U.S. patent application number 12/277266 was filed with the patent office on 2009-03-26 for pulmonary delivery of polyene antifungal agents.
This patent application is currently assigned to Nektar Therapeutics. Invention is credited to Marc S. Gordon, Sandeep Kumar, Razaq Sarwar, Michael Weickert, Bing Yang.
Application Number | 20090081302 12/277266 |
Document ID | / |
Family ID | 22977001 |
Filed Date | 2009-03-26 |
United States Patent
Application |
20090081302 |
Kind Code |
A1 |
Weickert; Michael ; et
al. |
March 26, 2009 |
PULMONARY DELIVERY OF POLYENE ANTIFUNGAL AGENTS
Abstract
The present invention provides spray-dried polyene compositions
for oral inhalation to the lung. The polyene antifungal
compositions demonstrate superior aerosol properties, do not
exhibit appreciable degradation of the polyene upon spray-drying,
and are useful in the treatment and prophylaxis of both pulmonary
and systemic fungal infections.
Inventors: |
Weickert; Michael; (Belmont,
CA) ; Gordon; Marc S.; (Sunnyvale, CA) ;
Kumar; Sandeep; (Sunnyvale, CA) ; Yang; Bing;
(Redwood City, CA) ; Sarwar; Razaq; (Fremont,
CA) |
Correspondence
Address: |
NEKTAR THERAPEUTICS
201 INDUSTRIAL ROAD
SAN CARLOS
CA
94070
US
|
Assignee: |
Nektar Therapeutics
San Carlos
CA
|
Family ID: |
22977001 |
Appl. No.: |
12/277266 |
Filed: |
November 24, 2008 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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10032239 |
Dec 21, 2001 |
7473433 |
|
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12277266 |
|
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60257613 |
Dec 21, 2000 |
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Current U.S.
Class: |
424/489 ;
424/46 |
Current CPC
Class: |
A61K 9/0075 20130101;
A61P 31/10 20180101; A61K 9/1688 20130101 |
Class at
Publication: |
424/489 ;
424/46 |
International
Class: |
A61K 9/72 20060101
A61K009/72; A61K 9/14 20060101 A61K009/14; A61P 31/10 20060101
A61P031/10 |
Claims
1. A method for preparing a spray-dried polyene powder suitable for
oral inhalation to the lung, said method comprising: (i) dissolving
a polyene antifungal compound in an acidified solvent to form an
acidic polyene-containing solution, and (ii) spray drying said
polyene-containing solution to form an inhaleable dry powder
containing no more than about 10% polyene degradation products and
characterized by an emitted dose greater than 60%.
2. The method of claim 1, wherein said dry powder produced in step
(ii) contains no more than about 5% polyene degradation
products.
3. The method of claim 1, wherein said solvent comprises acidified
alcohol.
4. The method of claim 3, wherein said solvent comprises acidified
methanol or ethanol.
5. The method of claim 1, wherein the pH of said acidified solution
in step (i) ranges from about 3.5 to 5.
6. The method of claim 1, wherein the polyene is dissolved in said
acidified solution to an extent greater than about 1 mg/ml.
7. The method of claim 6, wherein the polyene is dissolved in said
acidified solution to an extent greater than about 2 mg/ml.
8. The method of claim 6, wherein the polyene is dissolved in said
acidified solution to an extent greater than or equal to about 3
mg/ml.
9. The method of claim 1, wherein prior to said spray drying step,
the acidic polyene-containing solution is maintained at a
temperature below 25.degree. C.
10. The method of claim 9, wherein prior to said spray drying step,
the acidic polyene-containing solution is maintained at a
temperature below about 8.degree. C.
11. The method of claim 9, wherein the acidic polyene-containing
solution comprising a feed solution is maintained at a temperature
below 25.degree. C. during said spray drying step.
12. The method of claim 9, wherein the acidic polyene-containing
solution comprising a feed solution is maintained at a temperature
below 8.degree. C. during said spray drying step.
13. The method of claim 1, wherein said polyene comprises
amphotericin or nystatin.
14. The method of claim 1, further comprising the step of
dissolving a pharmaceutically acceptable excipient in said
acidified solvent to form a solution comprising said excipient and
said polyene.
15. The method of claim 14, wherein said excipient is leucine or
trileucine.
16. The method of claim 1, wherein said polyene-containing solution
comprises dissolved solids and wherein at least about 60% by weight
of the dissolved solids comprises said polyene.
17. The method of claim 16, wherein at least about 70% by weight of
the dissolved solids comprises said polyene.
18. The method of claim 17, wherein said polyene-containing
solution is substantially absent additional excipients or
stabilizers.
19. The method of claim 1, wherein said polyene-containing solution
is absent lipid or polymeric encapsulating agents.
20. The method of claim 1, wherein said inhaleable dry powder
comprises particles characterized by a MMAD of less than about 5
microns.
21. A method for preparing a spray-dried polyene powder suitable
for oral inhalation to the lung, said method comprising: (i)
suspending a polyene antifungal compound in an aqueous solvent to
form a suspension, (ii) wet milling the suspension from (i) to form
a wet-milled suspension, and (iii) spray drying the wet milled
suspension to produce an inhaleable dry powder containing no more
than about 10% polyene degradation products and characterized by an
emitted dose greater than about 60%.
22. The method of claim 21, wherein said wet milling step comprises
homogenizing to form a homogenized suspension.
23. The method of claim 21, wherein said aqueous solvent in step
(i) further comprises an excipient.
24. The method of claim 21, further comprising mixing the wet
milled suspension in step (ii) with either a solid excipient or an
aqueous solution comprising an excipient prior to spray drying.
25. The method of claim 23 or claim 24, wherein said excipient is
selected from the group consisting of leucine, trileucine, and
buffers.
26. The method of claim 25, wherein said excipient is a buffer that
is sodium citrate or sodium phosphate.
27. The method of claim 21, further comprising the step of exposing
the powder to moisture either during or after said spray
drying.
28. The method of claim 27, comprising the step of exposing the
powder formed in step (iii) to moisture in an amount effective to
provide a powder having a moisture content ranging from about 4% to
about 10% by weight.
29. The method of claim 27, wherein said spray drying step further
comprises spray drying said suspension using a drying gas
comprising an amount of water sufficient to form a spray dried
powder having a moisture content ranging from about 4% to about 10%
by weight.
30. The method of claim 27, wherein said exposing step comprises
exposing or maintaining the powder at a relative humidity above
about 10%.
31. The method of any one of claims 27-30, wherein said exposing
step is effective to reduce the MMAD of the spray dried powder from
that of a spray dried powder prepared in the absence of said
exposing step.
32. The method of claim 21, wherein the concentration of said
polyene in the suspension in step (i) ranges from about 1 mg-mL to
about 100 mg/mL.
33. The method of claim 32, wherein the concentration of said
polyene in the suspension in step (i) ranges from about 5 mg-mL to
about 20 mg/mL.
34. The method of claim 21, further comprising the step of
aerosolizing the dry powder formed in step (iii).
35. The method of claim 27, further comprising, after said exposing
step, aerosolizing the dry powder formed in (iii).
36. The method of claim 21, wherein the inhaleable dry powder
formed in step (iii) is further characterized by an MMAD of less
than about 5 microns.
37. The method of claim 27, wherein the inhaleable dry powder,
after exposure to moisture, is further characterized by an MMAD of
less than about 5 microns.
38. The method of claim 37, wherein the inhaleable dry powder,
after exposure to moisture, is further characterized by an MMAD of
less than about 3.5 microns.
39. The method of claim 21, wherein said polyene is amphotericin or
nystatin.
40. A dry powder produced by the method of claim 1.
41. A dry powder produced by the method of claim 21.
42. A spray-dried powder composition suitable for oral inhalation
to the lung comprising a therapeutically effective amount of a
polyene antifungal compound, wherein the composition comprises no
more than about 10% polyene degradation products and is
characterized by an emitted dose greater than about 60%.
43. The powder composition of claim 42, containing no more than
about 5% polyene degradation products.
44. The powder composition of claim 42, wherein the powder
comprises particles having an MMAD of less than about 5
microns.
45. The powder composition of claim 44, wherein the powder
comprises particles having an MMAD of less than about 3.5
microns.
46. The powder composition of claim 42, which is
non-proteinaceous.
47. The powder composition of claim 42, wherein said polyene is
nystatin or amphotericin B.
48. The powder composition of claim 42, wherein said polyene is
non-encapsulated.
49. The powder composition of claim 48, wherein said polyene is
non-liposome or non-polymer encapsulated.
50. The powder composition of claim 42 substantially comprising
neat polyene.
51. The powder composition of claim 42, further comprising a
pharmaceutically acceptable excipient.
52. The powder composition of claim 51, wherein said excipient is
selected from the group consisting of buffers, leucine, and
trileucine.
53. The powder composition of claim 51, comprising at least about
30% by weight polyene.
54. The powder composition of claim 53, comprising at least about
50% by weight polyene.
55. The powder composition of claim 42, having a water content
greater than about 4% by weight.
56. The powder composition of claim 55, having a water content
ranging from about greater than 4% by weight to about 10% by
weight.
57. A spray-dried powder composition suitable for oral inhalation
to the lung comprising a therapeutically effective amount of a
polyene antifungal compound and a leucyl-containing excipient
comprising from 1 to 3 amino acid residues.
58. An aerosolized, spray-dried powder composition suitable for
oral inhalation to the lung comprising a therapeutically effective
amount of a polyene antifungal compound, wherein the composition
comprises no more than about 10% polyene degradation products and
is characterized by an emitted dose greater than about 60%.
59. A method for treating or preventing fungal infection in a
subject in need thereof, said method comprising administering to
said subject by oral inhalation a therapeutically effective amount
of a spray dried powder composition of claim 42 in aerosolized
form.
60. In a method for preparing an inhaleable spray-dried powder
comprising the steps of spray drying a solution or suspension
containing an active agent to form particles having a particular
MMAD, the improvement comprising exposing said powder either during
or after spray drying to moisture in an amount effective to reduce
the MMAD of the particles from that of the particles formed in the
absence of said exposing step.
Description
[0001] This application claims the benefit of priority of U.S.
Provisional Patent Application Ser. No. 60/257,613, the contents of
which is incorporated herein by reference in its entirety.
FIELD OF THE INVENTION
[0002] The present invention relates to spray-dried polyene
compositions, and to methods for making and administering such
compositions. In particular, the invention is directed to polyene
powder compositions which possess a number of notable features,
making them advantageous for oral inhalation to the lung for the
treatment and/or prophylaxis of pulmonary and systemic fungal
infections. The polyene is surprisingly stable (i.e., exhibits
minimal chemical degradation) upon spray-drying, and the resulting
powder possesses superior aerosol properties (low MMAD, excellent
dispersibility), even in the absence of stabilizing carriers or
excipients.
BACKGROUND OF THE INVENTION
[0003] Pulmonary fungal infections, which are associated with
significant levels of morbidity and mortality, represent a major
medical challenge. In recent years, the frequency and seriousness
of fungal infections has increased, due to increasing numbers of
organ transplantations, aggressive antineoplastic therapy regimens,
and patients suffering from immune diseases such as HIV. Fungal
infections of the lung, e.g., fungal pneumonia, allergic
bronchopulmonary aspergillosis and other infections caused by
Aspergillus, are typically treated by direct intracavitary
instillation, oral, intraperitoneal, or intrapleural
administration, or intravenous infusion of one or more antifungal
agents such as amphotericin B (St. Georgiev, V., Respiration,
59:291-302 (1992). Unfortunately, serious drawbacks exist with each
of these commonly employed routes of administration as described
more fully below.
[0004] Direct intracavitary instillation, an invasive procedure, is
usually accomplished by repeated transthoracic injections into the
cavity. Drawbacks of intracavitary administration can include poor
tolerance (the development of fever), risk of pheumothorax, and
relapse of infection in the cavity (Glimp, R A, et al., Arch Intern
Med, 143:303-308 (1983). In general, endobronchial administration
of antifungals has met with minimal success (Henderson, A H, et
al., Thorax, 23:519-523 (1968)). Oral formulations tend to be
absorbed very poorly from the gastrointestinal tract, and like
intravenous therapy are limited by associated dose-dependent drug
toxicity, which (i) limits the intravenous dose that can be
administered, and (ii) can result in unpleasant or even life
threatening complications such as nephrotoxicity and normochromic
anemia. Some of the disadvantages of intravenous therapy using
conventional antifungal formulations have been addressed by the
development of liposomal compositions such as ambisome (a liposomal
formulation of amphotericin B), which, when administered by
injection, does not display serious toxicity such as renal tubular
damage, and allows the administration of doses which exceed those
used in conventional formulations (Hay, R J in Recent Progress in
Antifungal Chemotherapy. New York, Marcel Dekker, 1992
(323-332)).
