Note : Les descriptions sont présentées dans la langue officielle dans laquelle elles ont été soumises.
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TITLE OF INVENTION
Sparing Paclitaxel by the Use of Hyaluronan.
FIELD OF INVENTION
This invention relates to the use of forms of hyaluronan for use to deliver
to a patient the highly lipophilic therapeutic compound paclitaxel for example
sold under the trade mark Taxol and in one aspect, to deliver effective dosage
amounts of paclitaxel to the patient in need thereof which amount of paclitaxel is
present in an effective dosage amount which is less than the usual dosage
amount used today when treating a patient having cancer. In one aspect, this
10 invention relates to the use of forms of hyaluronan to deliver amounts of thehighly lipophilic therapeutic compound paclitaxel in such amount which would
today not be normally expected to provide effective treatment to a patient and
which amount becomes effective for treating the patient when combined with a
form of hyaluronan.
15 BACKGROUND OF INVENTION
PCT Application No. WO 91 /04058 discloses the use of forms of
hyaluronan for delivery of medicines and therapeutic agents for the treatment ofdiseases and conditions of patients. The treatments were for underperfused
tissue and pathological tissue (see p. 24 of the PCT Application).
Among the medicines and therapeutic agents suitable for delivery by
hyaluronan are lipophilic therapeutic compounds such as chemotherapeutic
agents to treat the underperfused and pathological tissue including tumours.
While the usual dosage amounts known at the time of application WO 91/04058
of the agent are successfully administered by the use of hyaluronan, larger dosage
25 amounts (excess amounts) of medicines (such as NSAIDS) may also be
successfully used to treat patients without the usual deleterious side effects
expected. The amount of hyaluronan substantially reduces the significant
deleterious side effects of for example the NSAIDS when administered together
even when substantially larger dosage amounts of NSAIDS are used.
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SUMMARY OF INVENTION
Applicants have now discovered that substantially lesser amounts than
the usual dosage amounts of the highly lipophilic therapeutic compound
paclitaxel may be successfully administered to treat patients and such lesser
amounts will be effective dosage amounts in the treatment if administered with
a form of hyaluronan to treat underperfused tissue and/or pathological tissue
(which express excess hyaluronan receptors [more than normal tissue]) such as
tumours. This unexpected finding provides the benefit to the patient of
successful treatment of pathological tissue expressing excess hyaluronan
10 receptors, while receiving much less than the usual accepted dosage amount ofthe highly lipophilic agent paclitaxel. As paclitaxel is toxic, the patient is not
administered as much as previously. The toxic side-effects of paclitaxel on the
patient and particularly his/her liver is very much reduced. Additionally as
paclitaxel is expensive (in the order of about $14,000/kg. of TaxolTM) and the
15 usual dosage of 175mg/m2 administered intravenously over 3 hours every three
weeks is reduced substantially, the costs of treatment are substantially reduced.
Thus, paclitaxel for example paclitaxel diphenhydromine is "spared" (the
amount being used in one dosage is substantially reduced from that normally
used by itself).
It is therefore an object of this invention to provide improved
formulations comprising "spared" amounts of the highly lipophilic
antineoplastic agent paclitaxel for the treatment of cancer.
It is a further object of the invention to provide methods of treatment of
patients using the spared amounts of the agent paclitaxel in dosage amounts of
25 formulations comprising forms of hyaluronan and which spared amounts of the
agent are effective amounts for successful treatment of patients having for
example cancer when administered with forms of hyaluronan and which
amount of the agent paclitaxel would not otherwise be an effective amount if
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administered without hyaluronan. Such spared amounts of the medicine
paclitaxel may be as much as about 4 - 6 times less than the usual amount of
paclitaxel administered alone and still be effective when combined with
hyaluronan. For example, where TaxolTM (paclitaxel) is involved, the amount of
TaxolTM (paclitaxel) "spared" may be at least 3 times less (and as much as 4 - 6times less) than the usual dosage amount of the agent when administered
without hyaluronan.
The accepted amount of paclitaxel usually given to a patient is in the order
of 175mg/m2. In accordance with this invention, an effective amount of
10 paclitaxel sold under the trade mark TaxolTM may be in the order of 55mg/m2 or
less (such as 28mg/m2) when paclitaxel is administered with the form of
hyaluronan and such amount is now found to be effective in the treatment of
patients suffering from cancer.
The form of hyaluronan may comprise amounts between 10mg and
15 1000mg or more in each dosage amount. As hyaluronan is not toxic, even
greater amounts of hyaluronan may be used in each dosage for example up to
3000mg per dosage.
The dosages are administered to a patient as required - for example,
dosages per week for the duration of treatment. The dosages may be
20 administered by injection, intravenously or directly into the tumour.
