Note: Descriptions are shown in the official language in which they were submitted.
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TA~AN~ CO~ lON AND METHQD
Field of the I~
The present invention relates to formulations of taxol and related taxane compounds,
5 which have improved safety, solubility and stability characteristics, and to methods of preparing
such formulations.
R~r~ .lC~S
Arbuck, S.G.. and Blaylock, B.A., in TAXOL: SCIENCE AND APPLICATIONS, (Suffness,10 M., Ed.) CRC Press, New York, NY, pp. 379-415 (1995).
Straubinger, R.M., in TAXOL: SC1PNCE AND APPLICATIONS, (Suffness, M., Ed.) CRC
Press, New York, NY, pp. 237-258 (1995).
n ~- ~v~ ,d of the ll~ liu~
Taxol, also known as paclitaxel, is a compound e~rr~rted from the bark of the western
yew, Ta~zcs brevifolia. Much attention has been drawn to taxol for use as an a.~ e~ ic
agent. Taxol has shown good response rates in the llGaLlll~ of ovarian and breast cancer pa-
tients who did not respond to cisplatin or vinca alkaloid therapy. Taxol is also being rx;~ rd
for treating a variety of other cancers, such as m~ nnrn~ Iymphoma and lung cancer.
A ma~or problem ~C~ofi~t~cl with taxol is its low solubility in aqueous solveMs.
Because taxol lacics r~ l groups that are ionizable in a ph~ ;r~lly acceptable range,
manipulation of pH does not enhance solubility. Producing salts or adding charged complexing
agents are also inapplicable (SLla~ill~ , 1995, p. 238). ~orm~ ting taxol in a bioco~"~aLible
carrier has thus been a challenge throughout its thc.a~ ic development.
In the search for taxol form~ tiQnc having improved solubility and toxicity properties,
a number of ph~ll,.~r~..lir~l vehicles have been invçstig~e~. Gene}ally, such vehicles have
inr~ cl a cosolvent, such as ethanol, di",~,Ll.~l:,ulfoxide (DMSO) or low molecular weighe
polyethylene glycol (e.g., PEG 400), with or without an oil or :iulrdc~ additive such as a
polyoxyethylene sorbitan fatty acid ester (e.g., "TWEEN 80", also known as polysorbate-80),
30 polyethoxylated castor oil (e.g., "CREMOPHOR EL"), soybean oil, or triacetin. However,
these formulations have suffered from either poor solubility, particularly foliowing dilution into
saline solution for intravenous ~ ion~ or from high toxicity, due to the oil or
~u~racL~IL. In particular, the ~ll" ,i~ dLion of "TWEEN-80" in amounts necessary to solubilize
taxol at high concentration is associated with pleural effusions and edema, and "CREMOPHOR
35 EL" can produce serious or fatal hy~ cn~iLi~/ity (Straubinger, 1995, pp. 241 and 244).
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There is therefore a need for formulations of taxol having reduced toxicity while
i"j"g high stability for long term storage.
of the Inven~ion
The present invention includes, in one aspect, a taxane storage solution for
ph~rm~rellrir~l use. The storage solution comprises (a) a taxane compound in a pharmaceu-
tically pure form, ~b) a polyoxyethylene sorbitan fatty acid monoester, (c) polyethoxylated
castor oil, and (d) ethanol. In the solution, the monoester and polyethoxylated castor oil are
present together in amounts effective to reduce the toxicity of the solution relative to the
toxicity observed when either the polyoxyethylene sorbitan fatty acid monoester or polyeLIl~ yl-
ated castor oil is used in the absence of the other. The pH of the storage solution is pl~r~ably
between about 1 and 8. The taxane compound is preferably taxol or clocer~
In a pl~rell~d embodiment, the solution additionally includes a low molecular weight
polyethylene glycol, such as PEG 300.
The solution may additionally include a pharm~reutic:~lly acc~:~JLal)le acid as a buffering
agent, wherein the pH is .~ i..rd between about 4 and about 6.
In a pi~rc;-~,d embodiment, the storage solution includes 4 mg/ nL to 8 mg/mL of a
taxane, such as taxol, 20 to 30% (v:v) polyethoxylated castor oil, 5 to 15% (v:v) polyoxyethyl-
ene ~20) sorbitan mono-oleate, 15 to 30% (v:v) ethanol, and 40 to 60% (v:v) low molecular
20 weight polyethylene glycol.