[0005] Oral or intravenously administered systemic antifungals for
treating respiratory infections suffer from an added
disadvantage--the uncertainty of drug penetration into the lung
tissue and infected secretions. This is important since effective
drug therapy for lower respiratory tract infections depends upon
not only the susceptibility of the infecting microorganisms, but
upon the attainment of effective antifungal concentrations in the
lung tissue and mucus. In an attempt to address this problem,
inhalation therapy using nebulizer-generated aerosols has been
investigated, using antifungals such as amphotericin B (Beyer, J.,
et al., Infection, 22:2, 143 (1994); Calvo, V., et al., Chest,
115:5 (1999); Dubois J., et al., Chest, 108:3, 750-753 (1995);
Diot, P. et al., Eur Respir J., 8:1263-1268 (1995)).
[0006] Aerosolized pharmaceutics for inhalation can be delivered in
a variety of different forms, including nebulized sprays,
pressurized powder formulations, and non-pressurized powder
formulations. Several drawbacks exist for both liquid and
pressurized formulations. Disadvantages of liquid formulations
include the chemical instability of certain active agents in
solution (polyenes in particular), the high potential for
microorganism contamination, and the use of cumbersome liquid
nebulizers. Pressurized powder formulations containing a compressed
gas or liquefied gas propellant have the disadvantages of employing
ozone depleting agents in the case of CFCs, or green house gases in
the case of HFCs. Further, liquid gas propellant typically cannot
accommodate the quantities of antifungal agent required to achieve
high levels of fungistatic/fungicidal activity locally at the site
of infection. Pressurized powder formulations can also exhibit a
high level of variability in the dose that is delivered to the
lungs, due the inability of patients to consistently coordinate the
firing of the inhaler to generate the aerosol with the appropriate
cycling of the inhalation. Achieving adequate solubilization or
suspension of antifungal agents such as the polyene, amphotericin
B, in the liquefied gas propellant can also be problematic.
[0007] Thus, in view of the problems noted above using conventional
antifungal therapies, it would be desirable to provide an
inhaleable, non-pressurized antifungal dry powder for localized
delivery to the lung, for both the treatment of pulmonary fungal
infections and for therapy of systemic fungous diseases. Inhaleable
dry powder formulations can provide high concentrations of
antifungal agent in the lung with negligible concentrations in the
blood and body tissues. Moreover, by utilizing a topical
administration route, most of the toxicities that are associated
with systemic antifungal agents (including nephrotoxicity,
convulsions, fever, and chills, among others) can be minimized or
avoided. Inhalation of antifungals using a dry powder inhaler
maximizes the convenience and speed of administration, and
overcomes the disadvantages of alternative inhalation therapies as
described above.
[0008] Unfortunately, the development of chemically stable dry
powders of an antifungal agent such as amphotericin B that also
possess the physical properties necessary for aerosolization (e.g.,
high dispersibilities which remain stable over time, appropriate
aerodynamic size) remains a technical challenge.
SUMMARY OF THE INVENTION
[0009] The present invention is directed to inhaleable, spray dried
powder formulations of polyene antibiotics. While polyenes such as
amphotericin are highly effective antifungal compounds, they also
possess very low solubilities in water and in conventional organic
solvents such as chloroform. Thus, formulation of these compounds
outside of dry mixing is extremely difficult. Although the
solubility of the polyene, amphotericin, can be increased under
extreme conditions of pH, such conditions typically lead to
significant levels of degradation of drug and are usually
considered undesirable for the formation of powders for direct
administration to the lung. Thus, the inventors were faced with the
challenge of trying to find conditions for spray drying the highly
insoluble drug, amphotericin, that (i) did not promote high levels
of degradation of drug, (ii) were economically practical, and (iii)
resulted in the formation of aerosolizable particles suitable for
inhalation. While finding a solution to one of these problems was
rather straightforward, arriving at a spray drying method in which
all of these factors were balanced to produce a chemically stable
and highly dispersible powder was a technical challenge.
[0010] In an effort to address these problems, the present
invention provides methods for spray drying polyene antifungal
agents that result in the formation of chemically stable yet highly
dispersible powders. That is to say, the antifungal powders of the
invention have excellent aerosol characteristics, such that they
are reproducibly prepared and can be efficiently administered by
inhalation to the lung, while exhibiting good chemical and physical
stability.
[0011] In one aspect, the present invention provides a method for
preparing a spray dried polyene, such as amphotericin B or
nystatin, for oral administration to the lung. The method includes
the steps of dissolving a polyene antifungal agent in an acidified
solvent and spray drying the polyene solution to form an inhaleable
powder containing no more than about 10% polyene degradation
products and characterized by an emitted dose of greater than
60%.
[0012] In one embodiment of this aspect of the invention, the
acidified solvent comprises an acidified alcohol such as methanol
or ethanol, aqueous methanol, or aqueous ethanol. In yet another
embodiment, the pH of the acidified solvent ranges from about 3.5
to about 5. In a preferred embodiment of the invention, the polyene
is dissolved in the acidified solvent to an extent greater than
about 1 mg/mL or preferably to an extent greater than about 2-3
mgs/mL.
[0013] In yet another embodiment of the method, the acidic
polyene-containing solution is maintained at a temperature below
25.degree. C. prior to and/or during spray drying. In a preferred
embodiment, the acidic polyene-containing solution is maintained at
a temperature below 8.degree. C. and even more preferably below
0.degree. C. prior to and/or during spray drying.
[0014] In yet another embodiment of the method, the
polyene-containing solution to be spray dried contains at least
about 50% polyene by weight based upon the total dissolved solids
content of the solution.
[0015] In yet another embodiment of the method, the
polyene-containing solution is absent lipid or polymeric
encapsulating agents.
[0016] In yet another aspect, provided is a method for preparing a
spray dried polyene powder for oral inhalation to the lung in which
a polyene antifungal compound is suspended in an aqueous solvent to
form a suspension, which is then wet milled, and spray dried. The
resulting inhaleable powder contains no more than about 10% polyene
degradation products (and typically less than that) and is
characterized by an emitted dose greater than about 60%.
[0017] In one particular embodiment of this aspect of the
invention, the antifungal compound is homogenized to form a
homogenized suspension prior to spray drying.
[0018] In yet another embodiment of this aspect of the invention,
the spray dried powder is exposed to moisture prior to packaging
(i.e., either during or post spray drying) to decrease or maintain
the aerodynamic diameter of the particles preferably below about 5
microns.
[0019] In one particular embodiment, the powder is spray dried
using a wet drying gas, such as wet air, argon or nitrogen.
Alternatively, the spray dried powder is exposed, post spray
drying, to an amount of moisture sufficient to form a powder having
a moisture content ranging from about 3% to about 10% by
weight.
[0020] In a preferred embodiment, the spray dried powder is exposed
or maintained at a relative humidity greater than about 5% prior to
packaging.
[0021] In yet another embodiment, the spray dried powder is
aerosolized in a current of air.
[0022] Also provided are polyene dry powders produced by either of
the above methods.
[0023] In yet another aspect, the invention provides a spray-dried
powder composition suitable for oral inhalation to the lung
comprising a therapeutically effective amount of a polyene
antifungal compound, where the composition comprises no more than
about 10% polyene degradation products and is characterized by an
emitted dose greater than about 60%.
[0024] In one particular embodiment of the invention, the spray
dried powder is composed of particles having an MMAD of less than
about 5 microns, and preferably less than about 3.5 microns.
[0025] In yet another embodiment, the spray dried powder is
non-proteinaceous.
[0026] In yet another embodiment of this aspect of the invention,
the polyene is non-encapsulated.
[0027] In one particular embodiment, the spray dried polyene powder
substantially comprises neat polyene antifungal compound (i.e.,
lacks additional additives and/or excipients). In an alternative
embodiment, the spray dried powder composition further comprises a
pharmaceutically acceptable excipient. Preferred excipients are
buffers, leucine and trileucine.
[0028] In yet another embodiment, the spray dry powder is composed
of at least about 30% by weight polyene.
[0029] In yet another aspect, provided herein in a method for
preparing an inhaleable spray dried powder which includes the steps
of spray drying a solution or suspension containing an active agent
to form particles having a particular MMAD, is an improvement
wherein the powder is exposed, either during or after spray drying,
to moisture in an amount effective to reduce the MMAD of the
particles over the MMAD of the particles formed in the absence of
such an exposing step. This method is particularly applicable to
powders comprising one or more molecular components having a large
percentage of water binding sites.
[0030] The invention further encompasses aerosolized powder
compositions as described above.
[0031] The invention further encompasses a method for treating or
preventing fungal infection in a mammalian subject in need thereof
by administering to the subject by oral inhalation a spray-dried
polyene antifungal powder as described herein.
[0032] These and other objects and features of the invention will
become more fully apparent when the following detailed description
is read in conjunction with the accompanying figures and
examples.
BRIEF DESCRIPTION OF THE FIGURES
[0033] FIG. 1. The FIGURE is a plot of MMAD as a function of
moisture content of spray dried, neat Amphotericin B powders, the
details of which are provided in Example 8. Solid circles represent
powders in which moisture content was varied by exposure to
controlled environments of different relative humidity (6-40%) and
empty triangles represent powders in which moisture content was
varied by changing the spray drying conditions.
DETAILED DESCRIPTION OF THE INVENTION
I. Definitions
[0034] The following terms as used herein have the meanings
indicated.
[0035] "Antifungal compound" refers to any compound or its
pharmaceutically acceptable salt having fungistatic and/or
fungicidal properties.
[0036] "Polyene" refers to an organic compound containing a series
of double bonds that are typically, but not necessarily,
conjugated.
[0037] "Pharmaceutically acceptable salt" includes, but is not
limited to, salts prepared with inorganic acids, such as chloride,
sulfate, phosphate, diphosphate, hydrobromide, and nitrate salts,
or salts prepared with an organic acid, such as malate, maleate,
fumarate, tartrate, succinate, ethylsuccinate, citrate, acetate,
lactate, methanesulfonate, benzoate, ascorbate,
para-toluenesulfonate, palmoate, salicylate and stearate, as well
as estolate, gluceptate and lactobionate salts. Similarly salts
containing pharmaceutically acceptable cations include, but are not
limited to, lithium, sodium, potassium, barium, calcium, aluminum,
and ammonium (including substituted ammonium). Pharmaceutically
acceptable salts of antifungal compounds have the same general
pharmacological properties as the parent compound from which they
are derived.
[0038] A "pharmaceutically acceptable ester" of a carboxylic
acid-containing antifungal compound is a hydrolyzable ester having
the same general pharmacological properties as the acid from which
it is derived. Such esters include unsubstituted and substituted
alkyl, aryl and phosphoryl esters. Non-limiting examples of
pharmaceutically-acceptable esters include, for example, isopropyl,
tertiarybutyl, 2-chloroethyl, 2,2,2-trichloroethyl,
2,2,2-trifluoroethyl, p-toluenesulfonylethyl, glycyl, sarcosyl,
benzyl, phenyl, 1,2-hexanoylglyceryl, para-nitrophenyl, 2,2
dimethyl-1,3-dioxolene-4-methyl, isopentenyl, o-carbomethoxyphenyl,
piraloyloxymethylsalicylyl, diethylamidophosphoryl,
pivaloyloxymethyl, acyloxymethyl, propionyloxymethyl,
isobutyryloxymethyl, dodecyl, octadecyl, and
isopropyloxymethyl.
[0039] "Relative pulmonary bioavailability" is the percentage of an
antifungal dose (for the treatment of systemic fungal disease) that
is deposited in the lungs that is absorbed and enters the blood of
a mammal relative to the percent that is absorbed into the blood
from an intravenous injection site. Representative model systems
for determining relative pulmonary bioavailabilities include rat,
rabbit, and monkey. The antifungal compositions of the invention
are, in one respect, characterized by a relative pulmonary
bioavailability of at least about 3% in plasma or blood, with
relative pulmonary bioavailabilities generally ranging from about
5% to about 20% or greater. Relative pulmonary bioavailability may
be estimated by measuring absorption from direct intratracheal
administration or by inhalation of an antifungal composition.
