U.S. patent application number 17/466328 was filed with the patent office on 2022-06-23 for method of preparing fluorine-18 labeled cabozantinib and its analogs.
The applicant listed for this patent is Exelixis, Inc.. Invention is credited to Patrick L. Chow, David J. Donnelly, Benjamin J. Henley, Wei Xu.
Application Number | 20220194902 17/466328 |
Document ID | / |
Family ID | |
Filed Date | 2022-06-23 |
United States Patent
Application |
20220194902 |
Kind Code |
A1 |
Xu; Wei ; et al. |
June 23, 2022 |
Method of Preparing Fluorine-18 Labeled Cabozantinib and Its
Analogs
Abstract
The present invention relates to a method of preparing
Cabozantinib (Cyclopropane-1,1-dicarboxylic acid
[4-(6,7-di-methoxy-quinolin-4-vloxy)-phenyl]amide
(4-fluoro-phenyl)amide) and .sup.18F labeled Cabozantinib.
Inventors: |
Xu; Wei; (Danville, CA)
; Donnelly; David J.; (Doylestown, PA) ; Chow;
Patrick L.; (Lawrenceville, NJ) ; Henley; Benjamin
J.; (Mount Laurel, NJ) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Exelixis, Inc. |
Alameda |
CA |
US |
|
|
Appl. No.: |
17/466328 |
Filed: |
September 3, 2021 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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15500312 |
Jan 30, 2017 |
11124481 |
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PCT/US2015/043195 |
Jul 31, 2015 |
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17466328 |
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62031471 |
Jul 31, 2014 |
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International
Class: |
C07D 215/233 20060101
C07D215/233; C07D 215/22 20060101 C07D215/22 |
Claims
1. A method for preparing a compound of Formula I: ##STR00049## or
pharmaceutically acceptable salt thereof wherein: each of R.sup.1
and R.sup.2 is independently alkoxy or haloalkoxy; R.sup.3 is H, F,
Cl, I, or Br; and R.sup.4 is F, Cl, I or Br; comprising: i)
reacting a compound of Formula 8 with a compound of Formula 9 in
the presence of a coupling reagent and a tertiary amine base and an
aprotic polar solvent to generate a compound of Formula I
##STR00050##
2. The method of claim 1, wherein the coupling reagent is selected
from the group consisting of: N,N'-dicyclohexylcarbodiimide (DCC),
N,N'-diisopropylcarbodiimide (DIC),
1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDCI),
hydroxybenzotriazole (HOBt), 1-hydroxy-7-azabenzotriazole (HOAt),
benzotriazole-1-yl-oxytripyrrolidinophosphonium hexafluorophosphate
(BOP reagent), benzotriazol-1-yl-oxytripyrrolidinophosphonium
hexafluorophosphate (PyBOP),
1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium
3-oxid hexafluorophosphate (HATU),
O-(benzotriazol-1-yl)-N,N,N'N'-tetramethyluronium tetrafluoroborate
(TBTU), N,N,N',N'-tetramethyl-O-(1H-benzotriazol-1-yl)uranium
hexafluorophosphate (HBTU),
O-[(ethoxycarbonyl)cyanomethylenamino]-N,N,N',N'-tetramethyluronium
tetrafluoroborate (TOTU), and
(1-cyano-2-ethoxy-2-oxoethylidenaminooxy)dimethylamino-morpholino-carbeni-
um hexafluorophosphate (COMU), or a combination thereof.
3. The method of claim 2, wherein the coupling reagent is
1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium
3-oxid hexafluorophosphate (HATU) or
benzotriazol-1-yl-oxytripyrrolidinophosphonium hexafluorophosphate
(PyBOP).
4. The method of claim 3, wherein the tertiary amine base is
selected from the group consisting of diisopropylethyl amine
(DIPEA), triethyl amine (TEA), N-methyl imidazole, pyridine,
4-(dimethylamino)pyridine (DMAP), 3,4-lutidine, 4-methoxypyridine,
N-methylmorpholine (NMO), 1,4-diazabicycle[2.2.2]octane (DABCO),
and 1,8-diazacycloundec-7-ene (DBU), or a combination thereof.
5. (canceled)
6. The method of claim 1, wherein the tertiary amine base is
diisopropylethyl amine (DIPEA).
7. The method of claim 1, wherein the aprotic polar solvent is
selected from the group consisting of acetonitrile, diethyl ether,
diisopropyl ether, 2-methoxyethyl ether, 1,2-dimethoxyethane,
tert-butyl methyl ether, tetrahydrofuran, 1,4-dioxane, benzene,
toluene, .alpha.,.alpha.,.alpha.-trifluorotolune, cyclohexane,
methylcyclohexane carbon tetrachloride, methylene chloride,
N,N-dimethylformamide, dimethyl sulfoxide, and
N-methyl-2-pyrrolidone or a combination thereof.
8. (canceled)
9. The method of claim 7, wherein the aprotic solvent is
N,N-dimethylformamide, dimethyl sulfoxide or a combination
thereof.
10. The method of claim 9, wherein the reaction is heated with
about 10 Watts to about 50 Watts of microwave radiation.
11. (canceled)
12. The method of claim 10, wherein the reaction is heated with
about 10 Watts to about 20 Watts of microwave radiation.
13. The method of claim 12, wherein the reaction is heated to about
20.degree. C. to about 100.degree. C.
14. The method of claim 13, wherein the reaction is heated to about
85.degree. C.
15. A method for preparing a compound of Formula I: ##STR00051## or
pharmaceutically acceptable salt thereof wherein: each of R.sup.1
and R.sup.2 is independently alkoxy or haloalkoxy; R.sup.3 is H, F,
Cl, Br, or I; and R.sup.4 is F, Cl, Br, or I; comprising: i)
reacting the compound of Formula 8 with a chlorinating or
brominating agent and a base to generate compound of Formula 8a,
wherein X is chloro or bromo: ##STR00052## and ii) reacting the
compound of Formula 8a with a compound of Formula 9 in the presence
of a base to generate the compound of Formula I: ##STR00053##
16. The method of claim 15, wherein the chlorinating or brominating
agent comprises thionyl chloride, thionyl bromide, oxalyl chloride,
phosphorus pentachloride or phosphorus trichloride.
17. The method of claim 16, wherein the chlorinating agent is
oxalyl chloride.
