Background of tha Invention
U.S. Patent o . 7 ,989, 473 and 8 , 178, 127 disclose stable
preparations of N-ethyl-N-phenyl-1 ,2-dihydro-4-hydroxy-S-chloro-leth
l-2-oxoquinoline-3-carboxamide (CAS Number 248281-8 4-7) , also
known as laquinimod (laq.j . Laquinimod has been shown in U.S.
Patent No. 6,077,851 to be effective in the acute experimental
autoimmune encephalomyelitis (aEAE) model. U.S. Patent No.
6,077,851 discloses the synthesis o f laquinimod and the preparation
o f its sodium salt. U.S. Patent No. 6,875,869 discloses an
additional synthesis process o f laquinimod.
PCT International Application Publication No. SO 2005/074899
discloses pharmaceutical compositions comprising laquinimod sodium.
U.S. Patent No. 7,589,208 discloses the aqueous solubility of Li,
Na, Ca, Cu, Zn, Fe and n salts of laquinimod and the
experimental preparation of Na, Ca, Fe(III), Li and Zn salts of
laquinimod.
The subject invention provides a Laquinimod amine salt, which is
laquinimod meglumine, laquinimod choline hydroxide, laquinimod Llysine
or laquinimod monoethanolamine .
Brief Description of the Figures
Fig, 1 : XRD di fractogram for amorphous LAQ Meglumine salt of
Laquinimod batch 1 with spin.
Fig. 2 : XRD di ractogram for amorphous LAQ Meglumine salt of
Laquinimod batch 3 with spin.
Fig. 3 : XRD d if ractogram for amorphous LAQ Meglumine salt of
Laquinimod batch 4 with spin.
Fig. A : DSC thermogram scan o f Meglumine salt o f Laquinimod (batch
3), 4.2640mg, Method: 2S-250C OC/ in 40ml/min 2 (QC-Tech-cr)
25. 0-250. 0°C 100°C/min N2 40.0ml/min.
Fig. 4B : DSC thermogram scan of Meglumine salt o f Laquinimod (batch
1), 2.6140mg, Method: 25-300C lOC/min 2 (QC-Tech) 25. 0-300. 0°C
100°C/iriin N2 40.0ml/min, Peaks: (1) Integral -20.33 m Normalized -
7.78 Jg "' Peak 60.46°C Left limit 48.5S°C Right limit 80.96°C, (2)
Integral -46.07mJ Normalized -17.62Jg Peak 152.84°C Lef limit
147.84°C Right limit 158.48°C.
Fig. 4C : DSC thermogram scan o f Meglumine salt o f Laquinimod (batch
A), 4.1230mg, Method: 25-300C lOC/min 40ml/min N2 (QC-tech) 25.0-
300. 0°C 10.00°C/min 2 40.0ml/min, Peak: Integral -203.80mJ
Normalized -49.3 / Jg~ Peak 61.95°C Left limit 42.91'C Right limit
109.95°C.
Fig . 5A : Solid state C-NMR spectrum for amorphous Meglumine salt
o f Laquinimod in the 0-180 ppm range (batch 1 ) 1 .3ppm/l .IHz,
Peaks: 171.6212, 161.5944, 142.4736, 128.2495, 119.3887, 113.5591,
109.3619, 72.0529, 54.3312, 44.3044, 34.5108, 29.8472, 13.5245.
Fig . 5B : Solid state C- M R spectrum for amorphous Meglumine salt
o f Laquinimod in the 100-180 ppm range (batch 1),
100.93ppm/12693.06Hz, Peaks: 171.6212, 168.4172, 161.5944, 142.4736,
128.2495, 119.3887, 113.5591, 109.3619.
Fig. 6A : DSC thermogram scan of Choline salt o f Laquinimod (batch
5), 3.1820mg, Method: 25-300C lOC/min 40ml/min N2 (QC-tech) 25.0-
300. °C 10.00°C/min N2 40.0ml/min, Peak: Integral -204.59mJ
Normalized -64,30Jg Peak 196.50°C Left limit 183.85°C Right limit
201 .00°C.
Fig. 6B : DSC thermogram scan of Choline salt of Laquinimod (batch
6), 3.4960mg, Method: 25-350C lOC/min 40ml/min 2 (QC-tech-cr)
25. 0-350. 0°C 10.00°C/min 2 40.0ml/min, Peaks: (1) Integral -2.36mJ
Normalized - .67Jg * Peak 123 .62 °C Left limit 103 .92 °C Right limit
133 . 1 °C, (2) Integral -885.2 m Normalized -253 .22Jg Peak
188.58°C Left limit 134.97°C Right limit 242.75°C, (3) Integral -
468.82mJ Normalized -134.10Jg Peak 258.48 'C Left limit 242.77°C
Right limit 336. 61°C.
Fig . 6C : DSC thermogram scan of Choline salt o f Laquinimod (batch
6 ) , 6.0640mg, Method: 25-350C lOC/min 40ml/min 2 (QC-tech-cr)
25. 0-350. 0°C 10.00°C/min N2 40.0ml/min, Peaks: (1) Integral -
584 .87mJ Normalized -96,45Jg Peak 134 .13°C Left limit 3 1 .32°C
Right limit 160.71°C, (2) Integral -253.79mJ Normalized -41.85Jg _
Peak 185.53°C Left limit 160.88°C Right limit 191 .78°C, (3)
Integral -644 .96mJ Normalized -106 .36Jg Peak 216 .28°C Left limit
191.78°C Right limit 236.59°C, (4) Integral -105.19mJ Normalized -
17.35Jg Peak 258.56°C Left limit 250.07°c Right limit 275.58°C,
(5) Integral -2 .85m Normalized -0.47Jg Peak 276.58°C Left limit
275.76°C Right limit 286.53°C
Fig . 6D : DSC thermogram scan of Choline salt of Laquinimod (batch
7 , 3.7440mg, Method: 25-350C lOC/min 40ml/min N2 (QC-tech-cr)
25. 0-350. 0°C 10.00°C/min 2 40.0ml/min, Peaks: (1) Integral -
868.62mJ Normalized -232.00Jg" Peak 188.70°C Left limit 140.72°C
Right limit 240.20°C, (2) Integral -381.62mJ Normalized -101.93Jg"
Peak 254.86°C Left limit 240.49°C Right limit 329.23°C.
Fig. 6E : DSC thermogram scan of Choline salt of Laquinimod (batch
8), 4.0420mg, Method: 25-350C lOC/min 40ml/min N2 (QC-tech-cr)
25. 0-350. 0°C 10.00°C/min N2 40 .Oml/min, Peaks: (1) Integral -
104.57mJ Normalized -25.87Jg Peak 156.68°C Left limit 106.47°C
Right limit 185.97°C, (2) Integral -333.47mJ Normalized -82.50Jg
Peak 221.69°C Left limit 167.42°C Right limit 241.49°C, (3)
Integral -244 .65mJ Normalized -80.53Jg Peak 2S5.86°C Left limit
2 43 .32 C Right limit 292 .37°C.
Fig. : X D dif ractogram for crystalline Laquinimod Choline salt
of Laquinimod (batch 5 with spin}, Temp 25. 0°C, Step: .050°C,
Integration Time l.OOOsec, Range: 2.000-40.000° Cont. Scan Rate:
3.000 [Vmin] Vertical Scale Unit: [CPS] Horizontal Scale Unit
[d e l .
Fi . 8 : XP.D d i ractogram for crystalline Laquinimod Choline salt
o f Laquinimod (batch 8 with spin). Temp 25.0°C, Step: -050°C,
Integration Time l.OOOsec, Range: 2.000-40.000° Cont. Scan Rate:
3.000 [°/min] Vertical Scale Unit: [CPS] Horizontal Scale Unit
[deg] .
Fig. 9A : Solid state C- MR spectrum for crystalline Choline salt
o f Laquinimod in the 0-180ppm range (batch 5), Peaks: 169.7495,
168.9855, 160.7157, 143.6476, 142.6524, 131.6536, 130.1416,
127.9094, 127.1402, 122.1125, 118.8875, 114.1395, 109.5352, 68.0921,
57.4369, 53.6739, 44.5251, 28.2553, 13.7011.
Fi . 9 : Solid state C-NMR spectrum for crystalline Choline salt
o f Laquinimod in the 100-180ppm range (batch 5 , Peaks: .169.7495,
168.9855, 160.7157, 143.6476, 142.6524, 131.6536, 130.1416,
127.9094, 127.1402, 122.1125, 118.8875, 114.1395, 109.5352.
Fig. 10A : DSC thermogram scan o f Lysine salt o f Laquinimod (batch
9 ) , 4.0560mg, Method: 25-350C lOC/min 40ml/min 2 (QC-tech-cr)
25. 0-350. 0°C 10.00°C/min N2 40.0ml/min, Peaks: (1) Integral -
645.89mJ Normalize -159.24Jg Peak 66.74°C Left limit 25.84°C
Right limit 147.72°C, (2) Integral 20.84mJ Normalized 5.14Jg Peak
159.80°C Left limit 150.91°C Right limit 161.08°C, (3) Integral -
140.93mJ Normalized -34.75Jg" Peak 166.74°C Left limit 162.41°C
Right limit 161.57°C, (4) Integral -33.38mJ Normalized -8.23Jg
Peak 1.68°0 Left limit 181.71°C Right limit 200.00°C.
