U.S. patent application number 10/834541 was filed with the patent office on 2005-03-03 for formulation to render an antimicrobial drug potent against organisms normally considered to be resistant to the drug.
Invention is credited to Doty, Mark J., Kipp, James E., Papadopoulos, Pavlos George, Rabinow, Barrett, Rebbeck, Christine, Sun, Chong-Son, White, Randy, Wong, Joseph Chung Tak.
Application Number | 20050048126 10/834541 |
Document ID | / |
Family ID | 46302012 |
Filed Date | 2005-03-03 |
United States Patent
Application |
20050048126 |
Kind Code |
A1 |
Rabinow, Barrett ; et
al. |
March 3, 2005 |
Formulation to render an antimicrobial drug potent against
organisms normally considered to be resistant to the drug
Abstract
The present invention relates to compositions of submicron- to
micron-size particles of antimicrobial agents. More particularly
the invention relates to a composition of an antimicrobial agent
that renders the agent potent against organisms normally considered
to be resistant to the agent. The composition comprises an aqueous
suspension of submicron- to micron-size particles containing the
agent coated with at least one surfactant selected from the group
consisting of: ionic surfactants, non-ionic surfactants,
biologically derived surfactants, and amino acids and their
derivatives. The particles have a volume-weighted mean particle
size of less than 5 .mu.m as measured by laser diffractometry.
Inventors: |
Rabinow, Barrett; (Skokie,
IL) ; White, Randy; (Wodbury, MN) ; Sun,
Chong-Son; (Lake Forest, IL) ; Wong, Joseph Chung
Tak; (Gumee, IL) ; Kipp, James E.; (Wauconda,
IL) ; Doty, Mark J.; (Grayslake, IL) ;
Rebbeck, Christine; (Algonquin, IL) ; Papadopoulos,
Pavlos George; (Antioch, IL) |
Correspondence
Address: |
Baxter International Inc.
Route 120 and Wilson Road (RLP-30)
Round Lake
IL
60073
US
|
Family ID: |
46302012 |
Appl. No.: |
10/834541 |
Filed: |
April 29, 2004 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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10834541 |
Apr 29, 2004 |
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10270268 |
Oct 11, 2002 |
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10270268 |
Oct 11, 2002 |
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10246802 |
Sep 17, 2002 |
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10246802 |
Sep 17, 2002 |
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10035821 |
Oct 19, 2001 |
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10246802 |
Sep 17, 2002 |
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10021692 |
Dec 12, 2001 |
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10021692 |
Dec 12, 2001 |
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09953979 |
Sep 17, 2001 |
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09953979 |
Sep 17, 2001 |
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09874637 |
Jun 5, 2001 |
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60466354 |
Apr 29, 2003 |
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60258160 |
Dec 22, 2000 |
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Current U.S.
Class: |
424/489 ;
514/254.07; 514/383 |
Current CPC
Class: |
A61K 9/1688 20130101;
A61K 31/496 20130101; A61K 9/5015 20130101; A61K 9/5138 20130101;
A61K 31/495 20130101; A61K 9/5042 20130101; A61K 9/5031 20130101;
A61P 31/10 20180101; A61K 9/10 20130101; A61K 9/146 20130101; A61K
9/5036 20130101; A61K 9/5192 20130101; A61K 9/14 20130101; A61K
9/145 20130101; A61K 9/5123 20130101; A61K 9/5146 20130101 |
Class at
Publication: |
424/489 ;
514/254.07; 514/383 |
International
Class: |
A61K 031/496; A61K
031/4196; A61K 009/14 |
Claims
What is claimed is:
1. A composition of an antimicrobial agent that renders the agent
potent against organisms normally considered to be resistant to the
agent, the composition comprising an aqueous suspension of
submicron- to micron-size particles containing the agent coated
with at least one surfactant selected from the group consisting of:
ionic surfactants, non-ionic surfactants, biologically derived
surfactants, and amino acids and their derivatives, wherein the
particles have a volume-weighted mean particle size of less than 5
.mu.m as measured by laser diffractometry.
2. The composition of claim 1, wherein the particles have a
volume-weighted mean particle size of less than 2 .mu.m as measured
by laser diffractometry.
3. The composition of claim 1, wherein the particles have a
volume-weighted mean particle size of less than about 1 .mu.m as
measured by laser diffractometry.
4. The composition of claim 1, wherein the particles have a
volume-weighted mean particle size of from about 150 nm to about 1
.mu.m as measured by laser diffractometry.
5. The composition of claim 1, wherein the antimicrocidal agent is
an antifungal agent.
6. The composition of claim 5, wherein the antimicrocidal agent is
a triazole antifungal agent.
7. The composition of claim 6, wherein the triazole antifungal
agent is selected from the group consisting of: itraconazole,
ketoconazole, miconazole, fluconazole, ravuconazole, voriconazole,
saperconazole, eberconazole, genaconazole, clotrimazole, econazole,
oxiconazole, sulconazole, terconazole, tioconazole, and
posaconazole.
8. The composition of claim 1, wherein the antimicrocidal agent is
itraconazole.
9. The composition of claim 1, wherein the ionic surfactant is
selected from the group consisting of: anionic surfactants,
cationic surfactants, zwitterionic surfactants, and combinations
thereof.
10. The composition of claim 9, wherein the anionic surfactant is
selected from the group consisting of: alkyl sulfonates, alkyl
phosphates, alkyl phosphonates, potassium laurate, triethanolamine
stearate, sodium lauryl sulfate, sodium dodecylsulfate, alkyl
polyoxyethylene sulfates, sodium alginate, dioctyl sodium
sulfosuccinate, phosphatidylserine, phosphatidylinositol,
diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinosine,
phosphatidic acid and its salts, sodium carboxymethylcellulose,
cholic acid and other bile acids and salts thereof.
11. The composition of claim 10, wherein the bile acid is selected
from the group consisting of cholic acid, deoxycholic acid,
glycocholic acid, taurocholic acid, and glycodeoxycholic acid.
12. The composition of claim 9, wherein the anionic surfactant is a
phospholipid.
13. The composition of claim 12, wherein the phospholipid is
natural or synthetic.
14. The composition of claim 12, wherein the phospholipid is
pegylated.
15. The composition of claim 8, wherein the cationic surfactant is
selected from the group consisting of: quaternary ammonium
compounds, such as benzalkonium chloride, cetyltrimethylammonium
bromide, lauryldimethylbenzylammonium chloride, acyl carnitine
hydrochlorides, alkyl pyridinium halides, or aliphatic amines.
16. The composition of claim 9, wherein the zwitterionic surfactant
is a phospholipid.
17. The composition of claim 16, wherein the phospholipid is
natural or synthetic.
18. The composition of claim 16, wherein the phospholipid is
pegylated.
19. The composition of claim 1, wherein the nonionic surfactant is
selected from the group consisting of: glyceryl esters,
polyoxyethylene fatty alcohol ethers (Macrogol and Brij),
polyoxyethylene sorbitan fatty acid esters (Polysorbates),
polyoxyethylene fatty acid esters (Myij), sorbitan esters (Span),
glycerol monostearate, polyethylene glycols, polypropylene glycols,
cetyl alcohol, cetostearyl alcohol, stearyl alcohol, aryl alkyl
polyether alcohols, polyoxyethylene-polyoxypropylene copolymers
(poloxamers), poloxamines, methylcellulose, hydroxymethylcellulose,
hydyoxypropylcellulose, hydroxypropylmethylcellul- ose,
noncrystalline cellulose, polysaccharides including starch and
starch derivatives such as hydroxyethylstarch (HES), polyvinyl
alcohol, and polyvinylpyrrolidone.
20. The composition of claim 1, wherein the biologically derived
surfactant is selected from the group consisting of: albumin,
casein, other proteins and polysaccharides.
21. The composition of claim 20, wherein the polysaccharide is
selected from the group consisting of starches, heparin and
chitosans.
22. The composition of claim 1, wherein the amino acid is selected
from the group consisting of: leucine, alanine, valine, isoleucine,
lysine, aspartic acid, glutamic acid, methionine, tyrosine and
phenylalanine.
23. The composition of claim 1, wherein the amino acid derivative
is an amide, an ester, or a polypeptide.
24. The composition of claim 1, wherein the surfactant is a bile
salt.
25. The composition of claim 24, wherein the bile salt is
deoxycholate.
26. The composition of claim 1, wherein the surfactant is a
polyalkoxyether.
27. The composition of claim 26, wherein the polyalkoxyether is
Poloxamer 188.
28. The composition of claim 1, wherein the surfactant is
hydroxyethylstarch.
29. The composition of claim 1, wherein the surfactant is
polyethylene-660-hydroxystearate.
30. The composition of claim 1, wherein the surfactant is
albumin.
31. The composition of claim 1, wherein the surfactant is a
phospholipid.
32. The composition of claim 1, wherein the aqueous medium further
comprises a pH adjusting agent.
33. The composition of claim 32, wherein the pH adjusting agent is
selected from the group consisting of: hydrochloric acid, sulfuric
acid, phosphoric acid, acetic acid, lactic acid, succinic acid,
citric acid, tris(hydroxymethyl)aminomethane, meglumine, sodium
hydroxide, and amino acids.
34. The composition of claim 33, wherein the amino acid is selected
from the group consisting of: glycine, arginine, lysine, alanine,
methionine, valine, asparagine, tyrosine, proline, serine,
isoleucine, tryptophan, phenylalanine, threonine, cysteine,
glutamine, aspartic acid, glutamic acid, histidine, taurine and
leucine.
35. The composition of claim 1, further comprising an osmotic
pressure adjusting agent.
36. The composition of claim 35, wherein the osmotic pressure
adjusting agent is selected from the group consisting of: glycerin,
monosaccharides, disaccharides, trisaccharides, and sugar
alcohols.
37. The composition of claim 36, wherein the monosaccharide is
dextrose.
38. The composition of claim 36, wherein the disaccharide is
selected from the group consisting of sucrose, maltose and
trehalose.
39. The composition of claim 36, wherein the trisaccharide is
raffinose.
40. The composition of claim 36, wherein the sugar alcohol is
mannitol or sorbitol.
41. The composition of claim 1, wherein the antimicrobial agent is
present is an amount of from about 0.01% to about 50% w/v.
42. The composition of claim 1, wherein the antimicrobial agent is
present in an amount of from about 0.05% to about 30% w/v.
43. The composition of claim 1, wherein the antimicrobial agent is
present in an amount of about 0.1% to about 20% w/v.
44. The composition of claim 1, wherein the surfactant is present
in an amount of from about 0.001% to about 5% w/v.
45. The composition of claim 1, wherein the surfactant is present
in an amount of from about 0.005% to about 5% w/v.
46. The composition of claim 1, wherein the surfactant is present
in an amount of from about 0.01% to about 5% w/v.
47. The composition of claim 1 is administered by a route selected
from the group consisting of: parenteral, oral, buccal,
periodontal, rectal, nasal, pulmonary, and topical.
48. The composition of claim 1 is administered by a route selected
from the group consisting of intravenous, intramuscular,
intracerebral, subcutaneous, intradermal, intralymphatic,
pulmonary, intraacticular, intrathecal, and intraperitoneal.
49. The composition of claim 1, wherein the aqueous medium is
removed to form dry particles.
50. The composition of claim 49, wherein the method of removing the
aqueous medium is selected from the group consisting of:
evaporation and lyophilization.
51. The composition of claim 49, wherein the method of removing the
aqueous medium is by lyophilization.
52. The composition of claim 49, wherein the dry particles are
formulated into an acceptable pharmaceutical dosage form.
53. The composition of claim 52, wherein the pharmaceutical dosage
form is selected from the group consisting of: parenteral
solutions, tablets, capsules, suspensions, creams, lotions,
emulsions, pulmonary formulations, topical formulations, controlled
or sustained release formulations, and tissue specific targeted
delivery formulations.
54. The composition of claim 1, wherein the composition is
frozen.
55. A composition of an antimicrobial agent that renders the agent
potent against organisms normally considered to be resistant to the
agent, the composition comprising an aqueous suspension of
submicron- to micron-size particles of itraconazole coated with at
least one surfactant, and an osmotic pressure adjusting agent,
wherein the nanoparticles having a volume-weighted mean particle
size of less than 5 .mu.m as measured by laser diffractometry, and
wherein the itraconazole is present in an amount of from about
0.01% to about 50% w/v, and the surfactant is present in an amount
of from about 0.001% to about 5%.
