U.S. patent application number 14/796920 was filed with the patent office on 2016-06-16 for methods of treating cancer with high potency lipid-based platinum compound formulations administered intraperitoneally.
The applicant listed for this patent is Insmed Incorporated. Invention is credited to Jin K. LEE, Vladimir MALININ, Roman PEREZ-SOLER, Walter R. PERKINS, Frank G. Pilkiewicz, Yiyu ZOU.
Application Number | 20160166608 14/796920 |
Document ID | / |
Family ID | 40642217 |
Filed Date | 2016-06-16 |
United States Patent
Application |
20160166608 |
Kind Code |
A1 |
Pilkiewicz; Frank G. ; et
al. |
June 16, 2016 |
METHODS OF TREATING CANCER WITH HIGH POTENCY LIPID-BASED PLATINUM
COMPOUND FORMULATIONS ADMINISTERED INTRAPERITONEALLY
Abstract
One aspect of the invention relates to methods of treating
cancer in a patient comprising administering intraperitoneally to a
patient in need thereof a cancer treating effective amount of a
composition comprising a lipid-complexed platinum compound wherein
the concentration of the platinum compound of the lipid-complexed
platinum compound composition is greater than about 1.2 mg/ml.
Another aspect of the invention relates to lipid-complexed platinum
compound compositions where the concentration of the platinum
compound is greater than about 1.2 mg/ml.
Inventors: |
Pilkiewicz; Frank G.;
(Princeton Junction, NJ) ; PEREZ-SOLER; Roman;
(New York, NY) ; ZOU; Yiyu; (Bronx, NY) ;
PERKINS; Walter R.; (Neshanic Station, NJ) ; LEE; Jin
K.; (Belle Mead, NJ) ; MALININ; Vladimir;
(Plainsboro, NJ) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Insmed Incorporated |
Bridgewater |
NJ |
US |
|
|
Family ID: |
40642217 |
Appl. No.: |
14/796920 |
Filed: |
July 10, 2015 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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12122191 |
May 16, 2008 |
9107824 |
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14796920 |
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11592754 |
Nov 3, 2006 |
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12122191 |
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60734474 |
Nov 8, 2005 |
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Current U.S.
Class: |
424/450 ;
424/489; 424/649 |
Current CPC
Class: |
A61K 31/555 20130101;
A61K 33/24 20130101; A61P 35/04 20180101; A61K 31/282 20130101;
A61K 9/0019 20130101; A61K 9/1617 20130101; A61K 47/6911 20170801;
A61K 9/127 20130101 |
International
Class: |
A61K 33/24 20060101
A61K033/24; A61K 9/127 20060101 A61K009/127; A61K 47/48 20060101
A61K047/48; A61K 9/16 20060101 A61K009/16 |
Claims
1. A method of treating cancer in a patient comprising
administering intraperitoneally to a patient in need thereof a
cancer treating effective amount of a composition comprising a
lipid-complexed active platinum compound, wherein the lipid
complexed active platinum compound has a lipid to drug (L/D) ratio
of less than about 1 by weight.
2. The method of claim 1, wherein the lipid to drug ratio is about
0.10 to about 1 by weight.
3. The method of claim 2, wherein the lipid to drug ratio is about
0.10 to about 0.50 by weight.
4. The method of claim 3, wherein the lipid to drug ratio is about
0.15 to about 0.45 by weight.
5. The method of claim 4, wherein the lipid to drug ratio is about
0.20 to about 0.40 by weight.
6. The method of claim 5, wherein the lipid to drug ratio is about
0.2 by weight.
7. The method of claim 1, wherein lipid-complexed active platinum
compound has an average volume-weighted diameter of about 0.5 to
about 20 microns.
8. The method of claim 6, wherein the average volume-weighted
diameter is about 1 to about 15 microns.
9. The method of claim 7, wherein the average volume-weighted
diameter is about 2 to 10 microns.
10. The method of claim 1, wherein the concentration of the active
platinum compound in the composition is greater than about 1.2
mg/ml.
11. The method of claim 10, wherein the platinum compound
concentration is about 1.2 to about 20 mg/ml.
12. The method of claim 12, wherein the platinum compound
concentration is about 1.5 to about 5 mg/ml.
13. The method of claim 1, further comprising a liposome.
14. The method of claim 13, wherein the liposome comprises an
active platinum compound.
15. The method of claim 14, wherein the lipid-complexed active
platinum compound contains about 70 to about 100% of the total
active platinum compound in the composition.
16. The method of claim 15, wherein in the lipid-complexed active
platinum compound contains about 75 to about 99% of the total
active platinum compound.
17. The method of claim 16, wherein the lipid-complexed active
platinum compound contains about 80 to about 90% of the total
active platinum compound.
18. The method of claim 14, wherein the liposome contains about 0
to about 30% of the total active platinum compound in the
composition.
19. The method of claim 18, wherein the liposome contains about 0.5
to about 25% of the total active platinum compound.
20. The method of claim 19, wherein the liposome contains about 1
to about 20% of the total platinum compound.
21. The method of claim 13, wherein the lipid-complexed active
platinum compound contains about 0.1 to about 5% of the total lipid
in the composition,
22. The method of claim 21, wherein the lipid-complexed active
platinum compound contains about 0.25 to about 3% of the total
lipid.
23. The method of claim 13, wherein the liposome contains about 75
to about 99.5% of the total lipid in the composition.
24. The method of claim 23, wherein the liposome contains about 80
to about 95% of the total lipid
25. The method of claim 13, wherein the lipid-complexed active
platinum compound has a lipid to platinum compound ratio of about
0.10 to about 0.50.
26. The method of claim 14, wherein the liposome has a lipid to
active platinum compound ratio of about 100:1 to about 400:1 by
weight.
27. The method of claim 26, wherein the lipid to active platinum
compound ratio is about 200:1 to 400:1 by weight.
28. The method of claim 1, wherein the active platinum compound is
selected from the group consisting of cisplatin, carboplatin
(diammine(1,1-cyclobutanedicarboxylato)-platinum(II)), tetraplatin
(ormaplatin)
(tetrachloro(1,2-cyclohexanediamine-N,N')-platinum(IV)), thioplatin
(bis(O-ethyldithiocarbonato)platinum(II)), satraplatin, nedaplatin,
oxaliplatin, heptaplatin, iproplatin, transplatin, lobaplatin,
cis-aminedichloro(2-methylpyridine) platinum, JM118
(cis-amminedichloro (cyclohexylamine)platinum(II)), JM149
(cis-amminedichloro(cyclohexylamine)-trans-dihydroxoplatinum(IV)),
JM216 (bis-acetato-cis-amminedichloro(cyclohexylamine)
platinum(IV)), JM335 (trans-amminedichloro
(cyclohexylamine)dihydroxoplatinum(IV)), (trans, trans,
trans)bis-mu-(hexane-1,6-diamine)-mu-[diamine-platinum(II)]bis[dia-
mine(chloro) platinum(II)]tetrachloride, and mixture thereof.
29. The method of claim 30, wherein the active platinum compound is
cisplatin.
30. The method of claim 1, wherein the lipid-complexed active
platinum compound comprises a phosphatidyl choline.
31. The method of claim 30, wherein the phosphatidyl choline is
DPPC.
32. The method of claim 31, wherein the lipid-complexed active
platinum compound further comprises a sterol.
33. The method of claim 30, wherein the sterol is cholesterol.
34. The method of claim 30, wherein the lipid-complexed active
platinum compound does not comprise a negatively charged
phospholipid.
35. The method of claim 30, wherein the lipid complexed active
platinum compound comprises DPPC and cholesterol in a ratio of
about 1:1 to about 5:1 by weight.
36. The method of claim 35, wherein the lipid complexed active
platinum compound comprises DPPC and cholesterol in a ratio of
about 2:1 to about 4:1 by weight.
37. The method of claim 36, the lipid complexed active platinum
compound comprises DPPC and cholesterol in a ratio of about 2.25:1
by weight.
38. The method of claim 1, wherein the cancer is selected from the
following: melanoma, testis (germ cell), osteosarcoma, soft tissue
sarcoma, thyroid cancer, colon cancer, ovarian cancer, cancer of
the kidney, breast cancer, colorectal cancer, prostate cancer,
bladder cancer, uterine cancer, lung cancer, stomach cancer, liver
cancer, spleen cancer, endometrial, or squamous cell carcinomas of
the head and neck.
39. The method of claim 1, wherein the cancer is ovarian
cancer.
40. The method of claim 1, wherein the cancer is colon cancer.
41. The method of claim 1, wherein the patient is a human.
42. The method of claim 1, wherein the composition is administered
to the patient at least once every three weeks.
43. The method of claim 1, wherein the amount of platinum compound
in the composition is 60 mg/m.sup.2 or greater, 100 mg/m.sup.2 or
greater, 140 mg/m.sup.2 or greater, or 180 mg/m.sup.2 or
greater.
44. The method of claim 1, wherein the amount of platinum compound
in the composition is 100 mg/m.sup.2 or greater, and the
composition is administered to the patient at least once every
three weeks.
Description
RELATED APPLICATIONS
[0001] This application claims the benefit of priority to U.S.
patent application Ser. No. 11/592,754, filed on Nov. 3, 2006,
which claims the benefit of priority to U.S. Provisional Patent
Application No. 60/734,474, filed Nov. 8, 2005, and which are both
hereby incorporated by reference in their entirety.
INTRODUCTION
[0002] Parenteral routes of administration involve injections into
various compartments of the body. Parenteral routes include
intravenous (iv), i.e. administration directly into the vascular
system through a vein; intra-arterial (ia), i.e. administration
directly into the vascular system through an artery;
intraperitoneal (ip), i.e. administration into the abdominal
cavity; subcutaneous (sc), i.e. administration under the skin;
intramuscular (im), i.e. administration into a muscle; and
intradermal (id), i.e. administration between layers of skin. The
parenteral route is preferred over oral ones in many occurrences.
For example, when the drug to be administered would partially or
totally degrade in the gastrointestinal tract, parenteral
administration is preferred. Similarly, where there is need for
rapid response in emergency cases, parenteral administration is
usually preferred over oral.
[0003] Regional delivery of chemotherapy into the peritoneal space
via ip administration has been found to be a safe and effective
treatment for locally recurrent cancers such as, for example,
ovarian and colon cancers.
[0004] The concept of the intraperitoneal administration of
antineoplastic agents in the management of cancers such as ovarian
cancer has attracted the interest of numerous investigators. In
fact, alkylating agents, the first cytotoxic drugs to be introduced
into clinical practice, were initially examined for intraperitoneal
delivery in the early 1950s. Markman M., Cancer Treat Rev., 1986,
13, 219-242.
[0005] However, it was not until the late 1970s that both the
problems and potential of regional drug administration in the
treatment of ovarian cancer began to be thoroughly explored.
Markman M., Cancer Treat Rev., 1986, 13, 219-242; Markman M.,
Semin. Oncol., 1991, 18(suppl 3), 248-254. An important event in
the development of a rational strategy for the examination of
intraperitoneal drug delivery was the publication of a now-classic
paper by Dedrick et al., from the National Cancer Institute where,
for the first time, a sound pharmacokinetic rationale for this
approach in the management of ovarian cancer was presented. Dedrick
R L, Myers C E, Bungay P M et al., Cancer Treat. Rep., 1978, 62,
1-9.