Fig. 10B : DSC thermogram scan o f Lysine salt o f Laquinimod (batch
10) , 3.2900mg, Method: 25-350C lOC/min 40ml/min N2 (QC-tech-cr)
25. 0-350. 0°C 10.00°C/min N2 40.0ml/min, Peaks: (1) Integral -
415.98mJ Normalized -126.44Jg ~ Peak 19Q.02°C Left limit 153.59°C
Right limit 201 . 7 C , 2 Integral - .86m J Normalized - 46 .16 g
Peak 224 .32"C Left limit 201.57°C Right limit 255. 37°C.
Fig. IOC : DSC thermogram scan of Lysine salt o f Laquinimod (batch
11), 4.16O0mg, Method: 2S-350C IOC/win 40m / in 2 (QC-tech-cr)
25. 0-350. 0°C 1 .00°C/min 2 40 .Om m in, Peaks: (1) Integral
293 . mJ Normalized -70.4 6Jg Peak 89.26°C Left limit 36 .30 °C
Right limit 136. 8 C , (2) Integral 159.89mJ Normalized 38.43Jg
Peak 166.08°C Left limit 145.39°C Right limit 171 .19°C, (3)
Integral -294.74mJ Normalized -70.85Jg Peak 237.62°C Left limit
177.87°C Right limit 265.80°C.
Fig. 11 : XRD dif fractogram for crystalline L-Lysine salt of
Laquinimod (batch 10 with spin). Temp 25 .0°C, Step: .050 °C,
Integration Time l.OOOsec, Range: 2.000-40.000° Cont. Scan Rate:
3.000 [°/min] Vertical Scale Unit: [CPS] Horizontal Scale Unit
[deg] .
Fig. 12A : DSC thermogram scan of Monoethanolamine salt of
Laquinimod (batch 14) 3.9820mg, Method: 25-350C lOC/min 40ml/min N2
(QC-tech-cr) 25. 0-350. 0°C 10.00°C/min N2 40. Oml/min, Peaks: (1)
Integral -37 6.96 Normalized -94.67Jg ' Peak 94.34°C Left limit
61 .55°C Right limit 115. 82°C, (2) Integral -292 .55mJ Normalized -
73 .4 g" Peak 146.21°C Left limit il7.27"C Right limit 170.96°C.
Fig. 12B : DSC thermogram scan of Monoethanolamine salt of
Laquinimod (batch 15) , 3.1960mg, Method: 25-350C lOC/min 40ml/min
N2 (QC-tech-cr) 25. 0-350. 0°C 10.00°C/min 2 40. Oml/min, Peaks: (1)
Integral -333.93mJ Normalized -104.48Jg Peak 151.94°C Left limit
129.50°C Right limit 155.29°C, (2) Integral -213.73mJ Normalized -
66.87Jg Peak 160.40°C Left limit 155.29°C Right limit 184.07°C.
Fig. 13 : XRD dif fractogram for crystalline Monoethanolamine salt of
Laquinimod (batch 4), Temp 25.0°C, Step: .050°C, Integration Time
l.OOOsec, Range: 2.000-40.000° Cont. Scan Rate: 3.000 [Vmin]
Vertical Scale Unit: [CPS] Horizontal Scale Unit [deg] . The peak
at 28.5 degrees two theta assign to addition of Silicon.
g g . 1 : XRD dif fractogram for crystalline Monoethanolainine salt of
L inimo (batch 5), Temp 25.0°C, Step: .050°C, Integration Time
l.OOOsec, Range: 2.000-40.000° Cont. Scan Rate: 3.000 [°/min]
Vertical Scale Unit: [CPS] Horizontal Scale Unit [ eg ] . The
shoulder at 28.5 degrees two theta assign to addition of Silicon.
Detailed Description of the Invention
The subject invention provides a Laquinimod amine salt, which is
laquinimod meglumine, laquinimod choline hydroxide, laquinimod -
lysine or laquinimod monoethanolamine .
In one embodiment, the laquinimod amine salt is laquinimod
meglumine. In another embodiment, the laquinimod meglumine is
isolated. In another embodiment, the laquinimod meglumine is
characterized by a DSC thermogram as shown in figure 4A, B and C .
In another embodiment the laquinimod meglumine is characterized
by a solid-state C NMR spectrum with broad peak at 60-77, broad
peak at 122-134, peak at 142.2 and 171.3 ± 0.2 ppm. In yet
another embodiment the laquinimod meglumine is characterized by a
solid state C NMR as shown in figures 5A and 5B.
In one embodiment, the laquinimod amine salt is laquinimod choline
hydroxide. In another embodiment, the laquinimod choline hydroxide
is isolated. In another embodiment, the laquinimod choline
hydroxide is characterized by a DSC thermogram as shown in figures
6 , B , C , D and E . In another embodiment, the laquinimod choline
hydroxide is characterized by a powder XRD pattern with
characteristic peaks at 10.1°, 11.8°, 13.4°, 14.4° and 16. ° 2-
theta + 0.2. In another embodiment, the laquinimod choline
hydroxide is characterized by a powder XRD pattern with
characteristic peaks at 19.3°, 21.2°, 22.7°, 24.8° and 27.6° 2-
theta ± 0.2. In another embodiment, the laquinimod choline
hydroxide is characterized by a powder XRD pattern as shown in
figures 7 and 8 . In another embodiment, the laquinimod choline
hydroxide is characterized by a solid-state C NMR spectrum with
peaks at about 122.1, 127.2, 142.7 and 169.7 ± 0.2 ppm. In another
embodiment, the laquinimod choline hydroxide is characterized by a
solid-state JC NMR spectrum having chemical shifts differences
between the signal exhibiting the lowest chemical shift and another
in the chemical shift range of 100 to 180 ppm of about 12.6, 17.7,
33.2 60.2 and + 0.1 ppm. In another embodiment, the laquinimod
choline hydroxide is characterized by a solid state C NMR as
shown in figures 9A and B . In another embodiment, the laquinimod
choline hydroxide s in crystalline form. In yet another embodiment,
the laquinimod choline hydroxide is in amorphous form.
In one embodiment, the laquinimod amine salt is laquinimod L-lysine .
In another embodiment, the laquinimod L-lysine salt is isolated. In
another embodiment, the laquinimod L-lysine is characterized the
DSC thermogram as shown in figures 0A , B and C . In another
embodiment, the laquinimod L-lysine is characterized by a powder
XRD pattern with characteristic peaks at 5.6°, 9.0°, 11.7°, 13.0°
and 15.9° 2-theta ± 0.2. In another embodiment, the laquinimod Llysine
is characterized by a powder XRD pattern with characteristic
peaks at 17.9°, 18.9°, 21.1°, 22.5° and 23.0° degrees 2-theta ± 0.2.
In another embodiment, the laquinimod L-lysine is characterized by
a powder XRD pattern as shown in figure 11. In another embodiment,
the laquinimod L-lysine is in crystalline form. In yet another
embodiment, the laquinimod L-lysine is in amorphous form.
In one embodiment, the laquinimod amine salt is laquinimod
monoethanolamine . In another embodiment, the laquinimod
monoethanol amine is isolated. In another embodiment, the laquinimod
monoethanolamine is characterized by a DSC thermogram as shown in
figure 12A. In another embodiment, the laquinimod monoethanolamine
is characterized by a powder XRD pattern with characteristic peaks
at 6.5°, 14.4°, 17.9°, 18.7° and 20.6° 2-theta ± 0.2. In another
embodiment, the laquinimod monoethanolamine is characterized by a
powder XRD pattern with characteristic peaks at 17.1°, 19.4°, 22.3°,
23.3° and 24.8° degrees 2-theta ± 0.2. In another embodiment, the
laquinimod monoethanolamine is characterized by a powder XRD
pattern as shown in figure 13. In another embodiment, the
laquinimod monoethanolamine is characterized by a DSC thermogram as
shown in figure 12B. In another embodiment, the laquinimod
monoethanolamine is characterized by a powder XRD pattern with
characteristic peaks at 8.2°, 9.8°, 11.2°, 13.2° and 17.9° 2-theta
± 0.2. In another embodiment, the laquinimod monoethanolamine is
characterized by a powder XRD pattern with characteristic peaks at
18.6°, 20. °, 22.9°, 24.3° and 26.2° 2-theta + 0.2. In another
embodiment, the laquinimod monoethanolamine is characterized by a
powder XRD pattern as shown in figure 14. In another embodiment,
the laquinimod monoethanolamine is in crystalline form. In yet
another embodiment, the laquinimod monoethanolamine is in amorphous
form.