18. The method of claim 17, wherein the base is selected from the
group consisting of potassium carbonate, sodium carbonate, sodium
bicarbonate, triethyl amine (TEA), diisopropyl ethyl amine (DIPEA),
pyridine, N,N-dimethylamino-4-pyridine (DMAP), and
N-methylmorpholine (NMO), or a combination thereof.
19. (canceled)
20. The method of claim 18, wherein the base is potassium
carbonate.
21. The method of claim 20, wherein the reaction is maintained at a
temperature ranging from about 20.degree. C. to about 40.degree.
C.
22. The method of claim 21, wherein the reaction is maintained at a
temperature ranging from about 20.degree. C. to about 25.degree.
C.
23-42. (canceled)
43. A method for preparing a compound of Formula 1a: ##STR00054##
or pharmaceutically salt thereof wherein: R.sup.4 is F; the method
comprising: i) reacting a compound of Formula 10 with a
chlorinating agent to generate a compound of Formula 11:
##STR00055## ii) coupling the compound of Formula 11 with a
compound of Formula 23 in the presence of a base to generate a
compound of Formula 12: ##STR00056## iii) coupling the compound of
Formula 12 with a compound of Formula 13 in the presence of a
coupling agent to generate a compound of Formula 14: ##STR00057##
iv) saponifying the compound of Formula 14 in the presence of a
base to generate a compound of Formula 15: ##STR00058## v) reacting
the compound of Formula 15 with a halogenating reagent to generate
a compound of Formula 15a: ##STR00059## wherein X is chloro or
bromo; and vi) reacting the compound of Formula 15a with a compound
of Formula 9 to generate a compound of Formula 1a: ##STR00060##
44. (canceled)
Description
PRIORITY CLAIM
[0001] This application claims priority to U.S. Application Ser.
No. 62/031,471, filed Jul. 31, 2014. The entire contents of the
aforementioned application are incorporated herein by
reference.
FIELD OF THE INVENTION
[0002] The present invention relates to a method of preparing
Cabozantinib (Cyclopropane-1,1-dicarboxylic acid
[4-(6,7-dimethoxy-quinolin-4-yloxy)-phenylo]amide
(4-fluoro-phenyl)amide) and .sup.18F-labeled Cabozantinib.
BACKGROUND OF THE INVENTION
[0003] Molecular imaging provides a non-invasive assessment of
biological and biochemical processes in living subjects. The use of
positron emission tomography (PET) has the potential to enhance the
understanding of a potential drug during preclinical and clinical
drug development. This information would be especially important in
determining whether a potential drug reaches its target tissue in
challenging environments such as glioblastoma multiforme (GEM)
brain tumors.
[0004] Cabozantinib (Cyclopropane-1,1-dicarboxylic acid
[4-(6,7-dimethoxy-quinolin-4-yloxy)-phenylo]amide
(4-fluoro-phenyl)amide, (1) is a multitargeted kinase inhibitor
with inhibitory activity against vascular endothelial growth factor
receptor-2 (VEGFR-2) (IC.sub.50 of 0.035 nM), tyrosine kinase MET
(IC.sub.50 of 1.3 nM), receptor tyrosine kinase encoded by
rearranged during transfection (RET) proto-oncogene (IC.sub.50 4
nM), and c-KIT (stem-cell factor) (IC.sub.50 of 4.6 nM).
##STR00001##
[0005] In cellular assays, Cabozantinib inhibits phosphorylation of
receptors VEGFR-2, RET, and MET, as well as c-KIT, with IC.sub.50
values of 1.9, 7.8. 5.0, and 42 nM. respectively. Cabozantinib
inhibits MET and VEGFR-2 phosphorylation in tumor models in vivo
and demonstrates potent antimetastatic, antitumor, and
antiangiogenic activity in preclinical models. The vascular
endothelial growth factor (VEGF) and the hepatocyte growth factor
(HGF) are potent mediators of angiogenesis. Angiogenesis is the
formation of new blood vessels and is one of the key requirements
of tumor growth during cancer progression. Recent studies suggest
that activation of VEGF through the VEFGR-2 and the HGF receptor
kinase MET, play synergistic roles in tumor progression.
[0006] In 2012, the FDA approved Cabozantinib as the L-malate salt
(COMETRIQ.RTM., Exelixis, Inc.) for the treatment of patients with
progressive metastatic medullary thyroid cancer (MTC) and is
currently being evaluated in patients with glioblastoma multiforme.
Over expression of MET and VEGFR-2 have shown to correlate with
poor prognosis in GBM, one of the most common and aggressive brain
tumors.
[0007] The synthesis of Cabozantinib (1) has been previously
described in International Patent Application publication No. WO
2005/030140 filed Sep. 9, 2004, the contents of which are
incorporated herein by reference in its entirety. There remains a
need for new processes for synthesizing Cabozantinib and
isotopically labeled Cabozantinib, [.sup.18F]-Cabozantinib, in a
minimum number of steps and concomitant high yields.
SUMMARY OF THE INVENTION
[0008] These and other needs are met by the present invention which
is directed to a method for synthesizing Cabozantinib by
incorporating the fluoroaniline moiety as the last step of the
synthesis. In some embodiments,
1-(4-(6,7-dimethoxyquinolin-3-yloxy)phenylcarbamoyl)
cyclopropanecarboxylic acid is coupled with 4-fluoroaniline or
[.sup.18F]-fluoroaniline.
[0009] Thus, in one aspect, the present invention provides a method
for generating a compound of Formula I:
##STR00002##
or a pharmaceutically acceptable salt thereof, wherein each of
R.sup.1 and R.sup.2 is independently alkoxy or haloalkoxy; R.sup.3
is H, F, Cl, I or Br; and R.sup.4 is F, .sup.18F, Cl, I or Br;
comprising:
[0010] i) reacting a compound f Formula 8 with a compound of
Formula 9 in the presence of a coupling reagent to generate a
compound of Formula I:
##STR00003##
[0011] In some implementations, the coupling reagent is selected
from the group consisting of N,N'-dicyclohexylcarbodiimide (DCC),
N,N'-diisopropylcarbodiimide (DIC),
1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDCI),
hydroxybenzotriazole (HOBt), 1-hydroxy-7-azabenzotriazole (HOAt),
benzotriazole-1-yl-oxytripyrrolidinophosphonium hexafluorophosphate
(BOP reagent), benzotriazol-1-yl-oxytripyrrolidinophosphonium
hexafluorophosphate (PyBOP),
1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium
3-oxid hexafluorophosphate (HATU),
O-(benzotriazol-1-yl)-N,N,N'N'-tetramethyluronium tetrafluoroborate
(TBTU), N,N,N',N'-Tetramethyl-O-(1H-benzotriazol-1-yl)uranium
hexafluorophosphate (HBTU),
O-[(ethoxycarbonyl)cyanomethylenamino]-N,N,N',N'-tetramethyluronium
tetrafluoroborate (TOTU), and
(1-cyano-2-ethoxy-2-oxoethylidenaminooxy)dimethylarnino-morpholino-carben-
ium hexafluorophosphate (COMU) or a combination thereof.