[0006] Cisplatin--cis-diamine-dichloroplatinum (II)--is one of the
more effective anti-tumor agents used in the systemic treatment of
cancers. This chemotherapeutic drug is highly effective in the
treatment of tumor models in laboratory animals and in human
tumors, such as endometrial, bladder, ovarian and testicular
neoplasms, as well as squamous cell carcinoma of the head and neck
(Sur, et al., 1983 Oncology 40(5): 372-376; Steerenberg, et al.,
1988 Cancer Chemother Pharmacol. 21(4): 299-307). Cisplatin is also
used extensively in the treatment of lung carcinoma, both SCLC and
NSCLC (Schiller et al., 2001 Oncology 61(Suppl 1): 3-13). Other
active platinum compounds (defined below) are useful in cancer
treatment.
[0007] Like other cancer chemotherapeutic agents, active platinum
compounds such as cisplatin are typically highly toxic. The main
disadvantages of cisplatin are its extreme nephrotoxicity, which is
the main dose-limiting factor, its rapid excretion via the kidneys,
with a circulation half life of only a few minutes, and its strong
affinity to plasma proteins (Freise, et al., 1982 Arch Int
Pharmacodyn Ther. 258(2): 180-192).
[0008] Attempts to minimize the toxicity of active platinum
compounds have included combination chemotherapy, synthesis of
analogues (Prestayko et al., 1979 Cancer Treat Rev. 6(1): 17-39;
Weiss, et al., 1993 Drugs. 46(3): 360-377), immunotherapy and
entrapment in liposomes (Sur, et al., 1983; Weiss, et al., 1993).
Antineoplastic agents, including cisplatin, entrapped in liposomes
have a reduced toxicity, relative to the agent in free form, while
retaining antitumor activity (Steerenberg, et al., 1987; Weiss, et
al., 1993).
[0009] Cisplatin, however, is difficult to efficiently entrap in
liposomes or lipid complexes because of the bioactive agent's low
aqueous solubility, approximately 1.0 mg/ml at room temperature,
and low lipophilicity, both of which properties contribute to a low
bioactive agent/lipid ratio.
[0010] Liposomes and lipid complexes containing cisplatin suffer
from another problem--stability of the composition. In particular,
maintenance of bioactive agent potency and retention of the
bioactive agent in the liposome during storage are recognized
problems (Freise, et al., 1982; Gondal, et al., 1993; Potkul, et
al., 1991 Am J Obstet Gynecol. 164(2): 652-658; Steerenberg, et
al., 1988; Weiss, et al., 1993) and a limited shelf life of
liposomes containing cisplatin, on the order of several weeks at
4.degree. C., has been reported (Gondal, et al., 1993 Eur J Cancer.
29A(11): 1536-1542; Potkul, et al., 1991).
[0011] Alberts et al. have shown that as compared with iv
cisplatin, ip cisplatin significantly improves survival and has
significantly fewer toxic effects in patients with stage III
ovarian cancer and residual tumor masses of 2 cm or less. Alberts
D. S. et al., New England Journal of Medicine, 1996, 335(26),
1950-5. However, ip cisplatin has several disadvantages such as no
improvement in nephrotoxicity which is the dose-limiting
toxicity.
[0012] Additionally, both preclinical and clinical data have firmly
established that any benefits associated with employing the
intraperitoneal route of drug delivery in the treatment of ovarian
cancer are limited to a relatively well-defined small subset of
patients with this malignancy. Markman M., Cancer Treat Rev., 1986,
13, 219-242; Markman M., Semin. Oncol., 1991, 18(suppl 3), 248-254;
Markman M, Reichman B, Hakes T et al., J. Clin. Oncol., 1991, 9,
1801-1805. For example, in a series of patients treated at the
Memorial Sloan-Kettering Cancer Center (MSKCC) with combination
cisplatin-based therapy as salvage treatment of advanced ovarian
cancer, 32% (17/50) of individuals whose largest residual tumor
mass measured .ltoreq.1 cm in maximum diameter at the initiation of
ip therapy achieved a surgically documented complete response,
compared to only 5% (2/39) of patients with at least one tumor mass
>1 cm in maximum diameter. Markman M, Reichman B, Hakes T et
al., J. Clin. Oncol., 1991, 9, 1801-1805. Clearly more is needed
than just direct routes of administration to overcome the
increasingly deleterious effects of cancer.
[0013] In addition to cisplatin, a number of other antineoplastic
agents have been examined for safety and potential efficacy when
delivered by the ip route as salvage treatment of ovarian cancer.
These include carboplatin, paclitaxel, mitoxantrone, doxorubicin,
mitomycin-C, 5-fluorouracil, methotrexate, thiotepa, recombinant
interferon-.alpha., recombinant interferon-.gamma., interleukin 2
and tumor necrosis factor. Markman M., Cancer Treat Rev., 1986, 13,
219-242; Markman M., Semin. Oncol., 1991, 18(suppl 3), 248-254;
Markman M, Reichman B, Hakes T et al., J. Clin. Oncol., 1991, 9,
1801-1805; Markman M., Regional antineoplastic drug delivery in the
management of malignant disease. Baltimore: The Johns Hopkins
University Press, 1991; Berek J. S., Markman M., Int. J. Gynecol.
Cancer, 1992, 1, 26-29; Markman M, Berek J. S., Int. J. Gynecol.
Cancer, 1992, 1, 30-34; Alberts D. S., Liu P. Y., Hannigan E. V. et
al., Proc. Am. Soc. Clin. Oncol., 1995, 14, 273a; Rowinsky E. K.,
Donehower R. C., N. Engl. J. Med., 1995, 332, 1004-1014.
Combination regimens have also been explored.
[0014] Despite the advances made with ip administration of platinum
compounds, the dose limiting toxicity and low drug level in
targeted tissues of platinum compounds make most therapies fail to
improve patients' life-expectancy. It would be advantageous to
develop a platinum compound composition with potency higher than
its aqueous solubility limit at room temperature. High potency
lipid complexed platinum compound compositions would reduce the
liquid dose volume for a given dose, consequently reducing the
dosing time (duration).
SUMMARY OF THE INVENTION
[0015] It is an object of the present invention to provide a method
of treating cancer comprising administering platinum compounds as
part of a high potency lipid-based composition with lower sub-acute
toxicity, in some cases by as much as two times, than when the
platinum compound is administered without the lipid-based
composition.
[0016] It is also an object of the present invention to provide a
high potency lipid-based platinum compound composition wherein the
potency is higher than the aqueous solubility of the platinum
compound at room temperature.
[0017] It is also an object of the present invention to treat
cancer using a high potency lipid-based platinum compound
composition to reduce the volume of composition that has to be
administered to achieve the same level of effectiveness.
[0018] It is also an object of the present invention to treat
cancer by introducing platinum compounds as high potency lipid
based compositions regionally to bypass gastrointestinal
degradation that is often associated with oral administration.
[0019] The subject invention results from the realization that high
potency lipid-based platinum compound compositions presented herein
can be effectively administered intraperitoneally.
[0020] In one embodiment, the present invention relates to a method
of treating cancer in a patient comprising administering
intraperitoneally to a patient in need thereof a cancer treating
effective amount of a composition comprising a lipid-complexed
active platinum compound, wherein the lipid complexed active
platinum compound has a lipid to drug ratio of less than about 1 by
weight, e.g. about 0.10 to 1, wherein the lipid-complexed active
platinum compound comprises at least one lipid and at least one
active platinum compound.
[0021] In some embodiments, wherein lipid-complexed active platinum
compound has an average volume-weighted diameter of about 0.5 to
about 20 microns.
[0022] In some embodiments, the composition further comprises a
liposome. The liposome may comprise at least one lipid, and may
further comprise at least one active platinum compound.
[0023] In some embodiments, the composition administered in the
aforementioned method has a concentration of the platinum compound
greater than about 1.2 mg/ml. In a further embodiment the platinum
compound concentration is about 3 mg/ml. In a further embodiment
the platinum compound concentration is about 5 mg/ml.
[0024] In a further embodiment the present invention relates to the
aforementioned method, wherein the cancer is selected from the
following: melanoma, testis (germ cell), osteosarcoma, soft tissue
sarcoma, thyroid cancer, colon cancer, ovarian cancer, cancer of
the kidney, breast cancer, colorectal cancer, prostate cancer,
bladder cancer, uterine cancer, lung cancer, stomach cancer, liver
cancer, spleen cancer, endometrial, or squamous cell carcinomas of
the head and neck. In a further embodiment the cancer is ovarian or
colon cancer.
[0025] In a further embodiment the present invention relates to the
aforementioned method, wherein the patient is a human. In a further
embodiment, the composition comprising a lipid-complexed platinum
compound is administered to the patient at least once every three
weeks. In a further embodiment, the composition is administered to
the patient at least twice every three weeks. In a further
embodiment, the composition is administered to the patient at least
three times every three weeks.
[0026] In a further embodiment, the amount of platinum compound in
the composition is 60 mg/m.sup.2 or greater, 100 mg/m.sup.2 or
greater, 140 mg/m.sup.2 or greater, or 180 mg/m.sup.2 or greater.
In a further embodiment, the amount is 100 mg/m.sup.2 or greater,
and the composition is administered to the patient at least once
every three weeks.
[0027] These embodiments of the present invention, other
embodiments, and their features and characteristics, will be
apparent from the description, drawings and claims that follow.
BRIEF DESCRIPTION OF THE DRAWINGS
[0028] FIG. 1 depicts the large decrease in toxicity of ip
administration of lipid-complexed cisplatin (L-CDDP-ip) as compared
to ip administration of cisplatin (CDDP-ip).
[0029] FIG. 2 depicts the increased amount of cisplatin from
lipid-complexed cisplatin in the blood stream when administered
intraperitoneally as compared to free cisplatin administered
intraperitoneally.
[0030] FIG. 3 depicts the increased amount of cisplatin in the
blood stream from lipid-complexed cisplatin when administered
intraperitoneally as compared to free cisplatin administered
intraperitoneally.
[0031] FIG. 4 depicts the increased amount of cisplatin from
lipid-complexed cisplatin in the kidney as compared to free
cisplatin.
[0032] FIG. 5 depicts the higher amount of cisplatin from
lipid-complexed cisplatin in the liver as compared to free
cisplatin when administered intraperitoneally.
[0033] FIG. 6 depicts the higher amount of cisplatin from
lipid-complexed cisplatin in the lung as compared to free cisplatin
when administered intraperitoneally.
[0034] FIG. 7 depicts the increased amount of cisplatin from
lipid-complexed cisplatin in the spleen when administered
intraperitoneally as compared to free cisplatin administered
intraperitoneally.
[0035] FIG. 8 depicts the blood/kidney concentration ratio of
platinum from lipid-complexed cisplatin administered
intraperitoneally and free cisplatin administered
intraperitoneally.
[0036] FIG. 9 depicts the increase in blood urea nitrogen (BUN)
levels when free cisplatin is delivered either intravenously or
intraperitoneally compared to lipid-complexed cisplatin
administered by either method.
[0037] FIG. 10 depicts the survival rate for mice with implanted
viable human ovarian cancer cells line SK-OV.sub.3-ip1 after ip
administration of free cisplatin and lipid-complexed cisplatin.
[0038] FIG. 11 depicts the survival rate for mice with implanted
viable L1210 tumor cells after ip administration of lipid-complexed
cisplatin, non-cyclic temperature cisplatin liposomes, and soluble
cisplatin.
DETAILED DESCRIPTION OF THE INVENTION
Definitions
[0039] For convenience, before further description of the present
invention, certain terms employed in the specification, examples
and appended claims are collected here. These definitions should be
read in light of the remainder of the disclosure and understood as
by a person of skill in the art. Unless defined otherwise, all
technical and scientific terms used herein have the same meaning as
commonly understood by a person of ordinary skill in the art.
[0040] The term "bioavailable" is art-recognized and refers to a
form of the subject invention that allows for it, or a portion of
the amount administered, to be absorbed by, incorporated to, or
otherwise physiologically available to a subject or patient to whom
it is administered.