56. A composition of particles of an antimicrobial agent that
renders the agent potent against organisms normally considered to
be resistant to the agent, the composition prepared by a method
comprising the steps of: (i) dissolving the antimicrobial agent in
a water-miscible first solvent to form a solution; (ii) mixing the
solution with a second solvent which is aqueous to define a
pre-suspension; and (iii) adding energy to the pre-suspension to
form particles having an average effective particle size of less
than 5 .mu.m; wherein the solubility of the antimicrobial agent is
greater in the first solvent than in the second solvent, and the
second solvent comprising one or more surfactants selected from the
group consisting of: nonionic surfactants, ionic surfactants,
biologically derived surfactants, and amino acids and their
derivatives.
57. A method for rendering an antimicrobial agent potent against
organisms normally considered to be resistant to the agent, the
method comprising formulating the agent as an aqueous suspension of
submicron- to micron-size particles containing the agent coated
with at least one surfactant selected from the group consisting of:
ionic surfactants, non-ionic surfactants, biologically derived
surfactants, and amino acids and their derivatives, wherein the
particles have a volume-weighted mean particle size of less than 5
.mu.m as measured by laser diffractometry.
58. A method for treating a subject infected with an organism
normally considered to be resistant to an antimicrobidcidal agent,
the method comprising the step of administering the agent to the
subject, wherein the agent is formulated as an aqueous suspension
of submicron- to micron-size particles containing the agent coated
with at least one surfactant selected from the group consisting of:
ionic surfactants, non-ionic surfactants, biologically derived
surfactants, and amino acids and their derivatives, wherein the
particles have a volume-weighted mean particle size of less than 5
.mu.m as measured by laser diffractometry.
Description
CROSS REFERENCE TO RELATED APPLICATIONS
[0001] This application claims priority from provisional
application Ser. No. 60/466,354, filed on Apr. 29, 2003. This
application is also a continuation-in-part of U.S. application Ser.
No. 10/270,268, filed on Oct. 11, 2002, which is a
continuation-in-part of U.S. patent application Ser. No. 10/246,802
filed Sep. 17, 2002 (which is a continuation-in-part of U.S. patent
application Ser. No. 10/035,821 filed Oct. 19, 2001), and a
continuation-in-part of U.S. patent application Ser. No. 10/021,692
filed Dec. 12, 2001, both of which are continuations-in-part of
U.S. patent application Ser. No. 09/953,979 filed Sep. 17, 2001,
which is a continuation-in-part of U.S. patent application Ser. No.
09/874,637 filed Jun. 5, 2001, which claims priority from
provisional Application Ser. No. 60/258,160 filed Dec. 22, 2000,
all of which are incorporated herein by reference and made a part
hereof.
FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT
[0002] Not applicable.
BACKGROUND OF THE INVENTION
[0003] 1. Technical Field
[0004] The present invention relates to compositions of
antimicrobial agents. More particularly the invention relates to
formulations of an antimicrobial agent which render the drug potent
against organisms normally considered to be resistant to the
agent.
[0005] 2. Background Of The Invention
[0006] Based upon in vitro microbicidal sensitivity tests, the
level of an antimicrobial drug considered effective against a
particular organism may be determined. This is referred to as the
MIC (minimum inhibitory concentration) of the drug. On the other
hand, safety studies will determine the amount of drug that can be
safely given to a patient or test animal. This maximal amount of
drug that can be dosed will determine the maximal biological
exposure to the host animal, normally measured by the area under
the curve (AUC) of the plot of drug concentration vs. time, the
peak height of the plot of drug concentration vs. time, tissue
levels vs. time, etc. The instantaneous tissue or plasma level of
the in vivo experiment can be compared with the MIC value to
determine relative efficacy of the attainable drug levels in the
biological fluids. The actual comparison must be corrected for
plasma protein binding, inasmuch as only the free drug level is the
important parameter because it is in this state that the drug is
freely diffusible to cross biological membranes.
[0007] As a result of such analysis, clinical literature has been
established specifying what drugs can be used generally for certain
strains of organisms, or more precisely, for certain strains of
organisms with MIC values below certain levels. As an example, the
antifungal agent itraconazole is not considered effective for
strains of Candida albicans with MIC>8 for this drug (e.g., for
C. albicans strain c43 (ATCC number 201794), MIC.sub.80=16 .mu.g/ml
for SPORANOX.RTM. itraconazole). These strains of Candida albicans
are considered to be resistant to itraconazole. This presupposes
the standard dosing level of this drug that can be
administered.
[0008] However, if a method were available to substantially
increase the amount of the antimicrobial drug (e.g., itraconazole)
that could be administered, than it might be possible to treat
infections hithertofore considered untreatable by this agent. Such
a method is available through formulation of the drug as a
nanosuspension. Submicron sized drug crystals stabilized by a
surfactant coating have been found, in some cases, not to dissolve
immediately upon injection into the blood stream. Instead, they are
captured by fixed macrophages of the spleen and liver. From this
sanctuary, the drug will be slowly released over a prolonged period
of days. This is in contrast to conventionally solubilized drugs,
which when injected, decrease in blood concentration at a much
faster rate.
[0009] An example of an antimicrobial agent which is conventionally
formulated to increase the solubility of the drug is the triazole
antifungal agent itraconazole (FIG. 2). Itraconazole is effective
against systemic mycoses, particularly aspergillosis and
candidiasis. New oral and intravenous preparations of itraconazole
have been prepared in order to overcome bioavailability problems
associated with a lack of solubility. For example, the
bioavailability of itraconazole is increased when it is formulated
in hydroxypropyl-beta-cyclodextrin, a carrier oligosaccharide that
forms an inclusion complex with the drug, thereby increasing its
aqueous solubility. The commercial preparation is known by the
tradename SPORANOX.RTM. Injection and was originated by JANSSEN
PHARMACEUTICAL PRODUCTS, L.P. The drug is currently manufactured by
Abbott Labs and distributed by Ortho Biotech, Inc.
[0010] Intravenous itraconazole may be useful in selected clinical
situations. Examples are achlorhydria in AIDS patients, an
inability to effectively absorb oral medications due to concurrent
treatments with other drugs, or in critical-care patients who
cannot take oral medications. The current commercial product,
SPORANOX.RTM. Injection, is made available in 25 mL glass vials
that contain 250 mg of itraconazole, with 10 g of
hydroxypropyl-beta-cyclodextrin (referenced as "HPBCD"). These
vials are diluted prior to use in 50 mL of 0.9% saline. The
resulting cyclodextrin concentration exceeds 10% (w/v) in the
reconstituted product. Although HPBCD has been traditionally
regarded as safe for injection, high concentrations, such as 10%,
have been reported in animal models to induce significant changes
to endothelial tissues (Duncker G.; Reichelt J., Effects of the
pharmaceutical cosolvent hydroxypropyl-beta-cyclodextrin on porcine
corneal endothelium. Graefe's Archive for Clinical and Experimental
Ophthalmology (Germany) 1998, 236/5, 380-389).
[0011] Other excipients are often used to formulate poorly
water-soluble drugs for intravenous injection. For example,
paclitaxel (Taxol.RTM., produced by Bristol-Myers Squibb) contains
52.7% (w/v) of Cremophor.RTM. EL (polyoxyethylated castor oil) and
49.7% (v/v) dehydrated alcohol, USP. Administration of
Cremophor.RTM. EL can lead to undesired hypersensitivity reactions
(Volcheck, G. W., Van Dellen, R. G. Anaphylaxis to intravenous
cyclosporine and tolerance to oral cyclosporine: case report and
review. Annals of Allergy, Asthma, and Immunology, 1998, 80,
159-163; Singla A. K.; Garg A.; Aggarwal D., Paclitaxel and its
formulations. International Journal of Pharmaceutics, 2002, 235/
1-2, 179-192).
[0012] The present invention discloses a composition which renders
antimicrobial drugs more effective on the basis of their physical
and biological properties than in their unformulated state or in
their existing formulations. The approach used is to formulate the
antimicrobial agents as nanosupensions. This permits using of the
improved formulation to treat microbes conventionally thought to be
resistant to the unformulated drug. Conventional formulation
approaches attempt to enhance solubility or bioavailability only.
Such methods include pH change, modification of the salt form, use
of organic modifiers, or cyclodextrin. The approach disclosed in
the present invention involves altering the pharmacokinetic
characteristic of the drug, permitting far greater dosing,
resulting in improved efficacy over and above what can be
accomplished by improving solubility and bioavailability only.
Acute toxicity tests have demonstrated that much more drug, when
formulated as a nanosuspension, can be administered to animals.
More of the drug is therefore available at the target organ to
exert efficacy.
SUMMARY OF THE INVENTION
[0013] The present invention relates to a composition of an aqueous
suspension of submicron- to micron-size particles of an
antimicrobial agent that renders the agent potent against organisms
normally considered to be resistant to the agent. The composition
includes an aqueous suspension of submicron- to micron-size
particles containing the agent coated with at least one surfactant
selected from the group consisting of: ionic surfactants, non-ionic
surfactants, biologically derived surfactants, and amino acids and
their derivatives. The particles have a volume-weighted mean
particle size of less than 5 .mu.m as measured by light scattering
(HORIBA) or by microscopic measurements. More preferably the
particles should be less than about 1 micron and most preferably
from about 150 nm to about 1 micron or any range or combination of
ranges therein.
[0014] The present invention is suitable for pharmaceutical
use.
[0015] In an embodiment of the invention, the antimicrobial agent
is an antifungal agent. In a preferred embodiment, the antifungal
agent is a triazole antifungal agent. In yet another embodiment of
the invention, the triazole antifungal agent is selected from
itraconazole, ketoconazole, miconazole, fluconazole, ravuconazole,
voriconazole, saperconazole, eberconazole, genaconazole,
clotrimazole, econazole, oxiconazole, sulconazole, terconazole,
tioconazole, and posaconazole. In a preferred embodiment of the
invention, the antifungal agent is itraconazole.
[0016] Suitable surfactants for coating the particles in the
present invention can be selected from ionic surfactants, nonionic
surfactants, biologically derived surfactants, or amino acids and
their derivatives.
[0017] In a further preferred embodiment, the composition of the
present invention is prepared by a microprecipitation method which
includes the steps of: (i) dissolving in the antifingal agent in a
first water-miscible first solvent to form a solution; (ii) mixing
the solution with a second solvent which is aqueous to define a
pre-suspension; and (iii) adding energy to the pre-suspension to
form particles having an average effective particle size of less
than 5 .mu.m; more preferably less than about 1 micron, and most
preferably from about 150 nm to about 1 micron or any range or
combination of ranges therein, wherein the solubility of the
antifungal agent is greater in the first solvent than in the second
solvent, and the first solvent or the second solvent comprising one
or more surfactants selected from the group consisting of: nonionic
surfactants, ionic surfactants, biologically derived surfactants,
and amino acids and their derivatives.
[0018] The present invention also relates to a method of rendering
an antimicrobial agent potent against organisms normally considered
to be resistant to the agent by formulating the agent as an aqueous
suspension of submicron- to micron-size particles containing the
agent coated with at least one surfactant selected from the group
consisting of: ionic surfactants, non-ionic surfactants,
biologically derived surfactants, and amino acids and their
derivatives.
[0019] The present invention further relates to a method of
treating infection of a subject by organisms normally considered to
be resistant to an antimicrobial agent by administering the agent
to the subject formulated as an aqueous suspension of submicron- to
micron-size particles containing the agent coated with at least one
surfactant selected from the group consisting of: ionic
surfactants, non-ionic surfactants, biologically derived
surfactants, and amino acids and their derivatives.
[0020] These and other aspects and attributes of the present
invention will be discussed with reference to the following
drawings and accompanying specification.
