aminoglycoside
derivative and a biofilm modifyer which is a Macrolide. The
composition of the invention can be in the form of a dry powder or
in a liquid form, such as a suspension or a solution, or
combination thereof. [0036] It is another object of the present
invention to provide a composition for inhalation, active against
biofilm, consisting in the combination of an aminoglycoside and a
macrolide, wherein the concentration (weight/weight) of each active
ingredient is high i.e. superior to 10% of said dry powder
composition, preferably superior to 15%, more preferably superior
to 20% of said composition. [0037] It is an object of the present
invention to provide a composition for inhalation wherein the ratio
(weight/weight) of the aminoglycoside and the macrolide is
comprised between 0.2 and 5, preferably between 0.5 and 3, more
preferably 0.8 and 2. [0038] It is another object of the invention
to provide a composition for inhalation wherein the total amount of
active drugs (antibiotic+biofilm modifyer) is comprised between 1
and 50 mg. [0039] It is another object of the present invention to
provide a composition for direct administration to the lungs
containing an antimicrobial agent and a biofilm modifyer where the
antimicrobial is an aminoglycoside derivative chosen from
Tobramycin, Kanamycin, Streptomycin, Gentamicin, Amikacin,
Apramycin, Arbekacin, Bekanamycin, Astromycin, Dihydrostreptomycin,
Framycetin, Neomycin, Netilmicin, Isepamicin, Kanamycin,
Micronomicin, Sisomicin or their salts and derivatives. [0040] It
is another object of the present invention to provide a biofilm
modifyer selected from the group of macrolides such as
erythromycin, Clarithromycin, Azithromycin, Roxithromycin,
Erythromycin, Telithromycin, Dirithromycin, Flurithromycin,
Josamycin, Kitasamycin, Midecamycin, Dalfopristin, Oleandomycin,
Midecamycin, Pristinamycin, Rokitamycin, Spiramycin, Tilmicosin,
Troleandomycin, Tylosin, Virginiamycin, or their salts and
derivatives [0041] It is another object of the present invention to
administer the composition of the present invention as an aerosol
or a dry powder, using a generator system which is a single dose or
a mulitdoses inhaler; dry powder inhalers are often referred to as
DPI. [0042] It is another object of the present invention to
provide a composition for inhalation containing an antiinfective
agent and a biofilm modifyer, further containing acceptable
pharmaceutical excipients selected from the group consisting of
carbohydrates or derivatives, lipids derivatives, or other carriers
but also containing sequestring (chelating agents), antioxidants,
stabilizers, buffering agents, surfactants. [0043] It is another
object of the present invention to provide a composition for
inhalation containing an antiinfective agent and a biofilm modifyer
wherein the carrier is a carbohydrate selected from the group of
sucrose, lactose, monohydrate, lactose anhydrous dextrose or a
combination thereof. [0044] It is another object of the present
invention to provide a composition for inhalation containing an
antiinfective agent and a biofilm modifyer wherein the excipients
used are lipid derivatives such as cholesterol, phospholipid
derivatives, fatty acid derivatives or mixtures thereof [0045] It
is another object of the present invention to provide a composition
for inhalation containing an antiinfective agent and a biofilm
modifyer and a chelating agents such as edetic acid, citric acid,
malic acid, or a salt thereof [0046] It is another object of the
present invention to provide a composition for inhalation
containing an antiinfective agent and a biofilm modifyer and a
antioxidant agent such as derivatives of cysteine, ascorbic acid or
derivatives, tocopherol derivatives, propylgallate, parabens
derivatives, etc. . . . [0047] It is also an object of the
invention to provide a composition for inhalation efficient against
bacterial biofilm consisting in a combination of an antiinfectious
agent which is a aminoglycoside and a biofilm modifyer which is a
macrolide, further comprising other therapeutically active agents
such as mucolytic, antiinflammatory, and bronchodilator. [0048] It
is another object of the present invention to provide a composition
for direct administration to the lungs comprising a combination of
antimicrobial which is an aminoglycoside derivative and a biofilm
modifyer which is a macrolide, allowing to decrease the
side-effects and/or the drug-drug interaction, and/or allowing to
increase the compliance and/or the efficacy.