[0041] The term "cancer treating effective amount" as used herein
refers to the amount of lipid-complexed platinum compound
composition effective for the treatment of cancer. In one
embodiment the cancer treating effective amount of lipid-complexed
platinum compound composition is typically about 100 mg/m.sup.2 for
ip delivery in a human.
[0042] The term "CDDP" stands for cis diamminedichloroplatinum
which is used interchangeably with "cisplatin."
[0043] The term "hydrophobic matrix carrying system" is a
lipid/solvent mixture prepared during the solvent infusion process
described below.
[0044] The term "intraperitoneal" or "intraperitoneally" or "ip" as
used herein refers to administration of a therapeutic agent, such
as, for example, an antineoplastic compound, such as a platinum
compound, to the peritoneal cavity of a patient. The term
"peritoneal cavity" as used herein refers to the serous membrane
lining the abdominopelvic walls and investing the viscera.
[0045] The term "L-CDDP" stands for a lipid complexed composition
of cis diamminedichloroplatinum and is used interchangeably with
"lipid-complexed cisplatin."
[0046] The terms "lipid-complexed platinum compound" as used herein
refers to a composition comprising a lipid and a platinum compound.
In some embodiments, the lipid complexed active platinum compound
comprises a lipid bilayer, where the lipid bilayer encapsulates or
entraps the platinum compound. Exemplary lipid-complexed platinum
compounds are described in U.S. patent application Ser. No.
12/027,752, filed Feb. 7, 2008, which is hereby incorporated by
reference in its entirety.
[0047] The term "mammal" is known in the art, and exemplary mammals
include humans, primates, bovines, porcines, canines, felines, and
rodents (e.g., mice and rats).
[0048] A "patient," "subject" or "host" to be treated by the
subject method may mean either a human or non-human animal.
[0049] The term "pharmaceutically-acceptable salts" is
art-recognized and refers to the relatively non-toxic, inorganic
and organic acid addition salts of compounds, including, for
example, those contained in compositions of the present
invention.
[0050] The term "solvent infusion" is a process that includes
dissolving one or more lipids in a small, preferably minimal,
amount of a process compatible solvent to form a lipid suspension
or solution (preferably a solution) and then adding the solution to
an aqueous medium containing bioactive agents. Typically a process
compatible solvent is one that can be washed away in a aqueous
process such as dialysis. The composition that is cool/warm cycled
is preferably formed by solvent infusion. Alcohols are preferred as
solvents, with ethanol being a preferred alcohol.
[0051] "Ethanol infusion," is a type of solvent infusion that
includes dissolving one or more lipids in a small, preferably
minimal, amount of ethanol to form a lipid solution and then adding
the solution to an aqueous medium containing bioactive agents. A
"small" amount of solvent is an amount compatible with forming
liposomes or lipid complexes in the infusion process.
[0052] The term "therapeutic agent" is art-recognized and refers to
any chemical moiety that is a biologically, physiologically, or
pharmacologically active substance that acts locally or
systemically in a subject. Examples of therapeutic agents, also
referred to as "drugs", are described in well-known literature
references such as the Merck Index, the Physicians Desk Reference,
and The Pharmacological Basis of Therapeutics, and they include,
without limitation, medicaments; vitamins; mineral supplements;
substances used for the treatment, prevention, diagnosis, cure or
mitigation of a disease or illness; substances which affect the
structure or function of the body; or pro-drugs, which become
biologically active or more active after they have been placed in a
physiological environment.
[0053] The term "therapeutic index" is an art-recognized term which
refers to the ratio of a quantitative assessment of toxicity to a
quantitative assessment of efficacy of a drug, e.g.
LD.sub.50/ED.sub.50 in the case of animals. The term "LD.sub.50" is
art recognized and refers to the amount of a given toxic substance
that will elicit a lethal response in 50% of the test organisms.
This is sometimes also referred to as the median lethal dose. The
term "ED.sub.50" is art recognized and refers to the median
effective dose.
[0054] The term "treating" is art-recognized and refers to curing
as well as ameliorating at least one symptom of any condition or
disease.
Introduction
[0055] Typical human parenteral dosage amounts with current
commercial cisplatin solutions (e.g. Platinol from Bristol Myers
Squibb) are about 100 mg/m.sup.2 (unit dose) administered once
every 3 weeks intravenously for a single drug therapy. Dose
limiting factors include of course unwanted side effects (e.g.
renal toxicity, severe vomiting, etc.). With higher potency
platinum compound compositions, it would take less time to
administer the platinum compound solution because less volume would
have to be administered to achieve an equal amount of platinum
compound. For example, a 100 mg/m.sup.2 dosage with a human body
surface of 1.6 to 2 m.sup.2 (for the sake of this example, lets use
2 m.sup.2) would require administering 200 mg of platinum compound
at one time. Using current commercial cisplatin solutions (1 mg/ml,
the solubility limit at room temperature) requires infusing 200 ml
of cisplatin solution into the body. At 5.times. the potency (5
mg/ml) only 1/5 the volume or 40 ml would need to be infused, which
also means the process would take 1/5 the amount of time.
[0056] Presented herein are compositions of a platinum compound
that exceed their ordinary solubility limitations of 1 mg/ml and
methods of treating cancer therewith. The greater potency of the
platinum compound compositions is achieved by preparing the
platinum compound in compositions comprising a lipid-complexed
compound. The high potency lipid-complexed platinum compound
compositions of the present invention and methods of treating
cancer therewith also benefit from the fact that the
lipid-complexed compositions decrease the sub-acute toxicity of the
platinum compound allowing even higher doses of platinum to be
administered.
Compositions
[0057] The lipids used in forming lipid complexes and liposomes for
ip delivery of an antineoplastic agent may be synthetic,
semi-synthetic or naturally-occurring lipids, including
phospholipids, tocopherols, sterols, fatty acids, glycoproteins
such as albumin, negatively-charged lipids and cationic lipids.
[0058] Liposomes are completely closed lipid bilayer membranes
containing an entrapped aqueous volume. Liposomes used for the
parenteral delivery of an antineoplastic compound may be
unilamellar vesicles (possessing a single membrane bilayer) or
multilamellar vesicles (onion-like structures characterized by
multiple membrane bilayers, each separated from the next by an
aqueous layer). The bilayer is composed of two lipid monolayers
having a hydrophobic "tail" region and a hydrophilic "head" region.
The structure of the membrane bilayer is such that the hydrophobic
(nonpolar) "tails" of the lipid monolayers orient toward the center
of the bilayer while the hydrophilic "heads" orient towards the
aqueous phase.
[0059] Liposomes and lipid complexes can be produced by a variety
of methods (for a review, see, e.g., Cullis et al. (1987)).
Bangham's procedure (J. Mol. Biol. (1965)) produces ordinary
multilamellar vesicles (MLVs). Lenk et al. (U.S. Pat. Nos.
4,522,803, 5,030,453 and 5,169,637), Fountain et al. (U.S. Pat. No.
4,588,578) and Cullis et al. (U.S. Pat. No. 4,975,282) disclose
methods for producing multilamellar liposomes having substantially
equal interlamellar solute distribution in each of their aqueous
compartments. Paphadjopoulos et al., U.S. Pat. No. 4,235,871,
discloses preparation of oligolamellar liposomes by reverse phase
evaporation.
[0060] Unilamellar vesicles can be produced from MLVs by a number
of techniques, for example, the extrusion of Cullis et al. (U.S.
Pat. No. 5,008,050) and Loughrey et al. (U.S. Pat. No. 5,059,421)).
Sonication and homogenization cab be so used to produce smaller
unilamellar liposomes from larger liposomes (see, for example,
Paphadjopoulos et al. (1968); Deamer and Uster (1983); and Chapman
et al. (1968)).
[0061] The original liposome preparation of Bangham et al. (J. Mol.
Biol., 1965, 13:238-252) involves suspending phospholipids in an
organic solvent which is then evaporated to dryness leaving a
phospholipid film on the reaction vessel. Next, an appropriate
amount of aqueous phase is added, the mixture is allowed to
"swell", and the resulting liposomes which consist of multilamellar
vesicles (MLVs) are dispersed by mechanical means. This preparation
provides the basis for the development of the small sonicated
unilamellar vesicles described by Papahadjopoulos et al. (Biochim.
Biophys, Acta., 1967, 135:624-638), and large unilamellar
vesicles.
[0062] Techniques for producing large unilamellar vesicles (LUVs),
such as, reverse phase evaporation, infusion procedures, and
detergent dilution, can be used to produce liposomes. A review of
these and other methods for producing liposomes may be found in the
text Liposomes, Marc Ostro, ed., Marcel Dekker, Inc., New York,
1983, Chapter 1, the pertinent portions of which are incorporated
herein by reference. See also Szoka, Jr. et al., (1980, Ann. Rev.
Biophys. Bioeng., 9:467), the pertinent portions of which are also
incorporated herein by reference.
[0063] Other techniques that are used to prepare vesicles include
those that form reverse-phase evaporation vesicles (REV),
Papahadjopoulos et al., U.S. Pat. No. 4,235,871. Another class of
liposomes that may be used are those characterized as having
substantially equal lamellar solute distribution. This class of
liposomes is denominated as stable plurilamellar vesicles (SPLV) as
defined in U.S. Pat. No. 4,522,803 to Lenk, et al. and includes
monophasic vesicles as described in U.S. Pat. No. 4,588,578 to
Fountain, et al. and frozen and thawed multilamellar vesicles
(FATMLV) as described above.
[0064] A variety of sterols and their water soluble derivatives
such as cholesterol hemisuccinate have been used to form liposomes;
see specifically Janoff et al., U.S. Pat. No. 4,721,612, issued
Jan. 26, 1988, entitled "Steroidal Liposomes." Mayhew et al., PCT
Publication No. WO 85/00968, published Mar. 14, 1985, described a
method for reducing the toxicity of drugs by encapsulating them in
liposomes comprising alpha-tocopherol and certain derivatives
thereof. Also, a variety of tocopherols and their water soluble
derivatives have been used to form liposomes, see Janoff et al.,
PCT Publication No. 87/02219, published Apr. 23, 1987, entitled
"Alpha Tocopherol-Based Vesicles".
[0065] Liposomes can also be prepared by the methods disclosed in
copending U.S. patent applications: Ser. No. 10/383,004, filed Mar.
5, 2003; Ser. No. 10/634,144, filed Aug. 4, 2003; Ser. No.
10/224,293, filed Aug. 20, 2002; and Ser. No. 10/696,389, filed
Oct. 29, 2003, the specifications of which are incorporated herein
in their entirety.
[0066] Another method of preparing liposomes or lipid complexes is
the "solvent infusion" process. Solvent infusion is a process that
includes dissolving one or more lipids in a small, preferably
minimal, amount of a process compatible solvent to form a lipid
suspension or solution (preferably a solution) and then adding the
solution to an aqueous medium containing, for example, platinum
compounds. Typically a process compatible solvent is one that can
be washed away in an aqueous process such as dialysis. The
composition that is cool/warm cycled is preferably formed by
solvent infusion, with ethanol infusion being preferred.
[0067] The process for producing lipid-complexed platinum compound
compositions may comprise mixing a platinum compound with an
appropriate hydrophobic matrix and subjecting the mixture to one or
more cycles of two separate temperatures. The process is believed
to form active platinum compound associations.
[0068] In aqueous solution, cisplatin forms large insoluble
aggregates with a diameter of greater than a few microns. In the
presence of a amphipathic matrix system, such as a lipid bilayer,
cisplatin-lipid associations form. For example, the associations
may be formed in the internal aqueous space, the hydrocarbon core
region of a lipid bilayer, or the liposome interface or headgroup.