The form of hyaluronan may be selected from hyaluronan and
pharmaceutically acceptable non-toxic salts thereof such as sodium hyaluronate.
Many forms of hyaluronan may be suitable for use herein although those
preferred are those discussed hereafter. Molecular weights of forms of
25 hyaluronan less than about 750,000 daltons and preferably greater than 150,000
daltons (protein standard) in sterile water prepared having a viscosity for
intravenous administration are suitable.
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One specific form of pharmaceutical grade is a 1% sterile sodium
hyaluronate solution (50 ml vials) provided by Hyal Pharmaceutical Corporation
which has the following characteristics:
Tests Specifications
1. Container Description 150 mL Flint glass vial with
a red or gray rubber stopper
and an aluminum seal, 20
mm in size.
2. Product Description A clear, colourless, odourless,
transparent, slightly viscous
liquid.
3. Fill Volume 50.0 to 52.0 mL
4. pH 5.0 to 7.0 at 25 degrees C.
5. Specific Gravity 0.990 to 1.010 at 25 degrees C.
6. IntrinsicViscosity 4.5 to 11.0 dL/g
7. Molecular Weight 178,000 to 562,000 daltons
8. Sodium Hyaluronate Assay 9.0 to 11.0 mg/mL. Positive
and Identification
9. Particulate Matter No visible Particulate Matter
10. Sterility MeetsRequirements forSterility,
USP 23
11. Bacterial Endotoxins (LAL) Meets Requirements for Bacterial
Endotoxins, USP 23.
This pharmaceutical grade 1% sterile solution of hyaluronan may be made
25 from granules/powder having the following characteristics:
Tests Specifications
1. Description White or cream-coloured granules or
powder, odourless
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2. Identification (IR Spectrum) Must conform with the Reference
Standard Spectrum.
3. pH (1% Solution) Between 5.0 and 7.0 at 25 degrees C.
4. Loss on Drying NMT 10.0% at 102 degrees C. for 6
hours.
5. Residue on Ignition Between 15.0 and 19.0%
6. ProteinContent NMT 0.10%
7. Heavy Metals NMT 20 ppm (as per USP 23 p. 1727).
8. Arsenic NMT 2 ppm (as per USP 23, p. 1724).
9. Residual Solvents a) Acetone: NMT 0.1%
b) Ethanol: NMT 2.0%
c) Formaldehyde: NMT 100 ppm
10. Sodium Hyaluronate Assay 97.0 to 102.0% (dried basis)
11. Intrinsic Viscosity Between 10.0 to 14.5 deciliters per gram.
12. Molecular Weight Between 500,000 to 800,000 daltons
(,~AI~lllA~d using the Laurent Formula) (based on intrincis viscosity).
13. Total Aerobic Microbial Count NMT 50 microorganism/gram (as per
USP 23, p. 1684).
14. Test for Escherichia coli Escherichia coli is absent (as per USP
23, p. 1685).
15. Yeasts & Molds NMT 50 microorganisms/gram (as per
USP 23, p. 1686).
16. Endotoxins (LAL) NMT 0.07 EU/mg (as per USP 23, p.
1696).
A topical grade of hyaluronan may, in certain circumstances be suitable
and may be made from the following granules/powder which have the
following characteristics:
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Tests Specifications
1. Description White or cream-coloured granules or
powder, odourless
2. Identification (IR Spectrum) Must conform to the Reference Standard
Spectrum.
3. pH (1% Solution) Between 6.0 and 8.0 at 25 degrees C.
4. Loss on Drying NMT 10.0% at 102 degrees C. for 6
hours.
5. Residue on Ignition Between 15.0 and 19.0%
6. ProteinContent NMT 0.40%
7. Heavy Metals NMT 20 ppm (as per USP 23 p. 1727).
8. Arsenic NMT 2 ppm (as per USP 23, p. 1724).
9. Residual Solvents a) Acetone: NMT 0.1%
b) Ethanol: NMT 2.0%
c) Formaldehyde: NMT 100 ppm
10. Sodium Hyaluronate Assay 97.0 to 102.0% (dried basis)
11. Intrinsic Viscosity Between 11.5 to 14.5 deciliters per gram.12. Molecular Weight Between 600,000 to 800,000 daltons
(calculated using the Laurent Formula) (based on intrinsic viscosity).
13. Total Aerobic Microbial Count NMT 100 microorganism/gram (as per
USP 23, p. 1684).
14. Test for Staphylococcus aureus Staphylococcus aureus is absent (as per
USP 23, p. 1684).
15. Test for Pseudomonas aeruginosa Pseudomonas aeruginosa is absent (as
per USP 23, p. 1684).
16. Yeasts & Molds NMT 200 CFU/gram (as per USP 23, p.
1686) .
This topical grade may then be sterilized.