BRIEF DESCRIPTION OF THE DRAWINGS
[0021] FIG. 1 is the general molecular structure of a triazole
antifungal agent;
[0022] FIG. 2 is the molecular structure of itraconazole;
[0023] FIG. 3 is a schematic diagram of Method A of the
microprecipitation process used in the present invention to prepare
the suspension;
[0024] FIG. 4 is a schematic diagram of Method B of the
microprecipitation process used in the present invention to prepare
the suspension;
[0025] FIG. 5 is a graph comparing the pharmacokinetics of
SPORANOX.RTM. with Formulation 1 suspension of itraconazole of the
present invention, wherein ITC=plasma concentration of itraconazole
measured after bolus injection of Formulation 1 (80 mg/kg),
ITC-OH=plasma concentration of primary metabolite,
hydroxyitraconazole, measured after bolus injection of Formulation
1 (80 mg/kg), Total=combined concentration of itraconazole and
hydroxyitraconazole (ITC+ITC-OH) measured after bolus injection of
Formulation 1 (80 mg/kg), Sporanox-ITC=plasma concentration of
itraconazole measured after bolus injection of 20 mg/kg Sporanox
IV, Sporanox-ITC-OH=plasma concentration of primary metabolite,
hydroxyitraconazole, measured after bolus injection of 20 mg/kg
Sporanox IV, Sporanox-Total =combined concentration of itraconazole
and hydroxyitraconazole (ITC+ITC-OH) measured after bolus injection
of 20 mg/kg Sporanox IV;
[0026] FIG. 6 is a graph comparing the drug level for the rapidly
dissolving formulation, Form A, and the slow dissolving (macrophage
targeting) formulation, Form B, as determined in an in vitro
dissolution experiment; the drug level for Form A is much higher
than that attained by Form B;
[0027] FIG. 7 is a graph showing the comparison of results for body
weight over time for immuno-suppressed rats treated with
SPORANOX.RTM. Injection and Formulations 14288-1 and 14288-B;
[0028] FIG. 8 is a graph of kidney drug level vs. dose showing that
the greater dosing that could be administered permitted greater
drug levels to be manifested in the target organs, in this case,
the kidney;
[0029] FIG. 9 is a graph of fungal counts vs. kidney drug level
(N=nanosuspension; S=Sporanox IV solution) showing that the greater
drug levels in the target organ (the kidney) led to a greater kill
of the infectious organisms; and
[0030] FIG. 10 is a graph showing the mortality/moribundity profile
after daily or every other day dosing with antifungal drugs for 10
days in rats systemically infected with itraconazole resistant C.
albicans.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT
[0031] While this invention is susceptible of embodiment in many
different forms, there are shown in the drawing, and will be
described herein in detail, specific embodiments thereof with the
understanding that the present disclosure is to be considered as an
exemplification of the principles of the invention and is not
intended to limit the invention to the specific embodiments
illustrated.
[0032] The present invention relates to a composition of an
antimicrobial agent that renders the agent potent against organisms
normally considered to be resistant to the agent. The composition
comprises an aqueous suspension of submicron- to micron-size
particles containing the agent coated with at least one surfactant
selected from the group consisting of: ionic surfactants, non-ionic
surfactants, biologically derived surfactants, and amino acids and
their derivatives. The composition disclosed in the present
invention involves altering the pharmacokinetic characteristic of
the drug, permitting far greater dosing, resulting in improved
efficacy over and above what can be accomplished by improving
solubility and bioavailability only. Submicron sized drug crystals
stabilized by a surfactant coating have been found, in some cases,
not to dissolve immediately upon injection into the blood stream.
Instead, they are captured by fixed macrophages of the spleen and
liver. From this sanctuary, the drug can be slowly released over a
prolonged period of days. Acute toxicity tests have demonstrated
that much more drug, when formulated as a nanosuspension, can be
administered to animals or human beings. More of the drug is
therefore available at the target organ to exert efficacy.
[0033] The particles in the present invention have a
volume-weighted mean particle size of less than 5 .mu.m as measured
by light scattering (HORIBA) or by microscopic measurements. More
preferably the particles should be less than about 1 micron and
most preferably from about 150 nm to about 1 micron or any range or
combination of ranges therein. The composition can be administered
to a subject to treat infection by organisms normally considered to
be resistant to the agent.
[0034] The antimicrobial agent is preferably a poorly water soluble
organic compound. What is meant by "poorly water soluble" is that
the water solubility of the compound is less than 10 mg/ml, and
preferably, less than 1 mg/ml. A preferred class of antimicrobial
agent is an antifungal agent. A preferred antifungal agent is the
triazole antifungal agents having a general molecular structure as
shown in FIG. 1. Examples of triazole antifungal agents include,
but are not limited to: itraconazole, ketoconazole, miconazole,
fluconazole, ravuconazole, voriconazole, saperconazole,
eberconazole, genaconazole, clotrimazole, econazole, oxiconazole,
sulconazole, terconazole, tioconazole, and posaconazole. A
preferred antifungal agent for the present invention is
itraconazole. The molecular structure of itraconazole is shown in
FIG. 2.
[0035] The present invention is suitable for pharmaceutical use.
The compositions can be administered by various routes, including
but not limited to, intravenous, intracerebral, intrathecal,
intralymphatic, pulmonary, intraarticular, and intraperitoneal. In
an embodiment of the present invention, the aqueous medium of the
composition is removed to form dry particles. The method to remove
the aqueous medium can be any method known in the art. One example
is evaporation. Another example is freeze drying or lyophilization.
The dry particles may then be formulated into any acceptable
physical form including, but is not limited to, solutions, tablets,
capsules, suspensions, creams, lotions, emulsions, aerosols,
powders, incorporation into reservoir or matrix devices for
sustained release (such as implants or transdermal patches), and
the like.
[0036] If the particles do not have to be taken up by the
macrophages, the particles can be larger than 5 .mu.m (e.g., less
than 50 .mu.m, or less than 7 .mu.m) or less than 150 nm (e.g.,
less than 100 .mu.m). These particles can be administered by
various routes, including but not limited to parenteral, oral,
buccal, periodontal, rectal, nasal, pulmonary, transdermal, or
topical. Modes of parenteral administration include intravenous,
intra arterial, intrathecal, intraperitoneal, intraocular, intra
articular, intrathecal, intracerebral, intramuscular, subcutaneous,
and the like.
[0037] The aqueous suspension of the present invention may also be
frozen to improve stability upon storage. Freezing of an aqueous
suspension to improve stability is disclosed in the commonly
assigned and co-pending U.S. Pat. Application Ser. No. 60/347,548,
which is incorporated herein by reference and made a part
hereof.
[0038] In an embodiment of the present invention, the antimicrobial
agent is present in an amount preferably from about 0.01% to about
50% weight to volume (w/v), more preferably from about 0.05% to
about 30% w/v, and most preferably from about 0.1% to about 20%
w/v.
[0039] Suitable surfactants for coating the particles in the
present invention can be selected from ionic surfactants, nonionic
surfactants, biologically derived surfactants or amino acids and
their derivatives. Ionic surfactants can be anionic, cationic, or
zwitterionic.
[0040] Suitable anionic surfactants include but are not limited to:
alkyl sulfonates, alkyl phosphates, alkyl phosphonates, potassium
laurate, triethanolamine stearate, sodium lauryl sulfate, sodium
dodecylsulfate, alkyl polyoxyethylene sulfates, sodium alginate,
dioctyl sodium sulfosuccinate, phosphatidylglycerol,
phosphatidylinosine, phosphatidylinositol, diphosphatidylglycerol,
phosphatidylserine, phosphatidic acid and their salts, sodium
carboxymethylcellulose, cholic acid and other bile acids (e.g.,
cholic acid, deoxycholic acid, glycocholic acid, taurocholic acid,
glycodeoxycholic acid) and salts thereof (e.g., sodium
deoxycholate, etc.). As anionic surfactants, phospholipids may be
used. Suitable phospholipids include, for example,
phosphatidylserine, phosphatidylinositol, diphosphatidylglycerol,
phosphatidylglycerol, or phosphatidic acid and its salts.
[0041] Zwitterionic surfactants are electrically neutral but posses
local positive and negative charges within the same molecule.
Suitable zwitterionic surfactants include but are not limited to
zwitterionic phospholipids. Suitable phospholipids include
phosphatidylcholine, phosphatidylethanolamine,
diacyl-glycero-phosphoethanolamine (such as
dimyristoyl-glycero-phosphoethanolamine (DMPE),
dipalmitoyl-glycero-phosp- hoethanolamine(DPPE),
distearoyl-glycero-phosphoethanolamine (DSPE), and
dioleolyl-glycero-phosphoethanolamine(DOPE)). Mixtures of
phospholipids that include anionic and zwitterionic phospholipids
may be employed in this invention. Such mixtures include but are
not limited to lysophospholipids, egg or soybean phospholipid or
any combination thereof. The phospholipid, whether anionic,
zwitterionic or a mixture of phospholipids, may be salted or
desalted, hydrogenated or partially hydrogenated or natural
semisynthetic or synthetic. The phospholipid may also be conjugated
with a water-soluble or hydrophilic polymer to specifically target
the delivery to macrophages in the present invention. However,
conjugated phospholipids may be used to target other cells or
tissue in other applications. A preferred polymer is polyethylene
glycol (PEG), which is also known as the monomethoxy
polyethyleneglycol (mPEG). The molecule weights of the PEG can
vary, for example, from 200 to 50,000. Some commonly used PEG's
that are commercially available include PEG 350, PEG 550, PEG 750,
PEG 1000, PEG 2000, PEG 3000, and PEG 5000. The phospholipid or the
PEG-phospholipid conjugate may also incorporate a functional group
which can covalently attach to a ligand including but not limited
to proteins, peptides, carbohydrates, glycoproteins, antibodies, or
pharmaceutically active agents. These functional groups may
conjugate with the ligands through, for example, amide bond
formation, disulfide or thioether formation, or biotin/streptavidin
binding. Examples of the ligand-binding functional groups include
but are not limited to hexanoylamine, dodecanylamine,
1,12-dodecanedicarboxylate, thioethanol,
4-(p-maleimidophenyl)butyramide(MPB),
4-(p-maleimidomethyl)cyclohexane-carboxamide (MCC),
3-(2-pyridyldithio)propionate(PDP), succinate, glutarate,
dodecanoate, and biotin.
[0042] Suitable cationic surfactants include but are not limited to
quaternary ammonium compounds, such as benzalkonium chloride,
cetyltrimethylammonium bromide, lauryldimethylbenzylammonium
chloride, acyl camitine hydrochlorides, or alkyl pyridinium
halides, or long-chain alkyl amines such as, for example,
n-octylamine and oleylamine.
[0043] Suitable nonionic surfactants include: glyceryl esters,
polyoxyethylene fatty alcohol ethers (Macrogol and Brij),
polyoxyethylene sorbitan fatty acid esters (Polysorbates),
polyoxyethylene fatty acid esters (Myrj), sorbitan esters (Span),
glycerol monostearate, polyethylene glycols, polypropylene glycols,
cetyl alcohol, cetostearyl alcohol, stearyl alcohol, aryl alkyl
polyether alcohols, polyoxyethylene-polyoxypropylene copolymers
(poloxamers), poloxamines, methylcellulose, hydroxymethylcellulose,
hydroxypropylcellulose, hydroxypropylmethylcellulose,
noncrystalline cellulose, polysaccharides including starch and
starch derivatives such as hydroxyethylstarch (HES), polyvinyl
alcohol, and polyvinylpyrrolidone. In a preferred form of the
invention, the nonionic surfactant is a polyoxyethylene and
polyoxypropylene copolymer and preferably a block copolymer of
propylene glycol and ethylene glycol. Such polymers are sold under
the tradename POLOXAMER also sometimes referred to as
PLURONIC.RTM., and sold by several suppliers including Spectrum
Chemical and Ruger. Among polyoxyethylene fatty acid esters is
included those having short alkyl chains. One example of such a
surfactant is SOLUTOL.RTM. HS 15, polyethylene-660-hydroxystearate,
manufactured by BASF Aktiengesellschaft.
[0044] Surface-active biological molecules include such molecules
as albumin, casein, hirudin or other appropriate proteins.
Polysaccharide biologics are also included, and consist of but are
not limited to, starches, heparin and chitosans. Other suitable
surfactants include any amino acids such as leucine, alanine,
valine, isoleucine, lysine, aspartic acid, glutamic acid,
methionine, phenylalanine, or any derivatives of these amino acids
such as, for example, amide or ester derivatives and polypeptides
formed from these amino acids.
[0045] A preferred ionic surfactant is a bile salt, and a preferred
bile salt is deoxycholate. A preferred nonionic surfactant is a
polyalkoxyether, and a preferred polyalkoxyether is Poloxamer 188.
Another preferred nonionic surfactant is Solutol HS 15
(polyethylene-660-hydroxystearate). Still yet another preferred
nonionic surfactant is hydroxyethylstarch. A preferred biologically
derived surfactant is albumin.
[0046] In another embodiment of the present invention, the
surfactants are present in an amount of preferably from about
0.001% to 5% w/v, more preferably from about 0.005% to about 5%
w/v, and most preferably from about 0.01% to 5% w/v.