In one embodiment, the subject invention provides a pharmaceutical
composition comprising a laquinimod amine salt and at least one
pharmaceutical acceptable excipient. In another embodiment, the
pharmaceutical composition further comprising laquinimod acid. In
another embodiment, the pharmaceutical composition in which
laquinimod acid is present in an amount of less than 1.5% based on
the total laquinimod content of the pharmaceutical composition. In
another embodiment, the pharmaceutical composition is free of
laquinimod acid. In another embodiment, the pharmaceutical
composition further comprising the sodium salt o f laquinimod. In
yet another embodiment, the pharmaceutical composition is free of
the sodium salt of laquinimod.
In one embodiment, the subject invention provides a process for
manufacture of laquinimod amine salt comprising: a ) combining a
solution of amine with laquinimod acid to form a first mixture; b )
adding a solvent to the first mixture to form a second mixture; c )
removing liquid from the second mixture; and d ) recovering the
laquinimod amine. In another embodiment, said amine is selected
from the group consisting o f meglumine, choline hydroxide, L-lysine
and monoethanolamine. In yet another embodiment, the solvent added
in step b ) is selected from the group consisting of acetone,
methanol, ethanol and dioxane or combination thereof, and in step c
the liquid is removed at ambient temperature and at reduced
pressure .
In one embodiment, the subject invention provides a process for
manufacture of the pharmaceutical composition comprising: a )
obtaining laquinimod amine salt; and b ) admixing the laquinimod
amine salt with at least one pharmaceutical acceptable excipient.
In one embodiment, the subject invention provides a method for
treating a subject afflicted with a form o f multiple sclerosis or
clinical isolated syndrome comprising administering to the
subject a laquinimod amine salt pharmaceutical composition s o as
to thereby treat the subject.
In one embodiment, the subject invention provides a method for
alleviating a symptom o f multiple sclerosis in a subject
afflicted with a form o f multiple sclerosis comprising
administering to the subject a laquinimod amine salt
pharmaceutical composition so as to thereby alleviate the symptom
o f multiple sclerosis in the subject.
For the foregoing embodiments, each embodiment disclosed herein
is contemplated as being applicable to each of the other
disclosed embodiments.
Laquinimod can be administered in admixture with suitable
pharmaceutical diluents, extenders, excipients, or carriers
(collectively referred to herein as pharmaceutically acceptable
carriers) suitably selected with respect to the intended form of
administration and as consistent with conventional pharmaceutical
practices. The unit will be in a form suitable for oral
administration. Laquinimod can be administered alone but is
generally mixed with a pharmaceutically acceptable carrier, and
co-administered n the for o f a tablet or capsule, liposome, or
as an agglomerated powder. Examples of suitable solid carriers
include lactose, sucrose, gelatin and agar.
General techniques and compositions for making dosage forms
useful in the present invention are described-in the following
references: 7 Modern Pharmaceutics, Chapters 9 and 10 (Banker s
Rhodes, Editors, 1979) ; Pharmaceutical Dosage Forms: Tablets
(Lieberman et al., 1981) ; Ansel, Introduction to Pharmaceutical
Dosage Forms 2nd Edition (1976) ; Remington's Pharmaceutical
Sciences, 17th ed. (Mack Publishing Company, Easton, Pa., 1985);
Advances in Pharmaceutical Sciences (David Ganderton, Trevor
Jones, Eds., 1992) ; Advances in Pharmaceutical Sciences Vol 7 .
(David Ganderton, Trevor Jones, James McGinity, Eds., 1995);
Aqueous Polymeric Coatings for Pharmaceutical Dosage Forms (Drugs
and the Pharmaceutical Sciences, Series 36 (James McGinity, Ed.,
1989) ; Pharmaceutical Particulate Carriers: Therapeutic
Applications: Drugs and the Pharmaceutical Sciences, Vol 61
(Alain Rolland, Ed,, 1993) ; Drug Delivery to the Gastrointestinal
Tract (Ellis Horwood Books in the Biological Sciences. Series in
Pharmaceutical Technology; J . G . Hardy, S . S . Davis, Clive G .
Wilson, Eds.); Modern Pharmaceutics Drugs and the Pharmaceutical
Sciences, Vol. 40 (Gilbert S . Banker, Christopher T . Rhodes,
Eds.) . These references in their entireties are hereby
incorporated by reference into this application.
Te r s/Definitions
As used herein, unless stated otherwise, each of the following
terms shall have the definition set forth below.
"About" in the context of a numerical value or range means ±10%
of the numerical value or range recited or claimed unless the
standard error of the analytical measure used to obtain the
numerical value results in a greater deviation.
A "salt" is salt of the instant compounds which have been
modified by making acid or base salts of the compounds. The term
"pharmaceutically acceptable salt" in this respect, refers to the
relatively non-toxic, inorganic and organic acid or base addition
salts of compounds of the present invention.
As used herein, "pharmaceutically acceptable carrier" refers to a
carrier or excipient that is suitable for use with humans and/or
animals without undue adverse side effects (such as toxicity,
irritation, and allergic response) commensurate with a reasonable
benefit/risk ratio. It can be a pharmaceutically acceptable
solvent, suspending agent or vehicle, for delivering the instant
compounds to the subject.
By any range disclosed herein, it is meant that all hundredth,
tenth and integer unit amounts within the range are specifically
disclosed a s part of the invention. Thus, for example, 0.01 mg to
50 mg means that 0.02, 0.03 ... 0.09; 0.1, 0.2 ... 0.9; and 1 ,
2 ... 49 g u it amounts are included as embodiments of this
invention .
A characteristic of a compound refers to any quality that a
compound exhibits, e.g., peaks or retention times, as determined
by "* H nuclear magnetic spectroscopy, 'C nuclear magnetic
spectroscopy, mass spectroscopy, infrared, ultraviolet or
fluorescence spectrophotometry, gas chromatography, thin layer
chromatography, high performance liquid chromatography (HPLC) ,
elemental analysis, Ames test, dissolution, stability and any
other quality that can be determined by an analytical method.
Once the characteristics o f a compound are known, the information
can be used to, for example, screen or test for the presence of
the compound in a sample. Quantity or weight percentage of a
compound present in a sample can be determined by a suitable
apparatus, for example, a HPLC.
As used herein, an "isolated" compound is a compound isolated
from the crude reaction mixture following an affirmative act of
isolation. The act o f isolation necessarily involves separating
the compound from the other known components of the crude
reaction mixture, with some impurities, unknown side products and
residual amounts of the other known components of the crude
reaction mixture permitted to remain. Purification is an example
of an affirmative act of isolation.
A s used herein, a composition that is "free" o f a chemical entity
means that the composition contains, if at all, an amount of the
chemical entity which cannot be avoided following an affirmative
act intended to purify the composition by separating the chemical
entity from the composition. A composition which is "free" of a
laquinimod of a salt thereof, if present, as used herein, means
that the laquinimod or a salt thereof is a minority component
relative to the amount o f 5-HLAQ, by weight.
A s used herein, "stability testing" refers to tests conducted at
specific time intervals and various environmental conditions
(e.g., temperature and humidity) to see if and to what extent a
drug product degrades over its designated shelf life time. The
specific conditions and time of the tests are such that they
accelerate the conditions the drug product is expected to
encounter over its shelf life. For example, detailed requirements
of stability testing for finished pharmaceuticals are codified in
21 C.F.R §211.166, the entire content of which is hereby
incorporated by reference.
This invention relates to four amine salts of laquinimod which
were surprisingly found to significantly increase solubility over
known laquinimod salts and to be stable, and pharmaceutically
processable. Solubility, specifically aqueous solubility, is a
very important characteristic for pharmaceutical preparations, as
solubility increases bioavailability. However, solubility alone
is insufficient, as stability and pharmaceutical processability
also impact the acceptability of a salt form of a drug. Often
highly soluble salts are very hygroscopic and deliquescent under
atmospheric air. Processing and storage of such deliquescent
material is possible only in inert gas or by using dry rooms. The
major problem with certain highly soluble materials is analysis.
Fast water uptake by the salt can result in variable assay
results. Thus certain highly hygroscopic forms are unacceptable.
Free laquinimod is a weak acid (pKa- . ) and is almost insoluble
in water, having an intrinsic solubility of about 0.007 mg/ml.
Previously, sodium salt was the most soluble and stable known
salt of laquinimod prepared, with an aqueous solubility of 138
mg/ml (US Patent No. 7,589,208) .
Amine salts of certain active compounds have been shown to
increase solubility, as compared to the corresponding sodium salt.
However, there are many examples of compounds in the prior art
for which meglumine, choline hydroxide, L-lysine and
monoethanolamine salts did not substantially increase solubility.
S lu i l _t
Meglumine
In certain contexts meglumine salt has increased solubility a s
compared to the corresponding sodium salt. Meglumine salt o f
ibuprofen increased solubility a s compared to ibuprofen sodium
s fr , 100 mg/ml (Sigma Aldrich Website) to 1290 mg/ml (U.S.
Patent No. 5 ,028, 625) .