BRIEF DESCRIPTION OF THE DRAWINGS
[0049] FIG. 1 illustrates the effect of tobramycin (4 .mu.g/ml),
clarithromycin (100, 200 and 500 .mu.g/ml) and combinations of
tobramycin/clarithromycin (4/100 .mu.g/ml, 4/200 .mu.g/ml and 4/500
.mu.g/ml) on a 12 day biofilm of Pseudomonas aeruginosa.
DETAILED DESCRIPTION OF THE INVENTION
[0050] In the present patent, "biofilm modifyer" is defined as a
substance able to destroy, destructure, and disorganize the biofilm
and/or to prevent or slow down its formation.
[0051] For the purpose of the present invention, the terms
"antimicrobial", "antiinfective", "antibacterial", and "antibiotic"
are synonyms and refer to substances having a bacteriostatic and
for a bactericidal effect against a given pathogen micro-organism
(bacteria, fungi, virus).
[0052] The present invention discloses the increase of efficacy of
antimicrobial agents in respiratory infections associated with
biofilm by the synergetic combination of at least two active agents
consisting of a biofilm modifyer and an antibiotic, administered by
inhalation.
[0053] Basically, one antibiotic, the aminoglycoside, at least,
should be active against the bacteria contained in the biofilm,
while the macrolide shall act on the biofilm for instance by
disorganizing it, destructuring it, inhibiting the production of
alginate, etc. . . . . The present invention is useful to prevent
the formation of biofilm in patients but also to treat patients
with a formed biofilm.
[0054] The present invention more precisely consists of a
composition dry or liquid for inhalation comprising at least one
antibiotic from the aminoglycoside group and one antibiotic from
the macrolide group, the antibiotic from the macrolide family being
active against biofilms (=biofilm modifyer).
[0055] The antibiotics from the aminoglycoside group comprise, but
are not restricted to: Tobramycin, Kanamycin, Streptomycin,
Gentamicin, Amikacin, Apramycin, Arbekacin, Bekanamycin,
Astromycin, Dihydrostreptomycin, Framycetin, Neomycin, Netilmicin,
Isepamicin, Micronomicin, Sisomicin or their salts and derivatives.
(see Martindale, 33.sup.rd edition, page 111).
[0056] The macrolides from the macrolides group comprise but are
not restricted to: Clarithromycin, Azithromycin, Roxithromycin,
Erythromycin, Telithromycin, Dirithromycin, Flurithromycin,
Josamycin, Kitasamycin, Midecamycin, Dalfopristin, Oleandomycin,
Midecamycin, Pristinamycin, Rokitamycin, Spiramycin, Tilmicosin,
Troleandomycin, Tylosin, Virginiamycin, or their salts and
derivatives. (see Martindale, 33.sup.rd edition, page 112).
[0057] There are significant advantages to administer the present
composition of an aminoglycoside and a macrolide directly to the
lungs instead of their usual route of administration i.e. most
often intravenous for aminoglycoside and oral for macrolide. First,
the very significantly decrease of the systemic exposure leads to
the decrease of potentially very severe adverse effects of
aminoglycoside (nephrotoxicity and ototoxicity) and the mild
adverse effects of macrolides. Second, the inhalation avoids drug
interactions that may occur for some macrolides that are
metabolized through the cytochrome P450 3A4 (like clarithromycin).
Those interactions may again result in important adverse effects.
Third, the interaction with food is also avoided when the
composition is inhaled rather than swallowed. And last but not
least, the inhaled route produces very high local concentrations of
the drugs where needed.
[0058] The amount of each antibiotic and their respective ratio may
vary, depending to the nature of the bacterium to eradicate, the
kind of biofilm and the kind of infection to treat. The amount of
aminoglycoside will be, in every case, such as to provide, locally,
concentrations in aminoglycoside superior to its MIC (Minimal
Inhibitory Concentration) against the planktonic bacterium
considered. However, the preferred ratio (w/w)
aminoglycoside/macrolide in the present invention is 0.2 to 5,
preferably 0.5 to 3, more preferably 0.8 to 2.