During the warming cycle of the process, it is believed that
cisplatin is returned to solution at a greater rate in aqueous
regions of the process mixture than from the lipid-complex. As a
result of applying more than one cool/warm cycle, cisplatin
accumulates further into the lipid-complex. Without limiting the
invention to the proposed theory, experimentation indicates that
the cisplatin-lipid associations cause the immediate surroundings
of the interfacial bilayer region to be more hydrophobic and
compact. This results in a high level of entrapment of active
platinum compound as cooling and warming cycles are repeated.
[0069] The process comprises combining the platinum compound with a
hydrophobic matrix carrying system and cycling the solution between
a warmer and a cooler temperature. Preferably the cycling is
performed more than one time. More preferably the step is performed
two or more times, or three or more times. The cooler temperature
portion of cycle can, for example, use a temperature from about
-25.degree. C. to about 25.degree. C. More preferably the step uses
a temperature from about -5.degree. C. to about 25.degree. C. or
from about 1.degree. C. to about 20.degree. C. For manufacturing
convenience, and to be sure the desired temperature is established,
the cooler and warmer steps can be maintained for a period of time,
such as approximately from 5 to 300 minutes or 30 to 60 minutes.
The step of warming comprises warming the reaction vessel to from
about 4.degree. C. to about 70.degree. C. More preferably the step
of warming comprises heating the reaction vessel to about
45.degree. C. or to about 55.degree. C. The above temperature
ranges are particularly preferred for use with lipid compositions
comprising predominantly diphosphatidycholine (DPPC) and
cholesterol.
[0070] Another way to consider the temperature cycling is in terms
of the temperature differential between the warmer and the cooler
steps of the cycle. This temperature differential can be, for
example, about 25.degree. C. or more, such as a differential from
about 25.degree. C. to about 70.degree. C., preferably a
differential from about 40.degree. C. to about 55.degree. C. The
temperatures of the cooler and higher temperature steps are
selected on the basis of increasing entrapment of active platinum
compound. Without being limited to theory, it is believed that it
is useful to select an upper temperature effective substantially
increase the solubility of active platinum compound in the
processed mixture. Preferably, the warm step temperature is about
50.degree. C. or higher. The temperatures can also be selected to
be below and above the transition temperature for a lipid in the
lipid composition.
[0071] The temperatures appropriate for the method may, in some
cases, vary with the lipid composition used in the method, as can
be determined by ordinary experimentation. The temperatures of the
warming and cooling steps are selected on the basis of increasing
entrapment of active platinum compound. Without being limited by
any particular theory, it is believed that it is useful to select
an upper temperature effective substantially increase the
solubility of active platinum compound in the process mixture.
During repetitive cooling/heating, bioactive agents are solubilized
and crystallized repetitively. As soluble drug is cooled, some
portion enters complexes with the lipid while the remainder
precipitates. On subsequent heating, unencapsulated bioactive agent
that is crystallized becomes soluble again. Importantly, active
platinum compound that has been encapsulated in the lipid complex
substantially stays in the lipid complex during the heating and
cooling cycling (e.g. it leaks at such a slow rate that no
appreciable amount leaves the lipid complex during the heating
phase of this process).
[0072] For example, as the temperature is increased during the
warming step of the cycle, the active platinum compound, such as
cisplatin, dissolves. During the cooling step, the cisplatin in the
aqueous phase precipitates out of solution to a greater extent that
the cisplatin associated with the lipid bilayers, thereby
increasing the amount of lipid-associated cisplatin with each
heating and cooling cycle. Additionally, solubility of cisplatin is
highly temperature-dependent. Lowering 15.degree. in temperature of
a cisplatin solution decreases the soluble concentration by about
50%. In other words, solubility limiting concentration increases
with increasing temperature by about 3% per degree increase in
temperature of aqueous cisplatin. In addition, the aggregate
(crystal)-to-monomer transition temperature (solubilizing
temperature) is higher than the monomer-to-aggregate (crystal)
transition temperature (crystallizing temperature) by about 15 to
20.degree. C.
[0073] Transplatin solubility is poorer than cisplatin, but it is
also temperature-dependent. Lowering the temperature by about
15.degree. C. decreases the soluble concentration of transplatin by
about 50%. The aggregate (crystal)-to-monomer transition
temperature (solubilizing temperature) is higher than the
monomer-to-aggregate (crystal) transition temperature
(crystallizing temperature) by about 20 to 30.degree. C.
[0074] Experimental results strongly indicate that the physical
state of cisplatin is solid (aggregates) or lipid bound since the
concentration of cisplatin is much higher than the solubility
limit. Results further indicate that process does not require
freezing the compositions, but that cooling to temperature higher
than the freezing point of water is effective. Results further
indicated that an entrapment efficiency achieved by 3-cycles was
similar to that achieved by 6-cycles of cooling and warming cycles,
which indicated that 3 cycles of temperature treatment was
sufficient to achieve high levels of active platinum compound
entrapment.
[0075] Results further indicate that the process can be scaled-up
while increasing process efficiency in entrapping cisplatin. Thus,
the invention further provides processes that are conducted to
provide an amount adapted for total administration (in appropriate
smaller volume increments) of 200 or more mLs, 400 or more mLs, or
800 or more mLs. All else being the same, it is believed that the
larger production volumes generally achieve increased efficiency
over smaller scale processes. While such volume is that appropriate
for administration, it will be recognized that the volume can be
reduced for storage.
[0076] Results further indicate that the lipid-complexed cisplatin
made by the method of the invention can retain entrapped cisplatin
with minimal leakage for over one year. This is a further
demonstration of the uniqueness in the composition, indicating that
the cisplatin is bound within the liposome structure and not free
to readily leak out.
[0077] The process of the present invention may further comprise
separating the components of the product of the aforementioned
process. For example, in some embodiments, the process provides
both the aforementioned lipid-complexed active platinum compound
and the aforementioned liposome. In certain embodiments, the
portion of the product comprising the lipid-complexed active
platinum compound, referred to herein as "the heavy fraction" may
be separated from the portion comprising the liposome, referred to
herein as "the light fraction." Methods of separating include
allowing the heavy product to settle over a period of time, or
centrifuging the product.
[0078] The lipid to platinum compound ratio (L/D) seen in the
lipid-complexed platinum compounds used in the present invention
may be less than about 1 by weight. For example the L/D ratio can
be about 0.10 to 1 by weight, wherein the lipid-complexed active
platinum compound comprises at least one lipid and at least one
active platinum compound. In some embodiments, the lipid to drug
ratio is about 0.10 to about 0.50 by weight. In some embodiments,
the lipid to drug ratio is about 0.15 to about 0.45 by weight, and
in other embodiments, the lipid to drug ratio is about 0.20 to 0.40
by weight. In some embodiments, the lipid to drug ratio is about
0.2 by weight.
[0079] The lipid-complexed active platinum compound may have an
average volume-weighted diameter of about 0.5 to about 20 microns.
In some embodiments, the average volume-weighted diameter is about
1 to about 15 microns, or about 2 to about 10 microns. In other
embodiments, the average volume-weighted diameter is about 3, 4, 5,
or 6 microns.
[0080] In some embodiments, the concentration of the active
platinum compound in the composition is greater than about 1.2
mg/mL, for example about 1.2 to about 20 mg/mL. In other
embodiments, the concentration of the active platinum compound is
about 1.2 to 10 mg/mL, about 1.5 to about 5 mg/mL, about 2.0 to
about 4 mg/mL, or about 3.0 to 2.5 mg/mL. In other embodiments, the
concentration is about 2, about 3, or about 5 mg/mL.
[0081] In some embodiments, the composition comprising the
lipid-complexed active platinum compound further comprises a
liposome. As explained in greater detail in the examples below, the
liposome comprises at least one lipid. The lipid may be the same as
or different from the lipid in the lipid-complexed active platinum
compound. In some embodiments, the liposome further comprises an
active platinum compound, wherein the active platinum compound can
be the same as or different from the active platinum compound of
the lipid-complexed active platinum compound. The active platinum
compound may be entrapped in the liposome.
[0082] In some embodiments, the liposomes have an average diameter
of about 0.1 to about 1 micron, 0.1 to about 0.5 microns, about 0.2
to about 0.5 microns, or about 0.2 to about 0.3 microns.
[0083] When the lipid composition further comprises a liposome, the
lipid-complexed active platinum compound may contain about 70 to
about 100% of the total active platinum compound in the
composition. In other embodiments, the lipid-complexed active
platinum compound contains about 75 to about 99%, about 75 to about
95%, or about 80 to about 90% of the total active platinum compound
in the composition. In some embodiments, the liposome contains
about 0 to about 30% of the total active platinum compound in the
composition. In other embodiments, the liposome may contain about
0.5 to about 25%, about 1 to about 20%, or about 5 to 10% of the
total active platinum compound.
[0084] When the composition further comprises a liposome, the
lipid-complexed active platinum compound may contain about 0.1 to
about 5% of the total lipid in the composition. In some
embodiments, the lipid-complexed active platinum compound contains
about 0.25 to about 3%, or about 0.5 to about 2% of the total
lipid. In some embodiments, the liposome contains about 75 to about
99.5%, about 80 to about 95%, or about 85 to about 95% of the total
lipid in the composition.
[0085] When present in the composition, the liposome may have a
lipid to active platinum compound ratio of about 100:1 to about
400:1 by weight. In other embodiments, the lipid to active platinum
compound ratio of the liposome is about 200:1 to about 400:1, about
200: to 300:1 about 250:1 to 300:1 or about 250:1 by weight.
[0086] In some embodiments, the composition comprising a
lipid-complexed active platinum compound and a liposome has an
active platinum compound concentration of greater than about 1.2
mg/mL, for example, the concentration may be about 1.2 to about 20
mg/mL, about 1.2 to about 10 mg/mL, about 1.5 to about 5 mg/mL,
about 2.0 to about 4 mg/mL, or about 3.0 to 2.5 mg/mL. In other
embodiments, the concentration is about 2, about 3, or about 5
mg/mL.
[0087] An "active platinum" compound is a compound containing
coordinated platinum and having antineoplastic activity. Additional
active platinum compounds include, for example, carboplatin and
DACH-platinum compounds such as oxaliplatin. In certain
embodiments, the active platinum compounds in the composition is
selected from the group consisting of cisplatin, carboplatin,
oxaliplatin, iproplatin, tetraplatin, transplatin, JM118
(cis-amminedichloro(cyclohexylamine)platinum(II)), JM149
(cis-amminedichloro(cyclohexylamine)-trans-dihydroxoplatinum(IV)),
JM216 (bis-acetato-cis-amminedichloro(cyclohexylamine)platinum(IV))
and JM335
(trans-amminedichloro(cyclohexylamine)dihydroxoplatinum(IV)). In
some embodiments, the active platinum compound is cisplatin.