However, there are many examples o f when meglumine salt caused
only a slight increase in solubility a s compared t o the
corresponding sodium salt. U.S. Patent No. 7,105,512 discloses a
meglumine salt o f meloxicam having a n aqueous solubility o f 2.3
mg/ml a t p H 8.57, a s compared to 2.0 mg/ml for the sodium salt a t
p H 8.06. U.S. Patent No. 7,105,512 also discloses that, "the
meglumine salt o f meloxicam suffers from such problems that it
has considerably low solubility in water and that i f the amount
thereof dissolved in water is increased, the p H value o f the
resulting solution abruptly increases to such a n extent that the
p H value i s beyond the preferred range o f from 5 t o 9." (col. 2 ,
Ins. 18-23) . Similar data was disclosed in U.S. Patent No.
6,869,948. Triclosan meglumine salt solubility was 2 .43 mg/ml,
a s compared to 5.95 mg/ml for the sodium salt (Grove, 2003) . In
comparison, the meglumine laquinimod sa t solubility was 1050 o r
1330 mg/ml, depending o n preparation method and form (Table 6),
a s compared to 138 mg/ml for laquinimod sodium.
Meglumine salt has also in certain contexts increased solubility
a s compared t o the corresponding free compound. The solubilities
o f meglumine salts o f three amino nicotinic acid derivative
compounds were 2 .81, .478 and .210 mg/ml a s compared to the
corresponding free amino nicotinic acid derivative
compounds, .004, .008 and .002 mg/ml respectively (PCT
International Application Publication No. 2010/102826) . Solubility
o f meglumine salt o f ibuprofen was 1290 mg/ml a s compared to free
ibuprofen, .06 mg/ml (U.S. Patent No. 5,028,625) . However, these
solubility increases are all a t least a n order o f magnitude less
than the solubility increase achieved by the meglumine salt of
laquinimod which increased solubility from .007 i g /m l to 1050 or
1330 g/m , depending on preparation method and form (Table 6 .
Choline Hydroxide
In certain contexts choline hydroxide salt has increased
solubility as compared to the corresponding sodium salt. Choline
salt of nimesulide provided high solubility and at the same time
moderate solution alkalinity as compared to nimesulide sodium
salt (European Patent Application No. 0 691 17A1 ).
However, there are many examples of when choline salt did not
increase solubility or caused only a slight increase in
solubility as compared to the corresponding sodium salt. U.S.
Patent No. 7,572,789 discloses a sodium salt of diazoxide with a
solubility of 18.1 mg/ml at pH 7 . Choline diazoxide salt
solubility at the same pH was 41.5 mg/ml, only 2.3 times greater.
Dicholine salt was found to have only twice the aqueous
solubility of the disodium salt for the anti-cancer drug SNS-314
(Muller, 2009) . U.S. Patent No. 6,638,537 discloses choline
salicylate salt as an insoluble compound. In comparison, the
solubility of choline hydroxide salt of laquinimod was over 14
times greater than the sodium salt, 2000 or 2100 mg/ml depending
on preparation and form, which is considered unlimited solubility
(Table 6 ) .
Choline salt has also in certain contexts increased solubility as
compared to the corresponding free compound. The solubility of
nimesulide choline was 500 mg/ml as compared to free
nimesulide, .01 mg/ml (European Patent Application No. 0869117A1) .
The solubility of diclofenac choline was 250 mg/ml as compared to
free diclofenac, .001 mg/ml. However, these solubility increases
are all at least an order of magnitude less than the solubility
increase achieved by the choline salt of laquinimod which
increased solubility from .007 mg/ml to 2000 or 2100 mg/ml,
depending on preparation method and form (Table 6 ) .
L-lysine
Regarding the L-lysine salt, there are many examples of when -
lysine salt did not increase solubility or caused only a slight
increase in solubility as compared to the corresponding sodium
salt. L-lysine salt does not offer significant increases in
solubility for nimesulide or a CD80 antagonist ( - ( -Fluoro -3-
oxo-1 , 3-di hydro -pyrazolo [ ,3-c] cinnol in-2 -y )-N-(2,2-diflurooethyl
)-benzamide . L-lysine of nimesulide was found to be 7.5
mg/ml in water at pH 9.3 and .057 mg/ml at pH 6.8 (Piel, 1997) .
The solubility of the nimesulide-L- lysine salt represented only a
slight improvement over nimesulide free acid form and no
advantage over the corresponding sodium salt, which had a
solubility of less than 10 mg/ml {European Patent Application No.
0869117A1) . The L-lysine salt only provided a significant
increase in solubility when used in a cyclodextrin complex. The
L-lysine salt of the CD80 antagonist disclosed in PCT
International Application Publication No. WO 2007/096588 offered
no increase in solubility as compared to the sodium salt. In
comparison, L-lysine laquinimod salt solubility was 1000 or 1176
mg/ml depending on preparation and form (Table 6 ) , as compared to
138 mg/ml for laquinimod sodium.
L-lysine salt in some contexts increased solubility as compared
to the corresponding free compound. The aqueous solubility of the
nimesulide L-lysine salt discussed above was 7.5 mg/ml as
compared to free nimesulide, .01 mg/ml. The free solubility of
the CD80 antagonist discussed above was <.5 mg/ml as compared to
the L-lysine salt, <5 mg/ml. However, these solubility increases
are all at least two orders of magnitude less than the solubility
increase achieved by the L-lysine salt of laquinimod which
increased solubility from .007 mg/ml to 1000 or 1176 mg/ml,
depending on preparation method and form (Table 6 ) .
Monoethanolamine
Regarding the monoethanolamine salt, there are many examples of
when monoethanolamine salt did not increase solubility or caused
only a slight increase in solubility as compared to the
corresponding sodium salt. Monoethanolamine salt of triclosan was
found to have an aqueous solubility of 5.84 g / . Triclosan
sodium benzoate salt solubility was 5.95 mg/ml, only slightly
higher than the monoethanolamine salt (Grove 2003) . U.S. Patent
No. 4,948,805 disclosed diclofenac monoethanolamine salt as being
"practically insoluble." Sodium salt of Diclofenac was found to
have a solubility of 13.6 mg/ml p H 7.6) . In comparison,
Laquinimod monoethanoloamine salt solubility was 625 or 11 6
mg/ml depending on method o f preparation and form (Table 6 ) , as
compared to 138 mg/ml for laquinimod sodium.
Monoethanolamine salt in some contexts increased solubility as
compared to the corresponding free compound. The solubility of
the monoethanolamine salt of piroxicam was 126.2 mg/ml at pH 7.4
as compared to free piroxicam, .17 at pH 7.4 (Cheong, 2002)•.
Solubility of meloxicam monoethanolamine was 8.36 mg/ml at pH 7.4
as compared to free meloxicam, .74 mg/ml at pH 7.4 (Ki, 2007) .
However, these solubility increases are at least an order of
magnitude less than the solubility increase achieved by the
monoethanolamine salt of laquinimod which increased solubility
from .007 mg/ml to 625 or 1176 mg/ml, depending on preparation
method and form (Table 6 ) .
Stability and Form
It is important for pharmaceutical salts to be capable of being
prepared in a manner that results in a physically stable form.
However, the properties of salts are unpredictable with respect
to both qualitative effect and to magnitude. European Patent
Application No. 0869117A1 discloses that the choline salt of
nimesulide is a highly crystalline solid with a melting point of
133-135°C, but that choline salts of the amino nicotinic acid
derivative appeared as an oil or an amorphous solid. At the same
time, meglumine salt of the amino nicotinic acid derivative
compound is crystalline (PCT International Application Publication
No. 2010/102826) .
Additionally, solubility and hygroscopicity are generally
inversely related to the stability o f a salt. Low stability o f
certain hygroscopic solids is related to the presence of water.
The absorbed water can form a layer of saturated solution above
the surface of the solid particles. If susceptible, the material
dissolved in the solution is exposed to liquid-phase degradation
reactions and hydrolysis and oxidation that usually do not occur
in sold phase (so called solvent-mediated reactions) . Therefore
isolation, processing and storage of highly soluble salts is
generally problematic. The amine salts of laquinimod described
herein, however, have very high aqueous solubility but
demonstrate good physical stability when in contact with
atmospheric air. Crystallini y is also important to stability, as
an increase in amorphous nature can lead to increased
hygroscopicity and thus a possible decrease in stability (Chen,
2009) .
Tables 7A, 7B, 7C and 7D present the stability data for meglumine,
choline hydroxide, L-lysine and monoethanolamine salts,
respectively. All salts prepared in the experiments described
herein demonstrate good physical stability under test conditions
(re erator, atmospheric air) . A t the same time, preparation
method was observed to affect solid stability for 3 of 4 salts
prepared in this study. This is another example of the
unpredictability of salt properties. Furthermore, the meglumine
salt was only be synthesized in amorphous form, which would be
expected to cause stability problems. However, two out of three
samples of the laquinimod meglumine salt unexpectedly showed no
physical change after 10 months of storage. The important
conclusion is that all four salts could be prepared in
processable and stable forms.
All four amine salts of laquinimod have extremely good aqueous
solubility that is much higher than the sodium salt of laquinimod.