[0040] "Amino acid" refers to any compound containing both an amino
group and a carboxylic acid group, and is meant to encompass
pharmaceutically acceptable salts thereof. Although the amino group
most commonly occurs at the position adjacent to the carboxy
function, the amino group may be positioned at any location within
the molecule. The amino acid may also contain additional functional
groups, such as amino, thio, carboxyl, carboxamide, imidazole, etc.
The amino acid may be synthetic or naturally occurring, and may be
used in either its racemic, or optically active (D-, or L-) forms,
for example, as a single optically active enantiomer or as any
combination or ratio of enantiomers.
[0041] "Enhancer" refers to a compound that enhances the absorption
of an antifungal compound through mucosal membranes, e.g., of the
lung.
[0042] "Dry powder" refers to a powdered composition that contains
finely dispersed solid particles that are capable of (i) being
readily dispersed in an inhalation device and (ii) inhaled by a
subject so that a portion of the particles reach the lungs to
permit penetration into the alveoli. Such a powder is considered to
be "respirable" or suitable for pulmonary delivery. Unless
otherwise stated, a "dry powder composition for delivery to the
deep lung" is one that, when aerosolized, is administered as dry
powder particles. A dry powder in accordance with the invention is
preferably a non-liposomal powder. Additionally, a dry powder of
the invention is one that is preferably absent polymeric
encapsulating agents or polymeric matrices.
[0043] "Oligopeptides" are peptides comprising two to ten amino
acid residues (dimers to decamers).
[0044] "Peptide" as used herein is meant to encompass both
naturally occurring and artificially constructed polypeptides in
which individual amino acid units are linked together through the
standard peptide amide bond (the carboxyl group of one and the
amino group of another) and having a molecular weight between about
1,000 and about 6000.
[0045] "Protein" refers to a particular class of polypeptides
having molecular weights ranging from about 6000 to more than
1,000,000.
[0046] A "leucyl-containing excipient comprising from 1 to 5 amino
acid residues" includes the amino acid leucine, and oligomers
composed of from 2 to 5 amino acid residues, one or more of which
is leucine.
[0047] A powder comprising essentially "neat antifungal polyene" is
one substantially lacking any other excipients or additives besides
the antifungal compound, i.e., contains less than about 3% by
weight non-antifungal component(s), preferably less than about 2%
by weight non-antifungal component, more preferably less than 1% by
weight antifungal component, and in some cases comprises 100%
antifungal compound.
[0048] "Emitted dose" provides an indication of the delivery of a
dry powder from the mouthpiece of a suitable inhaler device after a
firing or dispersion event. More specifically, the ED is a measure
of the percentage of powder which is drawn out of a unit dose
package and which exits the mouthpiece of an inhaler device. The ED
is defined as the ratio of the dose delivered by an inhaler device
to the nominal dose (i.e., the mass of powder per unit dose placed
into a suitable inhaler device prior to firing). The ED is an
experimentally-determined parameter, and is typically determined
in-vitro using a device set up which mimics patient dosing. To
determine a ED value, a nominal dose of dry powder, typically in
unit dose form, is placed into a suitable dry powder inhaler (such
as that described in U.S. Pat. No. 5,785,049, assigned to Inhale
Therapeutic Systems, Inc.) which is then actuated, dispersing the
powder. The resulting aerosol cloud is then drawn by vacuum from
the device, where it is captured on a tared filter attached to the
device mouthpiece. The amount of powder that reaches the filter
constitutes the delivered dose. For example, for a 5 mg, dry
powder-containing dosage form placed into an inhalation device, if
dispersion of the powder results in the recovery of 4 mg of powder
on a tared filter as described above, then the ED for the dry
powder composition is: 4 mg (delivered dose)/5 mg (nominal
dose).times.100=80%. For homogenous powders, ED values provide an
indication of the delivery of therapeutic moiety (i.e., antifungal
compound) from an inhaler device after firing. Similarly for MDI
and nebulizer dosage forms, the ED corresponds to the percentage of
drug which is drawn from a dosage form and which exits the
mouthpiece of an inhaler device.
[0049] "Fine particle fraction" or "FPF" provides a measure of
aerosolized powder delivery efficiency from a unit dosage form
(e.g., a blister pack) to the deep lung, and is determined
experimentally using a short stack Anderson cascade impactor
operated at a vacuum of 28.3 liters per minute. The FPF is defined
as the total mass, in milligrams, of aerosolized powder having a
particle size less than 3.3 micrometers, relative to the mass of
powder contained in a unit dosage form, in milligrams, and
expressed as a percentage.
FPF = total aerosolized powder mass less than 3.3 .mu.m ( mg ) unit
dosage form fill mass ##EQU00001##
[0050] A "dispersible" powder is one having an ED value of at least
about 30%, preferably at least about 35%, more preferably at least
about 40%, and most preferably at least about 50%. Powders of the
present invention are highly disperisible, having ED values of at
least 60% or greater. Dispersibility, as used herein, refers to the
dispersibility of a dry powder in a gas stream (e.g., a stream of
air) unless otherwise indicated.
[0051] A dry powder composition suitable for "inhalation therapy",
is one which, when aerosolized, may be (i) readily dispersed in an
inhalation delivery device, and (ii) inspired through either the
mouth by a mammalian subject so that at least a portion of the
particles are absorbed into the lung.
[0052] A composition suitable for "oral pulmonary administration"
comprises particles at least a portion of which, when delivered via
inhalation by the mouth, reach the tissues of the lung, including
the deep lung.
[0053] "Mass median diameter" or "MMD" is a measure of mean
particle size, since the powders of the invention are generally
polydisperse (i.e., consist of a range of particle sizes). MMD
values as reported herein are determined by laser diffraction,
although any number of commonly employed techniques can be used for
measuring mean particle size (e.g., centrifugal sedimentation,
electron microscopy, light scattering).
[0054] "Mass median aerodynamic diameter" or "MMAD" is a measure of
the aerodynamic size of a dispersed particle. The aerodynamic
diameter is used to describe an aerosolized powder in terms of its
settling behavior, and is the diameter of a unit density sphere
having the same settling velocity, generally in air, as the
particle. The aerodynamic diameter encompasses particle shape,
density and physical size of a particle. As used herein, MMAD
refers to the midpoint or median of the aerodynamic particle size
distribution of an aerosolized powder determined by cascade
impaction.
[0055] "Pharmaceutically acceptable excipient or carrier" refers to
an excipient that may be included in the particles of the invention
and taken into the lungs in association with the particles with no
significant adverse toxicological side effects (e.g., toxicity,
irritation, and allergic response) to the subject, and particularly
to the lungs of the subject.
[0056] "Pharmacologically effective amount" or "physiologically
effective amount" of an antifungal powder is the amount of an
antifungal compound present in a particulate dry powder composition
as described herein that is needed to provide a therapeutic or
prophylactic level of antifungal agent, either in the bloodstream
or at the infected tissue site (depending upon the fungus to be
treated) when such composition is administered by inhalation over a
particular duration of time. The precise amount will depend upon
numerous factors, e.g., the particular antifungal(s) contained in
the powder, the potency of the antifungal compound employed, the
condition being treated, the delivery device employed, the physical
characteristics of the powder, intended patient use (e.g., the
number of doses administered per day), and patient considerations,
and can readily be determined by one skilled in the art, based upon
the information provided herein. Recommended dosage ranges will be
described in greater detail below.
[0057] "Bulk density" refers to the density of a powder prior to
compaction (i.e., the density of an uncompressed powder), and is
typically measured by a well-known USP method.
[0058] The "extent of degradation" of a polyene is the percentage
of polyene contained in the dry powder composition determined to be
chemically modified from the intact starting material, as
determined by a suitable chemical assay (e.g., NMR, HPLC, etc.);
100% of the polyene remaining chemically intact during the spray
drying process represents an extent of degradation of 0%.
Inhaleable Antifungal Compositions
[0059] The present invention provides compositions for the oral
pulmonary delivery of polyene antifungal compounds. These
compositions overcome many of the problems and inconveniences
encountered heretofore in administering antifungals, and
particularly polyene antifungals, by other routes (e.g., poor
absorption from the gastrointestinal tract, severe toxic side
effects, the requirement for hospitalization during intravenous
therapy, etc.) particularly for treating and/or preventing systemic
and/or pulmonary fungal diseases. The powder compositions described
herein (i) are readily dispersed by dry powder delivery devices
(i.e, demonstrate superior aerosol properties), (ii) exhibit good
physical stability during powder manufacture, processing, and
storage, and (iii) are reproducibly prepared with minimal
degradation of polyene. Inhaleable polyene antifungal compositions
in accordance with the invention are preferably dry powders (i.e.,
for use in dry powder inhalers (DPIs).
[0060] The dry powder compositions according to the present
invention generally include one or more antifungal compounds, one
being a polyene, and optionally a pharmaceutically acceptable
excipient. Dry powders composed of neat polyene antifungal agent
(i.e., respirable powders composed of one or more polyene
antifungal compounds and essentially lacking any additional
excipients or additives) and demonstrating good aerosol properties
have been prepared (see, e.g., Examples 1, 2, 5, 6, and 8). The
challenge facing the inventors was to balance the factors
influencing chemical degradation of the polyene antifungal compound
during liquid formulation preparation and spray-drying (polyenes
are prone to chemical degradation, especially at elevated
temperatures) with those affecting aerodynamic particle size and
particle dispersibility. Surprisingly, the inventors have prepared
powders in which all of these factors are optimized--extent of
degradation of polyene, high dispersibilities, and small
aerodynamic particle sizes. (See the Examples). Moreover, it is
unusual to prepare powders characterized by both high emitted dose
values and low aerodynamic particle sizes, since these factors
often work in opposing directions. That is to say, particles having
good dispersibilities are often characterized by large aerodynamic
sizes, since larger primary particles, which exhibit less cohesive
force, tend to agglomerate less and thus disperse better.
[0061] Moreover, the preparation of antifungal powders having
superior aerosol properties, as characterized by high ED values and
small aerodynamic sizes, in the absence of additives or dispersing
agents for improving aerosol properties, is particularly surprising
(Yamashita, C., et al., Respiratory Drug Delivery VI, p. 483-485
(1998)), particularly for polyene fungal powders where the
antifungal powder is non-proteinaceous.
[0062] Specific components of antifungal dry powders suitable for
delivery to the lung will now be described.
A. Antifungal Compounds
[0063] Antifungal compounds for use in the dry powders of the
invention are those having fungistatic or fungicidal properties
when administered by oral inhalation to the lung. Preferred
antifungal compounds are those characterized by a polyene chemical
structure, such as amphotericin B, nystatin, hamycin, natamycin,
pimaricin, and ambruticin, and pharmaceutically acceptable
derivatives and salts thereof. Preferred for use in the present
invention are amphotericin B and nystatin.
[0064] Other suitable antifungal compounds for use in the powders
of the invention include acrisocin, aminacrine, anthralin,
benanomicin A, benzoic acid, butylparaben, calcium unidecyleneate,
candicidin, ciclopirox olamine, cilofungin, clioquinol,
clotrimazole, ecaonazole, flucanazole, flucytosine, gentian violet,
griseofulvin, haloprogin, hamycin, ichthammol, iodine,
itraconazole, ketoconazole, miconazole, nikkomycin Z, potassium
iodide, potassium permanganate, pradimicin A, propylparaben,
resorcinol, sodium benzoate, sodium propionate, sulconazole,
terconazole, tolnaftate, triacetin, unidecyleneic acid,
monocyte-macrophage colony stimulating factor (M-CSF) and zinc
unidecylenate. Of these, preferred are candicidin, clotrimazole,
econazole, fluconazole, griseofulvin, hamycin, itraconazole,
ketoconazole, miconazole, sulconazole, terconazole, and
tolnaftate.
[0065] In one particular embodiment of the invention, the
antifungal dry powder comprises an antifungal other than an
azole-containing antifungal agent (e.g., the powder does not
contain azoles such as clotrimazole, econazole, fluconazole,
itraconazole, ketoconazole, or miconazole).