In another aspect, the invention includes a method of treating a cancer condition in a
m~mm~ n subject. In the method, there is provided a taxane storage solution in accordance
with the description above. The storage solution is diluted with a diluent suitable for intrave-
nous ~ l dlion, to produce a dilute taxane solution. The solution is then ~lmini.~trred to
25 the subject in a pha~ r~ ir~lly acc~l~le amount effective to inhibit cancer growth in the
subject. In ~ler~ ;d embodiments, the method is used to treat ovarian cancer or breast cancer.
The invention also includes a method of pi~,~d~ g a taxane solution for hllldvellu-ls
;..I...i~i~l . dLion. In the m~thod, a taxane storage solution of the type described above is diluted
with a diluent suitable for h~lldv~nuu~7 ~ ",il~ ~ dlion' to produce a dilute taxane solution. The
30 dilute taxane solution may be a-lmini.ctered in a method of treating cancer, as noted above.
In another aspect, the invention includes a ph~rm~rell~ic~l vehicle for delivering a non-
po}ar drug, such as taxol, to a subject. The vehicle includes a polyoxyethylene sorbitan fatty
acid monoester, and polyethoxylated castor oil. The monoester and polyethoxylated castor oil
are present in arnounts effective to reduce the toxicity of the vehicle relative to the toxicity
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observed when either the monoester or the polyethoxylated castor oil is used in the absence of
the other. The vehicle is useful when a solubilizing agent is ~ecess,..y to dissolve a non-polar
drug in solution, and where using the polyoxyethylene sorbitan fatty acid monoester without
~ the polyethoxylated castor oil, or oil without the monoester, produces toxic effects which limit
5 the amount of drug that can be ~ f.cd In a plcr.~ d embodiment, the vehicle addi-
~' tionally includes a low molecular weight polyethylene glycol, such as PEG 300. The invention
also includes a drug composition comprising a non-polar drug in a vehicle of the type just de-
scribed.
These and other objects and features of the invention are described more fully below.
Detailed lDes~ lion of the Invention
I. Definition~
As used herein, the terms below are inter~ cl to have the following m~ning~.
By "taxane" is meant any compound (a) having the 6-8-6 fused ring backbone of taxol,
in~hl-ling additional substitn~ontc or bonding n~scess~tly for taxol activity (e.g., 9-ketone or 9-
hydroxyl, 4,5-oxetane ring, 4-acetoxy, and 2-benzoyloxy; see also Chapter 13 on taxane
structure-activity relationships in TAXOL: SCIENCE AND APPLICATIONS, cited above, particularly
page 339), and (b) which inhibits depolylllc~iGa~ion of microtubules. Exemplary taxane com-
pounds are taxol (paclit~ l) and docataxel ("TAXOTERE").
By "polyoxyethylene sorbitan fatty acid m(mf~est~or~ is meant a compound having a
sorbitan core (1,4-sorbitol cyclic ether), wherein the 2, 3, and 5-hydroxyl groups of the
sorbitan core are each dcliv~ cd with one or more ethylene oxide monol-lel~., and the 6-
hydroxyl of the core is derivatized with one or more ethylene oxide monomers which ~~. ."i.,~le
with a fatty acid ester group. The number of ethylene oxide monomers in the compound will
generally be between 10 and 50, and plercl~bly between 10 and 30. An exemplary
polyoxyethylene sorbitan fatty acid monoester is "TWEEN 80", also known as polyoxyethylene
(20) sorbitan mono-oleate, wherein "(20)" inrlie~tes that the total number of ethylene oxide
monomers ~tt~rh~l to the sorbitan core is 20.
By "fatty acid" is meant a C-16 to C-22 carboxylic acid which may be entirely aliphatic
or may contain one or more carbon-carbon double bonds. Exemplary fatty acids include
palmitic acid (C-16), stearic acid (C-18), and oleic acid (cis-9-oct~-i.oc~noic acid).