[0059] The amount of macrolide agent inhaled shall be high enough
to affect, in some way, the biofilm. It has to be noted that as the
effect of macrolide derivatives on the biofilm is mediated through
a non-antibacterial mechanism. Therefore, the amounts required to
destroy the biofilm by inhalation may be significantly lower than
the one needed for antiinfective activity pre os. Also importantly,
the macrolide derivative does not need to possess an antiinfective
activity against the targeted microorganism to acts on the biofilm.
Nevertheless, it is a another object of the present invention to
provide a composition containing high concentrations (or amounts)
of each of the aminoglycoside derivative and of the macrolide
derivative i.e. at least more than 10%, preferably more than 15%,
and more preferably more than 20% of the dry powder composition. It
is indeed particularly interesting to achieve high lung doses of
those therapeutic agents with the minimum amounts of inhalations
because it makes the administration easier and more importantly
increase the patient's compliance. It also decreases the nominal
dose of each active ingredient and thus the adverse effects linked
to these actives. In the present invention, the dry powder inhaler
also provides a high Fine Particle Dose (FPD) and Fine Particle
Function (FPF) when tested in vitro on a Multistage Liquid Impinger
(MIA, Eur. Pharm, 5.sup.th Edition, chapter 2.9.18). The FPD and
FPF are the parameters that predict in vivo lung deposition.
Briefly, the FPF (%) is defined as the fraction (expressed of in
percent) of the nominal dose presenting a diameter inferior to 5
.mu.m (maximum diameter of particles to be able to reach the lungs)
and the Fine Particle Dose (FPD) is the amount (in mg) per inhaled
unit dose composition presenting a diameter inferior to 5
.mu.m.
[0060] High lung deposition of each active ingredient from the
composition of the present invention will achieve high local
concentrations of the antibiotic (generally 5 to 20 times above the
Minimal Inhibitory Concentration or MIC) in order to kill the
pathogens and high local concentrations of the biofilm modifyer
agent in order to destroy or destructure rapidly the biofilm.
[0061] The DPI composition of the present invention provides with a
FPF of at least 15% of each active ingredient in comparison to the
nominal dose, preferably superior to 20%, more preferably superior
to 35%.
[0062] The preferred ratio (w/w) between the active ingredients
(aminoglycoside+macrolides) and the inactive ingredients in dry
powder composition of the invention, is comprised from 0.2 to 90,
preferably from 0.3 to 5, more preferably 0.4 to 2. Alternatively,
the compositions may be free of excipient (100% of active
drugs).
[0063] In a preferred embodiment of the present invention, both
antimicrobial are present under the form of a dry powder for
inhalation agents and are administered in a fixed combination
through inhalation. Said dry powder compositions may be formulated
as a single dose composition i.e. a composition to be filled
individually in capsules or blisters, or as a multidose composition
i.e. a composition filled in a device equipped with a reservoir
containing several doses and a metering dose system.
[0064] The dry powder composition of the present invention
preferably contains the aminoglycoside derivative in a micronized
form and the macrolide derivative in a micronized form. For the
purpose of the present invention, "micronized" means an average
particle size inferior to 20 .mu.m, preferably inferior to 10 .mu.m
and more preferably inferior to 5 .mu.m when measured by laser
diffraction for instance. The dry powder composition of the present
invention may contain more than one antibiotic and more than one
biofilm modifyer
[0065] The dry powder composition may further contain other
excipients like buffering agents, surfactants, lubricants,
chelating agents or antioxydants, aminoacids. When carbohydrate is
used as main inactive ingredient, it has a role of carrier. Then,
the preferred process is for manufacturing the composition of the
invention is a dry blending of the micronized active ingredients
with the non-micronized carrier. In case of use of a non-micronized
carrier, said carrier has preferably a mean particle size comprised
between 50 and 250 .mu.m, preferably between 80 and 200 .mu.m, more
preferably between 100 and 160 .mu.m. The preferred main carrier is
anhydrous lactose or lactose monohydrate but other
mono-disaccharide such as dextrose, xylitol, mannitol, saccharose
etc., may be used. Mixtures of two or more carriers may also be
used as well as mixtures a carrier with other kinds of excipients
(lubricants, surfactants, antioxidants, etc.).