[0066] In yet another embodiment of the invention, the components
(i.e., the antifungal compound and optional carriers/excipients)
comprise a homogeneous spray-dried powder rather than an admixture
of dry powder or particulate components. In yet another embodiment,
the antifungal compound (e.g., polyene) is not protected by
sequestration within liposomes or by a polymeric encapsulating
agent, or by a discrete coating layer, but rather, is directly
exposed to the harsh conditions of spray-drying during formation of
the spray dried antifungal powder. That is to say, the compositions
of the present invention typically are those in which the polyene
is non-encapsulated. Additionally, the compositions of the
invention are typically absent substantial amounts of polymer
additives, such as those employed in sustained release-type
formulations or non-biological polymers. For example,
representative formulations will typically contain no more than
about 10% by weight polymer, more preferably no more than about 5%
by weight polymer, and even more preferably no more than about 3%
polymer additive.
[0067] Compositions in accordance with the invention comprise a
therapeutically effective amount of an antifungal compound,
preferably a polyene such as amphotericin B. The amount of
antifungal contained in the powder particles will be that amount
necessary to pulmonarily deliver a therapeutically or
prophylactically effective amount (i.e., a fungistatic and/or
fungicidal amount) of the antifungal compound over the course of a
particular dosing regimen at the site of infection. In practice,
this will vary widely depending upon the particular antifungal
compound and its corresponding potency, the particular fungal
condition to be treated or prevented and its severity, the intended
patient population, and similar considerations, and can be readily
determined by one skilled in the art. A notable advantage of the
respirable compositions described herein is their ability, in the
case of pulmonary fungal infections, to be delivered directly to
the site of infection--the lung. This allows for the administering
of lower overall doses of antifungal than are typically
administered orally or intravenously to achieve the same level of
fungistatic or fungicidal action, which can also reduce the
potential for adverse side effects in the patient. Moreover, due to
the highly dispersible nature of the antifungal powders of the
invention, losses to the inhalation device are minimized, meaning
that more of the powder dose is actually delivered to the
patient.
[0068] Dry powder formulations will generally comprise from about
0.1% to 100% by weight of a polyene, preferably from about 5% to
about 100% by weight polyene, more preferably from about 20% to
about 100% by weight polyene, and most preferably will comprise
greater than about 30% by weight polyene. Particular powders of the
invention are those comprising one of the following percentages by
weight polyene antifungal compound: 1%, 3%, 5%, 10%, 15%, 20%, 25%,
30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%,
95%, or 100%. Particularly preferred are dry powder compositions
containing at least about 30% to 50% or more polyene. The
antifungal powders are particularly useful for antifungals that are
administered to the lung in doses of from 1 mg/day to 200 mg/day,
preferably 5 mg/day to 100 mg/day.
B. Excipients
[0069] In the compositions of the invention, an antifungal
compound, preferably a polyene, is optionally but not necessarily
combined with one or more pharmaceutical excipients that are
suitable for respiratory and pulmonary administration. Such
excipients may serve simply as bulking agents when it is desired to
reduce the active agent concentration in the powder that is being
delivered to a patient. Preferred are excipients that can also
serve in one or more of the following capacities: (i) improve the
dispersibility and aerosol performance of a powder within a powder
dispersion device in order to provide more efficient and
reproducible delivery of the compound, (ii) improve the handling
characteristics of the powder (e.g., flowability and consistency)
to facilitate manufacturing and filling into unit dosage forms, and
(iii) improve chemical and/or physical stability. In particular,
the excipient materials can often function to optimize the residual
moisture content, hinder excessive moisture uptake, influence
particle size, the degree of aggregation, particle surface
properties (i.e., rugosity), ease of inhalation, and targeting of
the resultant particles to the lung tissue including the deep
lung.
[0070] Alternatively, and in one preferred embodiment of the
present invention, the antifungal, preferably a polyene, may be
formulated in an essentially neat form, wherein the composition
contains antifungal particles within the requisite size range and
substantially free from other biologically active components,
pharmaceutical excipients, and the like.
[0071] Alternatively, the antifungal may be formulated in an
essentially neat form but complexed with a deoxycholate salt, by
adding sodium deoxycholate or another deoxycholate salt to form an
aqueous solution, wherein the composition contains antifungal
particles within the requisite size range and substantially free
from other biologically active components, pharmaceutical
excipients, and the like. The sodium deoxycholate enhances
solubilization of the antifungal agent, amphotericin B in
particular, in water at a pH that is especially conducive to good
chemical stability and yet, surprisingly, produces particularly
dispersible powders (see, for example, Example 2).
[0072] Pharmaceutical excipients and additives useful in the
present composition include but are not limited to proteins,
peptides, amino acids (which are preferably non-acylated or
non-sulfonated), lipids (which, if employed are typically not
encapsulating agents, i.e., liposomes), and carbohydrates (e.g.,
sugars, including monosaccharides, disaccharides, trisaccharides,
tetrasaccharides, and oligosaccharides; derivatized sugars such as
alditols, aldonic acids, esterified sugars and the like; and
polysaccharides), which may be present singly or in combination.
Also preferred are excipients having glass transition temperatures
(Tg), above about 35.degree. C., preferably above about 45.degree.
C., more preferably above about 55.degree. C. Illustrative
excipients suitable for use in the compositions described herein
include those described in Inhale Therapeutic Systems'
International Patent Application No. WO 98/16207.
[0073] Exemplary protein excipients include serum albumin such as
human serum albumin (HSA), recombinant human albumin (rHA),
gelatin, casein, and the like. Polypeptides and proteins suitable
for use in the dry powder composition of the invention are provided
in Inhale Therapeutic Systems' International Patent Publication No.
WO96/32096. HSA is a preferred proteinaceous excipient, and has
been shown to increase the dispersibility of dry powders for
aerosolized delivery to the lungs (WO 96/32096, ibid). However, as
shown by the Examples contained herein, the powders of the
invention display good dispersibilities, even in the absence of
dispersibility-enhancing agents such as HSA and the like.
[0074] Representative amino acid/polypeptide components, which may
optionally function in a buffering capacity, include alanine,
glycine, arginine, betaine, histidine, glutamic acid, aspartic
acid, cysteine, lysine, leucine, isoleucine, valine, methionine,
phenylalanine, aspartame, threonine, tyrosine, tryptophan and the
like. Preferred are amino acids and peptides that can also function
as dispersibility-enhancing agents. Amino acids falling into this
category include hydrophobic amino acids such as leucine (leu),
valine (val), isoleucine (isoleu), norleucine, tryptophan (try)
alanine (ala), methionine (met), phenylalanine (phe), tyrosine
(tyr), histidine (his), and proline (pro). One particularly
preferred amino acid is the amino acid, leucine. Leucine, when used
in the formulations described herein includes D-leucine, L-leucine,
racemic leucine, and combinations of D- and L-leucine at any ratio.
Dispersibility enhancing peptides for use in the invention include
dimers, trimers, tetramers, and pentamers composed of hydrophobic
amino acid components such as those described above, e.g.,
di-leucine and tri-leucine. Further examples include di-valine,
di-isoleucine, di-tryptophan, di-alanine, and the like, tri-valine,
tri-isoleucine, tri-tryptophan, etc.; mixed di- and tri-peptides,
such as leu-val, isoleu-leu, try-ala, leu-try, etc., and
leu-val-leu, val-isoleu-try, ala-leu-val, and the like, and
homo-tetramers or pentamers such as tetra-alanine and
penta-alanine. Preferred are amino acids and oligomers containing
from 1-5 amino acid residues, and more preferably containing 1-3
amino acid residues and containing at least one (i.e., one or more)
leucyl-residue. Illustrative dimers and trimers for use in the
antifungal compositions of the invention are provided in
International Patent Application No. PCT/US00/09785, entitled, "Dry
Powder Compositions Having Improved Dispersivity", the disclosure
of which is incorporated herein by reference in its entirety.
[0075] Preferred are compositions comprising at least 10% by weight
of a leucyl-containing excipient, e.g., leucine, dileucine or
trileucine, and more preferably at least 25% to 30% by weight of a
leucyl-containing excipient as described above. Representative
powder compositions in accordance with the invention comprise one
of the following percentages by weight excipient, preferably a
leucyl-containing excipient such as leucine, dileucine or
trileucine: 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%,
60%, 65%, 70%, 75%, or 80% or more excipient. Representative
formulations are provided in the Examples.
[0076] Carbohydrate excipients suitable for use in the invention
include, for example, monosaccharides such as fructose, maltose,
galactose, glucose, D-mannose, sorbose, and the like;
disaccharides, such as lactose, sucrose, trehalose, cellobiose, and
the like; trisaccharides such as melezitose and raffinose;
polysaccharides, maltodextrins, dextrans, starches, and the like;
and alditols, such as mannitol, xylitol, maltitol, lactitol,
xylitol, sorbitol (glucitol), myoinositol and the like. Preferred
carbohydrate excipients for use in the present invention include
mannitol, trehalose, and raffinose.
[0077] The dry powder compositions may also include a buffer or a
pH adjusting agent; typically, the buffer is a salt prepared from
an organic acid or base. Representative buffers include organic
acid salts such as salts of citric acid, ascorbic acid, gluconic
acid, carbonic acid, tartaric acid, succinic acid, acetic acid;
phthalic acid, Tris, and tromethamine hydrochloride. Commonly used
inorganic acids/buffers include hydrochloric acid, sulfuric acid,
boric acid, carbonic acid and phosphoric acid. Preferred buffers
for use in the compositions of the invention are citrate and
phosphate buffer.
[0078] Additionally, the formulations of the invention may include
small amounts of polymeric excipients/additives such as
polyvinylpyrrolidones, derivatized celluloses, such as
hydroxymethylcellulose, hydroxyethylcellulose, or
hydroxypropylmethylcellulose, Ficolls (a polymeric sugar),
hydroxyethylstarch, dextrates (e.g., cyclodextrins, such as
2-hydroxypropyl-.beta.-cyclodextrin and
sulfobutylether-.alpha.-cyclodextrin), polyethylene glycols,
polyamino acids (e.g., polyleucine, polyglutamic acid), pectin,
generally where such polymers are present as powder additives
rather than as encapsulating or coating agents or as components of
a polymeric matrix.
[0079] The composition of the invention may also optionally contain
flavoring agents, salts (e.g., sodium chloride), sweeteners,
antioxidants, antistatic agents, surfactants (e.g., polysorbates
such as "TWEEN 20" and "TWEEN 80"), lecithin, oleic acid,
benzalkonium chloride, sorbitan esters, lipids (e.g.,
phospholipids, fatty acids), steroids (e.g., cholesterol), and
chelating agents (e.g., EDTA). Other pharmaceutical excipients
and/or additives suitable for use in the compositions of the
invention are listed in "Remington: The Science & Practice of
Pharmacy", 19.sup.th ed., Williams & Williams, (1995), and in
the "Physician's Desk Reference", 52.sup.nd ed., Medical Economics,
Montvale, N.J. (1998), and "Handbook of Pharmaceutical Excipients",
(3rd Ed.), Vol. 3, Arthur H. Kibbe (Ed.), Ainley Wade, Paul J.
Weller (1999), the disclosures of which are herein incorporated by
reference.
[0080] The spray dried solid compositions in accordance with the
invention may be crystalline, amorphous (i.e., glassy), or a
mixture of both forms. Preferred are solid compositions that,
irrespective of their percent crystallinity, are stable with
respect to this percentage over time.
III. Preparing the Antifungal Formulations
[0081] Dry powder antifungal formulations of the invention are
preferably prepared by spray drying. In general, spray drying is a
process which combines a highly dispersed liquid and a sufficient
volume of a hot gas to produce evaporation and drying of the liquid
droplets to produce a powder. The preparation or feedstock can be a
solution, suspension, slurry, or colloidal dispersion that is
atomizable. Spray drying of an antifungal formulation is carried
out, for example, as described generally in the Spray Drying
Handbook, 5.sup.th ed., (1991), in European Patent Application EP
520 748 A1, in Inhale Therapeutics Systems' International Patent
Publications, WO 97/41833 and WO 96/32149, or as described in
International Patent Publication WO 99/16419, the contents of which
are incorporated herein by reference.