[0012] Another aspect of the present invention provides a method
for generating a compound of Formula I or a pharmaceutically
acceptable salt thereof wherein each of R.sup.1 and R.sup.2 is
independently alkoxy or haloalkoxy; R.sup.3 is H, F, Cl, I or Br;
and R.sup.4 is F, .sup.18F, Cl, I or Br; comprising:
[0013] i) reacting a compound of Formula 8 with a compound of
Formula 9 in the presence of a coupling reagent and heating the
reaction with microwave radiation to generate a compound of Formula
I:
##STR00004##
[0014] In some embodiments, heating the reaction during the
coupling reaction using microwave radiation results in shorter
reaction times compared to the coupling reaction without microwave
heating. In some embodiments, microwave radiation can be applied to
the coupling reaction in an amount ranging from about 10 watts to
about 20 watts, thereby generating a higher yield of a compound of
Formula I as compared to the yield of a compound of Formula I in
the absence of microwave heating.
[0015] Another aspect of the present invention provides a method
for synthesizing a compound of Formula I, the method
comprising:
[0016] i) reacting a compound of Formula 8 with a halogenating
agent to generate an acid halide compound of Formula 8a, followed
by reacting the acid halide compound of Formula 8a with a compound
of Formula 9 in the presence of a base to generate the compound of
Formula I:
##STR00005##
[0017] In some implementations, the base can include: potassium
carbonate, sodium carbonate, sodium bicarbonate, triethyl amine
(TEA), diisopropyl ethyl amine (DIPEA), pyridine,
N,N-dimethyiamino-4-pyridine (DMAP), and N-methylmorpholine (NMO),
or combination thereof.
[0018] In another aspect, the present invention provides a method
for generating a compound of Formula 9a, wherein R.sup.4 is
.sup.18F, the method comprising:
[0019] i) reacting a compound of Formula 18 with a fluorinating
reagent to generate a compound of Formula 22a:
##STR00006##
and
[0020] ii) reducing the compound of Formula 22a to generate a
compound of Formula 9a:
##STR00007##
[0021] In some implementations, the fluorinating reagent is
K[.sup.18F] bound to a cryptand. In some embodiments, suitable
cryptand compounds can include:
1,10-diaza-4,7,13,16.21,24-hexaoxa-1,10-diazabicyclo[8.8.8]hexac-
osane (Kryptofix.RTM. 2.2.2. or Kryptofix 2.2.2 or Kryptofix 222).
In some implementations, an exemplary fluorinating reagent is
Kryptofix 2.2.2/K.sup.18[F].
BRIEF DESCRIPTION OF THE DRAWINGS
[0022] FIG. 1 depicts the solid phase extraction (SPE) purification
of 1,4-[.sup.18F]-fluoronitrobenzene.
[0023] FIG. 2 depicts a radio-HPLC chromatogram showing co-elution
of [.sup.18F]-Cabozantinib co-injected with non-radioactive
reference standard.
[0024] FIG. 3 depicts Positron Emission Tomography (PET) images of
mice injected with [.sup.18F]-Cabozantinib.
[0025] FIG. 4 depicts the radiometabolite analysis of
[.sup.18F]-Cabozantinib in mouse plasma.
[0026] FIG. 5 depicts the biodistribution analysis of
[.sup.18F]-Cabozantinib in mice.
DETAILED DESCRIPTION OF THE INVENTION
[0027] As used herein, the following definitions shall apply unless
otherwise indicated.
[0028] As used in the present specification, the following words
and phrases are generally intended to have the meanings as set
forth below, except to the extent that the context in which they
are used indicates otherwise or they are expressly defined to mean
something different.
[0029] When chemical structures are depicted or described, unless
explicitly stated otherwise, all carbons are assumed to have
hydrogen substitution to conform to a valence of four. For example,
in the structure on the left-hand side of the schematic below there
are nine hydrogens implied. The nine hydrogens are depicted in the
right-hand structure. Sometimes a particular atom in a structure is
described in textual formula as having a hydrogen or hydrogens as
substitution (expressly defined hydrogen), for example,
--CH.sub.2CH.sub.2--. It is understood by one of ordinary skill in
the art that the aforementioned descriptive techniques are common
in the chemical arts to provide brevity and simplicity to
description of otherwise complex structures.
##STR00008##
[0030] If a group "R" is depicted as "floating" on a ring system,
as for example in the formula:
##STR00009##
then, unless otherwise defined, a substituent "R" may reside on any
atom f the ring system, assuming replacement of a depicted,
implied, r expressly defined hydrogen from one of the ring atoms,
so long as a stable structure is formed.
[0031] "Alkoxy" or "alkoxyl" refers to the group --O-alkyl, for
example including from one to eight carbon atoms of a straight,
branched, cyclic configuration, unsaturated chains, and
combinations thereof attached to the parent structure through an
oxygen atom. Examples include methoxy, ethoxy, propoxy, isopropoxy,
cyclopropyloxy, cyclohexyloxy and the like. Lower-alkoxy refers to
groups containing one to six carbons.
[0032] "Alkyl" is intended to include linear, branched, or cyclic
hydrocarbon structures and combinations thereof, inclusively. For
example, "C.sub.8 alkyl" may refer to an n-octyl, iso-octyl,
cyclohexylethyl, and the like. Lower alkyl refers to alkyl groups
of from one to six carbon atoms. Examples of lower alkyl groups
include methyl, ethyl, propyl, isopropyl, butyl, s-butyl, t-butyl,
isobutyl, pentyl, hexyl and the like. Higher alkyl refers to alkyl
groups containing more than eight carbon atoms. Exemplary alkyl
groups are those of C.sub.20 or below. Cycloalkyl is a subset of
alkyl and includes cyclic hydrocarbon groups of from three to
thirteen carbon atoms. Examples of cycloalkyl groups include
c-propyl, c-butyl, c-pentyl, norbornyl, adamantyl and the like. In
this application, alkyl refers to alkanyl, alkenyl, and alkynyl
residues (and combinations thereof); it is intended to include
cyclohexylmethyl, vinyl, allyl, isoprenyl, and the like. Thus when
an alkyl residue having a specific number of carbons is named, all
geometric isomers having that number of carbons are intended to be
encompassed; thus, for example, either "butyl" or "C.sub.4 alkyl"
is meant to include n-butyl, sec-butyl, isobutyl, r-butyl,
isobutenyl and but-2-yne radicals; and for example, "propyl" or
"C.sub.3 alkyl" each include n-propyl, propenyl, and isopropyl.