[0066] The dry powder composition of the invention may contain, in
addition to the main non micronized carrier described hereinabove,
a second carrier which can be non-micronized or micronized. When
this second carrier is micronized, the preferred mean particle size
measured by laser diffraction is inferior to 20 .mu.m, preferably
inferior to 10 .mu.m. The second carrier can be the same chemical
entity as the main carrier or a different one.
[0067] The dry powder composition obtained by dry blending may
further comprise excipients aimed to improve the stability of the
composition, the flowability of the powder or the lung deposition
of both active ingredients.
[0068] Another composition of the invention may contain in addition
to the micronized aminoglycoside and the micronized macrolide, a
lipid derivative or a mixture of different lipid derivatives as
excipients. In this case, the preferred process consists in the
spray-drying the active ingredients together with the lipid. The
spray-drying process requires the use of a liquid in which the
active ingredients and excipients are solubilized or in suspension.
The solution or suspension is homogeneized and then spray-dried to
obtain a particles in the required mean particle range i.e. <10
.mu.m, preferably inferior to 5 .mu.m. This spray-drying process is
a well known in the pharmaceutical industry and a specific process
to obtain dry powder composition may, for instance be found in EP 1
674 085 A1.
[0069] The preferred lipid excipients are either phospholipids
including anionic phospholipids, cationic phospholipids,
zwitterionic phospholipids and neutral phospholipids such as for
example phosphatidylcholine, phosphatidylglycerol,
phosphatidyl-inositol, phospatidyl-serine, or non-phospholipids
such as glycerol esters (like glycerol monostearate, glycerol
behenate), fatty alcohols (preferably with C16 or more), fatty
acids (preferably with C16 or more), ethers of fatty aclohols,
esters of fatty acids, hydrogenated oils, polyoxyethylenated
derivatives and sterols like cholesterol and its derivatives.
Mixtures of two or more lipid derivatives may also be used.
Preferably, a combination of a phospholipid with cholesterol or a
cholesterol derivative may be used in compositions of the present
invention.
[0070] The lipid excipients may also be combined to other lipidic
or non lipidic excipients like carbohydrate, surfactant, lubricant,
antioxidant, chelating agent.
[0071] The dry powder composition of the present invention may
additionally contain one or more chelating agent. The chelating
agent useful for the present invention may include edetic acid
(EDTA) or a salt thereof, but other chelating agent such as citric
acid, malic acid or their salts may be used. The chelating agent
will preferably be present at a concentration (w/w) ranging from
0.01% to 5% of the final dry powder composition. Combinations of
more than one chelating agents may also be used.
[0072] The dry powder composition of the present invention may
additionally contain one or more antioxidant agent. Examples of
antioxidants that can be used include derivatives of cysteine like
acetylcystein and its salts, glutathion, carbocystein derivatives
or ascorbic acid, derivatives of tocopherol, propylgallate, BHA,
BHT.
[0073] It is to be noted that the presence of either a chelating
agent or an antioxidant agent, or both, may further increase the
beneficial effect on the biofilm and may consequently result in a
better efficiency that the contribution of aminoglycoside and
macrolide without these agents.
[0074] In a second preferred embodiment, the composition can be in
the form of a liquid, comprising a carrier and both antibiotics
(macrolide and aminoglycoside) in suspension and/or solution
therein. Nebulizer solutions can be formulated in a similar way to
injectable Macrolide solutions well-known in the art. The liquid
carrier is advantageously water, or any pharmaceutically acceptable
solvent, such as ethanol, dimethylsulfoxide, glycerol, propylene
glycol, and mixtures thereof. The antibiotics in the liquid
compositions of the present invention shall be present in the same
amount ranges as defined supra for the dry powder compositions.
Example 1
In Vitro Demonstration of the Activity of Micronized
Tobramycin+Micronized Clarithromycin on Pseudomonas aeruginosa
Biofilm
[0075] Biofilms of Pseudomonas aeruginosa strain PY O.sub.1 were
formed according to the methods described by Ceri et al, the
calgary biofilm device: new technology for rapid determination of
antibiotic susceptibilities of bacterial biofilms, Journal of
clinical microbiology, pp. 1771-1776, 1999 and Abdi-Ali et al,
bactericidal activity of various antibiotics against
biofilm-producing Pseudomonas aeruginosa, International Journal of
Antimicrobial Agents 27, 196-200, 2006.