[0088] The platinum compounds that may be used in the composition
for the aforementioned method include any compound that exhibits
the property of preventing the development, maturation, or spread
of neoplastic cells. Non-limiting examples of platinum compounds
include cisplatin, carboplatin
(diammine(1,1-cyclobutanedicarboxylato)-platinum(II)), tetraplatin
(ormaplatin)
(tetrachloro(1,2-cyclohexanediamine-N,N')-platinum(IV)), thioplatin
(bis(O-ethyldithiocarbonato)platinum(II)), satraplatin, nedaplatin,
oxaliplatin, heptaplatin, iproplatin, transplatin, lobaplatin,
cis-aminedichloro(2-methylpyridine) platinum, JM118
(cis-amminedichloro (cyclohexylamine)platinum(II)), JM149
(cis-amminedichloro(cyclohexylamine)-trans-dihydroxoplatinum(IV)),
JM216 (bis-acetato-cis-amminedichloro(cyclohexylamine)
platinum(IV)), JM335
(trans-amminedichloro(cyclohexylamine)dihydroxoplatinum(IV)), and
(trans, trans,
trans)bis-mu-(hexane-1,6-diamine)-mu-[diamine-platinum(II)]bis[dia-
mine(chloro) platinum(II)]tetrachloride. In another embodiment the
platinum compound is cisplatin. Depending on the environment,
cisplatin may exist in a cationic aquated form wherein the two
negatively charged chloride atoms have been displaced by two
neutral water molecules. Because the aquated form of cisplatin is
cationic, anionic lipids such as glycerols help to stabilize the
lipid-complexed composition, but may also hinder release on the
cisplatin. The non-aquated, neutral form of cisplatin is more
difficult to stabilize but has different release kinetics. It is
considered an advantage of the present invention that in certain
embodiments the lipid-complexed cisplatin compositions comprise
neutral cisplatin and neutral lipids. Because of the equilibrium
between neutral, non-aquated cisplatin and cationic, aquated
cisplatin, one may favor neutral, non-aquated cisplatin by
preparing a composition with a low pH and high NaCl concentration.
In this embodiment a substantial amount of the cationic, aquated
form of cisplatin would not form until the neutral, non-aquated
cisplatin was delivered into the interior of a cell.
[0089] In certain embodiments, the active platinum compound is
selected from the group consisting of cisplatin, carboplatin,
oxaliplatin, iproplatin, tetraplatin, transplatin, JM118
(cis-amminedichloro(cyclohexylamine)platinum(II)), JM149
(cis-amminedichloro(cyclohexylamine)-trans-dihydroxoplatinum(IV)),
JM216 (bis-acetato-cis-amminedichloro(cyclohexylamine)platinum(IV))
and JM335
(trans-amminedichloro(cyclohexylamine)dihydroxoplatinum(IV)). In
some embodiments, the active platinum compound is cisplatin,
transplatin, carboplatin, or oxaliplatin, while in other
embodiments, the active platinum compound is cisplatin.
[0090] In other embodiments, other therapeutic agents may be used
with the platinum compounds. The other therapeutic agents may have
antineoplastic properties. Non-limiting examples of antineoplastic
compounds include altretamine, amethopterin, amrubicin, annamycin,
arsenic trioxide, asparaginase, BCG, benzylguanine, bisantrene,
bleomycin sulfate, busulfan carmustine, cachectin, chlorabucil,
2-chlorodeoxyadenosine, cyclophosphamide, cytosine arabinoside,
dacarbazine imidazole carboxamide, dactinomycin, daunomycin,
3'-deamino-3'-morpholino-13-deoxo-10-hydroxycarminomycin,
4-demethoxy-3-deamino-3-aziridinyl-4-methylsulphonyl-daunorubicin,
dexifosfamide, dexamethasone, diarizidinylspermine,
dibromodulcitol, dibrospidium chloride,
1-(11-dodecylamino-10-hydroxyundecyl)-3,7-dimethylxanthine,
doxorubicin, elinafide, epipodophyllotoxin, estramustine,
floxuridine, fluorouracil, fluoxymestero, flutamide, fludarabine,
fotemustine, galarubicin, glufosfamide, goserelin, GPX100,
hydroxyurea, idarubicin HCL, ifosfamide, improsulfan tosilate,
isophosphamide, interferon alfa, interferon alfa 2a, interferon
alfa 2b, interferon alfa n3, interferon gamma, interleukin 2,
irinotecan, irofulven, leucovorin calcium, leuprolide, levamisole,
lomustine, megestrol, L-phenylalanie mustard, L-sarcolysin,
melphalan hydrochloride, mechlorethamine, MEN10755, mercaptopurine,
MESNA, methylprednisolone, methotrexate, mitomycin, mitomycin-C,
mitoxantrone, nimustine, paclitaxel, pinafide, pirarubicin,
plicamycin, prednimustine, prednisone, procarbazine, profiromycin,
pumitepa, ranimuistine, sertenef, streptozocin, streptozotocin,
tamoxifen, tasonermin, temozolomide, 6-thioguanine, thiotepa,
tirapazimine, triethylene thiophosporamide, trofosfamide, tumor
necrosis factor, valrubicin, vinblastine, vincristine, vinorelbine
tartrate, and zorubicin.
[0091] Also included as suitable platinum compounds used in the
methods of the present invention are pharmaceutically acceptable
addition salts and complexes of platinum compounds. In cases
wherein the compounds may have one or more chiral centers, unless
specified, the present invention comprises each unique racemic
compound, as well as each unique nonracemic compound.
[0092] In cases in which the platinum compounds have unsaturated
carbon-carbon double bonds, both the cis (Z) and trans (E) isomers
are within the scope of this invention. In cases wherein the
neoplastic compounds may exist in tautomeric forms, such as
keto-enol tautomers, such as
##STR00001##
each tautomeric form is contemplated as being included within this
invention, whether existing in equilibrium or locked in one form by
appropriate substitution with R'. The meaning of any substituent at
any one occurrence is independent of its meaning, or any other
substituent's meaning, at any other occurrence.
[0093] Also included as suitable platinum compounds used in the
methods of the present invention are prodrugs of the platinum
compounds. Prodrugs are considered to be any covalently bonded
carriers which release the active parent compound in vivo.
[0094] The lipids used in the composition of the aforementioned
method can be synthetic, semi-synthetic or naturally-occurring
lipids, including phospholipids, tocopherols, sterols, fatty acids,
glycolipids, negatively-charged lipids, cationic lipids. In terms
of phospholipids, they can include such lipids as egg
phosphatidylcholine (EPC), egg phosphatidylglycerol (EPG), egg
phosphatidylinositol (EPI), egg phosphatidylserine (EPS),
phosphatidylethanolamine (EPE), and phosphatidic acid (EPA); the
soya counterparts, soy phosphatidylcholine (SPC); SPG, SPS, SPI,
SPE, and SPA; the hydrogenated egg and soya counterparts (e.g.,
HEPC, HSPC), stearically modified phosphatidylethanolamines,
cholesterol derivatives, carotinoids, other phospholipids made up
of ester linkages of fatty acids in the 2 and 3 of glycerol
positions containing chains of 12 to 26 carbon atoms and different
head groups in the 1 position of glycerol that include choline,
glycerol, inositol, serine, ethanolamine, as well as the
corresponding phosphatidic acids. The chains on these fatty acids
can be saturated or unsaturated, and the phospholipid may be made
up of fatty acids of different chain lengths and different degrees
of unsaturation. In particular, the compositions of the
compositions can include DPPC, a major constituent of
naturally-occurring lung surfactant. Other examples include
dimyristoylphosphatidycholine (DMPC) and
dimyristoylphosphatidylglycerol (DMPG) dipalmitoylphosphatidcholine
(DPPC and dipalmitoylphosphatidylglycerol (DPPG)
distearoylphosphatidylcholine (DSPC and
distearoylphosphatidylglycerol (DSPG),
dioleylphosphatidyl-ethanolamine (DOPE) and mixed phospholipids
like palmitoylstearoylphosphatidyl-choline (PSPC) and
palmitoylstearolphosphatidylglycerol (PSPG), triacylglycerol,
diacylglycerol, seranide, sphingosine, sphingomyelin and single
acylated phospholipids like mono-oleoyl-phosphatidylethanolarnine
(MOPE).
[0095] In some embodiments, the lipid complexed active platinum
composition comprises a neutral phospholipid, such as a
phosphatidyl choline. In other embodiments, the phosphatidyl
choline is DPPC.
[0096] The cationic lipids used can include ammonium salts of fatty
acids, phospholids and glycerides. The fatty acids include fatty
acids of carbon chain lengths of 12 to 26 carbon atoms that are
either saturated or unsaturated. Some specific examples include:
myristylamine, palmitylamine, laurylamine and stearylamine,
dilauroyl ethylphosphocholine (DLEP), dimyristoyl
ethylphosphocholine (DMEP), dipalmitoyl ethylphosphocholine (DPEP)
and distearoyl ethylphosphocholine (DSEP), N-(2,
3-di-(9-(Z)-octadecenyloxy)-prop-1-yl-N,N,N-trimethylammonium
chloride (DOTMA) and 1,
2-bis(oleoyloxy)-3-(trimethylammonio)propane (DOTAP).
[0097] Negatively charged lipids include PGs, PAs, PSs and PIs. In
some embodiments, the lipid complexed active platinum composition
does not comprise a phosphatidyl serine (PS). In some embodiments,
the composition does not comprise a PG, PA, PS or PI. In other
embodiments, the composition is substantially free of negatively
charged or positively charged phospholipids. In some embodiments
the composition does not comprise any negatively charged
phospholipids.
[0098] In some embodiments, the lipid complexed active platinum
composition further comprises a sterol. The sterols can include,
cholesterol, esters of cholesterol including cholesterol
hemi-succinate, salts of cholesterol including cholesterol hydrogen
sulfate and cholesterol sulfate, ergosterol, esters of ergosterol
including ergosterol hemi-succinate, salts of ergosterol including
ergosterol hydrogen sulfate and ergosterol sulfate, lanosterol,
esters of lanosterol including lanosterol hemi-succinate, salts of
lanosterol including lanosterol hydrogen sulfate and lanosterol
sulfate. The tocopherols can include tocopherols, esters of
tocopherols including tocopherol hemi-succinates, salts of
tocopherols including tocopherol hydrogen sulfates and tocopherol
sulfates. The term "sterol compound" includes sterols, tocopherols
and the like. In some embodiments, the sterol is cholesterol.
[0099] In some embodiments, the lipid complexed active platinum
composition comprises DPPC and cholesterol in a ratio of about 1:1
to about 5:1 by weight. In other embodiments, composition comprises
DPPC and cholesterol in a ratio of about 2:1 to about 4:1 by
weight. In some embodiments, the composition comprises DPPC and
cholesterol in a ratio of about 2.25:1 by weight.
[0100] In some embodiments, the composition further comprises a
pharmaceutically acceptable carrier or diluent. The composition may
be comprised of an aqueous dispersion of the lipid complexed active
platinum compound. The composition may contain excipients and
salts/buffers to provide the appropriate osmolarity and pH. The
pharmaceutical excipient may be a liquid, diluent, solvent or
encapsulating material, involved in carrying or transporting any
subject composition or component thereof from one organ, or portion
of the body, to another organ, or portion of the body. Each
excipient must be "acceptable" in the sense of being compatible
with the subject composition and its components and not injurious
to the patient. Suitable excipients include trehalose, raffinose,
mannitol, sucrose, leucine, trileucine, and calcium chloride.
Examples of other suitable excipients include (1) sugars, such as
lactose, and glucose; (2) starches, such as corn starch and potato
starch; (3) cellulose, and its derivatives, such as sodium
carboxymethyl cellulose, ethyl cellulose and cellulose acetate; (4)
powdered tragacanth; (5) malt; (6) gelatin; (7) talc; (8)
excipients, such as cocoa butter and suppository waxes; (9) oils,
such as peanut oil, cottonseed oil, safflower oil, sesame oil,
olive oil, corn oil and soybean oil; (10) glycols, such as
propylene glycol; (11) polyols, such as glycerin, sorbitol, and
polyethylene glycol; (12) esters, such as ethyl oleate and ethyl
laurate; (13) agar; (14) buffering agents, such as magnesium
hydroxide and aluminum hydroxide; (15) alginic acid; (16)
pyrogen-free water; (17) isotonic saline; (18) Ringer's solution;
(19) ethyl alcohol; (20) phosphate buffer solutions; and (21) other
non-toxic compatible substances employed in pharmaceutical
compositions.
[0101] The present invention, in part, discloses methods of
treating cancer more effectively which may have lower
nephrotoxicity previously not disclosed. By using lipid-complexed
compositions and ip delivery, a more potent and efficient cancer
treatment is achieved.