[0033] As used herein, a "halogen" or "halo" group refers to
fluorine, chlorine, bromine or iodine.
[0034] "Haloalkoxy" refers to an alkoxy group which is substituted
with one or more halogen atoms.
[0035] Cryptands are a family of synthetic and polycyclic
multidentate ligands for a variety of cations. An exemplary
cryptand can include:
1,10-diaza-4,7,13,16,21,24-hexaoxabicyclo[8.8.8]hexacosane, which
is a [2.2.2]cryptand. Cryptands are commercially available under
the tradename Kryptofix.RTM.. Cryptands with nitrogen atom(s)
exhibit high affinity for alkali metal cations.
[0036] Adsorbents are materials that adsorb various chemicals
and/or gases. Examples of adsorbents include silica gel (chemically
inert. nontoxic, polar and stable up to 399.degree. C.) zeolites,
basic alumina, neutral alumina, octadecyl carbon chain (C18)-bonded
silica column, C8-bonded silica, cyano-bonded silica and
phenyl-bonded silica.
[0037] Ion exchange resins are ion exchangers that exchange ions
from a solution and a complex. They are either cation exchangers
that exchange positively charged ions (cations) or anion exchangers
that exchange negatively charged ions (anions). There are also
amphoteric exchangers that are able to exchange both cations and
anions simultaneously.
[0038] "Pharmaceutically acceptable salt" of a compound means a
salt that is pharmaceutically acceptable and that possesses the
desired pharmacological activity of the parent compound. It is
understood that the pharmaceutically acceptable salts are
non-toxic. Additional information on suitable pharmaceutically
acceptable salts can be found in Remington's Pharmaceutical
Sciences, 17th ed., Mack Publishing Company, Easton, Pa., 1985,
which is incorporated herein by reference or S. M. Berge, et al.,
"Pharmaceutical Salts," J. Pharm. Sci., 1977;66:1-19 both of which
are incorporated herein by reference in their entirety.
[0039] Examples of pharmaceutically acceptable acid addition salts
include those formed with inorganic acids such as hydrochloric
acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric
acid, and the like; as well as organic acids such as acetic acid,
trifluoroacetic acid, propionic acid, hexanoic acid,
cyclopentanepropionic acid, glycolic acid, pyruvic acid, lactic
acid, oxalic acid, maleic acid, malonic acid, succinic acid,
fumaric acid, tartaric acid, malic acid, citric acid, benzoic acid,
cinnamic acid, 3-(4-hydroxybenzoyl)benzoic acid, mandelic acid,
methanesulfonic acid, ethanesulfonic acid, 1,2-ethanedisulfonic
acid, 2 hydroxyethanesulfonic acid, benzenesulfonic acid,
4-chiorobenzenesulfonic acid, 2 naphthalenesulfonic acid, 4
toluenesulfonic acid, camphorsulfonic acid, glucoheptonic acid,
4,4'-methylenebis-(3-hydroxy-2-ene-1-carboxylic acid),
3-phenylpropionic acid, trimethylacetic acid, tertiary butylacetic
acid, lauryl sulfuric acid, gluconic acid, glutamic acid,
hydroxynaphthoic acid, salicylic acid, stearic acid, muconic acid,
p-toluenesulfonic acid, and salicylic acid and the like.
[0040] "Pharmaceutically acceptable acid addition salt" refers to
those salts that retain the biological effectiveness of the free
bases and that are not biologically or otherwise undesirable,
formed with inorganic acids such as hydrochloric acid, hydrobromic
acid, sulfuric acid, nitric acid, phosphoric acid, and the like, as
well as organic acids such as acetic acid, trifluoroacetic acid,
propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic
acid, malonic acid, succinic acid, fumaric acid, tartaric acid,
citric acid, benzoic acid, cinnamic acid, mandelic acid,
methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid,
salicylic acid and the like.
[0041] "Pharmaceutically acceptable base addition salts" include
those derived from inorganic bases such as sodium, potassium,
lithium, ammonium, calcium, magnesium, iron, zinc, copper,
manganese, aluminum salts and the like. Exemplary salts are the
ammonium, potassium, sodium, calcium, and magnesium salts. Salts
derived from pharmaceutically acceptable organic non-toxic bases
include, but are not limited to, salts of primary, secondary, and
tertiary amines, substituted amines including naturally occurring
substituted amines, cyclic amines and basic ion exchange resins,
such as isopropylamine, trimethylamine, diethylamine,
triethylamine, tripropylamine, ethanolamine,
2-dimethylaminoethanol, 2-diethylaminoethanol, dicyclohexylamine,
lysine, arginine, histidine, caffeine, procaine. hydrabamine,
choline, betaine, ethylenediamine, glucosamine, methylglucamine,
theobromine, purines, piperazine, piperidine, N-ethylpiperidine,
polyamine resins, and the like. Exemplary organic bases are
isopropylamine, diethylamine, ethanolamine, trimethylamine,
dicyclohexylamine. choline, and caffeine. (See, for example, S. M.
Berge, et al., "Pharmaceutical Salts," J. Pharm. Sci., 1977;66:1-19
which is incorporated herein by reference.)
[0042] "Metabolite" refers to the break-down or end product of a
compound or its salt produced by metabolism or biotransformation in
the animal or human body; for example, biotransformation to a more
polar molecule such as by oxidation, reduction, or hydrolysis, r to
a conjugate (see Goodman and Gilman, "The Pharmacological Basis of
Therapeutics" 8.sup.th Ed., Pergamon Press, Gilman et al. (eds),
1990 for a discussion of biotransformation). As used herein, the
metabolite of a compound of the invention or its salt may be the
biologically active form of the compound in the body. In ne
example, a prodrug may be used such that the biologically active
form, a metabolite, is released in viva. In another example, a
biologically active metabolite is discovered serendipitously, that
is, no prodrug design per se was undertaken. An assay for activity
of a metabolite of a compound of the present invention is known to
one of skill in the art in light of the present disclosure.