[0082] In attempting to prepare a chemically stable, dispersible
polyene dry powder for pulmonary administration, the inventors
arrived at two processing approaches that (i) minimized the extent
of degradation of polyene antibiotic, (ii) maintained a reasonable
concentration of polyene in the pre-spray dried liquid composition
(e.g., greater than about 1 mg/ml), and (iii) resulted in
dispersible powders.
[0083] In one approach, which typically provides a uniform
distribution of formulation components in the resulting,
spray-dried particles (meaning that each of the particles in the
final spray-dried formulation possesses substantially the same
chemical composition and distribution of components within the
particle), the feedstock comprises a polyene-containing solution.
Preferred solvents are water, alcohols such as methanol or ethanol,
and combinations thereof.
[0084] The challenge in utilizing this approach was to find a
solvent or solvent system in which the polyene was both reasonably
soluble (to an extent greater than 1 mg/mL) and relatively stable
(i.e., exhibited less than about 10% degradation upon dissolution
and spray drying). In the case of amphotericin B, which is
essentially insoluble in water except at extremes of pH (e.g., pHs
less than 3 and greater than 10), its solubility in water at
neutral pHs can be significantly enhanced by complexation with a
desoxycholate salt such as sodium desoxycholate or by adjustment of
pH.
[0085] Utilizing the first approach, the polyene antifungal is
first dissolved in water, optionally containing a physiologically
acceptable buffer and/or complexing agent and/or acid or base to
adjust the pH, as described above. The pH range of the resulting
solution is preferably between about 4 and 10. The aqueous
formulation may optionally contain additional water-miscible
solvents, such as acetone, alcohols and the like as described
above.
[0086] In yet another embodiment of the invention wherein the
antifungal compound is spray dried as a solution rather than as a
suspension, the antifungal compound (e.g., amphotericin B or
nystatin) is dissolved in acidified alcohol. Representative
alcohols are lower alcohols such as methanol, ethanol, propanol,
isopropanol, and the like. One preferred alcohol is methanol. As
shown in Example 4, extremes of pH, while improving the solubility
of the representative polyene, amphotericin B, were deleterious to
the chemical integrity of the drug, as shown in Table 6. Thus, the
challenge was to optimize both solubility and chemical stability of
the polyene. After extensive experimentation, an optimum range of
pHs from about 3 to 6, preferably from about 3.5 to 6, even more
preferably from about 3 to 5, and even more preferably from about 4
to 5, was determined for spray drying the polyene solutions of the
invention (Examples 4-6). One particularly preferred pH range is
from about 4.4 to 4.8, with pHs from about 4.8 to 6, and more
preferably from about 4.8 to 5 being most preferred.
[0087] In further exploring ways to further improve the chemical
stability of the polyene solutions, it was discovered unexpectedly
that, for the polyene solutions examined, temperature had very
little effect on solubility while having a profound effect on the
rate of degradation of polyene. Thus, it was determined that low
temperatures, e.g., below ambient or 25.degree. C., preferably
below about 8.degree. C., and even more preferably at 0.degree. C.
or below, significantly improved the chemical stability of the
solutions while not adversely impacting or significantly decreasing
the solubility of the polyene in the solvent employed. In this
respect, maintenance of the polyene solution at low temperatures
such as those described above prior to spray drying (i.e., during
and/or after dissolution) and/or as the feed solution, is effective
to improve the chemical stability of the polyene to thereby form
spray dried powders. Utilization of such conditions is typically
effective to reduce the extent of degradation of the polyene to
less than about 10%, and even more preferably less than about
5%.
[0088] Thus, particularly preferred conditions for spray drying a
polyene solution that are effective to produce solubilities of
polyene greater than about 1 mg/ml, more preferably greater than
about 2 mg/ml, and even more preferably greater than about 3 mg/ml
and maintain an extent of degradation of polyene of less than about
10%, are the utilization of acidified solvents such as methanol or
ethanol or aqueous combinations thereof at pHs ranging from about
3.5 to 5, while optionally utilizing low temperature conditions as
described herein for forming, and/or maintaining, and/or spray
drying such solutions (Example 7). Additionally, powders thus
formed were shown to possess good dispersibilities and aerodynamic
diameters (Example 7).
[0089] In a second preferred approach for preparing the spray dried
powders of the invention, a suspension of polyene is spray dried.
In this approach, the polyene (e.g., amphotericin B), which, as
supplied by the vendor, generally possesses a median particle size
of from about 8 to 13 microns, is first suspended in an aqueous
solvent such as water and subjected to wet milling. The wet milling
process is effective to reduce the particle size of the polyene,
typically to less than about 5 microns, and preferably to less than
about 3 microns. Most typically, drug particles (and any optional
undissolved excipient solids) are reduced to a size of about 1
micron or less during the process. Particle size reduction, and in
particular, wet milling, is employed in the present process since,
in the absence of such step, the spray dried particles will
typically possess particle sizes that are too big for effective
administration to the lung as a dry powder. Wet milling processes
that may be employed include homogenization (e.g., using a
pressurized spray type or ultrasonic homogenizer) or
microfluidization.
[0090] Suspensions will generally contain about 1 mg/mL to about
100 mg/mL polyene, preferably from about 5 to about 100 mg/mL
polyene, and even more preferably from about 5 to 20 mg/mL polyene.
The wet milling step is effective to decrease both the particle
size and distribution; the number of passes will typically range
from about 1 to 10, although a plateau or leveling off of particle
size reduction is typically achieved after about 3-5 passes.
Surprisingly, no detectable degradation of polyene was observed
during wet milling of the exemplary amphotericin suspensions of the
invention. One exemplary method of wet milling for use in the
method is homogenization (Example 8).
[0091] Optionally, one or more excipients as described above can be
included in the suspensions of the invention. Such excipients may
be added in either solution or dry form to the suspension prior to
wet milling. Alternatively, one or more excipients in either dry or
solution form may be added to the suspension after wet milling, or
added in solution form as a co-spray dry solution during the spray
drying step. Optionally, a buffer such as phosphate or citrate or
the like is added to the wet milled suspension to form a suspension
having near neutral pHs from about 6 to 8, or more preferably from
about 7 to 8.
[0092] Alternatively, the polyene may be dry milled prior to
suspension formation.
[0093] The aerosol properties of the spray dried powders,
particularly but not necessarily those spray dried from
suspensions, may be further improved by exposure of the spray dried
powders to moisture. Specifically, it has been found that by
exposing the spray dried powders, either during (e.g., using "wet"
drying air) or post spray drying, to environments of controlled
relative humidity, the aerodynamic diameters of such powders can be
reduced, preferably to less than about 5 microns MMAD (Example 8).
Preferably, the powders are exposed to a degree of moisture
sufficient to increase to moisture content of the powders to
greater or equal to about 3% or 3.5% by weight, with preferred
moisture content of the powders ranging from about 4% to about 10%
by weight. Preferred controlled relative humidity (RH) conditions
are RHs greater than about 5%, with values typically ranging from
about 5% to about 60%. While most spray dried powders are
advantageously handled and packaged into unit dosage forms under
dry conditions to optimize aerosol performance, aerosol performance
(and mass median aerodynamic diameter in particular) of the powders
of the invention was improved (i.e., decrease in MMAD) by
increasing rather than decreasing their moisture contents. While
not being bound by any theory, it appears that exposure of the
powders to moisture is effective to bind water to the high energy
sites on the surface of the polyene particles, to thereby decrease
the tendency of the particles to aggregate.
[0094] Alternatively, higher level moisture environments can be
achieved indirectly from the spray drying process itself by
employing one or more of the following approaches: lowering the
inlet temperature, e.g., to below about 80.degree. C., e.g., to
temperatures ranging from about 50.degree. to 80.degree. C., or
from about 60.degree. to 80.degree. C.; increasing the suspension
feed rate to greater than about 5 mL/min, for example to rates
greater than about 10 mL/min, or, as described above, by use of a
drying gas other than dry air, i.e., air or another inert gas at
higher controlled relative humidities than dry air. Note that the
exact operating conditions will vary, depending upon the particular
processing unit employed.
[0095] Optionally, in-line sonication may also be employed to
further reduce the particle size of the spray-dried polyene
compositions of the invention. For instance, the feed suspension
may be passed through a sonicator prior to atomization.
[0096] Additional spray drying processes which may be suitable for
preparing the spray-dried antifungal compositions of the invention
are described in U.S. Pat. Nos. 5,985,248; 5,976,574; 6,001,336,
and 6,077,543, all assigned to Inhale Therapeutics Systems, Inc.,
the contents of which are expressly incorporated herein by
reference.
[0097] Polyene-containing solutions/suspensions such as those
described above are spray dried in a conventional spray drier, such
as those available from commercial suppliers such as Niro A/S
(Denmark), Buchi (Switzerland) and the like, resulting in
dispersible, chemically stable antifungal dry powders. Optimal
conditions for spray drying the polyene antifungal
solutions/suspensions will vary depending upon the formulation
components, and are generally determined experimentally. The gas
used to spray dry the material is typically air, although inert
gases such as nitrogen or argon are also suitable. Moreover, the
temperature of both the inlet and outlet of the gas used to dry the
sprayed material is such that it does not cause significant
decomposition of the polyene antifungal in the sprayed material.
Such temperatures are typically determined experimentally, although
generally, the inlet temperature will range from about 50.degree.
C. to about 200.degree. C., more preferably from about 60.degree.
C. to about 150.degree. C., while the outlet temperature will range
from about 30.degree. C. to about 150.degree. C.
[0098] An antifungal dry powder in accordance with the invention
may also be prepared, although less preferably, by lyophilization,
vacuum drying, spray freeze drying, super critical fluid
processing, or other forms of evaporative drying. Such drying
procedures will preferably be accompanied by additional processing
steps, e.g., by blending, grinding or jet milling, to obtain an
antifungal dry powder having suitable chemical, physical and
aerosol properties suitable for administration into the deep
lung.
[0099] In some instances, it will be desirable to prepare dry
powder formulations possessing improved handling/processing
characteristics, e.g., reduced static, better flowability, low
caking, and the like, by preparing compositions composed of fine
particle aggregates, that is, aggregates or agglomerates of the
above-described dry powder particles, where the aggregates are
readily broken back down to the fine powder components for
pulmonary delivery, as described, e.g., Johnson, et al., U.S. Pat.
No. 5,654,007, Aug. 5, 1997, incorporated herein by reference.
Alternatively, the powders may be prepared by agglomerating the
powder components, sieving the materials to obtain the
agglomerates, spheronizing to provide a more spherical agglomerate,
and sizing to obtain a uniformly-sized product, as described, e.g.,
and in Ahlneck, C.; et al., International PCT Publication No.
WO95/09616, Apr. 13, 1995, incorporated herein by reference. The
dry powders of the invention may also be prepared by blending,
grinding or jet milling formulation components directly in dry
powder form.
IV. Characteristics of Antifungal Powder Formulations
[0100] The antifungal compositions of the invention are further
characterized by several features, most notably, the ability of the
aerosolized composition to reach the tissues of the lung and lower
respiratory tract. Passage of the particles into the lung
physiology is an important aspect of the present invention, since
the concentration of antifungal at the site of infection is an
important feature in the successful treatment of pulmonary fungal
infections. Indeed, certain physical characteristics of antifungal
dry powders, to be described more fully below, are important in
maximizing the efficiency of aerosolized delivery of such powders
to the deep lung.
[0101] Polyene antifungal dry powders are composed of particles
effective to penetrate into the alveoli of the lungs, that is,
having a mass median diameter (MMD) from about 0.1 to 20 .mu.m.
Typically, the MMD of the particles is less than about 10 .mu.m
(e.g., ranging from about 0.1 to 10 .mu.m), preferably less than
7.5 .mu.m (e.g., ranging from about 0.5 to 7 microns), and most
preferably less than 5 .mu.m, and usually being in the range of 0.1
.mu.m to 5 .mu.m in diameter, depending upon the specific
characteristics of the powder. In looking at representative powders
in accordance with the invention (see, for example, Examples 1 and
3), the powders of the invention are most preferably although not
necessarily characterized by extremely small particle sizes, of
less than about 2 microns MMD. Such particles are extremely
effective in targeting the lung when administered by
inhalation.
[0102] In some cases, an antifungal powder composition will also
contain non-respirable carrier particles such as lactose, where the
non-respirable particles are typically greater than about 40
microns in size.