The magnitude of these increases was unexpected as compared to
the more modest and unpredictable changes in solubility found for
corresponding salts in the prior art. All four salts can be
prepared in a manner that results in physically stable laquinimod
salts. Choline hydroxide, L-lysine and monoethanol am i e can be
synthesized in both crystalline and amorphous forms. Meglumine
was only be synthesized as an amorphous solid yet unexpectedly
did not have stability issues. This unpredictable balance of
properties is a significant advantage of these Laquinimod amine
salts.
This invention will be better understood by reference to the
Experimental Details which follow, but those skilled in the art
will readily appreciate that the specific experiments detailed
are only illustrative of the invention as described more fully in
the claims which follow thereafter.
Experimental Details
These examples describe the preparation and characterization of
four novel laquinimod amine salts.
Four pharmaceutically acceptable amine bases with pKa >9 were
used. Properties o f the bases are summarized in Table 1 .
Table 1 . Properties of the amine bases
List of Equipment :
X-Ray Diffraction (XRD)
Analysis was performed on ARL (SCINTAG) powder X-Ray
dif Tactometer model X'TRA equipped with a solid state detector.
Copper radiation o f 1.5418 A was used. Scanning parameters: range:
2-40 degrees two-theta; scan mode: continuous scan; step size:
0.05°, and a rate of 3 deg/min.
Solid state Carbon-13 Nuclear Magnetic Resonance Spectroscopy
( C-NMR) The CP/MAS C NMR measurements were made on a Bruker
Avance 500 NMR S/ B spectrometer in 4 -m ZrO; rotor . Magic angle
spinning (MAS) speed was 10 kHz. As used herein, the term " C NMR
chemical shifts" refers to the shifts measured under above
specified conditions. These shifts can slightly differ from
instrument to instrument and can be shifted either upfield or
downfield due to the different instrumental setup and calibration
used. Nevertheless, the sequence o f individual peaks remains
identical .
The Microscope used for morphology was a Nikon Eclipse, ME- 600
equipped with a DeltaPix camera.
Example 1 : Laquinimod Meglumine salt
Meglumine, United States Pharmacopeial Convention (USP) / European
Pharmacopeia (Ph. Eur) grade, manufactured by Merck and Co. Inc,
was used in the experiments described herein. The preparation
methods are summarized in Table 2 below and the chemical
structure of the salt is presented below.
Reaction products appeared as solids and were subjected
analysis and characterized.
Table 2 . Preparation of Meglumine salts of Laquinimod
1.1. Precipit ate meglumine salt from solution: Batch 1
.77g of meglumine was dissolved in 10ml water. 1.38g of
laquinimod acid was added by portions while stirring. The
resulting mixture was heated to 50° C , and then 4ml of water and
0.02g of meglumine were added. The resulting solution (cloudy,
pH=9-10) was cooled to +5°C and held at this temperature for
36hrs. No precipitation was observed.
The solution was evaporated under vacuum in a rotary evaporator
(bath T=30-45°C) . The residue ( 3 . 51g of colorless liquid) was
stirred with magnetic stirrer and dry acetone was added by
portions at ambient temperature.
After addition of 15 m l acetone, intensive precipitation was
observed. The total volume of acetone introduced was 30ml.
The precipitate was a colorless, sticky honey-like material which
was not filterable.
Isolation of meglumine salt by evapora tion :
The mixture was evaporated in a rotary evaporator under vacuum
(bath T=45°C) . The residue, a white solid foam, was broken down
with a spatula and dried at ambient temperature under high vacuum
(2 mbar .
The solid product, 1 .82g of white powder, was sampled.
Analysis and tests ;
Solid State (SS! NMR confirmed salt formation. and ,C NMR
confirmed salt structure along with traces of acetone.
XRD identified amorphous structure.
Differential scanning calorimeter (DSC) identified amorphous
structure .
Physical stability: A sample of powder was exposed to atmospheric
air at relative humidity (RH) = 38% in an open dish for 20 hours.
No signs of aggregation or deliquescence were observed.
Samples of powder were stored in sealed transparent glass vials
at +5°C for 6 months. No physical change was observed. The
samples were white flowable powder.
Aqueous solubility was 1330 mg/ml ¾0.
1.2. Salt precipitation from dry solvent: (Batch 2 )
0.77g of meglumine was added to 20ml of absolute methanol while
stirring. The solution was then heated. Complete solid
dissolution was observed at 37°C and then 1.39g of laquinimod
acid was added and dissolved.
The resulting solution was cooled to 1 °C and 40ml of dry acetone
was added while stirring. Turbidity developed and the mixture was
held in a refrigerator at +5°C overnight.
A tiny amount of precipitate was formed.
The residue was evaporated under vacuum in a rotary evaporator
(bath T~42°C), the residue (3 .52g of colorless syrup) was stirred
and 6.5ml of dry acetone was added at ambient temperature.
Precipitation was observed. Precipitate was colorless, sticky,
honey-like, and not filterable. .
The mixture was evaporated under vacuum in a rotary evaporator
(bath T=42°C) . The warm residue (3 .77g of colorless syrup) was
introduced by drops into 25ml of cold dry acetone at 7-8°C, on an
ice-water bath.
Sticky semi-solid precipitate was formed.
1.3. Salt precipitation by solvent exchange: (Batch 3 )
The mixture from the previous experiment was heated to 40°C and
evaporated under vacuum in a rotary evaporator (bath T=42 - 60°C) ,
the residue (2.10g of semi-solid product) was dissolved in 25ml
of absolute ethanol and evaporated under the same conditions.
The residue of the second evaporation (2.19g of solid foam) was
dissolved in 25ml of absolute ethanol and evaporated under the
same conditions.
The residue of the third evaporation ( .22g) was dissolved in 3ml
of absolute ethanol, and the solution was added to cold 1,4-
dioxane over a period of 45 minutes.
During the addition, the mixture was cooled on an ice-water bath
and stirred vigorously.
No product precipitation was observed. Only a part of the dioxane
was crystallized.
The mixture was evaporated under vacuum in a rotary evaporator at
ambient temperature. After evaporation of half of the solvent,
volume precipitation took place.
Sticky, semi-solid and not filterable precipitate (soft gum)
formed.
Isolation o f sa t by evaporation :
Evaporation was continued while heating (bath T=60°C) . The
residue, 2 .16g o f solid white foam, was dried under high vacuum
{2 rb a at ambient temperature.
The dry product, 1 .9Sg o f white powder was sampled.
Analysis a d tests:
XRD identified amorphous structure.
DSC identified amorphous structure and endotherm (endo} peaks at
48 80 C and 147-162°C.
Physical stability: Samples o f powder were stored in sealed
transparent glass vials at +5°C for 6 months. Physical change was
observed. The sample was lumped white powder, with signs of
deliquescence .
1.4. Iso lation o f salt by lyophilization: (Batch 4 )
0 .77g of meglumine was dissolved in 10ml o f water, .39g o f
laquinimod acid was added by portions while stirring. After
dissolution, the solution was transferred to a polypropylene (PP)
bottle with 2ml o f rinse water.
The solution was frozen at -18 °C and lyophilized at 0.11 mbar
(collector temperature -84 °C) over a period o f 22 hours.
After lyophilization was completed, the fragile solid cake (2.16g)
was broken up with a spatula. The resulting white powder was
sampled.
Analysis and tests:
H and C R confirmed structure.
XRD identified amorphous structure.
DSC identified amorphous structure and endo peaks at 42-109°C and
147-162°C.
Physical stability: Samples of the powder were stored in sealed
transparent glass vials at +5°C for 6 months. No physical change
was observed. The sample was a white powder with no signs o f
deliquescence .
Aqueous solubility was 1050 mq/ml H 0 .
Example 2 : haquinimod Choline salt
Choline hydroxide, 46% w . aqueous solution, supplied by Sigma-
Aldrich, was used in the experiments described herein.
The experiments are summarized in Table 3 below and the chemical
structure is presented below.
Reaction Products appeared as solids and were subjected to
analysis and characterized.
Table 3 . Preparation of Choline salts of Laquinimod
2.1. Salt precipitation by solvent exchange: (Batch 5 )
0.80g of choline hydroxide, 46% t ., was dissolved in 2ml of
water. 1.08g of Laquinimod acid was added by portions while
stirring. 1ml of water was (rinse) added and complete dissolution
was achieved.
20ml o f acetone was added to the resulting solution at ambient
temperature while stirring. No precipitation was observed. The
solution was evaporated under vacuum in a rotary evaporator (bath
T=30-70 °C) to dryness.
The residue (1.46g of colorless glassy solid) was dissolved in
20ml of dry acetone and stirred at ambient temperature. After 5
minutes slow precipitation was observed.
The mixture was held in a refrigerator overnight at +6°C and then
filtered .
Wet solid product was dried in a vacuum oven at 40°C up to
constant mass.
The dry product was 1.07g of white powder with a yield of 73.5%.
Analysis and tests :
and 'C NMR confirmed structure. Acid to base was ratio 1:1,
and no residual solvents were detected.
XRD identified crystalline structure.