[0043] Chemical structures and nomenclature disclosed and described
herein are derived from ChemDraw, version 11.0.1, CambridgeSoft
(Perkin Elmer) Cambridge, MA.
Commonly Used Abbreviations and Their Definitions
[0044] The following abbreviations and terms have the indicated
meanings throughout.
TABLE-US-00001 Abbreviation Definition BOP
Benzotriazole-1-yl-oxytripyrrolidinophosphonium hexafluorophosphate
br broad Calcd calculated Ci curie COMU
(1-Cyano-2-ethoxy-2-oxoethylidenaminooxy)dimethyl-
amino-morpholino-carbenium hexafluorophosphate .degree. C. degrees
Celsius d doublet dd doublet of doublets di de-ionized dt doublet
of triplets DBU Diazabicylo[5.4.0]undec-7-ene DCM Dichloromethane
(or) methylene chloride (or) CH.sub.2Cl.sub.2 DCC
N,N-Dicyclohexylcarbodiimide DIC Diisopropylcarbodiimide DIEA
N,N-Diisopropylethyl amine DMAP N-N-Dimethylamino-4-pyridine DMF
Dimethylformamide DMSO Dimethyl sulfoxide EDCl
1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide EtOAc Ethyl acetate g
gram(s) GBM Glioblastoma multiforme GBq gigabecquerel h or hr
hour(s) HATU 1-[Bis(dimethylamino)methylene]1H-1,2,3-
triazolo[4,5-b]pyridinium-3-oxid hexafluorophosphate HBTU
N,N,N',N'-Tetramethyl-O-(1H-benzotriazol-1- yl)uranium
hexafluorophosphate HGF Hepatocyte growth factor HOAt
1-Hydroxy-7-azabenzotriazole HOBt Hydroxybenzotriazole HPLC high
pressure liquid chromatography Kryptofix
1,10-Diaza-4,7,13,16,21,24-hexaoxa-1,10- 2.2.2
diazabicyclo[8.8.8]hexacosane LCMS liquid chromatography/mass
spectral analysis M Molar or molarity m Multiplet mCi millicurie Me
methyl mg milligram(s) MHz megahertz (frequency) min minute(s) mL
milliliter(s) mmoL millimole(s) MRI Magnetic resonance imaging MS
mass spectral analysis MTC medullary thyroid cancer NaOH Sodium
hydroxide NMO N-methylmorpholine NMR Nuclear magnetic resonance
spectroscopy PET Positron emission tomography Ph Phenyl PyBOP
Benzotriazol-1-yl-oxytripyrrolidinophosphonium hexafluorophosphate
q quartet RT room temperature Rt retention time s singlet SPE
solid-phase extraction t triplet TEA Triethyl amine TLC thin layer
chromatography TOTU O-[Ethoxycarbonyl]cyanomethyleneamino]-
N,N,N',N'-tetramethyluronium tetrafluoroborate THF Tetrahydrofuran
UV ultra-violet .mu.L microliter(s) .mu.M Micromolar VEGF vascular
endothelial growth factor VEGFR-2 vascular endothelial growth
factor receptor-2
Methods of Synthesis
[0045] In one aspect, the present invention provides a method for
generating a compound of Formula I:
##STR00010##
or a pharmaceutically acceptable salt thereof; wherein each of
R.sup.1 and R.sup.2 is independently alkoxy or haloalkoxy; R.sup.3
is H, F, Cl, I or Br; and R.sup.4 is F, .sup.18F, Cl, I or Br; the
method comprising:
[0046] i) reacting a compound of Formula 8 with a compound of
Formula 9 in the presence of a coupling reagent
##STR00011##
to generate a compound of Formula I.
[0047] In some implementations, the coupling reagent can include:
N,N'-dicyclohexylcarbodiimide (DCC), N,N'-diisopropylcarbodiimide
(DIC), 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDCI),
hydroxybenzotriazole (HOBt), 1-hydroxy-7-azabenzotriazole (HOAt),
benzotriazole-1-yl-oxytripyrrolidinophosphonium hexafluorophosphate
(BOP reagent), benzotriazol-1-yl-oxytripyrrolidinophosphonium
hexafluorophosphate (PyBOP),
1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium
3-oxid hexafluorophosphate (HATU),
O-(benzotriazol-1-yl)-N,N,N'N'-tetramethyluronium tetrafluoroborate
(TBTU), N,N,N',N'-tetramethyl-O-(1H-benzotriazol-1-yl)uranium
hexafluorophosphate (HBTU),
O-[(ethoxycarbonyl)cyanomethylenamino]-N,N,N',N'-tetramethyluronium
tetrafluoroborate (TOTU), and
(1-cyano-2-ethoxy-2-oxoethylidenaminooxy)dimethylamino-morpholino-carbeni-
um hexafluorophosphate (COMU) or a combination thereof.
[0048] In some embodiments, the coupling reaction comprising a
compound of Formula 8 and a compound of Formula 9 occurs in the
presence of a tertiary amine base. In some examples, the tertiary
amine base can include: diisopropylethyl amine (DIPEA), triethyl
amine (TEA), N-methyl imidazole, pyridine,
4-(dimethylamino)pyridine (DMAP), 3,4-lutidine, 4-methoxypyridine,
N-methylmorpholine (NMO), 1,4-diazabicycle[2.2.2]octane (DABCO),
and 1,8-diazacycloundec-7-ene (DBU), or a combination thereof.
[0049] In some implementations, the coupling reaction comprising a
compound of Formula 8 and a compound f Formula 9 occurs in the
presence of an aprotic solvent. In some examples, the aprotic
solvent can include: acetonitrile, diethyl ether, diisopropyl
ether, 2-methoxyethyl ether, 1,2-dimethoxyethane, tert-butyl methyl
ether, tetrahydrofuran, 1,4-dioxane, benzene, toluene,
.alpha.,.alpha.,.alpha.-trifluorotoluene, cyclohexane,
methylcyclohexane carbon tetrachloride, methylene chloride,
N,N-dimethylformamide, dimethyl sulfoxide, and
N-methyl-2-pyrrolidone or a combination thereof.
[0050] In some implementations, the coupling reaction comprising a
compound of Formula 8 and a compound of Formula 9 is performed at
temperatures ranging from about 20.degree. C. to about 100.degree.