Dosages
[0102] The dosage of any compositions of the present invention will
vary depending on the symptoms, age and body weight of the patient,
the nature and severity of the disorder to be treated or prevented,
the route of administration, and the form of the subject
composition. Any of the subject compositions may be administered in
a single dose or in divided doses. Dosages for the compositions of
the present invention may be readily determined by techniques known
to those of skill in the art or as taught herein.
[0103] In certain embodiments, the dosage of the subject compounds
will generally be in the range of about 0.01 ng to about 10 g per
kg body weight, specifically in the range of about 1 ng to about
0.1 g per kg, and more specifically in the range of about 100 ng to
about 50 mg per kg.
[0104] Dosage amounts are also commonly administered as mg/m.sup.2
which stands for milligrams of drug (e.g. platinum compound) per
body surface area. Generally, dosage amounts for platinum compounds
may be about 60 mg/m.sup.2 or greater, 100 mg/m.sup.2 or greater,
140 mg/m.sup.2 or greater, or 180 mg/m.sup.2 or greater. Dosage
amounts of about 140 mg/m.sup.2 or greater are generally considered
at the high end of tolerance, but an advantage of the present
invention is that the platinum compound is administered as part of
a lipid-complexed composition which decreases the sub-acute
toxicities of the platinum compound. It is therefore envisioned by
the inventors that higher than normal dosage amounts of platinum
compound may be administered to the patient without unwanted toxic
side effects. Higher dosages may lead to longer duration cycles
between dosages and greater convenience for the patient. For
example, dosage amounts are generally administered to the patient
once about every three weeks. If higher dosage amounts of platinum
compound can be administered safely to the patient then the cycle
time may be increased to once about every four, five, six, seven,
or even eight weeks. Longer cycle times means less trips to a care
facility for treatment and less times the patient would have to
undergo the administration process.
[0105] An effective dose or amount, and any possible affects on the
timing of administration of the composition, may need to be
identified for any particular composition of the present invention.
This may be accomplished by routine experiment as described herein,
using one or more groups of animals (preferably at least 5 animals
per group), or in human trials if appropriate. The effectiveness of
any subject composition and method of treatment or prevention may
be assessed by administering the composition and assessing the
effect of the administration by measuring one or more applicable
indices, and comparing the post-treatment values of these indices
to the values of the same indices prior to treatment.
[0106] The precise time of administration and amount of any
particular subject composition that will yield the most effective
treatment in a given patient will depend upon the activity,
pharmacokinetics, and bioavailability of a subject composition,
physiological condition of the patient (including age, sex, disease
type and stage, general physical condition, responsiveness to a
given dosage and type of medication), route of administration, and
the like. The guidelines presented herein may be used to optimize
the treatment, e.g., determining the optimum time and/or amount of
administration, which will require no more than routine
experimentation consisting of monitoring the subject and adjusting
the dosage and/or timing.
[0107] While the subject is being treated, the health of the
patient may be monitored by measuring one or more of the relevant
indices at predetermined times during the treatment period.
Treatment, including composition, amounts, times of administration
and formulation, may be optimized according to the results of such
monitoring. The patient may be periodically reevaluated to
determine the extent of improvement by measuring the same
parameters. Adjustments to the amount(s) of subject composition
administered and possibly to the time of administration may be made
based on these reevaluations.
[0108] Treatment may be initiated with smaller dosages which are
less than the optimum dose of the compound. Thereafter, the dosage
may be increased by small increments until the optimum therapeutic
effect is attained.
[0109] The use of the subject compositions may reduce the required
dosage for any individual agent contained in the compositions
(e.g., the antineoplastic compound) because the onset and duration
of effect of the different agents may be complimentary.
[0110] Toxicity and therapeutic efficacy of subject compositions
may be determined by standard pharmaceutical procedures in cell
cultures or experimental animals, e.g., for determining the
LD.sub.50 and the ED.sub.50.
[0111] The data obtained from the cell culture assays and animal
studies may be used in formulating a range of dosage for use in
humans. The dosage of any subject composition lies preferably
within a range of circulating concentrations that include the
ED.sub.50 with little or no toxicity. The dosage may vary within
this range depending upon the dosage form employed and the route of
administration utilized. For compositions of the present invention,
the therapeutically effective dose may be estimated initially from
cell culture assays.
EXEMPLIFICATION
Example 1
Method of Producing an Aqueous Cisplatin with Higher Potency than
its Aqueous Solubility Limit at Room Temperature
[0112] 1) At temperatures about 50-60.degree. C., cisplatin in 0.9%
sodium chloride solution at a level of 4 mg/ml and an ethanolic
solution of about 16 mg/ml DPPC and 8 mg/ml cholesterol at about
55.degree. C. are aseptically prepared. [0113] 2) The lipid
solution is infused into the cisplatin solution while mixing the
cisplatin solution. [0114] 3) After infusion, cisplatin/lipid
dispersion is cooled down to about 10.degree. C. and then warmed up
again to about 50-60.degree. C. for 15 min. [0115] 4) Step 3) is
repeated 2-3 times. [0116] 5) The dispersion is aseptically washed
with sterile 0.9% sodium chloride solution to remove residual
ethanol and un-associated cisplatin via 500,000 MW cut-off membrane
diafiltration unit.
[0117] After washing process, the dispersion provides about 1 mg/ml
cisplatin potency and concentrated to 3 mg/ml cisplatin and further
concentrated to 5 mg/ml cisplatin by aseptically removing two third
of the aqueous vehicle of 1 mg/ml product and four fifth of the
aqueous vehicle of 1 mg/ml product, respectively. The removal of
aqueous vehicle was carried out at a rate of about 100 ml/min by
diafiltration at about 20.degree. C. without compensating the
permeate with fresh sterile 0.9% sodium chloride solution.
Example 2
[0118] 70 mg DPPC and 28 mg cholesterol was dissolved in 1 mL
ethanol and added to 10 mL of 4 mg/mL cisplatin in 0.9% saline
solution.
[0119] (i) An aliquot (50%) of the sample was treated by 3 cycles
of cooling to 4.degree. C. and warming to 50.degree. C. The
aliquot, in a test tube, was cooled by refrigeration, and heated in
a water bath. The resulting unentrapped cisplatin (free cisplatin)
was washed by dialysis.
[0120] (ii) The remainder of the sample was not treated by
temperature cycles and directly washed by dialysis.
TABLE-US-00001 TABLE 1 Percentage entrapment of cisplatin with and
without cooling and warming cycles. Final Concentration of %
cisplatin, .mu.g/mL Entrapment Lipid-complexed cisplatin without 56
1.4 cooling and warming cycles lipid-complexed cisplatin after 360
9.0 cooling and warming cycles
Example 3
[0121] The rigidity of a membrane bilayer in lipid-complexed
cisplatin prepared with cool/warm cycling as described in Example 2
was measured by fluorescence anisotropy of diphenylhexatriene
(membrane probe) inserted in the hydrophobic core region of the
bilayer. [Ref. Jahnig, F., 1979 Proc. Natl. Acad. Sci. USA 76(12):
6361.] The hydration of the bilayers was gauged by the deuterium
isotope exchange effect on fluorescence intensity of TMA-DPH
(trimethylamine-diphenylhexatriene). [Ref. Ho, C., Slater, S. J.,
and Stubbs, C. D., 1995 Biochemistry 34: 6188.]
TABLE-US-00002 TABLE 2 Degree of hydration and rigidity of
liposomes, lipid-complexed cisplatin without and with cool/warm
cycling. Placebo Lipid-complexed Lipid-complexed (Liposomes
cisplatin cisplatin without without cooling & with cooling
& cisplatin) warming cycles warming cycles Degree bilayer 0.29
0.29 0.36 rigidity Degree of bilayer 1.13 1.15 1.02 hydration
Example 4
[0122] Density Characterization of the light and heavy fractions
was performed as follows. Samples were prepared as in the previous
example. At cooling the temperature of samples was 0.degree. C. The
temperature cycle was done by 15 min cooling and 15 min warming.
The starting cisplatin concentration was 4 mg/mL and free cisplatin
was removed by dialysis.
[0123] Density Gradient Analysis
[0124] Seven different batches of cisplatin lipid complex were used
for these experiments. Density gradients were formed using
Iodixanol (SIGMA (D1556, lot no. 025K1414)) as a dense media and
0.9% NaCl saline solution to keep osmolality close to normal 300
mOsM. First, about 5.5 mL saline was added to the centrifuge tube,
and then the same volume of heavy medium (1:1 mixture of Iodixanol
60% and saline) was layered on the bottom of the tube using a
syringe with a long needle. Gradients were formed using a BioComp
107ip Gradient Master at the settings: time=2:14 min, angle=79.0,
speed=17 rpm, and using the long tube cap. An aliquot of Cisplatin
Lipid Complex samples (1 mL) were placed on the top of the gradient
and centrifuged for 30 min at 30,000 rpm at 20.degree. C. After
centrifugation, the top 0.8-1.0 mL volume of clear liquid was
discarded, and the next 2 mL was collected representing the light
fraction. The light fraction is believed to contain liposomes,
wherein at least some of the liposomes are associated with
cisplatin. There was a detectable amount of free cisplatin in the
light fraction of nebulized samples, which was determined by
filtering through Centricon-30 filtering devices and subtracted
from the total cisplatin.
[0125] The rest of the media was removed, leaving only a small
yellow pellet on the bottom representing the dense (heavy)
fraction, which was subsequently dispersed in 2 mL solution of 75%
n-Propanol, 5% saline, 20% water. Cisplatin in the heavy fraction
was not completely soluble at this point. An aliquot of this
dispersion was taken for cisplatin determination. Another part of
the dispersion (1 mL) was mixed with equal volume of 60% n-Propanol
and centrifuged 5 min at 1000 rpm on an Eppendorf 5810R centrifuge
to settle undissolved cisplatin, and then 1 mL of clear supernatant
was used for HPLC lipid determination.
[0126] Cisplatin Concentration:
[0127] Cisplatin was measured by HPLC by separating cisplatin on
YMC-Pack NH2 column using 90% acetonitrile mobile phase and
measuring absorbance at 305 nm. Cisplatin standards and samples
were diluted in solution of 75% n-Propanol, 5% saline and 20%
water. Standards were used with cisplatin concentrations of 75, 50,
25, and 10 .mu.g/mL. Cisplatin peak retention time was usually
around 6.4 min.
[0128] Lipid Analysis by HPLC:
[0129] Lipids were analyzed by HPLC as follows: lipids were
separated on a Phenomenex Luna C8(2) column using binary gradient
mode. Mobile phase A: methanol 70%, acetonitrile 20%, water 10%,
ammonium acetate 0.1%, mobile phase B: methanol 70%, acetonitrile
30%, ammonium acetate 0.07%. Lipid standards and samples were
diluted in a solution of 60% n-Propanol, 40% water. Lipids were
detected by Sedex 55 Evaporative Light Scattering Detector. The
retention time for cholesterol was about 8 min, for DPPC about 10
min.
[0130] Results
[0131] Nine batches of Lipid-cisplatin complex were fractionated on
an Iodixanol density gradient as described in the Methods section.
All nine samples separated into a similarly positioned white band
of light fraction and a yellow pellet of dense fraction. 2 mL of
the light fraction were collected and the rest of the liquid was
removed. The remaining pellet was dispersed in 2 mL of 75%
n-Propanol. Cisplatin and lipid concentrations in each fraction
were measured by HPLC as described. The lipid/cisplatin ratio in
the dense fraction was very high so that both lipid and cisplatin
could not be solubilized in same solvent at high enough
concentration for the lipid analysis. For that reason, the
lipid-cisplatin mixture in 75% n-propanol solution was centrifuged
to remove the insoluble portion of the cisplatin, and the
supernatant was used as is for lipid HPLC analysis.