DSC identified melting point at 19 °C .
2.2. Salt isolation by lyophilization : (Batch 6 )
.39g Choline hydroxide 46% was dissolved in 5ml o f water. 3 . 4g
of laquinimod acid was added by portions. The solution was
stirred vigorously, at ambient temperature, until dissolution.
The yellowish clear solution was transferred to a PP bottle, was
frozen at -18°C and lyophilized at 0.10-0.12 rnbar (collector
temperature 84°C) over a period of 45 hours.
After lyophilization was completed, the solid cake product ( . 1g )
contained two fractions:
- brittle glassy solid (fraction A )
honey-like semi-solid material (fraction B )
The fractions were sampled separately.
Fraction B was collected and then exposed to atmospheric air over
the period of a week. Physical transformation was observed.
The wet fraction B material (1.79g) was dried under vacuum at
S0°C to constant weight. The result was 1.72g of dry product
(fraction B ) .
Analysis and tests :
Main fraction A
H and C NMR - confirmed structure.
XRD identified amorphous structure.
DSC identified amorphous structure and endo peaks at 42-109"C and
147-162°C.
Physical stability: Samples of the powder were stored in sealed
transparent glass vials at + °C for 6 months. Physical change was
observed. The sample was a yellowish semi-solid with signs of
deliquescence .
Dry fraction B
Appearance was a yellowish solid.
XRD identified crystalline structure.
DSC identified melting and degradation point at 195°C .
2.3. Salt isolation by solvent evaporation (Batch 7 )
2.44g of choline hydroxide 46% was dissolved in 5ml of absolute
methanol and then 3.24g of laquinimod acid was added by portions
while stirring over a 10 min period. After additional stirring
and complete dissolution, the solution was evaporated to dryness
in a rotary evaporator under vacuum, (bath T= 50-70°C) .
During the evaporation, honey-like residuum foamed a t bath T= 60-
70 °C . The foam solidified under vacuum at room temperature.
The residue, .19g of solid foam, was broken up with a spatula
and dried overnight under high vacuum (2mbar) at room temperature.
The dry product was 4.17g of white glassy powder.
Analysis and tests :
XRD identified amorphous structure.
DSC identified amorphous structure, and endo peaks and
degradation at T>185°C.
Physical stability: Samples of the powder were stored in sealed
transparent glass vials at +5°C for 6 months. No physical change
s observed. The sample was white powder with no signs of
deliquescence .
Aqueous solubility was >2100 mg/ml H 0 (unlimited solubility) .
2.4 Salt isolation by precipitation from dioxane-met hanol (Batch
8 )
2 . g o f choline hydroxide 46% was dissolved in 5 m l of absolute
methanol and then 3 . 4g of laquinimod acid was added by portions
while stirring over a period of 10 minutes. After additional
stirring and complete dissolution the solution was evaporated to
dryness in a rotary evaporator under vacuum, (bath T = 50-70°C) .
During the evaporation, the honey-like residuum foamed at bath T=
60-70 °C . The foam solidified under vacuum at room temperature.
The residue was .18g o f solid foam that was broken up with
spatula and dissolved in 3ml of absolute methanol and 50ml of 1 ,
4 dioxane while stirring over a period of 20 inin. During the
addition, precipitation occurred. The mixture was stirred for one
additional hour and filtered using a Buchner filter. The solid
cake was washed with dioxane and dried in a vacuum oven at 50 °C.
Dry product was 3 .31g of white powder, with a yield of 76.3%.
Analysis and tests:
XRD identified crystalline structure.
DSC identified crystalline structure and melting point at 189°C.
Physical stability: Samples of the powder were stored in sealed
transparent glass vials at +5°C for 6 months. No physical change
was observed. The sample was white powder with no signs of
deliquescence .
Aqueous solubility was 2000 mg/ml H20 (unlimited solubility) .
L-Lysine >98 , supplied by Sigma-Aldrich, was used in the
experiments described herein. Materials that appeared as solids
were subjected to analysis and characterized.
The experiments are summarized in Table 4 below and chemical
structure s presented below.
Table 4 . Preparation of L-Lysine salts o f Laquinimod
3.1 Isolation of salt from water-acetone: (Batch 9 )
1.Og of L-Lysine was dissolved in 1.8ml of water and 2.44g of
laquinimod acid was added to the solution while stirring. After
partial dissolution, a very viscous gel-like solution was formed
(pH=7) .
n additional 4,5ml of water and 0 .02g of L-Lysine were added
while stirring, resulting in a clear solution (pH=8) .
50ml of acetone was added to the solution at ambient temperature
while stirring continued. During the addition of acetone
precipitation occurred.
The stirring was stopped and precipitate settled to the bottom.
After 1 hour of settling, the precipitate formed a lower liquid
layer .
The mixture was transferred to an evaporation flask with an
additional 5 l of water (rinse) and evaporated in a rotary
evaporator under vacuum {bath T=45-50°C) . The residue, a
colorless liquid, was dissolved in 25 m l of acetone and
evaporation continued under the same conditions.
The residue, 3.48g of white so lid foam, was broken down with a
spatula and dried at ambient temperature under high vacuum (2
mbar) .
3.44g of the white powder solid product was sampled.
Analysis and tests :
and , JC NMR confirmed structure and identified traces of
acetone.
XRD identified amorphous structure.
DSC identified amorphous structure and degradation at T>140°C.
Physical stability: Samples of the powder were stored in sealed
transparent glass vials at +5°C for 6 months. No physical change
was observed. The sample was white flowable powder.
Aqueous solubility was 1176 mg/ml ¾ 0
3.2 Precipitation o f salt from methanol-acetone: (Batch 10)
l.Og of L-Lysine was dissolved in 30ml of absolute methanol and
2.44g of laquinimod acid was added to the solution while stirring.
About two thirds of the resulting solution was evaporated in a
rotary evaporator under vacuum. The residue, .9g of solution,
was stirred and 15ml of dry acetone was added at ambient
temperature. During the acetone addition solid precipitated. The
mixture was stirred for an additional half hour and then
introduced to a refrigerator (+5°C) for 3 hours.
The solid product was collected by filtration, washed with m l
dry acetone and dried in an oven at 40°C under vacuum.
Dry product was 3. 9g of white powder, with a yield of 92. " %
Analysis and tests:
and C NM confirmed structure and identified traces of
acetone .
XRD identified crystalline structure.
DSC identified crystalline structure and melting point at 190°C .
Physical stability: Samples of the powder were stored in sealed
transparent glass vials at +5°C for 6 months. No physical change
was observed. The sample was white powder.
Aqueous solubility was 1000 mg/ml H 0 .
3.3 Salt isolation by solvent evaporation : (Batch 11)
2 . 4g of laquinimod acid was added to a solution of l.Og of -
Lysine in 10ml of water at ambient temperature while stirring.
After partial dissolution of the solid, an additional 0.03g of LLysine
was added (pH=7-8) and then the mixture was heated to 45°C.
The resulting solution was evaporated in a rotary evaporator
under vacuum (bath T=65°C) .
During the evaporation, a syrup-like residuum foamed. The foam
was solidified under vacuum.
The residue, 3.35g of solid foam, was broken up with a spatula
and dried overnight in an oven under vacuum at 40 °C.
Dry product was 3.34g of off-white solid.
Analysis and tests :
and i C NMR confirmed structure.
XRD identified amorphous structure.
DSC identified amorphous structure and degradation at T>140°C.
Example 4 : Laquinimod Ethanolamine salt
Monoethanolamine (purity 99.7%), supplied by Sigina-Aldrich, was
used in the experiments described herein. Materials that appeared
as solids were subjected to analysis and characterized.
The experiments are summarized in Table 5 below and the chemical
structure is presented below.
Table 5 . Preparation of Ethanolamine salts of Laquinimod
Exp Preparation method Product
Solvent (ratio) LAQ:EA Conditions appearance
(Moles)
4.1 Acetoneiwater 1:1.02 Evaporation from acetone- White powder
(50:6) water
4.2 Water 1: 1 .02 Lyophilized from aqueous White powder
solution
4.3 1,4 dioxane 1:1.02 Precipitation from dioxane White powder
4.4 Acetone 1: 1.02 Precipitation from acetone White powder
4.1 Sa t isolat ion by solvent evaporation: (Batch 12)
0 .61g of ethanolamine and 3 .50g of laquinimod acid were dissolved
in 6ml of water followed by 50 ml of acetone. No precipitation
was observed.
The mixture was evaporated to dryness under vacuum in a rotary
evaporator .
During the evaporation a residuum foamed. The foam was solidified
under vacuum.
The residue, a solid glassy foam, was broken up with a spatula
and dried overnight in oven under vacuum at 0°C .
Dry product was 2 . 2g of white powder.
Analysis and tests:
H and 1'C NMR confirmed structure and identified traces of
acetone
XRD identified amorphous structure.
DSC identified amorphous structure, endo peaks at 40-100°C, and
decomposition at 140-170°C.