C., for example, from about 25.degree. C. to about 100.degree. C.
In some embodiments, the coupling reaction is performed at ambient
temperature, i.e. from about 20.degree. C. to about 25.degree. C.
In some embodiments, the coupling reaction comprising a compound of
Formula 8 and a compound of Formula 9 is performed at an elevated
temperature ranging from about 80.degree. C. to about 90.degree. C.
In other embodiments, the coupling reaction comprising a compound
of Formula 8 and a compound of Formula 9 is performed at a
temperature of about 85.degree. C. The times necessary for the
exemplified coupling reactions comprising a compound of Formula 8
and a compound f Formula 9, may vary with the identities of the
reactants, the solvent system and the chosen temperature.
[0051] In another embodiment, the present invention provides a
method for synthesizing a compound of Formula I:
##STR00012##
or a pharmaceutically acceptable salt thereof, wherein each of
R.sup.1 and R.sup.2 is independently alkoxy or haloalkoxy; R.sup.3
is H, F, Cl, I or Br; and R.sup.4 is F, .sup.18F, Cl, I or Br; the
method comprising:
[0052] i) reacting a compound of Formula 8 with a compound of
Formula 9 in the presence of a coupling reagent and applying
microwave radiation to the reaction to generate compound of Formula
I:
##STR00013##
[0053] In some embodiments, when the coupling reaction is heated
with microwave radiation, the coupling reaction times are shorter
than when performing the coupling reaction in the absence of
microwave heating. In some embodiments, the coupling reaction is
performed with applied microwave radiation at power levels ranging
from about 10 watts to about 50 watts. In other embodiments, the
coupling reaction is performed using microwave radiation heating
wherein the amount of microwave radiation ranges between about 10
watts to about 20 watts. The times necessary for the exemplified
coupling reactions comprising a compound f Formula 8 and a compound
of Formula 9, may vary with the identities of the reactants, the
solvent system and the chosen temperature.
[0054] In some implementations, the coupling reagent can include:
N,N'-dicyclohexylcarbodiimide (DCC), N,N'-diisopropylcarbodiimide
(DIC), 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDCI),
hydroxybenzotriazole (HOBt), 1-hydroxy-7-azabenzotriazole (HOAt),
benzotriazole-1-yl-oxytripyrrolidinophosphonium hexafluorophosphate
(BOP reagent), benzotriazol-1-yl-oxytripyrrolidinophosphonium
hexafluorophosphate (PyBOP),
1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium
3-oxid hexafluorophosphate (HATU),
O-(benzotriazol-1-yl)-N,N,N'N'-tetramethyluronium tetrafluoroborate
(TBTU), N,N,N',N'-tetramethyl-O-(1H-benzotriazol-1-yl)uranium
hexafluorophosphate (HBTU),
O-[(ethoxycarbonyl)cyanomethylenamino]-N,N,N',N'-tetramethyluronium
tetrafluoroborate (TOTU), and
(1-cyano-2-ethoxy-2-oxoethylidenaminooxy)dimethylamino-morpholino-carbeni-
um hexafluorophosphate (COMU, or a combination thereof.
[0055] In some implementations, the coupling reaction of a compound
of Formula 8 and a compound of Formula 9 in the presence of
microwave heating further comprises the addition of a tertiary
amine base. In some examples, the tertiary amine base can include:
diisopropylethyl amine (DIPEA), triethyl amine (TEA), N-methyl
imidazole, pyridine, 4-(dimethylamino)pyridine (DMAP),
3,4-lutidine, 4-methoxypyridine, N-methylmorpholine (NMO),
1,4-diazabicycle[2.2.2]octane (DABCO), and
1,8-diazacycloundec-7-ene (DBU), or a combination thereof.
[0056] In some implementations, the coupling reaction of a compound
of Formula 8 and a compound of Formula 9 in the presence of
microwave heating further comprises the addition of an aprotic
solvent. In some examples, the aprotic solvent can include:
acetonitrile, diethyl ether, diisopropyl ether, 2-methoxyethyl
ether, 1,2-dimethoxyethane, tert-butyl methyl ether,
tetrahydrofuran, 1,4-dioxane, benzene, toluene,
.alpha.,.alpha.,.alpha.-trifluorotolune, cyclohexane,
methylcyclohexane carbon tetrachloride, methylene chloride,
N,N-dimethylformamide, dimethyl sulfoxide, and
N-methyl-2-pyrrolidone, or a combination thereof.
[0057] In some implementations, the coupling reaction of a compound
of Formula 8 and a compound of Formula 9, is heated to a
temperature ranging from about 25.degree. C. to about 100.degree.
C., or from about 80.degree. C. to about 90.degree. C., or about
85.degree. C. using microwave radiation. The times necessary for
the exemplified coupling reactions comprising a compound of Formula
8 and a compound of Formula 9, may vary with the identities of the
reactants, the solvent system and the chosen temperature.
[0058] Another aspect of the present invention is directed to a
method for generating a compound of Formula I:
##STR00014##
or a pharmaceutically acceptable salt thereof wherein each of
R.sup.1 and R.sup.2 is independently alkoxy r haloalkoxy; R.sup.3
is H, F, CI, I or Br; and R.sup.4 is F, .sup.18F, CI, I or Br;
comprising:
[0059] i) reacting a compound of Formula 8 with a chlorinating or
brominating agent to generate acid halide 8a, wherein X is chloro
or bromo:
##STR00015##
and
[0060] ii) reacting the compound of Formula 8a with a compound of
Formula 9 in the presence of a base to generate a compound of
Formula (I) or a pharmaceutically acceptable salt thereof.
##STR00016##
[0061] In some implementations, the chlorinating or brominating
agent can include: thionyl chloride, thionyl bromide, oxalyl
chloride, phosphorus pentachloride and phosphorus trichloride. In
some embodiments, the chlorinating agent is oxalyl chloride.
[0062] In some implementations, the base can include: potassium
carbonate, sodium carbonate, sodium bicarbonate, triethyl amine
(TEA), diisopropylethyl amine (DIPEA), pyridine,
4-(dimethylamino)pyridine (DMAP), and N-methylmorpholine (NMO), or
a combination thereof. In some embodiments, the base is potassium
carbonate.
[0063] In some implementations, the above reactions can be
performed at temperatures ranging from about 20.degree. C. to about
40.degree. C. In some embodiments, each reaction is performed at
ambient temperature. The times necessary for the above exemplified
reactions may vary with the identities of the reactants, the
solvent system and the chosen temperature.