By a polyoxyethylene sorbitan fatty acid monoester and polyethoxylated castor oil being
"present together in amounts effective to reduce the toxicity of the solution relative to the
toxicity observed when either the polyoxyethylene sorbitan fatty acid monoester or polyethoxyl-
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ated castor oil is used in the absence of the other" is meant that the monoester and oil are
present together in amounts effective to reduce the toxicity of a taxane storage solution (after
dilution for intravenous a~ .alion~ relative to the toxicity that would be obtained if the
monoester/oil colllbil,aLion of the invention were replaced with molloe~L~,. compound alone or
oil compound alone in an amount sllffleient to achieve the same degree of solubilization of the
taxane compound as achieved by the monoester/oil combination.
By "low molecular weight polyethylene glycol" is meant polyethylene glycol (PEG)having an average molecular weight of 200 to 3000 daltons.
mm~ n subject" is intPn~ (l to have its traditional mP~ning, and ellco-~lpdsses cats,
dogs, sheep, horses, and particularly humans, for example.
II. Taxane Stora~e Solution
The present invention is directed to an hll~l ovt;d composition and method for delivering
high doses of taxanes to cancer patients using a vehicle with reduced toxicity. The invention
is based in part on the discuv~y that using a polyoxyethylene sorbitan fatty acid monoester in
coll~ alion with a polyethoxylated castor oil, as solubilizing agents for a taxane compound,
is ~rre.;Live to provide high taxol solubility and stability, but with reduced toxicity.
The storage solution of the invention in~l~ld-oc a taxane in pl~ c--l irzllly pure form,
which is solubilized at high cunc~llLl aLion using a polyoxyethylene sorbitan fatty acid monoester
and polyethoxylated castor oil in an ethanol base. Preferably, the taxane is present at a
cunc~ a~ion of between about 2 and about 20 mg/mL, and typically between about 4 and
about 8 mg/mL.
The monoester and polyethoxylated castor oil are present together in amounts effective
to reduce the toxicity of the solution relative to the toxicity observed when either the polyoxy-
ethylene sorbitan fatty acid monoester or the polyethoxylated castor oil is used in the absence
of the other. The polyethoxylated castor oil is from any pharm~relltir~lly acceptable source.
One suitable ~l~dLion is available from BASF (Wyandotte, MD) under the trademark"CREMOPHOR EL". Generally, the polyethoxylated castor oil is present at a concentration
of about 10 to about 40% (v:v), and preferably between about 20 to about 30%.
The sorbitan fatty acid monoester is generally present at a concentration of about 5 to
about 20% (v:v), plert;ldbly between about 5 and about 15 % . One preferred polyoxyethylene
sorbitan fatty acid monoester is "TWEEN 80".
The polyethoxylated castor oil and sorbitan fatty acid monoester together constitute a
total conc~llL,aLion in the storage solution of between about 15 to about 60%, preferably from
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about 25 to 45 % (v:v). In addition, the polyethoxylated castor oil and sorbitan monoester are
used in a ratio (oil:sorbitan monoester, v:v) of between about 0.5 to 6, preferably between
about 1.3 and 6, and more preferably between about 2 and 3. It shouid be noted that the poly-
ethoxylated castor oil and polyoxyethylene sorbitan fatty acid monoester serve not only to
5 enhance the solubility of the taxane, but also to enhance the anti-cancer potency of the taxane
when a~ d against tumor cells. According to an h~ JolldllL feature of the invention,
using the polyethoxylated castor oil and sorbitan monoester together results in lower toxicity
due to these components than would be expected if the oil is used without the monoester
compound or the monoester compound is used without the oil compound.
The storage solution of the invention may also include a low molecular weight polyeth-
ylene glycol (PEG) having an average molecular weight of 200 to about 3000 daltons,
preferably between about 200 and about 1000 daltons. The PEG plc;lJdldlion is preferably one
which is a liquid at a temperature above 15~C, e.g., having an average molecular weight of
between about 200 and about 1000 daltons, and pl~,feldbly beL~eell about 200 and about 500.
15 PEG is optionally also in~hlfle~ in the storage solution to improve the solubility and stability
of the taxane. Preferably, the level of PEG is between 10 and 60%, more plc;rtldbly between
about 40 and about 60%.