[0103] Antifungal powders of the invention are further
characterized by an aerosol particle size distribution less than
about 10 .mu.m mass median aerodynamic diameter (MMAD), and
preferably less than 5 .mu.m, and more preferably less than about
3.5 .mu.m. The above-described approaches for exposing the powders
to high moisture environments are particularly effective in
producing powders having MMAD values less than about 3.5 microns.
The mass median aerodynamic diameters of the powders will
characteristically range from about 0.5 to 5.0 .mu.m, preferably
from about 1.0 to 4.0 .mu.m MMAD, more preferably from about 1.0 to
3.5 .mu.m MMAD, and even more preferably from about 1.0 to 3.0
.mu.m. As demonstrated in the Examples, illustrative antifungal
powders having extremely small aerodynamic sizes, typically less
than 3.5 microns, and often less than 2.0 microns, have been
reproducibly prepared. This feature of the present powders is
particularly advantageous, since large particles having diameters
above 5 microns are typically removed by impaction in the upper
airways (nose, mouth, pharynx, trachea and large bronchi), while
those having aerodynamic sizes below 0.5 microns are generally
exhaled. Thus, the present particles are beneficial for inhalation
therapy due, in one aspect, to their ability to efficiently target
the lung without extensive deposition in the upper airways.
[0104] Dry powder compositions of the invention will generally have
a moisture content below about 15% by weight, usually below about
10% by weight, and preferably ranging from about 3.0% to about 10%
by weight.
[0105] The powders of the invention are further characterized as
relatively free-flowing rather than compacted solids.
[0106] The emitted dose or ED (sometimes also referred to as
delivered dose efficiency, DDE) of these powders is greater than
30% and usually greater than 40%. More typically, the emitted dose
of the antifungal powders of the invention is greater than 50%, and
is often greater than 60%. Even more preferably, the ED of an
antifungal powder is greater than 65%. Highly preferred are powders
having ED values greater than 50% to 60% and MMADs of less than
about 3.5 microns.
[0107] Powders of the invention will typically possess a bulk
density value ranging from about 0.05 to 10 gram/cubic centimeter,
preferably from about 0.05 to 5 gram/cubic centimeter, more
preferably from about 0.10 to 4.0 grams/cubic centimeter, even more
preferably from about 0.10 to 1 gram/cubic centimeter, even more
preferably from about 0.10-0.75 gram/cubic centimeter, and most
preferably from about 0.17 to 0.75 gram/cubic centimeter.
[0108] An additional measure for characterizing the overall aerosol
performance of a dry powder is the fine particle fraction (FPF),
which describes the percentage of powder having an aerodynamic
diameter less than 3.3 microns. Antifungal powder compositions are
particularly well suited for pulmonary delivery, and will possess
FPF values ranging from about 45%-90%. Such powders contain at
least about 45 percent of aerosol particle sizes below 3.3 .mu.m to
about 0.5 .mu.m and are thus are extremely effective when delivered
in aerosolized form, in (i) reaching the tissues of the lung, and,
in the case of treatment of systemic fungal infections, (ii)
reaching the alveolar region of the lung, followed by (iii)
diffusion to the interstitium and (iv) subsequent passage into the
bloodstream through the endothelium.
[0109] The particles of the invention also possess substantially
intact polyene, that is to say, the amount of polyene degradation
products is typically less than about 10% relative to the pre-spray
dried control, and more preferably is less than about 5%. In other
words, relative to the pre-spray dried starting material, the
polyene remains at least 90% chemically intact or pure upon spray
drying. Preferably, the spray dried powder contains at least 95%
pure or chemically intact polyene relative to the pre-spray dried
material.
[0110] The compositions described herein also possess good
stability with respect to aerosol performance over time.
V. Pulmonary Administration of the Powder
[0111] Dry powder formulations as described herein may be delivered
using any suitable dry powder inhaler (DPI), i.e., an inhaler
device that utilizes the patient's inhaled breath as a vehicle to
transport the dry powder drug to the lungs. Preferred are Inhale
Therapeutic Systems' dry powder inhalation devices as described in
Patton, J. S., et al., U.S. Pat. No. 5,458,135, Oct. 17, 1995;
Smith, A. E., et al., U.S. Pat. No. 5,740,794, Apr. 21, 1998; and
in Smith, A. E., et. al., U.S. Pat. No. 5,785,049, Jul. 28, 1998,
herein incorporated by reference. When administered using a device
of this type, the powdered medicament is contained in a receptacle
having a puncturable lid or other access surface, preferably a
blister package or cartridge, where the receptacle may contain a
single dosage unit or multiple dosage units. Convenient methods for
filling large numbers of cavities (i.e., unit dose packages) with
metered doses of dry powder medicament are described, e.g., in
Parks, D. J., et al., International Patent Publication WO 97/41031,
Nov. 6, 1997, incorporated herein by reference.
[0112] Also suitable for delivering the antifungal powders
described herein are dry powder inhalers of the type described, for
example, in Cocozza, S., et al., U.S. Pat. No. 3,906,950, Sep. 23,
1974, and in Cocozza, S., et al., U.S. Pat. No. 4,013,075, Mar. 22,
1977, incorporated herein by reference, wherein a pre-measured dose
of FSP dry powder for delivery to a subject is contained within a
hard gelatin capsule.
[0113] Other dry powder dispersion devices for pulmonary
administration of dry powders include those described, for example,
in Newell, R. E., et al, European Patent No. EP 129985, Sep. 7,
1988); in Hodson, P. D., et al., European Patent No. EP472598, Jul.
3, 1996; in Cocozza, S., et al., European Patent No. EP 467172,
Apr. 6, 1994, and in Lloyd, L. J. et al., U.S. Pat. No. 5,522,385,
Jun. 4, 1996, incorporated herein by reference. Also suitable for
delivering the antifungal dry powders of the invention are
inhalation devices such as the Astra-Draco "TURBUHALER". This type
of device is described in detail in Virtanen, R., U.S. Pat. No.
4,668,218, May 26, 1987; in Wetterlin, K., et al., U.S. Pat. No.
4,667,668, May 26, 1987; and in Wetterlin, K., et al., U.S. Pat.
No. 4,805,811, Feb. 21, 1989, all of which are incorporated herein
by reference. Other suitable devices include dry powder inhalers
such as Rotahaler.RTM. (Glaxo), Discus.RTM. (Glaxo), Spiros.TM.
inhaler (Dura Pharmaceuticals), and the Spinhaler.RTM. (Fisons).
Also suitable are devices which employ the use of a piston to
provide air for either entraining powdered medicament, lifting
medicament from a carrier screen by passing air through the screen,
or mixing air with powder medicament in a mixing chamber with
subsequent introduction of the powder to the patient through the
mouthpiece of the device, such as described in Mulhauser, P., et
al, U.S. Pat. No. 5,388,572, Sep. 30, 1997, incorporated herein by
reference.
[0114] An inhaleable antifungal composition may also be delivered
using a pressurized, metered dose inhaler (MDI), e.g., the
Ventolin.RTM. metered dose inhaler, containing a solution or
suspension of drug in a pharmaceutically inert liquid propellant,
e.g., a HFC, chlorofluorocarbon or fluorocarbon, as described in
Laube, et al., U.S. Pat. No. 5,320,094, Jun. 14, 1994, and in
Rubsamen, R. M., et al, U.S. Pat. No. 5,672,581 (1994), both
incorporated herein by reference.
[0115] Prior to use, a packaged antifungal dry powder is generally
stored under ambient conditions, and preferably is stored at a
temperature at or below about 25.degree. C., and relative humidity
(RH) ranging from about 30 to 60% or greater as described
above.
VI. Therapeutic Applications
[0116] The antifungal powders of the invention, when administered
pulmonarily, are particularly effective in the treatment of
respiratory fungal infections. The powders, when inhaled, penetrate
into the airways of the lungs and achieve effective concentrations
in the infected secretions and lung tissue, including the
epithelial lining fluid, alveolar macrophages, and neutrophils,
typically exceeding the MIC.sub.90s of most respiratory fungal
pathogens. Moreover, the doses of antifungal compound that are
administered pulmonarily are typically much less than those
administered by other routes and required to obtain similar
antifungal effects, due to the efficient targeting of the inhaled
powder directly to the site of fungal infection.
[0117] The powders of the present invention are useful in the
prophylaxis of pulmonary fungal infections, particularly for
immunocompromised patients, such as individuals undergoing
chemotherapy or radiation therapy for cancer, organ transplant
recipients, patients suffering from conditions that adversely
affect the immune system such as HIV, or any other condition which
predisposes a subject to pulmonary fungal infections. The powders
are also advantageous for use in the treatment of active pulmonary
fungal infections, such as aspergillosis (most commonly due to
Aspergillus fumigatus, Aspergillus flavus, Aspergillus niger,
Aspergillus nidulans, and Aspergillus terreus), coccidioidomycosis,
histoplasmosis, blastomycosis, and other fungal pathogens.
[0118] For prophylaxis, the amount per dose of antifungal agent is
that amount that is effective to prevent pulmonary infection by a
fungus is generally between about 0.01 mg/kg to about 5.0 mg/kg.
Preferably, the amount per dose of the antifungal (e.g., polyene)
that is administered by inhalation to a subject in need thereof is
typically from about 0.4 mg/kg to about 4.0 mg/kg, or even more
preferably from about 0.7 mg/kg to about 3.0 mg/kg. A powder in
accordance with the invention is administered to a subject in any
regimen which is effective to prevent pulmonary infection by a
fungus. Illustrative prophylactic regimes include administering an
antifungal dry powder as described herein 1 to 21 times per week
over a time course from 1 to 26 weeks, followed, if needed,
thereafter by administration once or twice weekly. A suitable
subject for receiving treatment as described herein is any
mammalian subject in need thereof, preferably such mammal is a
human.
[0119] For treating a subject suffering from a pulmonary fungal
infection, the amount per dose of antifungal agent administered by
oral inhalation is that amount which is effective to treat the
infection. The amount of antifungal agent for the treatment of
infection will generally be higher than that used for prevention,
and will typically range from about 0.01 mg/kg to 7.0 mg/kg.
Preferably, the amount administered will be from about 0.2 mg/kg to
about 6.0 mg/kg, and more preferably from about 0.8 mg/kg to about
5.0 mg/kg. In one exemplary treatment regimen, an antifungal powder
in accordance with the invention is administered 1 to 8 times
daily, preferably from 2-6 times daily, over a course of from about
7 to about 183 days.
[0120] In treating these respiratory fungal conditions, the powders
of the invention are typically administered in doses that are 3-10
or more times the MIC.sub.90 of the causative fungal pathogens;
these levels are safely achievable by inhalation. Generally, the
dose of antifungal compound delivered to a subject will be from
about 2 mg to about 400 mg daily, preferably from about 10 to 200
milligrams daily, depending upon the particular antifungal
compound, the condition being treated, the age and weight of the
subject, and the like. The antifungal powder, when administered via
a dry powder inhaler, is typically administered in unit dose form,
with unit dose sizes varying from about 2 milligrams to 250
milligrams, and more preferably from about 5 milligrams to 100
milligrams. From one up to about 10 unit doses are generally
administered daily during the course of therapy, although more
preferably a treatment regimen will consist of from one to about 8
unit doses daily by inhalation.
[0121] The compositions of the invention offer several notable
advantages: (i) increased antifungal protection at drug entry site,
(ii) elimination or minimization of undesirable side effects
without a concomitant increase in renal toxicity, (iii) result in
minimal or undetectable levels of antifungal compound in non-target
tissues, (iv) reduce the fungal burden in the respiratory tract to
thereby decrease chronic antigenic stimulation, reduce inflammatory
response, and ameliorate symptoms, (v) possibly reduce the long
term risk of progression or slow progression, and (vi) are
conveniently administered.
[0122] The disclosure of each publication, patent or patent
application mentioned in this specification is incorporated by
reference herein to the same extent as if each individual
publication, patent or patent application were specifically and
individually indicated to be incorporated by reference.
[0123] The following examples illustrate, but in no way are
intended to limit the scope of the present invention.
EXPERIMENTAL
Materials and Methods
[0124] The following materials were employed. The grades and
manufacturers are representative of many that are suitable.
Materials
Amphotericin B (Spectrum, New Brunswick, N.J.; Alapharma)
Nystatin (Spectrum, New Brunswick, N.J.)