[0064] In one aspect, the present invention provides a method for
generating the compound of Formula 9a, wherein R.sup.4 is .sup.18F,
the method comprising:
[0065] i) reacting a compound of Formula 18 with a fluorinating
reagent to generate a compound of Formula 22a:
##STR00017##
and
[0066] ii) reducing the compound of Formula 22a to generate a
compound of Formula 9a:
##STR00018##
[0067] In some implementations, the fluorinating reagent is
K[.sup.18F] bound to a cryptand. In some examples, a suitable
cryptand is
1,10-diaza-4,7,13,16,21,24-hexaoxa-1,10-diazabicyclo[8.8.8]hexacosane
(Kryptofix.RTM. 2.2.2. or Kryptofix 2.2.2 or Kryptofix 222). In
some embodiments, the fluorinating reagent is Kryptofix
2.2.2/K.sup.18[F].
[0068] In some implementations, the fluorination reaction occurs in
the presence of a polar aprotic solvent. In some embodiments, the
polar aprotic solvent can include: N,N-dimethylformamide, dimethyl
sulfoxide, acetonitrile, N-methyl-2-pyrrolidone, tetrhydrofuran,
and 1,4-dioxane, or a combination thereof.
[0069] In some implementations, the compound of Formula 9a is
purified by passing through a series of columns packed with
adsorbents and/or ion exchange resins. In some embodiments, the
adsorbents can include: silica gel, neutral alumina, basic alumina,
octadecyl carbon chain (C18)-bonded silica column, C8-bonded
silica, cyano-bonded silica, phenyl-bonded silica, or a combination
thereof In other exemplary embodiments, the ion exchange resin can
include acidic or cation-exchange resins, for example, cation
ion-exchange resins containing sulphonate anions.
[0070] In some embodiments, the crude reaction mixture containing
the compound of Formula 9a, is further purified by passing the
crude reaction mixture through three Sep-Pak cartridges connected
in a series. In the first step, the crude reaction mixture is
passed through a basic alumina Sep-Pak cartridge to remove
unreacted fluorinating reagent (for example, K[.sup.18F]F. In the
second step, the resultant eluate is passed through a SCX Sep-Pak
cartridge to remove unreacted trimethylanilinium salt (18). In the
final step, the enriched eluate containing the compound of Formula
9a, is passed through a C-18 Sep-Pak cartridge to remove the
reaction solvent (for example, dimethyl sulfoxide). The C-18
Sep-Pak cartridge is subsequently washed with 0.1N hydrochloric
acid to remove any residual cryptand (for example, Kryptofix.RTM.).
In some embodiments, the final purification of the compound of
Formula 9a is accomplished by eluting the compound of Formula 9a
though the C-18 Sep-Pak cartridge with methanol.
[0071] In some implementations, the reduction of the nitro group of
the compound of Formula 22 is performed in the presence of a metal
catalyst, an acid, and hydrogen. In some embodiments, the metal
catalyst is derived from palladium, platinum, rhodium, or nickel.
In other embodiments, the metal catalyst is palladium black. In
some embodiments, the acid is mineral acid. More particularly, the
mineral acid is phosphorus acid. In some embodiments, the reduction
of the nitro group of the compound of Formula 22 is performed at
temperatures ranging from about 25.degree. C. to about 80.degree.
C. In some embodiments, the reduction of the nitro group of the
compound of Formula 22 is performed at 60.degree. C.
[0072] In another aspect, the present invention provides a method
for generating a compound of Formula 1a, or a pharmaceutically
acceptable salt thereof:
##STR00019##
wherein, R.sup.4 is F or .sup.18F; the method comprising:
[0073] i) reacting a compound of Formula 10 with a chlorinating
agent to generate a compound of Formula 11:
##STR00020##
[0074] ii) coupling the compound of Formula 11 with a compound of
Formula 23 in the presence of a base to generate a compound of
Formula 12:
##STR00021##
[0075] iii) coupling the compound of Formula 12 with a compound of
Formula 13 in the presence of a coupling reagent to generate a
compound of Formula 14:
##STR00022##
[0076] iv) saponifying the compound of Formula 14 in the presence
of a base to generate a compound of Formula 15:
##STR00023##
and
[0077] v) coupling the compound of Formula 15 with a compound of
Formula 9 in the presence of a coupling reagent to generate a
compound of Formula la or a pharmaceutically acceptable salt
thereof:
##STR00024##
[0078] In another aspect, the present invention provides a method
for generating a compound of Formula 1a, or a pharmaceutically
acceptable salt thereof:
##STR00025##
wherein R.sup.4 is F or .sup.18F; the method comprising:
[0079] i) reacting a compound of Formula 10 with a chlorinating
agent to generate a compound of Formula 11:
##STR00026##
[0080] ii) coupling the compound of Formula 11 with a compound of
Formula 23 in the presence of a base to generate a compound of
Formula 12:
##STR00027##
[0081] iii) coupling the compound of Formula 12 with a compound of
Formula 13 in the presence of a coupling agent to generate a
compound of Formula 14:
##STR00028##
[0082] iv) saponifying a compound of Formula 14 in the presence of
a base to generate a compound of Formula 15:
##STR00029##
[0083] v) reacting the compound of Formula 15 with a halogenating
reagent to generate a compound of Formula 15a;
##STR00030##
wherein
[0084] X is chloro or bromo; and vi) reacting the compound of
Formula 15a with a compound of Formula 9 to generate a compound of
Formula 1a, or a pharmaceutically acceptable salt thereof:
##STR00031##
Illustrative Synthetic Schemes
[0085] Illustrative synthetic routes to prepare a compound of
Formula I shown and described herein are exemplary only and are not
intended, nor are they to be construed, to limit the scope of the
present invention in any manner whatsoever. Those skilled in the
art will be able to recognize modifications of the disclosed
synthetic schemes and to devise alternate routes based on the
disclosed examples provided herein; all such modifications and
alternate routes are within the scope of the claims.
##STR00032##
[0086] In illustrative Scheme 1, the compound of Formula 2 wherein
R.sup.1 and R.sup.2 are as defined above can be converted to a
compound of Formula 3, wherein LG represents a leaving group.