The storage solution may also optionally include a buffering agent which m:~int~in~ the
pH of the storage solution between about 1 and about 8, preferably between about 4 and about
20 6. Preferably, the buffering agent is ph~rm~ellti~lly acceptable acid, more ,l~ler~ldbly a carb-
oxylic acid, such as citric acid, acetic acid, maleic acid, succinic acid, lactic acid, ascorbic
acid, ghlt~mic acid, or aspartic acid. Preferably, the buffering agent is al~ydl~us citric acid.
The buffering agent may be present at a concentration of between about 2 and about 200 mM,
typically between about S and about 20 rnM. The rPm~infler of the storage solution is
25 preferably made up by ethanol. The storage solution preferably does not contain water.
The storage solution of the invention is prepared by any method suitable to solubilize
the taxane component, inrln~ling the use of sonication and heating. Exemplary methods for
plClldlillg solutions in accold~lce with the invention are provided in Example 1. The solution
may be stored at room ~ elalule7 and preferably at 4~C or lower. The solution is p~rt;l~ly
30 treated to remove particulate matter by passage through a filter Ill~lllbldlle, e.g., a 0.22 ~m
pore-size lll~ll.hldlle. The solution may also be purged with nitrogen gas to remove oxygen.
The stability properties of the storage solution of the invention are illustrated by the
studies described in Examples 2 and 3. In the study described in Example 2, aliquots of two
storage solutions in accofdallce with the invention were placed in an autoclave and heated under
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~le~uie at 250~C for 20 minutes. The samples were then diluted in acetonitrile and analy~d
by HPLC. No sign of taxol degradation was detf Ct~f~
In the study described in Example 3, sarnple solutions were incubated at 37~C for 12
weeks, and aliquots were periodically removed and tested by HPLC for degradation of taxol.
5 The sample solutions tested included Formulations 1 and 2 from Example l, as well as a solu-
tion containing taxol in a 1:1 mixture of polyethoxylated castor oil and ethanol (Formnl~tif)n
3). As can be seen from the results in Example 3, the taxol solutions in accoldallce with the
present invention are at least as stable as Formulation 3, with less than 2% degradation after
12 weeks.
According to another important feature of the invention, the storage solution of the
invention is compatible with dilution into standard so}utions for intravenous a~ dLion of
drugs. In the study described in Example 4, the formlll~tions from Example 1 were diluted in
normal saline (O.g% NaCl in water) by dilution factors of 1:5, 1:10, 1:25 and 1:50 and were
then f-x~",i"f-d for signs of L lecil,iL~lion or cloudiness after 1, 2, 4, 8, 24, and 48 hours. All
15 dilutions r~m~infd clear for the first 24 hours for both forrmll~tif)ns, and Formulation 1
l~...~i..rd clear for 48 hours. These results indicate that storage solutions in accordance with
the invention are suitable for intravenous :lflmini~tration
In the study described in Exarnple 5, the relative toxicities of the storage vehicle alone
(storage solution without taxol) were ~;ol-.pal~d with a vehicle collsi~lillg of a 1:1 mixture of
20 polyethoxylated castor oil and ethanol (Formnl~tion 3). In one experiment, groups of 2 or 3
mice were ~ lcd single dosages of test formnl~tions in lmflilllt~d form, and the mice
were monitored for 21 days for signs of intolerance of the ~fl~ cd dosages. Signs of
intoleranceincludedanyofthefollowing: (l)~ignffilf~ntweightloss(>20%),(2)piloerection,
(3) prolonged plo~lla~ion, and (4) death. The highest dosage volumes (MTD, m~ximnm toler-
25 ated dose) which could be ~ elf1 without causing signs of intolerance were recorded.As can be seen from the results in Example SA, the m~ximnm tolerated dose for formlll~tif)nc
in accordance with the present invention is twice that of the formulation which used
polyethoxylated castor oil alone, without sorbitan monoester.