[0125] Methanol dehydrated, HPLC grade (EM Industries, Gibbstown,
N.J.)
Sodium Hydroxide, 50% (J. T. Baker, Phillipsburg, N.J.)
L-Leucine (Aldrich)
Hydrochloric Acid (J. T. Baker, Phillipsburg, N.J.)
Sodium Deoxycholate (Spectrum, New Brunswick, N.J.)
Methods
Reverse Phase-HPLC
[0126] RP-HPLC analysis was performed on a Waters 2690 HPLC system
with a Waters Detector 996 (Skywalker, Inhale No. 101232). The
system was equipped with either a Vydac C-18 column or an YMC
ODC-AQ.TM. C-18 column. Samples were eluted with a gradient mobile
phases consisted of 10 mM sodium acetate buffer (mobile phase A) at
pH 5 and CAN (mobile phase B). The flow rate was maintained at 1.0
ml/min, the detector wavelength was 383 nm, and the injection
volume was 30 .mu.L (microliter).
Stability Samples
[0127] Blisters from the stability formulations were placed into
25.degree. C. and 40.degree. C. temperature stations (Temperature
stations: 40.degree. C./75RH: Environmental Specialties, Inc.;
25.degree. C./60RH: Environmental Specialties, Inc).
[0128] The blisters were sealed in an aluminum bag with desiccants.
Samples were removed from the temperature station at predetermined
time points.
EXAMPLE 1
Inhaleable Amphotericin B Dry Powder Formulations
[0129] A. Finding a Suitable Solvent for Spray Drying
[0130] The solubility of amphotericin and excipients/additives of
interest was determined in various solvents in an attempt to find a
solvent system suitable for spray drying (i.e., having a
sufficiently high vapor pressure) and capable of dissolving both
amphotericin and any added excipients at an extent greater than
about 10 mg/mL solvent. Although active agents can be spray-dried
as suspensions, having the formulation components dissolved in
solution provides resulting particles having a homogeneous
composition (i.e., when comparing one particle to another
particle)--that is to say, each particle in the composition has
approximately the same composition and distribution of formulation
components.
[0131] Amphotericin B is difficult to spray dry due to its poor
solubility in water at any pH where it is likely to have reasonable
stability (amphotericin is insoluble in water at pH 6 to 7).
Amphotericin B is soluble in water at pHs lower than 3 and higher
than 10, but only to a limited extent (0.2 mg/ml)--making the
volume of solvent required for spray drying under these conditions
too large to be reasonably handled. Thus, the following
solubilities were experimentally determined to find a single
(non-mixed) solvent system for spray drying representative
amphotericin B and nystatin formulations.
TABLE-US-00001 TABLE 1 Solubility of Representative Polyenes and
Excipients Solubility (mg compound/g solvent) Solvent Amphotericin
Nystatin Leucine DI Water 0.2 1.1 Not performed IPA 0.1 0.1 0.8
Methanol 1.5 11.0 2.6 Ethanol 0.2 0.8 1.2 Hexane 0.0 0.0 7.0
Acetone 0.2 0.1 1.7 Pet. Ether 0.0 0.0 0.8 Ethyl Ether 0.2 0.1 0.8
Chloroform 0.2 0.7 1.2 Isobutyl Alcohol 0.0 0.5 Not performed Meth.
Chloride 1.5 3.9 1.8 IP Acetate 0.0 0.1 0.7 THF 0.6 1.6 0.8
[0132] Neither water nor any pure pharmaceutically acceptable
solvent appeared to be able to jointly dissolve amphotericin B and
one particularly preferred excipient, leucine, at the desired
levels. However, when acidified with an acid such as hydrochloric
acid, methanol was effective in dissolving both amphotericin B and
leucine at acceptable levels, that is, at combined solids
concentrations of at least about 10 mg/ml.
[0133] B. Preparing Dry Powders
[0134] The antifungal and solid excipient (where employed) were
mixed with methanol at a 1% w/w solids concentration to form a
suspension. The pH of the medium was adjusted (acidified) with
hydrochloric acid while stirring continuously and/or sonicating to
facilitate solubilization of the components. The pH was adjusted
until all of the components were in solution. The goal was to
utilize the least acidic condition possible that resulted in
complete solubilization, to minimize the chemical destabilization
of the components in the solution. Sodium hydroxide was added to
further adjust the pH if needed. The resulting solution was then
spray dried.
[0135] All batches were spray dried using a modified Buchi 190 Mini
Spray Dryer supplied with nitrogen as the gas source and equipped
with an oxygen level sensor. The solution feed rate was 5
ml/minute, the inlet temperature was adjusted to obtain an outlet
temperature of approximately 80.degree. C., the drying gas flow
rate was about 18 SCFM, and the atomizing air was supplied at 0.5
to 1.5 SCFM, typically at a pressure of about 100 PSI.
[0136] The characteristics of each of the formulations prepared and
the characteristics of the resulting powders are provided in Table
2 below.
TABLE-US-00002 TABLE 2 Aerosol Characteristics of Illustrative
Spray Dried Polyene Powders Drug % Formulation, by pH of Resid. %
ED .+-. Particle weight percent Lot No. Solution Solvent* RSD MMAD
Morphology Amphotericin 1696-HS-35 5.1** 1.6 63 .+-. 6 1.9
Raisin-like 90% 1696-HS-36 3.1 3.0 63 .+-. 5 2.4 Mostly
Amphotericin + collapsed 10% L- hemispheres Leucine 75% 1696-HS-37
2.9 2.9 81 .+-. 6 1.9 Raisin-like Amphotericin + 25% L- Leucine
Drug Formulation, by MMD weight percent % < 3.3 .mu.m .mu.m
Amphotericin 70 0.6 90% 65 0.8 Amphotericin + 10% L- Leucine 75% 80
0.6 Amphotericin + 25% L- Leucine *Residual solvent content was
determined by thermogravimetric analysis. **The accuracy of this
result is in question; the true value may be lower.
[0137] Spray drying neat amphotericin B dissolved in acidified
methanol provided a powder having a good dispersibility (an emitted
dose of 63%) and a superior MMAD of 1.9 microns. The aerosol
properties of the neat formulation were surprising, particularly in
view of the (i) absence of stabilizing or dispersibility enhancing
excipients, and (ii) the non-protein nature of the active agent.
While proteins and polypeptides have been demonstrated to have
dispersibility-enhancing characteristics when employed in dry
powder formulations (U.S. Pat. No. 6,136,346), it is unusual to
spray-dry a non-proteinaceous active agent to form a highly
dispersible powder. The addition of 10% by weight leucine did not
materially change the characteristics of the powder, however, when
the amount of leucine contained in the powder was increased to 25%
by weight of the composition, a significant improvement in emitted
dose was achieved (from 63% to 81%), without compromising the MMAD
value. This was also surprising, since typically these two factors,
emitted dose and aerodynamic diameter, work in opposing fashions.
That is to say, generally, an increase or improvement in ED is
often accompanied by an undesirable increase in aerodynamic
diameter, since larger particles tend to agglomerate less and thus
disperse better. That is to say, it is generally thought that
larger particles tend to exhibit fewer cohesive forces due to the
inverse relationship between Van der Waals forces and particle
size, and also due to a decreased impact of electrostatic forces on
larger particles. Thus, it is unusual to prepare non-protein
containing dry powders which possess both excellent EDs (greater
than 50%, preferably greater than 60%, more preferably greater than
80%) and superior aerodynamic diameters (less than about 5 microns,
preferably less than about 3.5 microns, more preferably less than
about 3 microns, and even more preferably less than about 2
microns).
EXAMPLE 2
Inhaleable Amphotericin B Dry Powder Formulations Containing
Deoxycholate
[0138] A. Preparing Dry Powders
[0139] Sodium deoxycholate was dissolved in water. Amphotericin was
added to the sodium deoxycholate solution, and sonicated. 6 molar
sodium hydroxide was slowly added to the mixture while stirring
and/or sonicating, until the amphotericin was dissolved. The pH of
the resulting solution was adjusted (acidified) to 7.0-7.5, while
stirring, with 1.2 normal hydrochloric acid. The solution was
protected from light. The aim was to utilize the most neutral
solution possible that resulted in complete solubilization, to
minimize or essentially eliminate any chemical destabilization of
the components in the solution. The resulting solution was then
spray dried as detailed in Example 1.
[0140] The characteristics of each of the formulations prepared and
the characteristics of the resulting powders are provided in Table
3 below.
TABLE-US-00003 TABLE 3 Amphotericin B/ Sodium Deoxycholate Powder
Preparation: Formulation Parameters Quantity per Batch Batch
2242-AA- Batch 2242-AA- Ingredient 57 60 Amphotericin B 0.700 g
0.825 g Sodium 0.314 g 0.1848 g Deoxycholate DI Water 100 ml 165 ml
12 N and/or 1.2 N 0.925 ml 0.225 ml HCl 6 M NaOH 0.162 ml 0.225 ml
Amphotericin B With Sodium Deoxycholate: Powder Characteristics
Batch # 2242-AA-57 2242-AA-60 Molar Ratio Amphotericin B/Sodium 1.0
2.0 Deoxycholate Final Solution pH 7.5 7.3 Amphotericin B/Sodium
Deoxycholate ~7.0/3.1 ~5.0/1.1 Solution Concentration (mg/ml) % ED
.+-. % RSD 72 .+-. 4 75 .+-. 3 MMAD (.mu.m) 2.8 2.6 % <3.3 .mu.m
62 67 % Moisture Content (by TGA) 2.5 Not available
[0141] Spray drying a nearly neutral pH aqueous solution of
amphotericin B with sodium deoxycholate provided a powder having a
good dispersibility (an emitted dose of greater than 70%) and a
good MMAD of less than 3.0 microns
EXAMPLE 3
Inhaleable Dry Powder Formulations of Nystatin
[0142] The solubility of nystatin and leucine in various solvents
was explored to identify a solvent for preparing a spray-dried
powder of the invention; solubility results are provided in Example
1 above.
[0143] Dry powders were prepared as described in Example 1 above
using acidified methanol as the solvent. The characteristics of the
resulting powders are summarized below.
TABLE-US-00004 TABLE 4 Inhaleable Formulations of Nystatin:
Composition Characteristics % Drug pH of Residual % ED .+-.
Formulation Lot No. Solution Solvent RSD MMAD Morphology Nystatin
1696-HS-40 3.0 1.6 74 .+-. 4 1.6 Dimpled spheres 75% Nystatin +
1696-HS-42 3.9 1.8 79 .+-. 3 1.5 Highly dimpled 25% L- spheres
Leucine MMD Drug Formulation % < 3.3 .mu.m .mu.m Nystatin 84 0.8
75% Nystatin (w/w) 88 0.6 25% L-Leucine (w/w)
[0144] Spray drying neat nystatin dissolved in acidified methanol
yielded a powder with a good emitted dose of greater than 70% and a
superior MMAD of 1.6 microns. The addition of 25% leucine to the
formulation resulted in a nominal improvement in emitted dose to
79% without compromising the NMAD (1.5 microns). Again, the
superior aerosol properties of these powders, particularly the neat
powders, were surprising in view of the lack of dispersibility
enhancing agents such as proteins or polypeptides in the
formulation. An optimized formulation comprising 25% leucine was
identified.
EXAMPLE 4
Optimization of Low pH Solution Spray Drying Conditions for
Amphotericin
[0145] Exemplary amphotericin B solutions were prepared and their
solubilities and chemical stabilities were evaluated. The solutions
were spray dried, and the chemical stabilities of the spray dried
powders were also assessed.
TABLE-US-00005 TABLE 5 Composition of AmB formulations Total % of
Total Solid solid Formulation Description pH adjustment AmB SDC Leu
Na/Cl (%) Final pH R01013 AmB in Acidified to pH 96.5 0.0 0.0 3.5
1.0 4.0 MeOH 1.4 R01017 AmB in Acidified to pH 68.6 0 22.0 9.4 1.1
3.6 MeOH/Leu 1.0 *SDC: Sodium Deoxycholate
[0146] The solutions were spray dried at a feed rate of
approximately 5 mL min and atomization pressures ranging from about
80-150 psi. The batch sizes was 1.5 liters for both formulations,
with yields ranging from about 30-40%. Formulations R01011 and
R01013 were placed in the temperature stations after filling into
unit dosage forms (blisters). The chemical stability of AmB in both
R01013 and R01017 was very poor at initial time point (Table
6).