Physical stability:
• At room temperature:
A sample of white flowable powder was exposed to atmospheric air
in air-conditioned room at RH=25-30%, T=18-20°C.
After 4 hours there was no change.
After 2 4 hours there was yellowish aggregated powder.
• In refrigerator:
A sample of white flowable powder was stored in sealed
transparent glass vials at +5°C for 3 months. Physical change was
observed. The sample was a yellowish aggregated powder.
4.2 (Batch 13)
0.61g of ethanolamine and 3 . 0g of laquinimod acid were dissolved
in 10ml of water while stirring at ambient temperature. The
solution was transferred to a PP container and lyophilized at
0.014-0.010 mbar (collector temperature -79 to -81 °C) over a
period of 72 hours.
The product, 3.94g of solid white glassy cake, was broken up with
a spatula and the white powder was sampled.
Analysis and tests :
Ή and C NMR confirmed structure.
XRD identified amorphous structure.
DSC identified amorphous structure, endo at 40-100°C, and
decomposition at 140-170°C.
Physical stability: Sample of the powder was stored in sealed
transparent glass vials at +5 C for 3 months. The sample was an
off-white powder.
Aqueous solubility was 1176 mg/ H 0 .
4.3 Precipitation of salt from dioxane: (Batch 14)
0 . 1g o f ethanolamine was dissolved in 25ml of dry methanol, then
3.50g of laquinimod acid was added and dissolved while stirring.
The solution was evaporated under vacuum in a rotary evaporator
(bath T=65°C) and the obtained residue (4.38g of semi-solid
material) was dissolved in 10ml of warm 1,4 dioxane.
White solid precipitate was formed. An additional 15ml of dioxane
was introduced and the resulting slurry was stirred at ambient
temperature for 2 hours.
Solid product was collected by filtration, washed with 5ml of
dioxane and dried under vacuum in an oven at 40°C.
Dry product was .O g of white solid, ground to white flowable
powder .
Analysis and tests :
and C MR - Confirmed structure, and identified 1 mole of
dioxane solvate.
XRD identified crystalline structure and form I (solvate) .
DSC identified crystalline structure, melting point at 80-90°C,
and decomposition at T>120 C .
Physical stability:
• At room temperature:
A sample of white flowable powder was exposed to atmospheric air
in an air-conditioned room at RH-25-30% , T=18-20°C.
After 4 hours there was no change.
After 16 hours there was aggregated white powder.
» In refrigerator:
A sample of white flowable powder was stored in sealed
transparent glass vials at +5°C for 3 months. No physical change
was observed. The material appears to be white powder.
4.4 Precipitation of salt from acetone: (Batch 15)
0.61g of ethanolamine was dissolved in 20ml of dry methanol, and
then 3.50g of laquinimod acid was added and dissolved while
stirring .
The solution was evaporated to dryness under vacuum in a rotary
evaporator (bath T=55°C) .
The residue was dissolved in 30ml of warm dry acetone and
evaporated under the same conditions.
Then the residue ( . g of white glassy foam) was dissolved in
20ml of warm dry acetone and stirred at ambient temperature for 1
hour. White solid precipitate was formed and the mixture was
stored in a refrigerator at +5°C.
Solid product was collected by filtration and dried under vacuum
in an oven at 40°C.
Dry product was 3. 4g of white powder with a yield of 83%.
Analysis and tests :
and C MR - confirmed structure and identified traces of
acetone.
XRD identified crystalline structure, form II.
DSC identified crystalline structure and melting point at 150°C.
Physical stability:
• At room temperature:
A sample of white flowable powder was exposed to atmospheric air
in an air-conditioned room at RB=29~32% » T=18-20°C,
After 4 hrs there was no change; remained white powder.
After 25 hrs there was no change; remained white powder.
• In refrigerator
A sample of white flowable powder was stored in sealed
transparent glass vials at +5°C for 3 months. Small physical
change was observed. The material appears to be an off-white
powder .
Aqueous solubility - 625 mg/ l H 0 .
suits
Formation of salt
All four amines o f the invention formed highly soluble solid
salts with laquinimod acid in aqueous media and/or polar organic
solvent (methanol) .
The isolated compounds appeared as a white powder.
Crysta in ty of Laquinimod amine _salts
The meglumine salt of laquinimod prepared by the above-described
methods appeared as an amorphous solid.
The choline, lysine and monoethanolamine salts of laquinimod,
prepared by precipitation, were crystalline compounds. The
choline, lysine and monoethanolamine salts of laquinimod prepared
by lyophil ization and solvent evaporation were solid amorphous
materials .
An important parameter affecting the salt crystallization is the
presence of water in the crystallizing mixture. Even at low water
concentration the salts precipitate as oil or semi-solid
materials .
Solubility of Laquinimod amine salts
The amine base salts of the invention have an enhanced aqueous
solubility over the sodium salt (Table 6 ) .
Table 6 . Solubility o f Laquinimod salts
* U.S. Patent No. 7,589,208
** - unlimited solubility
The amine salts o f the invention were very soluble in water
according to the USP . The existing Na salt is only freely soluble
to soluble. Amorphous forms o f lysine and ethanolamine salts were
found more soluble than the crystalline analogs.
P_h S_ical. Stability o f Laquinimod amine salts
Table 7 . A Physical stability o f meglumine salt in glass vials at
+5°C, air atmosphere
Table 7.C Physical stability o f L-Lysine salt in glass vials
+5"C, air atmosphere
Sample: Batch 10 Batch 9
Time, month Sample appearance
0 White flowable powder White flowable powder
5 White flowable powder White flowable powder
9 White flowable powder White flowable powder
Table 7.D Physical stability o f ethanolami e salt in glass vials
at +5°C, air atmosphere
Meglumine salt: Batches 1 and 4 appeared as white flowable
powders with no sign o f physical change after 10 months o f
storage. In batch 3 physical change was observed after 5 months.
Choline salt : Batches 7 and 8 appeared as white powders with no
sign o f physical change after 9 months o f storage, batch 6 ,
prepared by lyophilization, developed color and deliquescence
occurred after 5 months.
_ _ *lt Batches 10 and 9 appeared as white flowable
powders with no sign o f physical change after 9 months o f storage
Ethanolamine salt: Batch 14 appeared as white flowable powder
with no sign of physical change after 7 months o f storage. In
batches 13 and 15 some changes were observed. In batch 13 slight
aggregation was developed after 7 month o f storage. In batch 15
color change was developed after 3 month. Amorphous material
prepared b y evaporation (batch 12) was less stable. Stronger
color development and aggregation were observed in this sample.
The presented data demonstrates that all four amine salts can be
prepared as processable and stable powders.
Obs ervat ion
Meglumine Salt:
Batch 1 was amorphous material prepared by vacuum evaporation
from an acetone : ater solution to dryness. The sample was white
flowable powder. Particles were highly aggregated and irregular
in size (300-1000 um) .
Batch 4 was amorphous material prepared by lyophilization from an
aqueous solution. Sample was white flowable powder. Particles
were aggregated and irregular. Most of the primary particles were
aggregates were 500-2000 .
Choline Salt:
Batch 7 was amorphous material prepared by evaporation to dryness
from a methanol solution. The sample was white powder. Particles
were irregular, 200-2000 in size.
Batch 8 was crystalline material prepared by precipitation from
dioxane :methanol . The sample was white powder. Particles were
aggregated rod-shaped crystals, 2-10 m in size.
L-Lysine Salt:
Batch 9 was amorphous material prepared by evaporation to dryness
from an acetone-water solution. The sample was white flowable
powder. Particles were irregular, 20-1000um in size.
Batch 10 was crystalline material prepared by precipitation from
methanol-acetone . The sample was white flowable powder. Particles
were highly aggregated rod-shape crystals, 5-10 in size.
Ethanolamine Salt:
Batch 12 was amorphous material prepared by evaporation to dryness,
from a water-acetone solution. The sample was white flowable powder.
Particles were irregular, 500-3000 in size.
Batc 13 was amorphous material prepared by lyophili zation of an
aqueous solution. Sample was white powder. Particles were irregular,
20 200 in size.
Batch 14 was crystalline material (solvate with dioxane, Form I),
prepared by precipitation from dioxane. The sample was white
flowable powder. Particles were aggregated rod-shaped crystals,
20-80 n size.
Batch 15 was crystalline material Form II) prepared by
precipitation from acetone. The sample was white flowable powder.
Particles were aggregated, irregular, rod and prism shaped
crystals, 2-101 in size.
te f r nc
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Absoption of Piroxicam via Salt Formation with
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7 , 1998.
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triclosan by solubilization, complexation and in situ salt
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2007/096588, published August 30, 2007.
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published September 16, 2010.