[0132] Results of the density gradient analysis are presented in
Table 4. L/D represents the ratio of lipid to cisplatin by weight.
The percentages presented are with respect to the total cisplatin
or lipid in the formulation. Lower section of the table shows
averages of lipid and cisplatin contents in each fraction derived
from all nine samples tested. Standard deviations (SD) are shown to
demonstrate consistency. These data demonstrate that the majority
of lipid (90.6% on average, +/-3.1%) is in the light fraction,
while only 0.87+/-0.09% lipid is in the dense fraction. The
majority of cisplatin (82.3+/-2.9%) is in the dense fraction, while
only 8.4+/-2.1% is in the light fraction. The lipid to drug ratio
(L/D) calculated for the total sample was an average of 22.7. The
same L/D ratio in separate fractions was as high as 255+/-47 for
the light fraction, and as low as 0.24+/-0.03 for dense fraction.
Results are summarized in Table 3.
TABLE-US-00003 TABLE 3 Distribution of cisplatin and lipids in the
light and dense fractions of Cisplatin Lipid Complex samples. Lipid
Lipid Cisplatin Cisplatin Batch mg/mL % total mg/mL % total L/D 8
total 62.2 2.47 25.2 8 Light fraction 58.0 93.3 0.20 8.2 285 8
Dense fraction 0.49 0.79 2.01 81.3 0.24 9 total 51.5 2.46 20.9 9
Light fraction 47.8 92.7 0.18 7.2 270 9 Dense fraction 0.44 0.85
1.96 79.7 0.22 10 total 55.2 2.57 21.5 10 Light fraction 46.7 84.7
0.20 7.7 237 10 Dense fraction 0.47 0.85 2.15 83.6 0.22 11 total
57.3 2.61 22.0 11 Light fraction 49.7 86.8 0.15 5.9 326 11 Dense
fraction 0.48 0.84 2.20 84.2 0.22 12 total 57.9 2.57 22.5 12 Light
fraction 51.6 89.1 0.18 6.9 290 12 Dense fraction 0.47 0.82 2.17
84.3 0.22 13 total 80.46 3.36 24.0 13 Light fraction 73.42 91.26
0.44 13.0 168 13 Dense fraction 0.82 1.01 2.58 76.7 0.32 14 total
71.19 3.41 20.9 14 Light fraction 64.82 91.06 0.29 8.7 220 14 Dense
fraction 0.65 0.92 2.76 81.1 0.24 15 total 66.92 2.83 23.7 15 Light
fraction 62.35 93.17 0.23 8.0 275 15 Dense fraction 0.66 0.99 2.45
86.5 0.27 16 total 68.5 2.86 23.9 16 Light fraction 64.0 93.48 0.28
9.8 228 16 Dense fraction 0.51 0.75 2.40 83.8 0.21 Average 22.7
Light fraction 90.6 8.4 255 +/-SD 3.1 2.1 47 Dense fraction 0.87
82.3 0.24 +/-SD 0.09 2.9 0.03
Example 5
Separation of Light and Dense Fractions
[0133] 30 mL of cisplatin lipid complex was mixed with 10 mL
iodixanol 30% in saline. This mixture was in half and 20 mL
portions were layered on the top of another 10 mL iodixanol 30% in
saline using two 50 mL centrifuge tubes. The samples were
centrifuged for 30 minutes at 4000 rpm at 5.degree. C. on an
Eppendorf 5810 centrifuge. Supernatant, containing a mixture of
light and heavy fractions, was discarded. The pellet, containing
the dense fraction of the cisplatin lipid composition, was gently
dispersed in 5 mL of saline. After determining the concentration of
cisplatin, the concentration was adjusted to make the concentration
2.7 mg/mL of cisplatin.
[0134] To obtain the light fraction, the cisplatin lipid complex
batch was allowed to settle by gravity at 5.degree. C. for 1 week.
The top portion of the sample, containing the light fraction, was
collected.
Example 6
[0135] Entrapment of cisplatin or transplatin in a lipid complex by
repetitive cooling/heating achieves a high drug/lipid ratio as
shown in Table 4.
TABLE-US-00004 TABLE 4 cisplatin transplatin Starting drug
concentration 5 mg/mL 1 mg/mL Lipids 25 mg/mL 5 mg/mL
(DPPC:Cholesterol = 7:3 wt) Temperature cycles 6 cycles 6 cycles
Final drug concentration 1.4 mg/mL 0.3 mg/mL (1.4% free) (6.0%
free) Recovery % 29% 26% Drug/Lipid 0.056 0.06
Example 7
[0136] Samples of the heavy fraction comprising lipid-complexed
cisplatin were diluted in filtered saline (NaCl 0.9%) at a ratio of
1:2000 and analyzed by an AccuSizer Optical Particle Sizer 780
using the following settings: injection loop volume 1 mL, Data
collection time 60 s, Detector LE 400-05SE summary mode, Minimum
diameter 0.05 microns. The detector used counts only particles 0.5
microns and larger. Four batches were analyzed. The distribution
plots show relative volumes occupied by particles of different
sizes. The particles in the range of 0.5 to 1 micron represent the
main distribution of the light fraction, the majority of which has
particle sizes less than 5 microns. The plots also show a large
peak at the right from 1 micron to 20 micron, with a median size of
about 8 to 10 microns.
Example 8
Comparison of Free Cisplatin and Lipid-Complexed Cisplatin
Formulation at 3 mg/ml Cisplatin Concentration Administered
Intraperitoneally to Rats
[0137] Male Sprague-Dawley rats were given free cisplatin at
dosages of 6 and 12 mg/kg, and lipid-complexed cisplatin
formulations at 3 mg/ml at dosages of 6, 12, and 18 mg/kg
intraperitoneally. Control groups comprised 6 rats and groups for
the test articles comprised 3 rats per group. Observation of
morbidity/mortality were conducted daily as were weight
measurements. Results are presented in Table 5.
TABLE-US-00005 TABLE 5 Rat study lethality summary for 3 mg/ml
lipid-complexed cisplatin formulation. Number Dead/Number Day of
Treatment Group/Dosage Treated Death Group 1 - Control (no
treatment) 0/6 Group 2 - 6 mg/kg lipid-complexed 0/3 cisplatin
formulation Group 3 - 12 mg/kg lipid-complexed 1/3 13 cisplatin
formulation Group 4 - 18 mg/kg lipid-complexed 1/3 8 cisplatin
formulation
Example 9
Reduction of Sub-Acute Toxicity of Cisplatin by iv or ip
Administration when Administered as a Lipid-Complexed
Formulation
[0138] ICR mice, male and female, 6-7 weeks old, were divided into
24 groups with 10 mice in each. Five mice were housed in each cage
with free access to standard mouse food and water. Each group of
mice was injected with lipid-complexed cisplatin formulations
prepared according to the following. The lipid-complexed cisplatin
formulation used here contained 1 mg/ml cisplatin, 16 mg/ml DPPC,
and 7.9 mg/ml cholesterol in 0.9% NaCl solution. An aliquot (50%)
of the sample was treated by 3 cycles of cooling to 4.degree. C.
and warming to 50.degree. C. The aliquot, in a test tube, was
cooled by refrigeration, and heated in a water bath. The resulting
unentrapped cisplatin (free cisplatin) was washed away by dialysis.
The lipid-complexed cisplatin in the form of liposomes were
injected through iv (tail vein) or ip route. The liposomes had a
mean diameter of about 0.39 .mu.m. The formulations, doses, and
administration routes are listed in Table 6.
TABLE-US-00006 TABLE 6 Dose and administration route for
lipid-complexed cisplatin sub-acute toxicity study. Formulation
Route Dose (mg Cisplatin/kg mouse) Lipid- ip 23 27 31 35 40 45 50
Complexed Cisplatin Cisplatin ip 9 12 15 18 21 24 27
[0139] Starting one week before the administration, body weights of
the mice were measured every two days until the end of the
experiment. The animals were observed daily and the death was
recorded. A curve of percent survival verses time (days) post
administration for each formulation with each injection route was
calculated (FIG. 1). The LD.sub.10, LD.sub.50 and LD.sub.90 of each
formulation under each injection route were estimated. The computer
fitted results are listed in Table 7.
TABLE-US-00007 TABLE 7 Lethal toxicity of lipid-complexed cisplatin
and cisplatin after ip and iv injection. Lethal Dose (mg
Cisplatin/kg mouse) Formulation Route LD.sub.10 LD.sub.50 LD.sub.90
Lipid- ip 22.4 29.5 38.9 Complexed Cisplatin Cisplatin ip 9.9 14.3
20.7
[0140] The result indicate that the sub-acute toxicity of ip
lipid-complexed cisplatin was 2-fold lower than ip cisplatin,
whereas not nearly as great of change was observed for iv
lipid-complexed cisplatin.
Example 10
Pharmacokinetics and Organ Distribution in Animals of ip and iv
Injected Lipid-Complexed Cisplatin and Cisplatin (Part I)
[0141] The mice (the same as from Example 2) were divided into 4
groups with 24 mice in each. They were injected with ip
lipid-complexed cisplatin (12 mg/kg), ip cisplatin (12 mg/kg), iv
lipid-complexed cisplatin (8 mg/kg), and iv cisplatin (8 mg/kg),
separately. The lipid-complexed cisplatin formulation were prepared
in the same manner as in Example 3. At each designed time point,
e.g., 2-5 min, 20 min, 40 min, 2 h, 8 h, 1 day, 2 days, and 3 or 5
days after injection, 3 mice from each group were anesthetized by
ip injection of 35-50 mg/kg of Nembutal, then the blood was drown
and heart, kidney, liver, lung, small intestine, and spleen were
resected and homogenized after adding 4-fold pure water. The
Platinum concentration in each sample was determined with AA
method. The content of Pt (.mu.g of Pt in 1 ml of blood or 1 gram
of tissue) was calculated and used for presenting the kinetic
characteristics of each formulation under two different
administration routes.
[0142] The results indicated that in the blood, the Cmax and AUC of
lipid-complexed cisplatin was 3- and 6-fold higher than that of
cisplatin, respectively (FIG. 2).
Example 11
Pharmacokinetics and Organ Distribution in Animals of ip and iv
Injected Lipid-Complexed Cisplatin and Cisplatin (Part II)
[0143] Sixty ICR mice (female, 7 weeks old) were divided into 4
groups. They received intraperitoneal or intravenous injection of
L-CDDP or CDDP, separately. The lipid-complexed cisplatin
formulation were prepared in the same manner as in Example 3. The
dose was 12 mg/kg for ip L-CDDP and 8 mg/kg for the rest of
treatment groups. At each designed time point, three to four mice
were anaesthetized with 70 mg/kg of Nembutal ip (e.g., 3, 20, and
40 min, and 2, 8, 24, 48, and 72 h). The blood was drawn from the
inferior vena cava. Organs including duodenum, kidney, liver, lung,
and spleen were resected from the mice. The blood and organ samples
were homogenized in distilled water (4-fold of the sample weight)
and digested with nitric acid. The platinum concentration in each
sample was measured by Inductively Coupled Plasma-Mass Spectrometer
(ICP-MS). The pharmacokinetics profiles (FIGS. 3-7, all Y-axes are
concentration of .mu.g platinum in one gram of tissue or fluid per
mg of injected dose) and parameters (Tables 4 and 5) of each
formulation were simulated and calculated. It was found that 1)
uptake of platinum in the spleen was significantly enhanced by the
liposomal formulation irrespective of the injection routes, the
AUCs and C.sub.max were 26.about.47-fold higher than those of CDDP,
p<0.0003; 2) with ip injection, the circulation AUC and
elimination t.sub.1/2 of L-CDDP were 4- and 15-fold (p<0.006)
higher than those of CDDP, but this phenomenon was not found after
iv injection; 3) the AUC and t.sub.1/2 of ip L-CDDP were also
higher than those of iv L-CDDP (ip vs iv: AUC, 1.9-fold, p=0.04;
t.sub.1/2, 5.7-fold, p=0.006); 4) the lung and liver platinum
uptake of iv L-CDDP were slightly higher than that of iv CDDP
(Lung: p=0.043; Liver: p=0.051); 5) the kidney platinum uptake of
L-CDDP was higher than that of CDDP (p=0.046). These results imply
that the formulation and administration route both play important
role on the PK and organ distribution of the drug, and ip L-CDDP
showed sustained release function.