[0047] In a preferred embodiment of the present invention, the
particles are suspended in an aqueous medium further including a pH
adjusting agent. Suitable pH adjusting agents include, but are not
limited to, hydrochloric acid, sulfuric acid, phosphoric acid,
monocarboxylic acids (such as, for example, acetic acid and lactic
acid), dicarboxylic acids (such as, for example, succinic acid),
tricarboxylic acids (such as, for example, citric acid), THAM
(tris(hydroxymethyl)aminomethane), meglumine (N-methylglucosamine),
sodium hydroxide, and amino acids such as glycine, arginine,
lysine, alanine, histidine and leucine. The aqueous medium may
additionally include an osmotic pressure adjusting agent, such as
but not limited to glycerin, a monosaccharide such as dextrose, a
disaccharide such as sucrose, a trisaccharide such as raffinose,
and sugar alcohols such as mannitol, xylitol and sorbitol.
[0048] In a preferred embodiment of the present invention, the
composition comprises an aqueous suspension of particles of
itraconazole present at 0.01 to 50% w/v, the particles are coated
with 0.001 to 5% w/v of a bile salt (e.g., deoxycholate) and 0.001
to 5% w/v polyalkoxyether (for example, Poloxamer 188), and
glycerin added to adjust osmotic pressure of the formulation.
[0049] In another preferred embodiment of the present invention,
the composition comprises an aqueous suspension of particles of
itraconazole present at about 0.01 to 50% w/v, the particles coated
with about 0.001 to 5% w/v of a bile salt (for example,
deoxycholate) and 0.001 to 5% polyethylene-660-hydroxystearate w/v,
and glycerin added to adjust osmotic pressure of the
formulation.
[0050] In another preferred embodiment of the present invention,
the composition comprises an aqueous suspension of itraconazole
present at about 0.01 to 50% w/v, the particles are coated with
about 0.001 to 5% of polyethylene-660-hydroxystearate w/v, and
glycerin added to adjust osmotic pressure of the formulation.
[0051] In still yet another preferred embodiment of the present
invention, the composition comprises an aqueous suspension of
itraconazole present at 0.01 to 50% w/v, the particles are coated
with about 0.001 to 5% albumin w/v.
[0052] The method for preparing the suspension in the present
invention is disclosed in commonly assigned and co-pending U.S.
Pat. Applications Ser. Nos. 60/258,160; 09/874,799; 09/874,637;
09/874,499; 09/964,273; 10/035,821, 60/347,548; 10/021,692;
10/183,035; 10/213,352; 10/246,802; 10/270,268; 10/270,267, and
10/390,333; which are incorporated herein by reference and made a
part hereof. A general procedure for preparing the suspension
useful in the practice of this invention follows.
[0053] The processes can be separated into three general
categories. Each of the categories of processes share the steps of:
(1) dissolving an antifungal agent in a water miscible first
organic solvent to create a first solution; (2) mixing the first
solution with a second solvent of water to precipitate the
antifungal agent to create a pre-suspension; and (3) adding energy
to the presuspension in the form of high-shear mixing or heat to
provide a stable form of the antifungal agent having the desired
size ranges defined above.
[0054] The three categories of processes are distinguished based
upon the physical properties of the antifungal agent as determined
through x-ray diffraction studies, differential scanning
calorimetry (DSC) studies or other suitable study conducted prior
to the energy-addition step and after the energy-addition step. In
the first process category, prior to the energy-addition step the
antifungal agent in the presuspension takes an amorphous form, a
semi-crystalline form or a supercooled liquid form and has an
average effective particle size. After the energy-addition step,
the antifungal agent is in a crystalline form having an average
effective particle size essentially the same as that of the
presuspension (i.e., from less than about 50 .mu.m).
[0055] In the second process category, prior to the energy-addition
step the antifungal agent is in a crystalline form and has an
average effective particle size. After the energy-addition step,
the antifungal agent is in a crystalline form having essentially
the same average effective particle size as prior to the
energy-addition step but the crystals after the energy-addition
step are less likely to aggregate.
[0056] The lower tendency of the organic compound to aggregate is
observed by laser dynamic light scattering and light
microscopy.
[0057] In the third process category, prior to the energy-addition
step the antifungal agent is in a crystalline form that is friable
and has an average effective particle size. What is meant by the
term "friable" is that the particles are fragile and are more
easily broken down into smaller particles. After the
energy-addition step the organic compound is in a crystalline form
having an average effective particle size smaller than the crystals
of the pre-suspension. By taking the steps necessary to place the
organic compound in a crystalline form that is friable, the
subsequent energy-addition step can be carried out more quickly and
efficiently when compared to an organic compound in a less friable
crystalline morphology.
[0058] The energy-addition step can be carried out in any fashion
wherein the pre-suspension is exposed to cavitation, shearing or
impact forces. In one preferred form of the invention, the
energy-addition step is an annealing step. Annealing is defined in
this invention as the process of converting matter that is
thermodynamically unstable into a more stable form by single or
repeated application of energy (direct heat or mechanical stress),
followed by thermal relaxation. This lowering of energy may be
achieved by conversion of the solid form from a less ordered to a
more ordered lattice structure. Alternatively, this stabilization
may occur by a reordering of the surfactant molecules at the
solid-liquid interface.
[0059] These three process categories will be discussed separately
below. It should be understood, however, that the process
conditions such as choice of surfactants or combination of
surfactants, amount of surfactant used, temperature of reaction,
rate of mixing of solutions, rate of precipitation and the like can
be selected to allow for any drug to be processed under any one of
the categories discussed next.
[0060] The first process category, as well as the second and third
process categories, can be further divided into two subcategories,
Method A, and B shown diagrammatically in FIG. 3 and FIG. 4,
respectively.
[0061] The first solvent according to the present invention is a
solvent or mixture of solvents in which the organic compound of
interest is relatively soluble and which is miscible with the
second solvent. Such solvents include, but are not limited to
water-miscible protic compounds, in which a hydrogen atom in the
molecule is bound to an electronegative atom such as oxygen,
nitrogen, or other Group VA, VIA and VII A in the Periodic Table of
elements. Examples of such solvents include, but are not limited
to, alcohols, amines (primary or secondary), oximes, hydroxamic
acids, carboxylic acids, sulfonic acids, phosphonic acids,
phosphoric acids, amides and ureas.
[0062] Other examples of the first solvent also include aprotic
organic solvents. Some of these aprotic solvents can form hydrogen
bonds with water, but can only act as proton acceptors because they
lack effective proton donating groups. One class of aprotic
solvents is a dipolar aprotic solvent, as defined by the
International Union of Pure and Applied Chemistry (IUPAC Compendium
of Chemical Terminology, 2nd Ed., 1997):
[0063] A solvent with a comparatively high relative permittivity
(or dielectric constant), greater than ca. 15, and a sizable
permanent dipole moment, that cannot donate suitably labile
hydrogen atoms to form strong hydrogen bonds, e.g. dimethyl
sulfoxide.
[0064] Dipolar aprotic solvents can be selected from the group
consisting of: amides (fully substituted, with nitrogen lacking
attached hydrogen atoms), ureas (fully substituted, with no
hydrogen atoms attached to nitrogen), ethers, cyclic ethers,
nitriles, ketones, sulfones, sulfoxides, fully substituted
phosphates, phosphonate esters, phosphoramides, nitro compounds,
and the like. Dimethylsulfoxide (DMSO), N-methyl-2-pyrrolidinone
(NMP), 2-pyrrolidinone, 1,3-dimethylimidazolidin- one (DMI),
dimethylacetamide(DMA), dimethylformamide(DMF), dioxane, acetone,
tetrahydrofuran (THF), tetramethylenesulfone (sulfolane),
acetonitrile, and hexamethylphosphoramide(HMPA), nitromethane,
among others, are members of this class.
[0065] Solvents may also be chosen that are generally
water-immiscible, but have sufficient water solubility at low
volumes (less than 10%) to act as a water-miscible first solvent at
these reduced volumes. Examples include aromatic hydrocarbons,
alkenes, alkanes, and halogenated aromatics, halogenated alkenes
and halogenated alkanes. Aromatics include, but are not limited to,
benzene (substituted or unsubstituted), and monocyclic or
polycyclic arenes. Examples of substituted benzenes include, but
are not limited to, xylenes (ortho, meta, or para), and toluene.
Examples of alkanes include but are not limited to hexane,
neopentane, heptane, isooctane, and cyclohexane. Examples of
halogenated aromatics include, but are not restricted to,
chlorobenzene, bromobenzene, and chlorotoluene. Examples of
halogenated alkanes and alkenes include, but are not restricted to,
trichloroethane, methylene chloride, ethylenedichloride (EDC), and
the like.
[0066] Examples of the all of the above solvent classes include but
are not limited to: N-methyl-2-pyrrolidinone (also called
N-methyl-2-pyrrolidone), 2-pyrrolidinone (also called
2-pyrrolidone), 1,3-dimethyl-2-imidazolidinone (DMI),
dimethylsulfoxide, dimethylacetamide, carboxylic acids (such as
acetic acid and lactic acid), aliphatic alcohols (such as methanol,
ethanol, isopropanol, 3-pentanol, and n-propanol), benzyl alcohol,
glycerol, butylene glycol (butanediol), ethylene glycol, propylene
glycol, mono- and diacylated monoglycerides (such as glyceryl
caprylate), dimethyl isosorbide, acetone, dimethylsulfone,
dimethylformamide, 1,4-dioxane, tetramethylenesulfone (sulfolane),
acetonitrile, nitromethane, tetramethylurea,
hexamethylphosphoramide(HMPA), tetrahydrofuiran (THF), dioxane,
diethylether, tert-butylmethyl ether (TBME), aromatic hydrocarbons,
alkenes, alkanes, halogenated aromatics, halogenated alkenes,
halogenated alkanes, xylene, toluene, benzene, substituted benzene,
ethyl acetate, methyl acetate, butyl acetate, chlorobenzene,
bromobenzene, chlorotoluene, trichloroethane, methylene chloride,
ethylenedichloride (EDC), hexane, neopentane, heptane, isooctane,
cyclohexane, polyethylene glycol (PEG, for example, PEG-4, PEG-8,
PEG-9, PEG-12, PEG-14, PEG-16, PEG-120, PEG-75, PEG-150),
polyethylene glycol esters (examples such as PEG-4 dilaurate,
PEG-20 dilaurate, PEG-6 isostearate, PEG-8 palmitostearate, PEG-150
palmitostearate), polyethylene glycol sorbitans (such as PEG-20
sorbitan isostearate), polyethylene glycol monoalkyl ethers
(examples such as PEG-3 dimethyl ether, PEG-4 dimethyl ether),
polypropylene glycol (PPG), polypropylene alginate, PPG-10
butanediol, PPG-10 methyl glucose ether, PPG-20 methyl glucose
ether, PPG-15 stearyl ether, propylene glycol
dicaprylate/dicaprate, propylene glycol laurate, and glycofurol
(tetrahydrofurfuryl alcohol polyethylene glycol ether). A preferred
first solvent is N-methyl-2-pyrrolidinone. Another preferred first
solvent is lactic acid.
[0067] The second solvent is an aqueous solvent. This aqueous
solvent may be water by itself. This solvent may also contain
buffers, salts, surfactant(s), water-soluble polymers, and
combinations of these excipients.
[0068] Method A
[0069] In Method A (see FIG. 3), the antimicrobial agent is first
dissolved in the first solvent to create a first solution. The
antimicrobial agent can be added from about 0.01% (w/v) to about
50% (w/v) depending on the solubility of the antimicrobial agent in
the first solvent. Heating of the concentrate from about 30.degree.
C. to about 100.degree. C. may be necessary to ensure total
dissolution of the antimicrobial agent in the first solvent.
[0070] A second aqueous solution is provided with one or more
surfactants added thereto. The surfactants can be selected from an
ionic surfactant, a nonionic surfactant or a biologically derived
surfactant set forth above.
[0071] It may also be desirable to add a pH adjusting agent to the
second solution such as sodium hydroxide, hydrochloric acid, tris
buffer or citrate, acetate, lactate, meglumine, or the like. The
second solution should have a pH within the range of from about 3
to about 11.
[0072] In a preferred form of the invention, the method for
preparing submicron sized particles of an antimicrobial agent
includes the steps of adding the first solution to the second
solution. The addition rate is dependent on the batch size, and
precipitation kinetics for the antimicrobial agent. Typically, for
a small-scale laboratory process (preparation of 1 liter), the
addition rate is from about 0.05 cc per minute to about 10 cc per
minute. During the addition, the solutions should be under constant
agitation. It has been observed using light microscopy that
amorphous particles, semi-crystalline solids, or a supercooled
liquid are formed to create a pre-suspension. The method further
includes the step of subjecting the pre-suspension to an annealing
step to convert the amorphous particles, supercooled liquid or
semicrystalline solid to a crystalline more stable solid state. The
resulting particles will have an average effective particles size
as measured by dynamic light scattering methods (e.g.,
photocorrelation spectroscopy, laser diffraction, low-angle laser
light scattering (LALLS), medium-angle laser light scattering
(MALLS), light obscuration methods (Coulter method, for example),
rheology, or microscopy (light or electron) within the ranges set
forth above).