[0076] PY O.sub.1: is a cystic Fibrosis clinical mucoid strain of
Pseudomonas aeruginosa received from the Erasme Hospital,
Brussels.
[0077] The determination of the minimal inhibitory concentration
(MIC) is performed according to the standard of NCCLS (NCCLS,
Methods for dilution Antimicrobial Susceptibility Tests for
bacteria that grow aerobically; approved standards, sixth edition,
M7-A6, vol. 23 no. 2 January 2003.
[0078] In the present experiment, the MIC of tobramycin,
clarithromycin and the combination of both antibiotics was first
determined on planktonic bacteria (=probacteria i.e. free bacteria
not included in a biofilm) to prove that there is no direct
additive effect of the active ingredient. The MIC of tobramycin for
Pseudomonas aeruginosa is 3.9 .mu.g/mi. The MIC of clarithromycin
for Pseudomonas aeruginosa could not be determined since the
results showed that the bacterium is not sensitive to this
antibiotic. The MIC of the combination of tobramycin and
clarithromycin is found to be around 3.9 .mu.g/ml. These results,
similar to the MIC value found for tobramycin alone demonstrate
that there is no additional antibiotic effect of clarithromycin on
planktonic Pseudomonas aeruginosa.
[0079] In a first attempt to measure the antibiotic activity (MIC)
of tobramycin on Pseudomonas aeruginosa when incorporated in a
biofilm, a culture of planktonic Pseudomonas aeruginosa was
prepared to produce a biofilm during a period of 24 hours. Upon
completion of the 24 hours the MIC of tobramycin was measured using
these cultures. Surprisingly, it was found that the Minimum
Inhibitory Concentration (MIC) of tobramycin on a 24 hours old
biofilm of Pseudomonas aeruginosa was similar to the activity of
tobramycin of planktonic bacteria. In other words, Tobramycin is
still active on such a biofilm and there is no need to add a
biofilm destroying/destructuring agent.
[0080] In a second experiment using the same modus operandi as
above, we measured the MIC of Tobramycin on a 12 day old
Pseudomonas aeruginosa biofilm. This situation is much closer to
the situation observed in vivo in chronic respiratory diseases like
Cystic Fibrosis. In this case, tobramycin was no longer active
against said Pseudomonas aeruginosa.
[0081] There exists thus a significant difference between a 1 day
old versus a 12 days old biofilm: it appears that a biofilm is a
living entity that evolutes from the native to the mature stage.
Also these experiments shall warn researchers that antibiotic
activity results obtained from species that form a biofilm may not
be taken in consideration unless the biofilm has had sufficient
time to form properly and results found in the literature have to
be taken with precaution.
Effects of Tobramycin, Clarithromycin and Combinations Thereof on
12-Day Biofilm of Pseudomonas aeruzinosa
[0082] After having shown that the number of Pseudomonas aeruginosa
within the biofilms was stable after 12 days, the products listed
in Table 1 were added to the media for the duration of 24 hours.
Thereafter the biofilm was rinsed three times with a 0.01M
phosphate buffer adjusted at pH 7.5 in order to remove all cells
not bound to the biofilm. The microplate was then placed on
ultrasonic bath at 35.degree. C. for 5 minutes to allow the
bacteria present in the biofilm to separate from such biofilm. A
bacterial count was then performed (number of coloning forming unit
CFU/ml). Each experience was done twice and the CFU counting was
also repeated twice/experience.
[0083] The results are shown in Table 1 and FIG. 1. It is concluded
that neither tobramycin 4 .mu.g/ml nor clarithromycin at 100, 200
and 500 .mu.g/ml alone are able to decrease the number of CFU/ml of
the 12-day biofilm versus the positive control.