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Other forms may be suitable such as one form of hyaluronic acid and/or
salts thereof (for example, sodium salt) may be an amount also supplied by Hyal
Pharmaceutical Corporation. One such amount is a 15 ml vial of Sodium
hyaluronate 20mg/ml (300mg/vial - Lot 2F3). The sodium hyaluronate fraction
is a 2% solution with a mean average molecular weight of about 225,000. The
amount also contains water q.s. which is triple distilled and sterile in accordance
with the U.S.P. for injection formulations. The vials of hyaluronic acid and/or
salts thereof may be carried in a Type 1 borosilicate glass vial closed by a butyl
stopper which does not react with contents of the vial.
The amount of hyaluronic acid and/or salts thereof (for example sodium
salt) may comprise hyaluronic acid and/or salts thereof having the following
characteristics:
a purified, substantially pyrogen-free fraction of hyaluronic acid
obtained from a natural source having at least one characteristic
selected from the group consisting of the following:
i) a molecular weight within the range of 150,000 - 225,000;
ii) less than about 1.25% sulphated mucopolysaccharides on a
total weight basis;
iii) less than about 0.6% protein on a total weight basis;
iv) less than about 150 ppm iron on a total weight basis;
v) less than about 15 ppm lead on a total weight basis;
vi) less than 0.0025% glucosamine;
vii) less than 0.025% glucuronic acid;
viii) less than 0.025% N-acetylglucosamine;
ix) less than 0.0025% amino acids;
x) a UV extinction coefficient at 257 nm of less than about
0.275;
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xi) a UV extinction coefficient at 280 nm of less than about 0.25;
and,
xii) a pH within the range of 7.3 - 7.9. Preferably, the hyaluronic
acid is mixed with water and the fraction of hyaluronic acid
fraction has a mean average molecular weight within the
range of 150,000 - 225,000.
Preferably this amount of hyaluronic acid comprises at least one
characteristic selected from the group consisting of the following characteristics:
i) less than about 1% sulphated mucopolysaccharides on a
total weight basis;
ii) less than about 0.4% protein on a total weight basis;
iii) less than about 100 ppm iron on a total weight basis;
iv) less than about 10 ppm lead on a total weight basis;
v) less than 0.00166% glucosamine;
vi) less than 0.0166% glucuronic acid;
vii) less than 0.016% N-acetylglucosamine;
viii) less than 0.00166% amino acids;
ix) a UV extinction coefficient at 257 nm of less than about 0.23;
x) a UV extinction coefficient at 280 nm of less than 0.19; and
xi) a pH within the range of 7.5 - 7.7
Other forms of hyaluronic acid and/or its salts may be chosen from other
suppliers, for example those described in prior art documents disclosing forms of
hyaluronic acid having lower molecular weights between about 150,000 daltons
and 750,000 daltons (protein standard) being prepared as for example, 1-2%
solutions in sterile water for intravenous administration. In addition, sodium
hyaluronate produced and supplied by LifeCoreTM Biomedical, Inc. having the
following specifications may be suitable (if sterile):
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Characteristics Specification
Appearance White to cream
colored particles
Odor No perceptible odor
Viscosity Average < 750,000 Daltons
Molecular Weight
UV/Vis Scan, 190-820nm Matches reference scan
OD, 260nm < 0.25 OD units
Hyaluronidase Sensitivity Positive Response
IR Scan Matches reference
pH, 10mg/g solution 6.2 - 7.8
Water 8% maximum
Protein < 0.3 mcg/mg NaHy
Acetate < 10.0 mcg/mg NaHy
Heavy Metals, maximum ppm
As Cd Cr Co Cu Fe Pb Hg N i
2.0 5.0 5.0 10.0 10.0 25.0 10.0 10.0 5.0
Microbial Bioburden None observed
Endotoxin < 0.07EU/mg NaHy
Biological Safety Testing Passes RabbitOcular
Toxicity Test
The following references teach hyaluronic acid, sources thereof and
processes of the manufacture and recovery thereof.
Canadian Letters Patent 1,205,031 (which refers to United States Patent
4,141,973 as prior art) refers to hyaluronic acid fractions having average
molecular weights of from 50,000 to 100,000; 250,000 to 350,000; and 500,000 to
730,000 and discusses processes of their manufacture
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- 10-
Where high molecular weight hyaluronic acid (or salts or other forms
thereof) is used, it must, prior to use be diluted to permit administration and
ensure no intramuscular coagulation. Recently, it has been found that large
molecular weight hyaluronic acid having a molecular weight exceeding about
1,000,000 daltons self-aggregates and thus, does not interact very well with HA
receptors. Thus, the larger molecular weight hyaluronic acid (such as HealonTM)
should be avoided.