[0073] The energy-addition step involves adding energy through
sonication, homogenization, counter current flow homogenization
(e.g., the Mini DeBEE 2000 homogenizer, available from BEE
Incorporated, NC, in which a jet of fluid is directed along a first
path, and a structure is interposed in the first path to cause the
fluid to be redirected in a controlled flow path along a new path
to cause emulsification or mixing of the fluid), microfluidization,
or other methods of providing impact, shear or cavitation forces.
The sample may be cooled or heated during this stage. In one
preferred form of the invention the annealing step is effected by
homogenization. In another preferred form of the invention the
annealing may be accomplished by ultrasonication. In yet another
preferred form of the invention the annealing may be accomplished
by use of an emulsification apparatus as described in U.S. Pat. No.
5,720,551 which is incorporated herein by reference and made a part
hereof.
[0074] Depending upon the rate of annealing, it may be desirable to
adjust the temperature of the processed sample to within the range
of from approximately -30.degree. C. to 100.degree. C.
Alternatively, in order to effect a desired phase change in the
processed solid, it may also be necessary to adjust the temperature
of the pre-suspension to a temperature within the range of from
about -30.degree. C. to about 100.degree. C. during the annealing
step.
[0075] Method B
[0076] Method B differs from Method A in the following respects.
The first difference is a surfactant or combination of surfactants
are added to the first solution. The surfactants may be selected
from ionic surfactants, nonionic surfactants, or biologically
derived as set forth above.
[0077] A drug suspension resulting from application of the
processes described in this invention may be administered directly
as an injectable solution, provided Water for Injection is used in
formulation and an appropriate means for solution sterilization is
applied. Sterilization may be accomplished by separate
sterilization of the drug concentrate (drug, solvent, and optional
surfactant) and the diluent medium (water, and optional buffers and
surfactants) prior to mixing to form the pre-suspension.
Sterilization methods include pre-filtration first through a 3.0
micron filter followed by filtration through a 0.45-micron particle
filter, followed by steam or heat sterilization or sterile
filtration through two redundant 0.2-micron membrane filters.
[0078] Optionally, a solvent-free suspension may be produced by
solvent removal after precipitation. This can be accomplished by
centrifugation, dialysis, diafiltration, force-field fractionation,
high-pressure filtration or other separation techniques well known
in the art. Complete removal of N-methyl-2-pyrrolidinone was
typically carried out by one to three successive centrifugation
runs; after each centrifugation the supernatant was decanted and
discarded. A fresh volume of the suspension vehicle without the
organic solvent was added to the remaining solids and the mixture
was dispersed by homogenization. It will be recognized by others
skilled in the art that other high-shear mixing techniques could be
applied in this reconstitution step.
[0079] Furthermore, any undesired excipients such as surfactants
may be replaced by a more desirable excipient by use of the
separation methods described in the above paragraph. The solvent
and first excipient may be discarded with the supernatant after
centrifugation or filtration. A fresh volume of the suspension
vehicle without the solvent and without the first excipient may
then be added. Alternatively, a new surfactant may be added. For
example, a suspension consisting of drug, N-methyl-2-pyrrolidinone
(solvent), Poloxamer 188 (first excipient), sodium deoxycholate,
glycerol and water may be replaced with phospholipids (new
surfactant), glycerol and water after centrifugation and removal of
the supernatant.
[0080] I. First Process Category
[0081] The methods of the first process category generally include
the step of dissolving the antimicrobial agent in a water miscible
first solvent followed by the step of mixing this solution with an
aqueous solution to form a presuspension wherein the antimicrobial
agent is in an amorphous form, a semicrystalline form or in a
supercooled liquid form as determined by x-ray diffraction studies,
DSC, light microscopy or other analytical techniques and has an
average effective particle size within one of the effective
particle size ranges set forth above. The mixing step is followed
by an energy-addition step and, in a preferred form of the
invention is an annealing step.
[0082] II. Second Process Category
[0083] The methods of the second processes category include
essentially the same steps as in the steps of the first processes
category but differ in the following respect. An x-ray diffraction,
DSC or other suitable analytical techniques of the presuspension
shows the antimicrobial agent in a crystalline form and having an
average effective particle size. The antimicrobial agent after the
energy-addition step has essentially the same average effective
particle size as prior to the energy-addition step but has less of
a tendency to aggregate into larger particles when compared to that
of the particles of the presuspension. Without being bound to a
theory, it is believed the differences in the particle stability
may be due to a reordering of the surfactant molecules at the
solid-liquid interface.
[0084] III. Third Process Category
[0085] The methods of the third category modify the first two steps
of those of the first and second processes categories to ensure the
antimicrobial agent in the presuspension is in a friable form
having an average effective particle size (e.g., such as slender
needles and thin plates). Friable particles can be formed by
selecting suitable solvents, surfactants or combination of
surfactants, the temperature of the individual solutions, the rate
of mixing and rate of precipitation and the like. Friability may
also be enhanced by the introduction of lattice defects (e.g.,
cleavage planes) during the steps of mixing the first solution with
the aqueous solution. This would arise by rapid crystallization
such as that afforded in the precipitation step. In the
energy-addition step these friable crystals are converted to
crystals that are kinetically stabilized and having an average
effective particle size smaller than those of the presuspension.
Kinetically stabilized means particles have a reduced tendency to
aggregate when compared to particles that are not kinetically
stabilized. In such instance the energy-addition step results in a
breaking up of the friable particles. By ensuring the particles of
the presuspension are in a friable state, the organic compound can
more easily and more quickly be prepared into a particle within the
desired size ranges when compared to processing an organic compound
where the steps have not been taken to render it in a friable
form.
[0086] In addition to the microprecipitation methods described
above, any other known precipitation methods for preparing
submicron sized particles or nanoparticles in the art can be used
in conjunction with the present invention. The following is a
description of examples of other precipitation methods. The
examples are for illustration purposes, and are not intended to
limit the scope of the present invention.
[0087] Emulsion Precipitation Methods
[0088] One suitable emulsion precipitation technique is disclosed
in the co-pending and commonly assigned U.S. Ser. No. 09/964,273,
which is incorporated herein by reference and is made a part
hereof. In this approach, the process includes the steps of: (1)
providing a multiphase system having an organic phase and an
aqueous phase, the organic phase having a pharmaceutically
effective compound therein; and (2) sonicating the system to
evaporate a portion of the organic phase to cause precipitation of
the compound in the aqueous phase and having an average effective
particle size of less than about 2 .mu.m. The step of providing a
multiphase system includes the steps of: (1) mixing a water
immiscible solvent with the pharmaceutically effective compound to
define an organic solution, (2) preparing an aqueous based solution
with one or more surface active compounds, and (3) mixing the
organic solution with the aqueous solution to form the multiphase
system. The step of mixing the organic phase and the aqueous phase
can include the use of piston gap homogenizers, colloidal mills,
high speed stirring equipment, extrusion equipment, manual
agitation or shaking equipment, microfluidizer, or other equipment
or techniques for providing high shear conditions. The crude
emulsion will have oil droplets in the water of a size of
approximately less than 1 .mu.m in diameter. The crude emulsion is
sonicated to define a microemulsion and eventually to define a
submicron sized particle suspension.
[0089] Another approach to preparing submicron sized particles is
disclosed in co-pending and commonly assigned U.S. Ser. No.
10/183,035, which is incorporated herein by reference and made a
part hereof. The process includes the steps of: (1) providing a
crude dispersion of a multiphase system having an organic phase and
an aqueous phase, the organic phase having a pharmaceutical
compound therein; (2) providing energy to the crude dispersion to
form a fine dispersion; (3) freezing the fine dispersion; and (4)
lyophilizing the fine dispersion to obtain submicron sized
particles of the pharmaceutical compound. The step of providing a
multiphase system includes the steps of: (1) mixing a water
immiscible solvent with the pharmaceutically effective compound to
define an organic solution; (2) preparing an aqueous based solution
with one or more surface active compounds; and (3) mixing the
organic solution with the aqueous solution to form the multiphase
system. The step of mixing the organic phase and the aqueous phase
includes the use of piston gap homogenizers, colloidal mills, high
speed stirring equipment, extrusion equipment, manual agitation or
shaking equipment, microfluidizer, or other equipment or techniques
for providing high shear conditions.
[0090] Solvent Anti-Solvent Precipitation
[0091] Suitable solvent anti-solvent precipitation technique is
disclosed in U.S. Pat. Nos. 5,118,528 and 5,100,591 which are
incorporated herein by reference and made apart hereof. The process
includes the steps of: (1) preparing a liquid phase of a
biologically active substance in a solvent or a mixture of solvents
to which may be added one or more surfactants; (2) preparing a
second liquid phase of a non-solvent or a mixture of non-solvents,
the non-solvent is miscible with the solvent or mixture of solvents
for the substance; (3) adding together the solutions of (1) and (2)
with stirring; and (4) removing of unwanted solvents to produce a
colloidal suspension of nanoparticles. The '528 patent discloses
that it produces particles of the substance smaller than 500 nm
without the supply of energy.
[0092] Phase Inversion Precipitation
[0093] One suitable phase inversion precipitation is disclosed in
U.S. Pat. Nos. 6,235,224, 6,143,211 and U.S. patent application No.
2001/0042932 which are incorporated herein by reference and made a
part hereof. Phase inversion is a term used to describe the
physical phenomena by which a polymer dissolved in a continuous
phase solvent system inverts into a solid macromolecular network in
which the polymer is the continuous phase. One method to induce
phase inversion is by the addition of a nonsolvent to the
continuous phase. The polymer undergoes a transition from a single
phase to an unstable two phase mixture: polymer rich and polymer
poor fractions. Micellar droplets of nonsolvent in the polymer rich
phase serve as nucleation sites and become coated with polymer. The
'224 patent discloses that phase inversion of polymer solutions
under certain conditions can bring about spontaneous formation of
discrete microparticles, including nanoparticles. The '224 patent
discloses dissolving or dispersing a polymer in a solvent. A
pharmaceutical agent is also dissolved or dispersed in the solvent.
For the crystal seeding step to be effective in this process it is
desirable the agent is dissolved in the solvent. The polymer, the
agent and the solvent together form a mixture having a continuous
phase, wherein the solvent is the continuous phase. The mixture is
then introduced into at least tenfold excess of a miscible
nonsolvent to cause the spontaneous formation of the
microencapsulated microparticles of the agent having an average
particle size of between 10 nm and 10 .mu.m. The particle size is
influenced by the solvent:nonsolvent volume ratio, polymer
concentration, the viscosity of the polymer-solvent solution, the
molecular weight of the polymer, and the characteristics of the
solvent-nonsolvent pair. The process eliminates the step of
creating microdroplets, such as by forming an emulsion, of the
solvent. The process also avoids the agitation and/or shear
forces.
[0094] pH Shift Precipitation
[0095] pH shift precipitation techniques typically include a step
of dissolving a drug in a solution having a pH where the drug is
soluble, followed by the step of changing the pH to a point where
the drug is no longer soluble. The pH can be acidic or basic,
depending on the particular pharmaceutical compound. The solution
is then neutralized to form a presuspension of submicron sized
particles of the pharmaceutcially active compound. One suitable pH
shifting precipitation process is disclosed in U.S. Pat. No.
5,665,331, which is incorporated herein by reference and made a
part hereof. The process includes the step of dissolving of the
pharmaceutical agent together with a crystal growth modifier (CGM)
in an alkaline solution and then neutralizing the solution with an
acid in the presence of suitable surface-modifying surface-active
agent or agents to form a fine particle dispersion of the
pharmaceutical agent. The precipitation step can be followed by
steps of diafiltration clean-up of the dispersion and then
adjusting the concentration of the dispersion to a desired level.
This process of reportedly leads to microcrystalline particles of
Z-average diameters smaller than 400 nm as measured by photon
correlation spectroscopy.
[0096] Other examples of pH shifting precipitation methods are
disclosed in U.S. Pat. Nos. 5,716,642; 5,662,883; 5,560,932; and
4,608,278, which are incorporated herein by reference and are made
a part hereof.