TABLE-US-00001 TABLE 1 Effect of tobramycin (4 .mu.g/ml),
clarithromycin (100, 200 and 500 .mu.g/ml) and combinations of
tobramycin/clarithromycin (4/100 .mu.g/ml, 4/200 .mu.g/ml and 4/500
.mu.g/ml) on a 12 day biofilm of Pseudomonas aeruginosa. T C C C
T/C T/C T/C 4 100 200 500 4/100 4/200 4/500 PC (.mu.g/ (.mu.g/
(.mu.g/ (.mu.g/ (.mu.g/ (.mu.g/ (.mu.g/ ml) ml) ml) ml) ml) ml) ml)
MIC (CFU .times. 10.sup.7) 16 25 14 10 7 3 1 5 (Low value of MIC is
desired.) PC: Positive control (Mueller-Hinton medium also called
CAMHB) T: Tobramycin C: Clarithromycin T/C: Combination
Tobramycin/Clarithromycin
[0084] To the contrary all the combinations of
Tobrarnycin/Clarithromycin were able to decrease the number of
CFU/ml originated from the biofilm of Pseudomonas aeruginosa with a
maximal effect being observed for the combination TOBRAMYCIN 4
.mu.g/ml+CLARITHROMYCIN 200 .mu.g/ml which shows a number of CFU/ml
of about 10.sup.7 while tobramycin alone at 4 .mu.g/ml shows a
number of CFU/ml of around 2.5.times.10.sup.8. This means a more
than 25 times decrease in the number of CFU/ml for the combination
versus the reference product tobramycin.
[0085] It can be seen that enhanced results are determined when at
least 100 .mu.g/ml clarithromycin is combined with tobramycin,
preferably at least 200 .mu.g/ml. The efficacy of the mixture
decreases somehow for concentration in clarithromycin grater than
500 .mu.g/ml.
Example 2
[0086] A dry powder composition for inhalation of tobramycin and
clarithromycin was formulated using micronized tobramycin supplied
by Teva Plantex (Israel). Clarithromycin was supplied by Teva
Plantex (Israel) in a non-micronized form. Clarithromycin was then
micronized using the micronizer MC-one.RTM. (JetpHarma,
Switerland). To obtain a product with particle size suitable to
reach the respiratory tract (i.e. 80% of particles inferior to 10
.mu.m, and 90% of particles inferior to 5 .mu.m when measured by
laser diffraction). The micronisation parameters were a pressure of
10 bars in the Venturi, a pressure of 8 bars in the ring and a
feeding rate of 5 g/minute. The mean particle size of the
micronized clarithromycin obtained (measured by laser diffraction)
was 1.6 .mu.m.
Manufacturing of DPI Composition:
[0087] 400 g of anhydrous lactose (100-160 .mu.m) were put in a
planetary mixer together with 50 g of micronized lactose
monohydrate. The two lactoses were blended at 40 rpm for 10
minutes. 200 g of micronized tobramycin and 100 g of micronized
clarithromycin were added to the mix of lactoses using the
"sandwich technique", i.e. by alternating the layer of lactoses and
the layer of active ingredients to obtain a final mix as
homogeneous as possible. The mix was blended for 10 minutes at a
speed of 40 rpm.
[0088] Samples were taken from this powder blend to assure both
active ingredients were homogeneously blended. 50 mg of the powder
mix was then filled into number 3 hydroxypropylmethylcellulose
(HPMC) capsules. These capsules are ready for use with a dry powder
inhaled device such as the MIAT monodose inhaler, or any other
suitable capsule based inhalation device.
Example 3
[0089] Edetic acid in an amount of 0.5% (weight/weight) was added
to the blend of example 2. The powder was thereafter filled in a
Miat multidose inhaler device.
Example 4
[0090] 400 g of anhydrous lactose (100-160 .mu.m) was introduced in
a planetary mixer and 150 g of micronized tobramycin, 150 g of
micronized clarithromycin and 20 g of N-acetylcysteinate lysine (as
antioxidant) were added using the "sandwich technique", i.e. by
alternating the layer of lactose and the layer of active
ingredients to obtain a final blend as homogeneous as possible. The
blend was mixed for 10 minutes at a speed of 40 rpm. The blend was
then filled in size 3 hard gelatine capsules (40 mg of
powder/capsule).
Example 5
[0091] 10 g of micronized tobramycin, 5 g of clarithromycin were
dissolved in a 80/20 (w/w) water/ethanol mixture. 300 mg of
Phospholipon 90H.RTM. and 1.2 g of cholesterol were added and
dissolved in said solution containing the active ingredients. The
solution was thereafter spray-dried to obtain a powder consisting
of micrometric spherical lipidic particles with a very high content
in active ingredients. This powder was filled in HPMC capsules for
inhalation (20 mg of powder/capsule).