Thus, according to an aspect of the invention, the invention provides a
composition comprising a therapeutically effective amount of paclitaxel with a
10 therapeutically effective amount of hyaluronic acid and/or a pharmaceuticallyacceptable salt thereof, together with a pharmaceutically-acceptable diluent
wherein the therapeutically effective amount of paclitaxel is at a significantlyreduced level than would be normally used (for example greater than three times
less). Thus the amount of paclitaxel reduced from that normally used, is
15 "spared" .
The invention also provides a method of treatment of a pathological
condition in a subject in need of such treatment, comprising the step of
administering an effective dose of a composition according to the invention to
said subject over such period as may be required. For example, the treatment
20 may continue for months. However in each instance the effective amount of the agent is "spared".
The invention also provides the use of a composition comprising a form
of hyaluronan and an effective dosage amount of paclitaxel for treating a
pathological condition (cancer) and wherein the therapeutically effective amount25 of paclitaxel is at a significantly reduced level than would be normally used (for
example greater than three times less).
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It will be clearly understood that the dose and route of administration will
depend upon the condition to be treated, and the attending physician will readily
be able to determine suitable doses and routes.
The subject to be treated may be a human (or may be an animal).
The invention further provides a method of preparing a composition of
the invention, comprising the step of combining such highly lipophilic
therapeutic compound paclitaxel with hyaluronan and/or a pharmaceutically-
acceptable salt thereof, and a diluent (such as sterile water) and put into a suitable
pharmaceutically acceptable tolerable form (which is of course, non-toxic). The
10 agent once again will be "spared". The inventors believe that the unique
attraction between hydrophobic patches of the hyaluronan and paclitaxel enable
the paclitaxel to be "spared".
While hyaluronan (HA) is generally hydrophilic, it has these hydrophobic
patches which Applicants believe permit the binding/association of the
15 hyaluronan with the paclitaxel. These hydrophobic patches are dispersed on the
molecule. Because of the dispersion and lack of knowledge of the exact positions,
it is not known if any particular compound combined with the hyaluronan
would be capable of being spared to provide effective dosages to the patient.
The invention will now be illustrated with reference to the following tests
20 which illustrate the invention.
The tumour model used in the tests was that described in detail on pages
59 - 60 of Fourth International Workshop - on Hyaluronan in Drug Delivery as
follows:
Colon-26 cells
Colon-26 cells, a gift from the Imperial Cancer Research Fund
(London), were maintained in antibiotic supplemented DMEM
containing 10% fetal calf serum at 37~C in 95% oxygen/5% carbon
dioxide.
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Tumour Induction
Anaesthetized BALB/c mice were subcutaneously implanted
with sponges (8 x 4 mm) and seeded with 106 Colon-26 cells three
days later by injection.
5 The tumours were allowed to establish for 4 days before dosing was initiated.
Dosing was intravenous (0.25ml/day) for 6 days of amounts as shown in the table
below and the tumours were excised 24 hours after the last dose. The data so farrelates to wet and dry weights of the tumour mass, and is shown in the following table:
Group n Wet Weight/mg Dry Weight/mg
Control PBS 7 470 15 84.5+4.2
Cremophor 9 514+ 39 94.4+ 7.8
Taxol 2.5mg/kg 6 482 37 81.7 9.2
Taxol 10mg/kg 8 464+ 30 73.9 + 6.4
HA7.5mg/kg 7 485 28 72.1 + 5.3~
Taxol 2.5mg/kg + 8 391 + 38~ 60.4 + 8.0~*
HA 7.5mg/kg
~ p<0.05
~ ~ p<0.01 comparison with Cremophor control.
HA = hyaluronan (sodium hyaluronate (EM) having a molecular weight
between 50,000 and 750,000 daltons (protein standard)
Taxol was solubilised in a mixture of 1:1 ethanol and cremophor EL.
15 Further dilution was in sterile PBS. The final cremophor concentration in allTaxol groups was 1.6%. A similar solution served as vehicle control (the
cremophor group in the table above).The HA alone should be compared with the
PBS control group. Numbers refer to dose in mg/kg.
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The dry weights are of particular interest. We have a dose response
reduction in dry weight with Taxol, and the combination of low dose Taxol with
HA gives the greatest suppression of all.
To note: the reduction in tumour weight caused by HA alone; this is
5 significant only against the Cremophor and not the PBS control. Nonetheless,
the data speaks for itself and demonstrates that HA had no detrimental effect ontumour growth. In fact it appears to suppress tumour growth possibly to the
same extent as the high dose Taxol. (See PCT application WO 95/30423). The
form of HA and TaxolTM may be administered in any suitable manner such as by
10 direct injection, intravenous administration etc.
As many changes can be made to the embodiments without departing
from the scope of the invention, it is intended that all material contained herein
be interpreted as illustrative of the invention and not in a limiting sense.