TABLE-US-00008 TABLE 8 Circulation AUC and t.sub.1/2. AUC (h
.mu.g/g/.mu.g) t.sub.1/2 (h) ip ip L-CDDP 37.60.sup.a 26.58.sup.b
CDDP 7.08 1.82 .sup.ap = 0.008 (L-CDDP vs CDDP); .sup.bp = 0.006
(L-CDDP vs CDDP) by two side Log rank test.
TABLE-US-00009 TABLE 9 Organ AUC and C.sub.max. ip AUC (h
.mu.g/g/.mu.g) C.sub.max (.mu.g/g/.mu.g) L-CDDP CDDP L-CDDP CDDP
Duodenum 5.49 8.32 0.30 0.45 Kidney 15.60.sup.c 8.61 0.46 0.46
Liver 29.99 28.99 0.71 0.71 Lung 26.90 29.00 0.73 1.61 Spleen
331.86.sup.e 10.45 11.32.sup.f 0.43 Heart 2.63 2.67 0.07 0.18 Two
side Log rank test was used to evaluate the significance of the
differences of L-CDDP versus CDDP in AUC or C.sub.max values.
Statistically significant pairs (p < 0.05) were labeled with a
superscript letter. Their p values are as follows: .sup.c, p =
0.046; .sup.d, p = 0.008; .sup.e, p = 0.0002; .sup.f, p = 0.0001;
.sup.g, p = 0.0003; .sup.h, p = 0.0002.
Example 12
Nephrotoxicity
[0144] ICR mice, 7 weeks old, female, were divided into 4 groups
with 3 to 4 mice in each. They were injected with maximum tolerated
dose (MTD) of L-CDDP or CDDP via iv or ip. The lipid-complexed
cisplatin formulation were prepared in the same manner as in
Example 3. Four days after the injection, the mice were euthanized
with Nembutal ip. The blood was drawn and the serum was isolated.
The blood urea nitrogen (BUN) was quantitatively measured with a
colorimetric method at Antech Diagnostics. Organs including
duodenum, heart, kidney, liver, lung, and spleen were resected from
the mice and fixed with 10% buffered Formalin. The fixed tissues
were processed with standard procedure for H and E staining. A
pathology expert Dr. Carman Tornos at the Memorial Sloan-Kettering
Cancer Center examined kidney tissues and gave a toxicity grade to
each kidney tissue sample. The grading was based on the general
pathology guidelines for kidney toxicity.
[0145] The pathological results demonstrate that irrespective of
administration routes, CDDP caused severe nephrotoxicity in more
than 50% mice receiving the treatment, but L-CDDP did not cause any
nephrotoxicity. The similar conclusion can be drawn from BUN test
(FIG. 9), where iv CDDP significantly increased BUN level by
6.8-fold compared with normal controls (p=0.008), and ip CDDP
caused much less BUN accumulation, only 2.1-fold increase compared
to normal controls (p=0.04), but L-CDDP injected by either route
did not cause BUN level elevation.
Example 13
Preclinical In Vivo Antitumor Activity of Lipid-Complexed Cisplatin
in a Murine L1210 Tumor Model
[0146] The purpose of this experiment is to assess the in vivo
antitumor activity of lipid-complexed cisplatin against a cavity
confined tumor (ascitic L1210 leukemia) by local ip administration.
Lipid-complexed cisplatin was compared to free cisplatin for viable
L1210 tumor cells. The test articles and materials are presented
below in Table 6. The lipid-complexed cisplatin was prepared in the
same manner as in Example 3.
TABLE-US-00010 TABLE 10 Test articles and materials. Test Articles
Cisplatin in 0.9% saline 1.0 mg/ml cisplatin Lipid-Complexed
Cisplatin 0.82 mg/ml cisplatin Test Animals B6D2F1/Hsd hybrid mice
from Harlan 5 male/group except control (9 male), 7 groups Tumor
Cells Viable L1210 tumor cells, One million per mouse, transplanted
in vivo
The procedure was as outlined below and summarized in Table 11:
[0147] 1. Day 0, inoculate 6 groups of 5 male mice and 1 group of 9
male control animals with one million viable L1210 cells per mouse
by ip injection. [0148] 2. Administer ascending doses of either
cisplatin solution or lipid-complexed cisplatin to groups of 5
animals. Cisplatin solution: dose ip at 3.0 and 4.5 mg/Kg on days
3, 7, and 11. Each dose level represents one group of five mice.
Lipid-complexed cisplatin: dose ip at 3.0, 4.5, 6.0 and 9.0 mg/Kg
on days 3, 7, and 11. Control group contains 9 untreated mice.
[0149] 3. Mice were monitored daily for deaths and or signs of
clinical illness. The date of euthanasia was recorded for the
purpose of experimental end-points. A total of 39 mice divided into
7 groups were studied. At the end point survival was assessed and
expressed as % T/C (percent median survival of treated group:
median survival of control group.)
TABLE-US-00011 [0149] TABLE 11 Procedural parameters Group Mice
Drug tested ip Dose Day 0 Day 3 Day 7 Day 11 1 9 male no treatment
control inoculate 2 5 male free cisplatin 3 mg/kg inoculate 1 mg/kg
1 mg/kg 1 mg/kg 3 5 male free cisplatin 4.5 mg/kg inoculate 1.5
mg/kg 1.5 mg/kg 1.5 mg/kg 4 5 male Lipid- 3 mg/kg inoculate 1 mg/kg
1 mg/kg 1 mg/kg complexed cisplatin 5 5 male Lipid- 4.5 mg/kg
inoculate 1.5 mg/kg 1.5 mg/kg 1.5 mg/kg complexed cisplatin 6 5
male Lipid- 6 mg/kg inoculate 2 mg/kg 2 mg/kg 2 mg/kg complexed
cisplatin 7 5 male Lipid- 9 mg/kg inoculate 3 mg/kg 3 mg/kg 3 mg/kg
complexed cisplatin
The results from the experiment are summarized in Table 12.
TABLE-US-00012 TABLE 12 Survival data as measured by % T/C. Group
Drug tested Dose Median survival* % T/C 1 no treatment Control 13.5
100 2 free cisplatin 3 mg/kg 26.5 196 3 free cisplatin 4.5 mg/kg
29.5 218 4 Lipid-complexed 3 mg/kg 19.5 144 cisplatin 5
Lipid-complexed 4.5 mg/kg 23.5 174 cisplatin 6 Lipid-complexed 6
mg/kg 25.5 189 cisplatin 7 Lipid-complexed 9 mg/kg 25.5 189
cisplatin *Median survival is defined as day of death for 50% of
mice with each group.
Day of death for 50% of mice within each group was determined and
an initial % Treated/Control (T/C) value was recorded in the above
table. At the optimal dose, the cisplatin in lipid-complexed
cisplatin does not lose any of its antitumor activity compared to
free cisplatin.
Example 14
Antitumor Activity of L-CDDP Against Human Ovarian Cancer
Xenograft
[0150] Nude mice, female, 6-7 weeks old, were intraperitoneally
inoculated with human ovarian cancer cell line SK-OV.sub.3-ip1
(1.5.times.10.sup.6 cells/mouse). One week after the inoculation,
the mice were randomly divided into 3 groups with 5 mice in each.
One group of mice was given single bolus ip injection of CDDP with
MTD (9 mg/kg) to mimic the current chemotherapy (positive control).
Another group was treated with single bolus ip injection of L-CDDP
with MTD (23 mg/kg). The lipid-complexed cisplatin formulation were
prepared in the same manner as in Example 3. The third group of
mice without treatment was used as negative control. The mice were
observed on a daily basis. Death of mice was recorded and the
increased lifespan (ILS) was calculated. Results are presented in
FIG. 10.
Example 15
Comparison of Lipid-Complexed Cisplatin Prepared by the Cyclic
Temperature Effusion Process and Non Cyclic Temperature Cisplatin
Liposomes
[0151] The lipid-complexed cisplatin prepared by the cyclic
temperature effusion process were prepared as in Example 3 and
contained 1.1 mg/ml cisplatin and 27 mg/ml total lipid. The non
cyclic temperature cisplatin liposomes were prepared according to
the following procedure. [0152] 1. DPPC (3.0 g) and cholesterol
(1.2 g) were co-dissolved in 20 mL of ethanol. [0153] 2. Cisplatin
(200 mg) was dissolved in 0.9% saline (200 ml). [0154] 3. The
lipid/ethanol solution was infused into the cisplatin solution as
it was being well-stirred (liposomes formed). [0155] 4. The
lipid-cisplatin suspension was dialyzed to wash away un-entrapped
cisplatin. [0156] 5. The resulting liposomal cisplatin contained
0.03 mg/ml total cisplatin (75% of total cisplatin was entrapped
and 25% was un-entrapped); the total lipid concentration was 21
mg/ml.
[0157] The mice were given equivalent amounts of cisplatin
containing therapeutics based on the amount of lipid instead of the
amount of cisplatin. This was necessary because in non cyclic
temperature cisplatin liposomes the lipid to cisplatin ratio is so
high that it is not possible to administer that much lipid
necessary to equal the amount of cisplatin in the lipid-complexed
formulations prepared as in Example 3.
[0158] Female DBA/2 mice (Charles Rivers) were used. Thirty (30)
mice were injected with 2.times.10.sup.6 L1210 cells ip on Day 0.
On day 1, the mice were weighed and randomized into 3 groups of 10
mice. On days 5, mice received a single bolus intraperitoneal
injection of soluble cisplatin (6 mg/kg), lipid-complexed cisplatin
(6 mg/kg, ip) or non cyclic temperature cisplatin liposomes (equal
lipid to lipid-complexed cisplatin, 0.2 mg/kg). Survival was
monitored. Mice were weighed daily after day 10. Mice that lost 20%
or greater of their starting weight were euthanized by CO.sub.2
inhalation. The date of their death was recorded on data sheets.
Median survival was calculated by Prism GraphPad.
[0159] The results of experiments where both types of cisplatin
formulations were administered intraperitoneally to mice with
implanted viable L1210 tumor cells are depicted in FIG. 11. There
was no significant difference between survival curves of mice that
received lipid-complexed cisplatin intraperitoneally and those that
received soluble cisplatin intraperitoneally (p=0.20). All survival
curves of cisplatin-treated groups were significantly different
from the mice that received non cyclic temperature cisplatin
liposomes (p=0.0035, and p<0.0001, respectively). The days of
median survival were 19 for lipid-complexed cisplatin, ip; 19.5 for
free cisplatin, ip; and 14 for non cyclic temperature cisplatin
liposomes, ip.
INCORPORATION BY REFERENCE
[0160] All of the patents and publications cited herein are hereby
incorporated by reference.
EQUIVALENTS
[0161] Those skilled in the art will recognize, or be able to
ascertain using no more than routine experimentation, many
equivalents to the specific embodiments of the invention described
herein. Such equivalents are intended to be encompassed by the
following claims.
* * * * *