TABLE-US-00006 TABLE 6 Chemical stability of AmB % AmB in sample T
= 1 month Formulation Description Rec. pH Pre-SD T = 0 25.degree.
C. 40.degree. C. R01013 AmB in 4.3 45 MeOH R01017 AmB/Leu 3.7 32 in
MeOH
EXAMPLE 5
Optimization of Low pH Solution Spray Drying Conditions for
Amphotericin
[0147] In an attempt to reduce the extent of chemical degradation
of amphotericin, while also finding solution conditions under which
amphotericin B was reasonably soluble (e.g., to an extent greater
than about 1 mg/mL), solubility experiments were conducted. The
experiments were performed at room temperature for the first 2 hour
while adjusting for solution pH. After the amphotericin B was
completely dissolved, the samples were transferred to a 4.degree.
C. refrigerator for further chemical stability analysis. Table 7
summarizes the formulation details and apparent solubilities at
specific pH conditions. Table 8 shows the chemical stability of
amphotericin B at 2 hours and 8 hours, as determined by
RP-HPLC.
TABLE-US-00007 TABLE 7 Apparent Solubility of AmB in MeOH
Formulations % of Total pH adj. Solid Total Na Cl AmB AmB Na/Cl
Solid Formulation pH (mg/mL) (mg/mL) (mg/mL) (mM) (%) (%) (mg/mL)
2452-36-3 4.4 0 0.07 2.92 3.16 97.6 2.4 3 2452-36-1 6.9 0 0 0.58
0.63 100 0 0.58 2452-36-2 9.6 0.01 0 0.95 1.02 98.8 1.2 0.96
TABLE-US-00008 TABLE 8 Chemical stability of Amphotericin in the
formulations AmB %, T = 2 hr AmB %, T = 8 hr Main peak Main peak
Formulation pH AmB (%) (%) AmB (%) (%) 2452-36-3 4.4 93 91.1 92
91.2 2452-36-1 6.9 98 91.8 98 91.7 2452-36-2 9.6 96 92.4 96 93
[0148] As shown in formulation 2452-36-3, the apparent AmB
solubility was 2.9 mg/mL, while the chemical stability was 92% at 8
hours. Based upon these favorable results (i.e., good solubility
and stability), this formulation was further spray dried at various
conditions as shown in Table 9. The solutions were spray dried
utilizing atomization pressures ranging from about 25-60 psi.
Chemical stability of the resulting powder was also evaluated.
TABLE-US-00009 TABLE 9 Spray drying condition and chemical
stability MAIN Feet rate AmB PEAK Samples pH (mL/min) (%) (%)
Pre-spray dry solution, 4.4 103 90.7 t = 0 Pre-spray dry solution,
4.4 103 90.9 t = 7 hr R01037 powder 4.4 5 78 70.8 R01038 powder 4.4
5 73 76.8 R01039 powder 4.4 7 70 72.1 R01040 powder 4.4 5 92
85.7
[0149] Thus, formulations were determined in which both solubility
and chemical stability of the amphotericin were at acceptable
levels. At pH 4.4 in methanol, amphotericin B exhibited an apparent
solubility of 2.9 mg/mL, and the solution chemical stability was
about 92% at 8 hours. Powders obtained by spray drying contained
92% intact amphotericin B when the outlet temperature was
maintained below 80.degree. C., at 50.degree. C. The experiment
also indicated that temperature is another key factor for improving
the stability of amphotericin B upon formulation and spray
drying.
EXAMPLE 6
Optimization of Low pH Solution Spray Drying Conditions for
Amphotericin
[0150] Additional solubility of amphotericin B and stability of the
representative formulation, R0140, was investigated. The solubility
of amphotericin at pH 5 and at pH 4.7 was investigated;
additionally, the chemical stability of amphotericin B at pH 4.8
was further examined. In these experiments, samples were placed in
an ice bath during the formulation preparation. Results are listed
in Table 10.
TABLE-US-00010 TABLE 10 Solubility and Stability of AmB in MeOH
Solubility AmB Main Peak Samples Temp. (mg/mL) (%) % AmB/MeOH, pH
4.7 In ice 3.4 AmB/MeOH, pH 5.0 In ice 3.0 AmB/MeOH, pH 4.8 In ice
3.0* 97 92.0 *AmB concentration in the formulation
[0151] Even at low temperatures below ambient such as around
0.degree. C. (ice bath) (solubility is known to decrease as
temperature decreases), the solubility of AmB is approximately 3
mg/mL at pH 5 and 3.4 mg/mL at pH 4.7. That is to say, both of
these are reasonable solubility levels of drug for spray drying.
Additionally, the formulation at pH 4.8 demonstrated good chemical
stability of AmB (97%) during the formulation preparation.
EXAMPLE 7
Chemical Stability of Amphotericin Versus Temperature
[0152] Solutions were prepared as follows. Amphotericin B (AmB) was
added to MeOH (3 mg AmB/mL) at either room temperature or in an ice
bath. Then 1N HCL was added to the solutions slowly with agitation
to adjust the solution pH to 4.8 upon complete dissolution of
amphotericin B. The chemical stability of AmB was determined by an
HPLC assay as a function of time over 24 hours. The results are
summarized in Table 11.
TABLE-US-00011 TABLE 11 Chemical stability of AmB as a function of
time in MeOH at pH 4.8 Time % AmB (hr) At Room Temperature In Ice
Bath 1.5 89 97 4 85 99 6 83 97 8 84 94 24 78 97
[0153] Although the solubility of AmB increased at lower pHs
(<pH 4.8), the chemical stability decreases. Using the same AmB
solution preparation procedures as described above, the chemical
stability of AmB at pH 3 and pH 4 in an ice bath is summarized
below.
TABLE-US-00012 TABLE 12 Chemical stability of AmB in MeOH at
different pHs in an ice bath Time % AmB Time % AmB (hr) pH 3 (hr)
pH 4 0.67 95 1 94 4 93 4 96 18.5 90 18.5 97 26 85 26 93
[0154] An AmB solution in MeOH at 3 mg/mL was spray dried; the
aerosol properties of the resulting AmB powders are provided
below.
TABLE-US-00013 TABLE 13 Aerosol Properties Formulation Lot # ED (%)
MMAD (?m) Neat AmB at pH 4.4 R01041 49 2.6
EXAMPLE 8
Spray Drying Homogenized Suspensions of Amphotericin and Moisture
Conditioning to Enhance Aerosol Performance
[0155] The following describes the successful utilization of a
suspension-based approach for preparing chemically stable,
dispersible, inhaleable dry powders of amphotericin B.
A. Particle Size Reduction by Homogenization--General Preparation
Method Employed
[0156] Amphotericin B (median particle size 8-13 microns) was
weighed and dispersed in water using a high shear mixer
(Ultraturax) to achieve a uniform suspension. The suspension was
passed through an Avestin C-5 homogenizer several times (1-5) at
high pressures (.about.25,000 psi) under ambient conditions to
reduce the particle size to less than about 1 micron, determined
using a Malvern Mastersizer. Pressures ranging from about 5,000 to
30,000 psi can be utilized. The concentration of amphotericin in
the suspension ranged from about 5-20 mg/mL, although suspensions
with concentrations of polyene ranging from about 1 mg/mL to about
100 mg/mL can be utilized.
[0157] The homogenized suspensions were then spray dried using
outlet temperatures ranging from about 60.degree. C. to 80.degree.
C., a feed rate of 5 mL/min, and atomizer pressures ranging from
60-100 psi. The feed solids content ranged from 0.3-1.0% (w/v).
HPLC analysis showed no degradation of Amphotericin B after
homogenization and spray drying compared to the unprocessed drug
(raw material) used as a control.
B. Formulations
[0158] Optional excipients were included in the formulations.
Excipients were added to the formulations using various approaches:
(i) an aqueous solution of excipient or solid excipient was added
to/mixed with the homogenized aqueous suspension of amphotericin B
followed by spray drying; or (ii) excipient was added to a
suspension of amphotericin B prior to homogenization. Excipients
included leucine, trileucine, raffinose, sodium citrate and sodium
phosphate. During some of the spray drying runs, in-line sonication
was employed to minimize aggregation of the particles.
C. Aerosol Properties of Homogenized, Spray Dried Powders.
TABLE-US-00014 [0159] TABLE 14 Composition MMAD (microns) Emitted
Dose, % 100% Amphotericin B 6.0 72 (R01131-1) 100% Amphotericin B
5.0 67 (R01131-3) Amphotericin B w/ 30% 4.0 Not done Raffinose
(R01196) Amphotericin B w/ 30% 3.9 Not done Leucine (R01198)
Amphotericin B w/ sodium 4.7 Not done phosphate (pH 7.4), (R01199)
100% Amphotericin B 5.55 61 (R01101) 100% Amphotericin B 4.72 71
(R01115) Amphotericin B with 30% 4.76 72 Leucine (R01102)
Amphotericin B with 30% 4.36 71 Trileucine (R01103)
D. Chemical Stability
[0160] Degradation of amphotericin B was essentially undetectable
both before and after homogenization. That is to say, amphotericin
B was chemically stable under these process conditions.
TABLE-US-00015 TABLE 15 Chemical stability of AmB % AmB % AmB in
Main Formulation Description (mg/mL) Formulation Peak Standard A
Mixing with Excipients 11.6 96.1 to prepare for R01101 to R01103
Standard B To prepare for R01104 12.8 95.8 R01101 Neat AmB 96.2
96.3 R01102 AmB with 30% Leu 72.3 96.4 R01103 AmB with 30% Trileu
75.3 95.5 R01104 AmB with 5% DSPC/ 89.3 96.1 Ca.sup.++
E. Effect of Moisture
[0161] Neat Amphotericin B Formulations with different levels of
moisture content were prepared by one of the two following methods:
[0162] (a) The spray dried powders were exposed to controlled
environments of different relative humidity (Range 6% to 40% RH)
[0163] (b) Aqueous suspensions of Amphotericin B were spray dried
under different conditions of outlet temperatures and feed rates to
generate powders with different residual moisture contents.
[0164] The aerosol properties of the powders were tested. The
following graph shows the correlation between MMAD and moisture
content of neat Amphotericin B powders. For powders in which
moisture was introduced by exposure to humidity after spray drying,
the MMAD decreases with increase in moisture content up to about
4.0-4.5%. Beyond 4.5% moisture content, the MMAD is independent of
moisture. A similar dependence of MMAD on moisture content is
observed for powders in which moisture content was increased by
spray drying under less aggressive conditions. However, the
threshold moisture content beyond which the MMAD becomes
independent of the moisture content appears to be about 3.5%. Thus,
a minimum moisture content is necessary for improvement in the
aerosol properties of spray dried powders containing Amphotericin
B. However, the threshold moisture content beyond which the aerosol
properties become independent of moisture content may differ
depending upon the process used.
[0165] Amphotericin B powders containing excipients also show
improvements in aerosol performance upon increase in moisture
content. Excipient--containing Amphotericin B powders were filled
into blisters under two different controlled relative humidity
conditions and tested for MMAD. The data (in the table below)
clearly shows that a significant reduction in MMAD can be achieved
by increasing the moisture content of Amphotericin B powders (both
in the absence and presence of excipients).
TABLE-US-00016 TABLE 16 Effect of moisture conditioning on MMAD of
neat and excipient- containing Amphotericin B dry powder
formulations MMAD of powder MMAD of powder filled filled at RH
<5.0% at 40% RH Formulation (microns) (microns) 100%
Amphotericin B 5.0 3.5 80% Amphotericin & 20% 3.6 2.9 sodium
phosphate 70% Amphotericin B & 30% 3.3 2.5 leucine 70%
Amphotericin B, 20% 3.1 2.3 sodium phosphate & 10% leucine
[0166] This finding is unexpected, since most spray dried powders
either show better aerosol properties in the absence of moisture or
are insensitive to moisture content. This phenomenon is likely due
to water binding to the high energy sites located either on the
surface of the Amphotericin B particles or within the crystal
lattice. Such an association of water with Amphotericin B particles
decreases their propensity to aggregate, thereby improving the
aerosol performance of the powders.
* * * * *