[0097] Infusion Precipitation Method
[0098] Suitable infusion precipitation techniques are disclosed in
the U.S. Pat. Nos. 4,997,454 and 4,826,689, which are incorporated
herein by reference and made a part hereof. First, a suitable solid
compound is dissolved in a suitable organic solvent to form a
solvent mixture. Then, a precipitating nonsolvent miscible with the
organic solvent is infused into the solvent mixture at a
temperature between about -10.degree. C. and about 100.degree. C.
and at an infusion rate of from about 0.01 ml per minute to about
1000 ml per minute per volume of 50 ml to produce a suspension of
precipitated non-aggregated solid particles of the compound with a
substantially uniform mean diameter of less than 10 .mu.m.
Agitation (e.g., by stirring) of the solution being infused with
the precipitating nonsolvent is preferred. The nonsolvent may
contain a surfactant to stabilize the particles against
aggregation. The particles are then separated from the solvent.
Depending on the solid compound and the desired particle size, the
parameters of temperature, ratio of nonsolvent to solvent, infusion
rate, stir rate, and volume can be varied according to the
invention. The particle size is proportional to the ratio of
nonsolvent:solvent volumes and the temperature of infusion and is
inversely proportional to the infusion rate and the stirring rate.
The precipitating nonsolvent may be aqueous or non-aqueous,
depending upon the relative solubility of the compound and the
desired suspending vehicle.
[0099] Temperature Shift Precipitation
[0100] Temperature shift precipitation technique, also known as the
hot-melt technique, is disclosed in U.S. Pat. No. 5,188,837 to
Domb, which is incorporated herein by reference and made a part
hereof. In an embodiment of the invention, lipospheres are prepared
by the steps of: (1) melting or dissolving a substance such as a
drug to be delivered in a molten vehicle to form a liquid of the
substance to be delivered; (2) adding a phospholipid along with an
aqueous medium to the melted substance or vehicle at a temperature
higher than the melting temperature of the substance or vehicle;
(3) mixing the suspension at a temperature above the melting
temperature of the vehicle until a homogenous fine preparation is
obtained; and then (4) rapidly cooling the preparation to room
temperature or below.
[0101] Solvent Evaporation Precipitation
[0102] Solvent evaporation precipitation techniques are disclosed
in U.S. Pat. No. 4,973,465 which is incorporated herein by
reference and made a part hereof. The '465 patent discloses methods
for preparing microcrystals including the steps of: (1) providing a
solution of a pharmaceutical composition and a phospholipid
dissolved in a common organic solvent or combination of solvents,
(2) evaporating the solvent or solvents and (3) suspending the film
obtained by evaporation of the solvent or solvents in an aqueous
solution by vigorous stirring. The solvent can be removed by adding
energy to the solution to evaporate a sufficient quantity of the
solvent to cause precipitation of the compound. The solvent can
also be removed by other well known techniques such as applying a
vacuum to the solution or blowing nitrogen over the solution.
[0103] Reaction Precipitation
[0104] Reaction precipitation includes the steps of dissolving the
pharmaceutical compound into a suitable solvent to form a solution.
The compound should be added in an amount at or below the
saturation point of the compound in the solvent. The compound is
modified by reacting with a chemical agent or by modification in
response to adding energy such as heat or UV light or the like to
such that the modified compound has a lower solubility in the
solvent and precipitates from the solution.
[0105] Compressed Fluid Precipitation
[0106] A suitable technique for precipitating by compressed fluid
is disclosed in WO 97/14407 to Johnston, which is incorporated
herein by reference and made a part hereof. The method includes the
steps of dissolving a water-insoluble drug in a solvent to form a
solution. The solution is then sprayed into a compressed fluid,
which can be a gas, liquid or supercritical fluid. The addition of
the compressed fluid to a solution of a solute in a solvent causes
the solute to attain or approach supersaturated state and to
precipitate out as fine particles. In this case, the compressed
fluid acts as an anti-solvent which lowers the cohesive energy
density of the solvent in which the drug is dissolved.
[0107] Alternatively, the drug can be dissolved in the compressed
fluid which is then sprayed into an aqueous phase. The rapid
expansion of the compressed fluid reduces the solvent power of the
fluid, which in turn causes the solute to precipitate out as fine
particles in the aqueous phase. In this case, the compressed fluid
acts as a solvent.
[0108] Other Methods for Preparing Particles
[0109] The particles of the present invention can also be prepared
by mechanical grinding of the active agent. Mechanical grinding
include such techniques as jet milling, pearl milling, ball
milling, hammer milling, fluid energy milling or wet grinding
techniques such as those disclosed in U.S. Pat. No. 5,145,684,
which is incorporated herein by reference and made a part
hereof.
[0110] Another method to prepare the particles of the present
invention is by suspending an active agent. In this method,
particles of the active agent are dispersed in an aqueous medium by
adding the particles directly into the aqueous medium to derive a
pre-suspension. The particles are normally coated with a surface
modifier to inhibit the aggregation of the particles. One or more
other excipients can be added either to the active agent or to the
aqueous medium.
EXAMPLE 1
Preparation of 1% Itraconazole Suspension with Deoxycholic acid
coating
[0111] Each 100 mL of suspension contains:
1 Itraconazole 1.0 g (1.0% w/v) Deoxycholic Acid, Sodium Salt,
Monohydrate 0.1 g (0.1% w/v) Poloxamer 188, NF 0.1 g (0.1% w/v)
Glycerin, USP 2.2 g (2.2% w/v) Sodium Hydroxide, NF (0.1 N or 1.0
N) for pH Adjustment Hydrochloric Acid, NF (0.1 N or 1.0 N) for pH
Adjustment Sterile Water for Injection, USP QS Target pH (range)
8.0 (6 to 9)
[0112] Preparation of Surfactant Solution (2 Liters) for
Microprecipitation
[0113] Fill a properly cleaned tank with Sterile Water for
Injection and agitate. Add the required amount of glycerin and stir
until dissolution. Add the required amount of deoxycholic acid,
sodium salt monohydrate and agitate until dissolution. If
necessary, adjust the pH of the surfactant solution with minimum
amount of sodium hydroxide and/or hydrochloric acid to a pH of 8.0.
Filter the surfactant solution through a 0.2 .mu.m filter.
Quantitatively transfer the surfactant solution to the vessel
supplying the homogenizer. Chill the surfactant solution in the
hopper with mixing.
[0114] Preparation of Replacement Solution
[0115] Preparation of 4 liters of replacement solution. Fill a
properly cleaned tank with WFI and agitate. Add the weighed
Poloxamer 188 (Spectrum Chemical) to the measured volume of water.
Begin mixing the Poloxamer 188/water mixture until the Poloxamer
188 has completely dissolved. Add the required amount of glycerin
and agitate until dissolved. Once the glycerin has completely
dissolved, add the required amount of deoxycholic acid, sodium salt
monohydrate and stir until dissolution. If necessary, adjust the pH
of the wash solution with the minimum amount sodium hydroxide
and/or hydrochloric acid to a pH of 8.0. Filter the replacement
solution through a 0.2 .mu.m membrane filter.
[0116] Preparation of Drug Concentrate
[0117] For a 2-L batch, add 120.0 mL of N-methyl-2-pyrrolidinone
into a 250-mL beaker. Weigh 2.0 g Poloxamer 188. Weigh 20.0 g of
itraconazole (Wyckoff). Transfer the weighed Poloxamer 188 to the
250 mL beaker with N-methyl-2-pyrrolidinone. Stir until dissolved,
then add the itraconazole. Heat and stir until dissolved. Cool the
drug concentrate to room temperature and filter through a
0.2-micron filter.
[0118] Microprecipitation
[0119] Add sufficient WFI to the surfactant solution already in the
vessel supplying the homogenizer so that the desired target
concentration is reached. When the surfactant solution is cooled,
start adding the drug concentrate into the surfactant solution with
continuous mixing.
[0120] Homogenization
[0121] Slowly increase the pressure of the homogenizer until the
operating pressure 10,000 psi has been reached. Homogenize the
suspension with recirculation while mixing. For 2,000 mL of
suspension at 50 Hz, one pass should require approximately 54
seconds. Following homogenization, collect a 20-mL sample for
particle size analysis. Cool the suspension.
[0122] Wash Replacement
[0123] The suspension is then divided and filled into 500-mL
centrifuge bottles. Centrifuge until clean separation of sediment
is observed. Measure the volume of supernatant and replace with
fresh replacement solution, prepared earlier. Quantitatively
transfer the precipitate from each centrifuge bottle into a
properly cleaned and labeled container for resuspension (pooled
sample). Resuspension of the pooled sample is performed with a high
shear mixer until no visible clumps are observed. Collect a 20-mL
sample for particle size analysis.
[0124] The suspension is then divided and filled into 500-mL
centrifuge bottles. Centrifuge until clean separation of sediment
is observed. Measure the volume of supernatant and replace with
fresh replacement solution, prepared earlier. Quantitatively
transfer the precipitate from each centrifuge bottle into a
properly cleaned and labeled container for resuspension (pooled
sample). Resuspension of the pooled sample is performed with a high
shear mixer until no visible clumps are observed. Collect a 20-mL
sample for particle size analysis.
[0125] Second Homogenization
[0126] Transfer the above suspension to the hopper of the
homogenizer and chill the suspension with mixing. Slowly increase
the homogenizer pressure until an operating pressure 10,000 psi has
been reached. Homogenize while monitoring the solution temperature.
Following homogenization, cool the suspension and collect three
30-mL samples for particle analysis. Collect the remaining
suspension in a 2-liter bottle.
[0127] Filling
[0128] Based on acceptable particle size determination testing
(mean volume-weighted diameter of 50 nm to 5 microns), collect 30
mL samples in 50 mL glass vials with rubber stoppers.
EXAMPLE 2
Preparation of 1% Itraconazole Nanosuspension with Phospholipid
Coating.
[0129] Each 100 mL of suspension contains
2 Itraconazole 1.0 g (1.0% w/v) Phospholipids (Lipoid E 80) 1.2 g
(1.2% w/v) Glycerin, USP 2.2 g (2.2% w/v) Sodium Hydroxide, NF (0.1
N or 1.0 N) for pH Adjustment Hydrochloric Acid, NF (0.1 N or 1.0
N) for pH Adjustment Sterile Water for Injection, USP QS Target pH
(range) 8.0 (7.5 to 8.5)
[0130] Preparation of Surfactant Solution (2 Liters) for
Microprecipitation
[0131] The surfactant solution is prepared in two phases. Phase 1
is dispersed phospholipids, whereas Phase 2 includes filtered
glycerin. The two fractions are combined prior to pH
adjustment.
[0132] Phase 1: Fill a properly cleaned vessel with approximately
700 mL of Sterile Water for Injection, USP (WFI) with agitation at
50-500 rpm. Increase the temperature of the filtrate to 50.degree.
C.-70.degree. C. and add the required amount of phospholipids with
mixing at 50-500 rpm until complete dispersion is achieved.
Document the time and temperature at which the phospholipids were
added and at which it was dispersed. Determine the total mixing
time required to disperse the phospholipids. Cool the surfactant
solution to 18.degree. C.-30.degree. C. prior to the addition of
glycerin.
[0133] Phase 2: Fill a properly cleaned vessel with approximately
700 m/L of WFI with agitation at 50-500 rpm. Add the required
amount of glycerin at 18.degree. C.-30.degree. C. and agitate at
50-500 rpm until dissolution.
[0134] Combined Phases: Filter the glycerin solution through a 0.2
.mu.m filter set-up into Phase 1 (at 18.degree. C.-30.degree. C.)
while mixing at 50-500 rpm. Volume is approximately 1.4 liters.
Record the pH of the surfactant solution. If necessary, adjust the
pH of the surfactant solution with a minimum amount of sodium
hydroxide and/or hydrochloric acid to a pH of 8.0.+-.0.5. Measure
the volume of the surfactant solution at 18.degree. C.-30.degree.
C. using a 2-L graduated cylinder.
[0135] Quantitatively transfer the surfactant solution to the
vessel supplying the homogenizer (Avestin C-160). Chill the
surfactant solution in the hopper with mixing at a speed with an
observable solution vortex until the temperature is not more than
10.degree. C.
[0136] Preparation of Replacement Solution (4 L)
[0137] The replacement solution is prepared in two phases. Phase 1
includes dispersed phospholipids, whereas Phase 2 includes filtered
glycerin. The two fractions are combined prior to pH
adjustment.