Similar results are obtained when the same formulations are ~ c;d in small
30 volumes/doses at 6 hour intervals for 5 days. Again, the m~ximllm tolerated cllmlll~ive dose
of formn1~tif)ns in accordance with the present h~ ioll is found to be twice that of the form-
ulation using polyethoxylated castor oil alone. These results show that the vehicle of the pres-
ent invention has lower inherent toxicity than when polyethoxylated castor oil is used without
sorbitan monoester, allowing greater f~ ntitif ~ of taxane to be ~flmini~t.ored, or alLelllaLiv~ly,
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the same amount of taxane as used before, but with reduced toxic side effects. The invention
therefore provides a signific~nt advantage over prior taxane formulations in which deleterious
side effects of the vehicle itself have limited the amount of taxane which could be ~ ed~
s
III. Tlt:d~ c,-l Method
In another aspect, the invention includes a method of treating a cancer condition in a
m:~mm~ n subject. In the method, there is provided a taxane storage solution in accordance
with the description above. The storage solution is diluted with a diluent suitable for intrave-
10 nous ~.l"~ .dlion, to produce a dilute taxane solution. The solution is then ~imini~frred tothe subject in a ph~ rel-tir~lly ~rcept~ble amount effective to inhibit cancer growth in the
subject.
The dilute taxane solution is ~ eled to treat any cancer condition in which the
taxane is effective to inhibit or destroy cancer growth. Such cancer conditions may include
15 ovarian cancer, breast cancer, bladder cancer, lung cancer, me!~nnm~ and lymphoma, for
example.
The diluent used in the method is any intravenous solution suitable for intravenous
.dLion. Typically, the diluent will include sodium chloride to establish a selected
physiological o~m~ ty7 e.g., 0.9% (w/v) sodium chloride). The diluent may additionally
20 include suitable supplements, such as glucose, and/or an antimicrobial agent such as penicillin
or tetracycline. The solution is preferably disy~l~.ed using a non-plasticized cullLdhler~ to
prevent le~rhing of pl~et;ci7.ors into the solution. The diluted taxane formulation is admin-
istered at a selected rate until the desired amount of drug has been ~rlmini.ctered. The
formulation is ~ ed periodically until remission has been achieved, or until it appears
25 that proliferation of the target cancer is inhibited. The formulation may also be ~flmini~tered
following surgery to inhibit l~;ull~nce of the cancer, for a time sllfflcient to indicate that the
cancer has been succes~fully removed.
Dosage ~e ,hllells for treating cancer patients with taxol and taxol derivatives are ~nown
in the art and are described, for example, in Arbuck and Blaylock (1995), which is
30 incorporated herein by lerel~llce.
It will be appreciated that use of the storage solution of the invention may be made in
colllbilldlion with any other anti-cancer regimen deemed ~lu~liate for the patient. For
example, the storage solution of the invention may be used in colllbilldlion with cisplatin,
edatrexate, L-buthionine sulfoxide, tiazofurin, gallium nitrate, doxorubicin, etoposide, or cyclo-
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phosphamide, for exarnple, or may be used in combination with radiation therapy. Further,
while the preceding discussion describes the advantages of the vehicle of the invention in terrns
of utility with taxol, the invention cont~-~plates use of the vehicle with other non-polar
taxol/taxane derivatives, such as docet~x~l, whether of synthetic or natural origin.
The following examples illustrate but are not int--n~lecl in any way to limit the invention.
Example 1
Taxol Forrmll~finns
For the studies described below, two formulations, were prepared in the following
10 proportions.
Formulation 1Formulation 2
PEG 300 20 rnL 25 rnL
Absolute Ethanol 10 mL 10 mL
A~ ydl~nls citric acid 100 mg 100 mg
"CREMOPHOR EL" 15 mL 10 mL
"TWEEN 80" 5 rnL 5 mL
Taxol 300 mg 300 mg
Final Volume: 50 rnL 50 rnL
To prepare the above formulations, the PEG 300, citric acid and ethanol (EtOH) were
rnixed with a high speed mixer or stir bar until the citric acid was completely dissolved. If
n~cf-Ssz~ry, the mixture was heated to 50~C or sonic~tP(l to complete dissolution. To the
25 mixture was then added "CREMOPHOR EL" and "TWEEN 80", and the resultant mixture was
stirred for 30 minutes with a high speed mixer. The ta%ol was then added, and mixing was
continued until the taxol was completely dissolved. The resulting solution was purged with dry
nitrogen and filtered through a 0.22 micron filter ("MILL~PACK" 200). In both formulations,
the final concellLlalion of taxol was 6 mg/rnL.