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Cyclodextrins and L-Lysine on the Aqueous Solubility of
Nimesulide Isolation and Characterization of Nimesulide-LLys
ine-Cyclodextrin Complexes", __ Sci_. 86(4) : 475-
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November 5 , 2012,
< w .sigmaaldrich .com,'catalog/product /FLUKA/ 1 892?lang=en& re
gion=US>
U.S. Patent No. 4,948,805, issued August 14, 1990 (Ziggiotti
U.S. Patent No. 5,028,625, issued July 2 , 1991 (Motola
a )
U.S Patent No. 6,077,851, issued June 20, 2000 (Bjork et
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U .S Patent No. 6,638,537 issued October 28, 2003 (Dennis et
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U.S Patent No. 6,869,948 issued March 22, 2005 (Bock et
al)
0 . Patent Mo. 7,105,512 issued September 12, 2006
(Mori zono et al)
U.S. Patent No. 7,589,208 issued September 15, 2009 (Jannson
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hat is claimed i :
1 . Laquinimod amine salt, which is laquinimod meglumine,
laquinimod choline hydroxide, laquinimod L-lysine or
laquinimod monoethanolamine .
2 . The laquinimod amine salt of claim 1 which is laquinimod
meglumine .
3 . The laquinimod meglumine of claim 2 which is isolated.
4 . The laquinimod meglumine according to claim 2 or 3 , wherein
the laquinimod meglumine is characterized by a DSC thermogram
as shown in figure 4A, B and C .
5 . The laquinimod meglumine according to any of claims 2-4,
wherein the laquinimod meglumine is characterized by a
solid-state JC NMR spectrum with broad peak at 60-77, broad
peak at 122-134, peak at 142.2 and 171.3 + 0.2 ppm.
6 . The laquinimod meglumine according to any of claims 2-5,
wherein the laquinimod meglumine is characterized by a solid
state JC NMR as shown in figures 5A and 5B.
7 . The laquinimod amine salt of claim 1 which is laquinimod
choline hydroxide.
8 . The laquinimod choline hydroxide of claim 7 which is
isolated.
9 . The laquinimod choline hydroxide according to claim 7 or 8 ,
wherein the laquinimod choline hydroxide is characterized by
a DSC thermogram as shown in figures 6A, B , C , D and E .
10. The laquinimod choline hydroxide according to any of claims
7-9, wherein the laquinimod choline hydroxide is
characterized by a powder XRD pattern with characteristic
peaks at 10.1°, 11.8°, 13.4°, 14.4° and 16.1° 2-theta + 0.2.
11. The laquinimod choline hydroxide according to any of claims
7-10, wherein the laquinimod choline hydroxide is
character zed by a powd X D pattern with characteristic
peaks at 19,3°, 21.2°, 22.7°, 24.8° and 27.6° 2-theta ± 0.2.
The laquinimod choline hydroxide according to any of claims
7-11, wherein the laquinimod choline hydroxide is
characterized by a powder XRD pattern as shown in figures 7
and 8 .
The laquinimod choline hydroxide according to any of claims
7-12, wherein the laquinimod choline hydroxide is
characterized by a solid-state ¾ NMR spectrum with peaks at
about 122.1, 127.2, 142.7 and 169.7 0.2 ppm.
The laquinimod choline hydroxide according to claims 7-13,
wherein the laquinimod choline hydroxide is characterized by
a solid-state ' 'c NMR spectrum having chemical shifts
differences between the signal exhibiting the lowest chemical
shift and another in the chemical shift range o f 100 to 180
ppm of about 12.6, 17.7, 33.2 60.2 and + 0.1 ppm.
The laquinimod choline hydroxide according to any of claims
7-14, wherein the laquinimod choline hydroxide is
characterized by a solid state C NMR as shown in figures 9A
and B .
The laquinimod choline hydroxide according to any of claims
7-15, wherein the laquinimod choline hydroxide is in
crystalline form.
The laquinimod choline hydroxide according to any of claims
7-15, wherein the laquinimod choline hydroxide is in
amorphous form.
The laquinimod amine salt of claim 1 which is laquinimod L
lysine .
The laquinimod L-lysine of claim 18 which is isolated.
The laquinimod L-lysine according to claim 18 or 19, wherein
the laquinimod L-lysine is characterized by the DSC
thermogram as shown in figures IDA, B and C .
21. The laquinimod L-lysine according to any of claims 18-20,
wherein the laquinimod L-lysine is characterized by a powder
XRD pattern with characteristic peaks at 5.6°, 9.0°, 11.7°,
13.0° and 15.9° 2-theta ± 0.2.
22. The laquinimod L-lysine according to any of claims 18-21,
wherein the laquinimod L-lysine is characterized by a powder
XRD pattern with characteristic peaks at 17.9°, 18.9°, 21.1°,
22.5° and 23.0° degrees 2-theta ± 0.2.
23. The laquinimod L-lysine according to any of claims 18-22,
wherein the laquinimod L-lysine is characterized by a powder
XRD pattern as shown in figure 11.
24. The laquinimod L-lysine according to any o f claims 18-23,
wherein the laquinimod L-lysine is in crystalline form.
25. The laquinimod L-lysine according to any of claims 18-23,
wherein the laquinimod L-lysine is in amorphous form.
26. The laquinimod amine salt of claim 1 which is laquinimod
monoethanolamine .
27. The laquinimod monoethanolamine of claim 26 which is
isol ate .
28. The laquinimod monoethanolamine of claim 26 or 27, wherein
the laquinimod monoethanolamine is characterized by a DSC
thermogram as shown in figure 12A.
29. The laquinimod monoethanolamine of any of claims 26-28,
wherein the laquinimod monoethanolamine is characterized by a
powder XRD pattern with characteristic peaks at 6.5°, 14.4°,
17.9°, 18.7° and 20.6° 2-theta ± 0.2.
30. The laquinimod monoethanolamine of any of claims 26-29,
wherein the laquinimod monoethanolamine is characterized by a
powder XRD pattern with characteristic peaks at 17.1°, 19.4°,
22.3°, 23.3° and 24.8° degrees 2-theta ± 0.2.
31. The laquinimod monoethanolamine of any of claims 26-30,
wherein the laquinimod monoethanolamine is characterized by a
powder XRD pattern as shown in figure 13.
32. The laquinimod monoethanolamine of any of claims 26-31,
wherein the laquinimod monoethanolamine is characterized by a
DSC thermogram as shown in figure 12B.
33. The laquinimod monoethanolamine of any of claims 26-32,
wherein the laquinimod monoethanolamine is characterized by a
powder XRD pattern with characteristic peaks at 8.2°, 9.8°,
11.2°, 13.2° and 17.9° 2-theta ± 0.2.
34. The laquinimod monoethanolamine of any of claims 26-33,
wherein the laquinimod monoethanolamine is characterized by a
powder XRD pattern with characteristic peaks at 18.6°, 20.4°,
22.9°, 24.3° and 26.2° 2-theta ± 0.2.
35. The laquinimod monoethanolamine according to any of claims
26-34, wherein the laquinimod monoethanolamine is
characterized by a powder XRD pattern as shown in figure 14.
36. The laquinimod monoethanolamine according to any of claims
26-35, wherein the laquinimod monoethanolamine is in
crystalline form.
37. The laquinimod monoethanolamine according to any o f claims
26-35, wherein the laquinimod monoethanolamine is in
amorphous form.
38. A pharmaceutical composition comprising the laquinimod amine
salt of any one of claims 1-37 and at least one
pharmaceutical acceptable excipient.
39. The pharmaceutical composition according to claim 38, further
comprising laquinimod acid.
40. The pharmaceutical composition according to any one of claims
38-39, wherein the laquinimod acid is present in an amount of
less than 1.5% based on the total laquinimod content of the
pharmaceuticai composition.
4 . The pharmaceut composition according to claim 38, which
is free of laquinimod acid.
42. The pharmaceutical composition according to any one of claims
38-41, further comprising the sodium salt of laquinimod.
43. The pharmaceutical composition according to any one of claims
38-41, which is free of the sodium salt of laquinimod.
44. A process for manufacture of laquinimod amine salt according
to any one of claims 1 to 43, comprising:
a ) combining a solution of amine with laquinimod acid to
form a first mixture;
b ) adding a solvent to the first mixture to form a second
mixture ;
c ) removing liquid from the second mixture; and
d ) recovering the laquinimod amine.
45. The process according to claim 44, wherein said amine is
selected from the group consisting of meglumine, choline
hydroxide, t ysine and m noethanola re .
46. The process according to claims 44 and 45, wherein the
solvent added in step b ) is selected from the group
consisting of acetone, methanol, ethanol and dioxane or
combination thereof, and wherein in step c ) the liquid is
removed at ambient temperature and at reduced pressure.
47. A process for manufacture of the pharmaceutical
composition according to any of claims 38-43, comprising:
a ) obtaining laquinimod amine salt; and
b ) admixing the laquinimod amine salt with at least one
pharmaceutical acceptable excipient.
48. A method for treating a subject afflicted with a form of
multiple sclerosis or clinical isolated syndrome comprising
administering to the subject the pharmaceutical composition
according to any one of claims 38-43 so as to thereby treat
the subject.
49. A method for alleviating a symptom of multiple sclerosis in
a subject afflicted with a form o f multiple sclerosis
comprising administering to the subject the pharmaceutical
composition of any one of claims 38-43 so as to thereby
alleviate the symptom of multiple sclerosis in the subject.