[0138] Phase 1: Fill a properly cleaned vessel with approximately
1.4 liters of WFI with agitation at 50-500 rpm. Increase the
temperature of the water to 50.degree. C.-70.degree. C. and add the
required amount of phospholipids with mixing at 50-500 rpm until
complete dispersion is achieved. Cool the surfactant solution to
18.degree. C.-30.degree. C. prior to the addition of glycerin.
[0139] Phase 2: Fill a properly cleaned vessel with approximately
1.4 L of WFI with agitation at 50-500 rpm. Add the required amount
of glycerin and agitate at 50-500 rpm until dissolution.
[0140] Combined Phases: Filter the glycerin solution through a 0.2
.mu.m filter set-up into Phase 1 (at 18.degree. C.-30.degree. C.)
while mixing at 50-500 rpm. Dilute to volume with Water for
Injection to 4.0 L using a graduated cylinder. Record the pH of the
wash solution. If necessary, adjust the pH of the wash solution
with the minimum amount sodium hydroxide and/or hydrochloric acid
to a pH of 8.0.+-.0.5.
[0141] Preparation of Drug Concentrate
[0142] For a 2-L batch, add 120.0 mL of N-methyl-2-pyrrolidinone
(Pharmasolve.RTM., ISP) into a 250-mL beaker. Weigh 20.0 g of
itraconazole (Wyckoff). Transfer the weighed itraconazole to the
250-mL beaker with NMP at NMT 70.degree. C. Maintain below
70.degree. C. and stir at 100-1000 rpm until dissolved. Cool the
drug concentrate to 18.degree. C.-30.degree. C. Filter the drug
concentrate through a prefilter and filter set-up. Use one
polypropylene prefilter SBPP and two 0.2 .mu.m filters at 15 psi
and ambient temperature. Transfer the drug concentrate to three
60-mL syringes and attach syringe needles to the luer connections
of the syringes. Using the syringes, determine the volume of drug
concentrate.
[0143] Microprecipitation
[0144] Add Water for Injection to the surfactant solution already
in the vessel supplying the homogenizer. The amount of water at
18.degree. C.-30.degree. C. added should be calculated as:
V=2,000 mL-Volume of Drug Concentrate-Volume of Surfactant
Solution
[0145] Mount each syringe needle assembly using a syringe pump.
Position the outlet of the needle on top of the vessel. When the
surfactant solution is not more than 10.degree. C., start adding
the drug concentrate into the surfactant solution with continuous
mixing at a speed needed to create a distinctive solution vortex.
The concentrate should be added so that the drops hit the point of
highest shear, at the bottom of the vortex. The rate of addition
should be approximately 2.5 mL/min.
[0146] Homogenization
[0147] An Avestin C160 homogenizer was used. Slowly increase the
pressure of the homogenizer until the operating pressure 10,000 psi
has been reached. Homogenize the suspension for 20 passes (18
minutes) with recirculation while mixing at 100-300 rpm and
maintaining the suspension temperature below 70.degree. C. For
2,000 mL of suspension at 50 Hz, one pass requires approximately 54
seconds. Following homogenization, collect a 20 mL sample in a 50
mL glass vial for particle size analysis. Cool the suspension to
not more than 10.degree. C.
[0148] Wash Replacement
[0149] The suspension is then divided and filled into 500-mL
centrifuge bottles. Set the speed for the centrifuge at 11,000 rpm
using the rotor SLA-3000, Superlite equivalent to approximately
20,434 g. The total centrifuge time is 60 min at not more than
10.degree. C. Measure the volume of supernatant and replace with
fresh replacement solution. Using spatula(s), quantitatively
transfer the precipitant from each centrifuge bottle into a
properly cleaned and labeled container for resuspension (pooled
sample). Resuspension of the pooled sample is performed with a high
shear mixing until no visible clumps are observed.
[0150] Second Washing and Centrifuging Step
[0151] The suspension is then divided and filled into 500-mL
centrifuge bottles. Set the speed for the centrifuge at 11,000 rpm
using the rotor SLA-3000, Superlite equivalent to approximately
20,434 g. The total centrifuge time is 60 min at not more than
10.degree. C. Measure the volume of supernatant and replace with
fresh replacement solution. Using spatula(s), quantitatively
transfer the precipitant from each centrifuge bottle into a
properly cleaned and labeled container for resuspension (pooled
sample). Resuspension of the pooled sample is performed under
high-shear mixing until no visible clumps are observed. Record the
pH of the suspension. If necessary, adjust the pH of the suspension
with the minimum amount sodium hydroxide and/or hydrochloric acid
to a pH of 8.0.+-.0.5.
[0152] Second Homogenization
[0153] Transfer the above suspension to the hopper of the
homogenizer. Chill the suspension with mixing until the temperature
is less than 10.degree. C. Slowly increase the pressure until an
operating pressure of 10,000 psi has been reached. Homogenize for
20 passes (18 minutes) while maintaining the solution temperature
below 70.degree. C. Following homogenization, cool the suspension
to less than 1.degree. C. and collect three 30-mL samples for
particle-size analysis. Collect the remaining suspension in a
2-liter bottle. Sparge the suspension with nitrogen gas for 10 min.
Ensure the nitrogen gas is filtered through a 0.2 .mu.m filter.
[0154] Filling
[0155] Based on acceptable particle size determination testing
(mean volume-weighted diameter of 50 to 1000 nm), collect 30-mL
samples in 50-mL glass vials with PTFE.RTM.-coated stoppers. Purge
the headspace of each vial with nitrogen prior to sealing.
EXAMPLE 3
Other Formulations of Itraconazole Suspensions
[0156] Other formulations of itraconazole suspensions with
different combinations of the surfactants can also be prepared
using the method described in Example 1 or Example 2. Table 1
summarizes the compositions of the surfactants of the various
itraconazole suspensions.
3TABLE 1 Summary of the compositions of the various 1% itraconazole
suspensions Formulation No. Surfactants in the formulation Amount*
1 Poloxamer 188 0.1% Deoxycholate 0.1% Glycerin 2.2% 2 Poloxamer
188 0.1% Deoxycholate 0.5% Glycerin 2.2% 3 Poloxamer 188 2.2%
Deoxycholate 0.1% Glycerin 2.2% 4 Poloxamer 188 2.2% Deoxycholate
0.5% Glycerin 2.2% 9 Solutol 0.3% Deoxycholate 0.5% Glycerin 2.2%
14331-1 Solutol 1.5% Glycerin 2.2% 14443-1 Albumin 5% 14
Phospholipid 2.2% Deoxycholate 0.5% Glycerin 2.2% Na.sub.2PO.sub.4
0.14% A6 Phospholipid 1.2% Glycerin 2.2% B Phospholipid 1.2%
Glycerin 2.2% N-methyl-2-pyrrolidinone trace C Phospholipid 1.2%
Glycerin 2.2% Lactic acid trace 14412-3 Phospholipid 1.2%
Hydroxyethyl starch 1.0% Glycerin 2.2% TRIS 0.06% *% by weight of
the final volume of the suspension (w/v)
EXAMPLE 4
Comparison of the Acute Toxicity Between Commercially Available
Itraconazole Formulation (sporanox.RTM.) and the Suspension
Compositions of the Present Invention
[0157] The acute toxicity of the commercially available
itraconazole formulation (SPORANOX.RTM.) is compared to that of the
various 1% itraconazole formulations in the present invention as
listed in Table 1. SPORANOX.RTM. is available from Janssen
Pharmaceutical Products, L.P. It is available as a 1% intravenous
(I.V.) solution solubilized by hydroxypropyl-.beta.-cyclodextrin.
The results are shown in Table 2 with the maximum tolerated dose
(MTD) indicated for each formulation.
4TABLE 2 Comparison of the acute toxicity of various formulations
of itraconazole Formulation Number Results and Conclusions SPORANOX
.RTM. I.V. LD.sub.10 = 30 mg/kg MTD = 20 mg/kg (slight ataxia) 1
MTD = 320 mg/kg; NOEL = 80 mg/kg Spleen obs.sup.b: 320 mg/kg Red
ears/feet: .gtoreq.160 mg/kg 2 MTD = 320 mg/kg Spleen obs.sup.b:
320 mg/kg Slight lethargy: 320 mg/kg Red urine: .gtoreq.80 mg/kg
Tail obs.sup.c: .gtoreq.40 mg/kg 3 MTD = 160 mg/kg; NOEL = 80 mg/kg
Spleen obs.sup.b: 320 mg/kg Red ears/feet: .gtoreq.160 mg/kg 4 MTD
= 160 mg/kg LD.sub.20 = 320 mg/kg Spleen obs.sup.b: 320 mg/kg
Slight lethargy: 320 mg/kg Red urine: .gtoreq.40 mg/kg Tail
obs.sup.c: .gtoreq.40 mg/kg 9 LD.sub.60 = 320 mg/kg; MTD = 160
mg/kg Spleen obs.sup.b: 320 mg/kg Tail obs: 320 mg/kg Red
ears/feet: .gtoreq.160 mg/kg Red urine: .gtoreq.40 mg/kg 14331-1
MTD = 40 mg/kg; NOEL = 40 mg/kg LD.sub.40 = 80 mg/kg 14443-1
LD.sub.40 = 80 mg/kg; NOEL = 40 mg/kg 14 MTD = 320 mg/kg; NOEL =
40-80 mg/kg Spleen obs.sup.b: 320 mg/kg Ataxia = 320 mg/kg Tail obs
= 320 mg/kg A6 MTD = 320 mg/kg; NOEL = 160 mg/kg Spleen obs.sup.b:
320 mg/kg B MTD = 320 mg/kg; NOEL = 80 mg/kg Spleen obs.sup.b: 160
mg/kg Red ears/feet: .gtoreq.160 mg/kg C MTD = 320 mg/kg; NOEL = 80
mg/kg Spleen obs.sup.b: .gtoreq.160 mg/kg Red ears/feet:
.gtoreq.160 mg/kg 14412-3 MTD = 320 mg/kg; NOEL = 80 mg/kg Spleen
obs.sup.b: .gtoreq.160 mg/kg .sup.acyclodextrin =
hydroxypropyl-.beta.-cyclodextrin .sup.bSpleen obs = Enlarged
and/or pale .sup.cTail obs = gray to black and/or necrosis
LD.sub.10 = Lethal dose resulting in 10% mortality LD.sub.40 =
Lethal dose resulting in 40% mortality LD.sub.50 = Lethal dose
resulting in 50% mortality NOEL = No effect level MTD = Maximum
tolerated dose
[0158] The data in Table 2 indicated that the animals tolerated a
much higher level of the antifungal agent itraconazole when
formulated in a nanosuspension than when formulated as a solution
with cyclodextrin. It may be thought that the reason for the
increased tolerability is associated with not using cyclodextrin.
However, cyclodextrin, by itself, at the levels used in Sporanox
would not cause the degree of toxicity observed. Rather, it is
believe, the reason lies in alteration of the pharmacokinetic
profile caused by the nanosuspension.
EXAMPLE 5
Pharmacokinetic Comparison of SPORANOX.RTM. vs. Suspension
Formulation of Itraconazole
[0159] Young adult, male Sprague Dawley rats were treated
intravenously (IV) via a caudal tail vein with a single injection
at a rate of 1 ml/min. with either SPORANOX.RTM. Injection, or
Formulations 1 and B at 20,40, and 80 mg/kg, or Formulations 3,14,
A6 and C at 80 mg/kg.
[0160] Following administration, the animals were anesthetized and
retro-orbital blood was collected at different time points (n=3).
The time points were as follows: 0.03, 0.25, 0.5, 1, 2, 4, 6, 8,
24, 48, 96, 144, 192, 288, and 360 hours (SPORANOX.RTM. Injection
only to 192 hours). Blood was collected into tubes with EDTA and
centrifuged at 3200 rpm for 15 minutes to separate plasma. The
plasma was stored frozen at -70.degree. C. until analysis. The
concentration of the parent itraconazole and the metabolite
hydroxy-itraconazole were determined by high-performance liquid
chromatography (HPLC). Pharmacokinetic (PK) parameters for
itraconazole (ITC) and hydroxy-itraconazole (OH-ITC) were derived
using noncompartmental methods with WinNonlin.RTM. Professional
Version 3.1 (Pharsight Corp., Mountain View, Calif.).
[0161] Table 3 provides a comparison of the plasma pharmacokinetic
parameters determined for each itraconazole formulation. Plasma
itraconazole was no longer detected at 24 hours for SPORANOX.RTM.