Exarnple 2
Te111PelaLUl~ Stabilitv
Samples of Forrnulations 1 and 2 (200 ,~bL each) were placed in 2 mL arnber vials,
which were then purged with nitrogen, stoppered using Teflon-coated rubber stoppers and
35 sealed with ~IIlminllm seals. The vials were placed in an autoclave and heated under pressure
at 250~F for 20 minutes. The sarnples were then diluted with HPLC-grade z~etonhrile (1:20)
and analyzed by HPLC on a Waters C8 Novapak colurnn (8 mrn I.D. x 10 cm, buffer A =
20% acetonitrile in water, 0.1% trifluoroacetic acid; buffer B = 80% ~cetonitri~e in water,
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0.1 % trifluoroacetic acid; isocratic gradient at 45 % B; detection at 230 nm). HPLC analysis
showed no sign of degradation of the taxol.
Example 3
S Co~ Li~e Lon~ Term Stability of Pormulations
Samples (200 ~L each) of Pormulations 1 and 2, and a formulation cont~ining taxol
(6 mg/mL) in a 1: 1 mixture of "CREMOPHOR EL" and ethanol (Formulation 3), were
placed in 2 mL amber vials which were then purged with nitrogen, sealed, and placed in a
heat chamber at 37~C. Samples (50 ,uL) were withdrawn at 1, 3, 6 and 12 weeks, diluted
10 with HPLC-grade acetonitrile (1:20) and analyzed by HPLC. The results were as follows:
Formulation,
Time (wks~ rc~.;e~ ge Taxol ~r.~ g
#1 #2 #3
100 100 100
6 98.8 98.8 98.7
12 98.7 98.8 97.8
Example 4
Stability of Taxol Formulations
Stock solutions in accol-lance with Fonn~ ti~ns 1 and 2 were diluted 1:5, 1:10,
1:25 and 1:50 in normal saline (0.9% NaCl in water) to give taxol concentrations of 1.2,
0.6, 0.24, and 0.12 mg/mL, respectively. The solutions were checked at 1~ 2, 4, 8, 24,
and 48 hours for signs of plc~ dLion or cloudiness.
All dilute solutions of Formulation 1 rPm~inPd clear after 48 hours, showing no
signs of cloudiness or ~ ildLion. All dilute solutions of Formulation 2 were clear after
24 hours, but all showed some pl~cil~iLaLion after 48 hours, with the 1:5 dilution of Formu-
lation 2 showing the most pleei~iLdlion.
ExamDle S
Col.l~ dLi~e Toxicities of Taxol Formulations
A. Toxicitv of Un~ihlted Samples. Samples of taxol Formulations 1, 2 and 3 were
tested in nn~lilllte-l form for acute toxicity in Balb/C mice. The samples were ~ d
intravenously, over a range of ~ ed volumes, to groups of 2 or 3 mice weighing 18-
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20 grams. The mice were then monitored for signs of intolerance for 21 days after
Lion. Signs of intolerance included any one of the following: (I) signific~n~
weight loss (>20%), (2) piloerection, (3) prolonged p~ ,LldLion, and (4) death. The results
are tabulated below, where MTD is the m~imnm tolerated dose expressed in units of
5 mL/kg.
FormulationMTD (mL/k~)Number of Mice
#1 5.0 2
#2 5.0 3
#3 < 2.5 3
B. Toxicitv Followin~ Lon~ Term A.llllil.i!~l.alion. Samples of Formulations 1, 2
and 3 were diluted 1:1 in normal saline and ~lmini~tered intravenously to Balb/C mice (18-
15 20 grams in weight), 4 times a day for 5 days. The mice were monitored for signs of
intolerance from the time ~lminictration was started until 21 days after ~l."i"i~ ion had
ceased. The m:~imnm tolerated c~lmlll~ive doses are tabulated below:
FormulationMTD. mL/k~Number of mice
#1 lO 5
#2 lO 5
#3 5.0 5
While the invention has been described with reference to specific methods and
embodirnents, it will be appreciated that various modifications may be made without
departing from the invention.