Example 6
[0092] 400 g of anhydrous lactose (100-160 .mu.m) was mixed in a
planetary mixer (40 rpm for 10 minutes) with 200 g of micronized
tobramycin and 200 g of micronized clarithromycin. 40 mg of the
blend obtained was filled into size 3 hydroxypropylmethylcellulose
capsules. This produced capsules each containing 10 mg of
micronized tobramycin and 10 mg of micronized clarithromycin that
may be used for inhalation
In Vitro Lung Deposition
[0093] The determination of the Fine Particle Fraction (FPF) i.e.
the fraction (expressed in percent) of the nominal dose presenting
a diameter inferior to 5 .mu.m (maximum diameter to reach the
lungs) and the Fine Particle Dose (FPD) i.e. the amount (in mg) per
capsule presenting a diameter inferior to 5 .mu.m, has been
performed on the capsules using the Axahaler device as powder
inhaler device. The in vitro lung deposition test was performed
using equipment and conditions as described in the European
Pharmacopoeia (5.sup.th edition, chapter 2.9.18 apparatus C). This
equipment consists of a Multistage Liquid Impinger (MLI) and was
operated with an air flow of 100 L/min during a period of time of
2.4 seconds to simulate inhalation capabilities of patients. The
quantification of the deposition of each drug on each stage of the
MLI was performed by HPLC equipped with a a Corona detector. The
results are presented in Table 2.
TABLE-US-00002 TABLE 2 FPF (%) and FPD (mg) obtained with the
compositions of example 6 (MLI 100 L/min) containing 10 mg of
tobramycin and 10 mg of clarithromycin/capsule (n = 3) MLI1 MLI2
(mg) (mg) MLI3 (mg) Mean (mg) SD Tobramycin (mg) Device 1.035 0.938
1.272 1.082 0.17 Throat 0.929 0.859 0.771 0.853 0.08 Stage 1 1.760
1.837 1.546 1.714 0.15 Stage 2 0.614 0.726 0.606 0.648 0.07 Stage 3
1.512 1.818 1.895 1.742 0.20 Stage 4 1.598 1.965 2.202 1.922 0.30
Filter 0.664 0.838 0.778 0.760 0.09 FPD (mg) 3.60 4.45 4.72 4.26
0.59 FPF (mg) 35.96 44.50 47.20 42.55 0.06 Clarithromycin (mg)
Device 1.155 1.048 1.433 1.212 0.20 Throat 1.025 1.134 1.223 1.127
0.10 Stage 1 1.338 1.566 1.352 1.419 0.13 Stage 2 0.664 0.845 0.680
0.729 0.10 Stage 3 1.715 1.891 2.078 1.895 0.18 Stage 4 1.162 1.482
1.533 1.392 0.20 Filter 0.478 0.598 0.574 0.550 0.06 FPD (mg) 3.14
3.76 3.98 3.63 0.44 FPF (mg) 31.37 37.56 39.85 36.26 0.04
[0094] The FPF of tobramycin and clarithromycin obtained are 42.5%
and 36.3% respectively. The FPD/capsule of tobramycin and
clarithromycin are 4.26 mg and 3.63 mg respectively. Those results
clearly demonstrate that the compositions of the invention allow to
reach very high lung deposition of both the antibiotic and the
biofilm modifyer. Such high lung deposition is suitable for use in
vivo. Indeed, the volume of epithelial liquid in the lung is
generally estimated at about 100 ml. and lung deposition results
show that each capsule of the composition of example 6 allows thus
to obtain a lung concentration of respectively 42.6 .mu.g/ml of
tobramycin and 36.3 genii of clarithromycin.
Example 6
[0095] Different compositions (Fl to F5) were manufactured using
the blending process as described in example 6.
TABLE-US-00003 mg/dry powder composition Active Ingredient F1 F2 F3
F4 F5 Tobramycin base 20 / 5 15 5 Amikacine / 15 5 / /
Clarithromycin 10 10 15 / 5 Azithromycin / / / 10 / Anhydrous
lactose 20 25 20 20 10 Total weight/composition 50 50 45 45 20
* * * * *