Injection at 5 mg/kg, at 48 hours for SPORANOX.RTM. Injection at 20
mg/kg, and at 96 hours for Formulations 1 and B. Plasma
hydroxy-itraconazole was initially detected at 0.25 hours for
SPORANOX.RTM. Injection and Formulations 1 and B. Plasma
hydroxy-itraconazole was initially detected at 0.25 hours for
SPORANOX.RTM. Injection at 5 and 20 mg/kg and Formulations 1 and ;B
at 20 mg/kg, Hydroxy-itraconazole was no longer detected at 48
hours for SPORANOX.RTM. Injection at 5 mg/kg, at 96 hours for
SPORANOX.RTM. Injection at 20 mg/kg, and at 144 hours for
Formulations 1 and B.
5TABLE 3 Comparison of Plasma Pharmacokinetic Parameters for
Sporanox and a Suspension Formulation After IV Administration in
Rats Dosage/Formulation PK Pa- 5 mg/kg 20 mg/kg 40 mg/kg 80 mg/kg
Analyte rameters Spor Spor 1 B 1 B 1 B A6 C 3 14 C max 2.42 13.12
30.41 9.10 119.16 10.20 446.33 15.20 39.72 53.19 365.09 68.15
(ug/ml) Itraconazole T max (h) 0.03 0.03 0.03 0.03 0.03 0.03 0.03
0.03 0.03 0.03 0.03 0.03 AUC 3.90 28.25 16.70 15.79 42.67 36.11
143.70 80.31 58.71 94.19 108.87 85.53 (0-.infin.) (ug .multidot.
h/ml) T.sub.1/2 (h) 2.75 5.36 14.36 14.54 23.95 20.49 25.89 28.63
54.02 33.75 38.46 31.17 CL (ul/h) 320.17 176.97 299.35 316.67
234.38 276.90 139.18 249.04 340.64 212.33 183.71 233.83 MRT (h)
2.57 4.48 13.29 15.32 24.37 28.76 27.45 52.84 58.21 46.85 31.21
41.93 Hydroxy-Itraconazole C max 0.38 0.78 0.40 0.44 0.61 0.69 1.03
0.48 0.32 0.56 0.52 0.51 (ug/ml) T max (h) 4.04 4.0 24 8 24 24 24
48 48 24.0 24.0 24.0 AUC 3.96 13.41 17.89 20.71 37.71 44.69 70.24
56.01 47.27 59.40 51.27 51.89 (0-.infin.) (ug .multidot. h/ml)
T.sub.1/2 (h) 7.98 5.89 15.50 18.06 22.27 28.12 23.21 36.45 60.87
38.84 50.29 25.50 MRT (h) 7.55 12.17 30.99 29.23 43.06 36.02 46.80
68.35 74.88 65.71 60.81 58.02
[0162] FIG. 5 compares the pharmacokinetics (PK) of SPORANOX.RTM.
with Formulation 1 suspension of itraconazole particles. Because,
as shown above, the present suspension formulation is less toxic
than Sporanox.RTM., it was administered at higher amounts in this
equitoxic experiment. Sporanox was dosed at 20 mg/kg and
Formulation 1 at 80 mg/kg. The SPORANOX.RTM. decreases in plasma
concentration relatively quickly, over 20 hours. The nanosuspension
plasma levels remain elevated for approximately 3-4 times longer.
The nanosuspension exhibits an initial minimum at 30 minutes in the
plasma level. This corresponds to a nadir in plasma concentration
due to sequestration of the drug nanocrystals by the macrophages of
the spleen and liver, thus temporarily removing drug from
circulation. However, the drug levels rebound quickly, as the
macrophages apparently release the drug into the circulation.
Furthermore, the nanosuspension drug is metabolized effectively, as
is shown by the PK curve for the hydroxy itraconazole metabolite.
The rate of appearance of the metabolite for the nanosuspension is
delayed, compared with the PK curve for the metabolite for the
SPORANOX.RTM. formulation. However, as with the case of the parent
molecule for the nanosuspension, the metabolite persists in
circulation for a much longer time than is the case with the
metabolite for the SPORANOX.RTM. formulation. When the AUC (area
under the blood concentration vs time curve) is normalized by the
dose, the nanosuspension is at least as bioavailable as
SPORANOX.RTM..
EXAMPLE 6
Acute Toxicity Of Fast Dissolving Nanosuspensions
[0163] Additional experiments were performed. Itraconazole
nanosuspensions were formulated differently, so as to dissolve much
more readily in blood. This was accomplished by making the
particles either smaller or amorphous, or both. These acute
toxicity of these formulations is described for formulation entries
14331-1 and 14443-1 in Table 1. In contrast to the slowly
dissolving nanosuspensions, the fast dissolving nanosuspension
caused death in the animals at much lower levels, similar to what
was found with SPORANOX.RTM.. Since these fast dissolving
nanosuspensions did not contain cyclodextrin, it is clear that this
excipient was not responsible for the toxicity. Rather the rapid
dissolution, resulting in immediate availability of the drug in the
blood was the causative factor. The drug level for the rapidly
dissolving formulation, Form A, is much higher than that attained
by the slow dissolving (macrophage targeting) formulation, Form B,
as determined in an in vitro dissolution experiment. This involved
a plasma simulating media consisting of 5% albumin/ Sorenson's
buffer. Results are shown in FIG. 6.
EXAMPLE 7
Antifungal Efficacy Studies
[0164] Normal and immuno-suppressed (prednisolone administered
twice daily on the day before and on the day of inoculation) rats
inoculated with 9.5.times.10.sup.6 or 3.times.10.sup.6 cfu C.
albicans/ml saline once intravenously were intravenously treated
with SPORANOX.RTM. Injection once daily for ten consecutive days,
with the first dose given 4 to 5 hours after inoculation.
SPORANOX.RTM. Injection rats were dosed at 5 or 20 mg/kg for the
first 2 days, then at 5 or 10 mg/kg for the remaining 8 days, due
to toxicity at 20 mg/kg after 2 days of dosing. Similarly,
immuno-suppressed rats inoculated with 1.times.10.sup.6.5 cfu C.
albicans/ml saline were intravenously treated with Formulation 1 or
B each at 20,40, or 80 mg/kg once every other day for ten days,
beginning the day of inoculation. The SPORANOX.RTM. Injection,
Formulation 1, and Formulation B treatment rats were terminated 11
days after the C. albicans inoculation and the kidneys were
collected, weighed and cultured for determination of C. albicans
colony counts and itraconazole and hydroxy-itraconazole
concentration. Kidneys were collected from untreated control rats
when a moribund condition was observed or when an animal had a 20%
body weight. In addition, body weights were measured periodically
during the course of each study.
[0165] Comparison of results for immuno-suppressed rats treated
with SPORANOX.RTM. Injection and Formulations 1 and B are shown in
Table 4 and FIG. 7. Daily SPORANOX.RTM. Injection treatment at
10-20 mg/kg appeared to be slightly more effective than daily
treatment with SPORANOX.RTM. Injection at 5 mg/kg. Based on kidney
colony counts, every other day dosing at 20 mg/kg of Formulation 1
or B appeared to be as effective as every day dosing with
SPORANOX.RTM. Injection at 20 mg/kg and possibly more effective
than SPORANOX.RTM. Injection at 5 mg/kg (i.e., the recommended
clinical dose), whereas the higher doses for both Formulation 1 and
B appeared to most effective, based on kidney colony counts (i.e.,
C. albicans not detected) and increased kidney itraconazole
concentration.
6TABLE 4 Mean C. albicans Colony Count and Itraconazole and
Hydroxy-Itraconazole Concentration in Kidney Concentration C.
albicans Titer in Kidney Count ITC OH-ITC Treatment (cfu/g)
Incidence (.mu.g/g) (.mu.g/g) No Treatment (3 .times. 10.sup.6
cfu/ml) 6.9 .times. 10.sup.4 6/6 -- -- SPORANOX .RTM., 5 mg/kg, (3
.times. 10.sup.6 cfu/ml) 96.5 6/6 1.2 1.5 SPORANOX .RTM., 10-20
mg/kg, (3 .times. 10.sup.6 cfu/ml) 12.4 4/6 8.5 8.0 No Treatment
(2.5 .times. 10.sup.6 cfu/ml) 3.5 .times. 10.sup.5 6/6 -- --
Formulation 1, 20 mg/kg, (2.5 .times. 10.sup.6 cfu/ml) 5.3 4/6 6.1
5.7 Formulation 1, 40 mg/kg, (2.5 .times. 10.sup.6 cfu/ml) 0 0/6
18.5 6.0 Formulation 1, 80 mg/kg, (2.5 .times. 10.sup.6 cfu/ml) 0
0/6 41.2 6.2 No Treatment (2.5 .times. 10.sup.6 cfu/ml) 8.0 .times.
10.sup.4 6/6 -- -- Formulation B, 20 mg/kg, (2.5 .times. 10.sup.6
cfu/ml) 8.9 4/6 2.5 2.5 Formulation B, 40 mg/kg, (2.5 .times.
10.sup.6 cfu/ml) 0 0/6 7.8 4.0 Formulation B, 80 mg/kg, (2.5
.times. 10.sup.6 cfu/ml) 0 0/6 21.3 4.6
[0166] In the examples above, a nanosuspension formulation of an
anti-fungal agent was shown to be less toxic than a conventional
totally soluble formulation of the same drug. Thus, more of the
drug could be administered without eliciting adverse effects.
Because the nanoparticles of the drug did not immediately dissolve
upon injection, they were trapped in a depot store in the liver and
spleen. These acted as prolonged release sanctuaries, permitting
less frequent dosing. The greater dosing that could be administered
permitted greater drug levels to be manifested in the target
organs, in this case, the kidney (FIG. 8). The greater drug levels
in this organ led to a greater kill of infectious organisms. (FIG.
9).
EXAMPLE 8
Resistant Strain Anti-fungal Efficacy Test
[0167] A lethal dose of a C. albicans strain c43 (ATCC number
201794) (MIC.sub.80=16 .mu.g/ml for SPORANOX.RTM. itraconazole;
8-16 for Vfend, and 0.1 for Cancidas) was administered to an
immunocompromised rat model (prednisolone qd). 24h later, test
groups (n=6) were treated q2d with 20,40, or 80 mg/kg NANOEDGE.TM.
itraconazole nanosuspension. Control groups included a no treatment
arm, Sporanox(D (10 mg/kg/d), Vfend.RTM. (10 mg/kg/d), and
Cancidas.RTM. (1 mg/kg/d). Treatment was continued for 10 days.
Survival and kidney cfu/g were assessed.
[0168] The number of surviving animals after 6 and 10 days, were
respectively: Sporanox (3,0), 20 and 40 mg/kg nanosuspension (5,3),
80 mg/kg nanosuspension (6,4), Vfend (0,0), Cancidas (0,0). FIG.
10.
[0169] It can be concluded that the greater dosing possible with
the itraconazole nanosuspension can effectively treat infections of
C. albicans strains, conventionally assumed to be resistant to
itraconazole, resulting in increased survival in an
immunocompromised rat model.
[0170] Current definitions of sensitive and resistant fungal
strains presume a specified dose of itraconazole that is
administered, using conventional dosage forms. Greater drug
loading, attendant with nanosuspension injections, may permit
treatment of what are currently considered itraconazole-resistant
C. albicans infections.
EXAMPLE 9
Prophetic Examples of Other Triazole Antifungal Agents
[0171] The present invention contemplates preparing a 1% suspension
of submicron- or micron size of a triazole antifungal agent using
the method described in Example 1 or Example 2 and the formulations
described in Example 3 with the exception that the antifungal agent
is a triazole antifungal agent other than itraconazole. Examples of
triazole antifungal agents that can be used include, but are not
limited to, ketoconazole, miconazole, fluconazole, ravuconazole,
voriconazole, saperconazole, eberconazole, genaconazole,
clotrimazole, econazole, oxiconazole, sulconazole, terconazole,
tioconazole, and posaconazole.
EXAMPLE 10
Prophetic Example of a Non-Triazole Antifungal Agent
[0172] The present invention contemplates preparing a 1% suspension
of submicron- or micron size non-triazole antifungal agent using
the method described in Example 1 or Example 2 and the formulations
described in Example 3 with the exception that the antifungal agent
is amphotericin B, nystatin, terbinafine, anidulafungin, or
flucytosine instead of itraconazole.
[0173] From the foregoing, it will be observed that numerous
variations and modifications may be effected without departing from
the spirit and scope of the invention. It is to be understood that
no limitation with respect to the specific apparatus illustrated
herein is intended or should be inferred. It is, of course,
intended to cover by the appended claims all such modifications as
fall within the scope of the claims.
* * * * *