Non-limiting examples of leaving groups that can be used include
halo groups (e.g., Cl, Br, or F) that can be added by halogenating
agents such as SOCl.sub.2, SO2Cl.sub.2, COCl.sub.2, PCl.sub.5,
POCl.sub.3, and the like. The reaction is advantageously carried
out under suitable reaction conditions. Non-limiting examples of
suitable reaction conditions in Scheme 1 can include the use of
suitable solvents. Non-limiting example of suitable solvents that
can be used during the halogenation of the compound of Formula 2
include a polar, aprotic solvent, such as CH.sub.3CN, DMF, and the
like, or mixtures thereof. In other embodiments, the chlorination
can be carried out using POCl.sub.3 in acetonitrile, COCl.sub.2 in
DMF, or SOCl.sub.2 in DMF. The addition of the chlorination agent
is advantageously carried out at a temperature ranging from about
60.degree. C. to about 90.degree. C. In another embodiment, the
addition of the chlorination agent can be carried out at a
temperature ranging from about 70.degree. C. to about 85.degree. C.
In another embodiment, the addition of the chlorination agent can
be carried out at a temperature ranging from about 74.degree. C. to
about 80.degree. C. The product can then be collected by filtration
and purified using standard techniques.
##STR00033##
[0087] In illustrative Scheme 2, a compound of Formula 3 wherein
R.sup.1 and R.sup.2 are as defined above, is reacted with a
substituted 4-aminophenol 4 wherein R.sup.3 is defined above, to
generate a compound of Formula 5. A non-limiting example of a
leaving group (LG) includes a halo group such as Cl, Br, or F.
Various compounds of Formula 4 are commercially available, such as
2-fluoro-4-aminophenol and 4-aminophenol. Also, the skilled artisan
would be able to make any variation of a compound of Formula 4
using commercially available starting materials and by using known
techniques to modify these commercially available starting
materials to yield various compounds within the scope of a compound
of Formula 4.
[0088] The Scheme 2 reaction in this embodiment is advantageously
carried out under suitable reaction conditions. Non-limiting
examples of suitable reaction conditions include using suitable
solvents such as polar solvents. Non-limiting examples of polar
solvents that can be used include tetrahydrofuran (THF),
dimethylacetamide (DMA), dimethylsulfoxide (DMSO),
dimethylformamide (DMF), ethyl acetate, N-methyl pyrrolidone (NMP),
propylene carbonate, and the like, or mixtures thereof. In another
embodiment, the polar solvent is dimethylacetamide (DMA). In
another embodiment, the polar solvent is dimethylsulfoxide (DMSO).
In another embodiment, the polar solvent is dimethylformamide
(DMF). In another embodiment, the polar solvent is ethyl acetate.
In another embodiment, the polar solvent is N-methyl pyrrolidone
(NMP). In another embodiment, the polar solvent is propylene
carbonate. In another embodiment, the solvent is a mixture of
solvents, such as a mixture comprising THF and DMA.
[0089] The reactant compounds of Formulas 3 and 4 can be added
together at a temperature ranging from about I0.degree. C. to about
30.degree. C., or alternatively, from about 15.degree. C. to about
28.degree. C., or alternatively, from about 20.degree. C. to about
25.degree. C. The mixture is then heated to a temperature ranging
from about 80.degree. C. to about 125.degree. C., or alternatively,
from about 95.degree. C. to about 110.degree. C., or alternatively,
from about 100.degree. C. to about 105.degree. C., and the selected
temperature is maintained until the reaction is complete.
[0090] Other non-limiting examples of suitable reaction conditions
in Scheme 2 include the use of a suitable base, such as a metal
hydroxide or a non-nucleophilic base. Examples of metal hydroxides
include sodium hydroxide or potassium hydroxide. Non-limiting
examples of non-nucleophilic bases that can be used include lithium
diisopropylamide, lithium tetramethylpiperidide, and alkali metal
alkoxides such as sodium tert-butoxide, potassium tert-butoxide,
sodium-pentoxide, and the like, or mixtures thereof. Preferably,
the base is sodium tert-butoxide or sodium tert-pentoxide. In one
embodiment, the base is sodium tert-pentoxide. Typically the sodium
tert-pentoxide is commercially available as 35 weight percent
solution of base in tetrahydrofuran, or as a 95 weight percent
solid reagent. Preferably, the sodium tert-pentoxide is a 95 weight
percent solid.
[0091] Typically, approximately 1.1 to 3.0 molar equivalents of
base are used relative the moles of compound of Formula 3 that are
used. More preferably, 1.3 to 2.5 molar equivalents of base are
used relative the moles of 3 that are used. More preferably. 1.5 to
2.2 molar equivalents of base are used relative to the moles of
compound of Formula 3 that are used. More preferably, 1.7 to 2.1
molar equivalents of base are used relative to the moles of
compound of Formula 3 that are used.
[0092] Typically, the amount of molar equivalents of amino phenol
that are used exceeds the molar equivalents of base that are used.
In one embodiment, 1.1 to 2 molar equivalents of amino phenol are
used relative to the molar equivalents of base that are used.
[0093] Once the reaction is substantially complete, the reaction
mixture can be cooled to a temperature ranging from about
10.degree. C. to about 25.degree. C. Precooled water can be charged
at a rate to maintain a temperature that ranges from about
5.degree. C. to about 35.degree. C. Alternatively, the precooled
water can be charged at a rate to maintain a temperature that
ranges from about 10.degree. C. to about 25.degree. C. As a
non-limiting example, the precooled water can be at a temperature
ranging from about 0.degree. C. to about 10.degree. C. As another
non-limiting example, the precooled water can be at a temperature
ranging from about 2.degree. C. to about 7.degree. C. The
precipitate can be collected by filtration under standard
conditions and purified by standard purification techniques.
##STR00034##
[0094] In illustrative Scheme 3, an amine compound of Formula 5
wherein R.sup.1 and R.sup.2 are as defined above, is coupled with
1-(methoxycarbonyl)cyclopropanecarboxylic acid 6 in the presence of
a coupling reagent to generate an amide compound of Formula 7.
Examples of suitable coupling reagents include:
N,N'-dicyclohexylcarbodiimide (DCC), N,N'-diisopropylcarbodiimide
(DIC), 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDCI),
hydroxybenzotriazole (HOBt), 1-hydroxy-7-azabenzotriazole (HOAt),
benzotriazole-1-yl-oxytripyrrolidinophosphonium hexafluorophosphate
(BOP reagent) benzotriazol-1-yl-oxytripyrrolidinophosphonium
hexafluorophosphate (PyBOP),
1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium
3-oxid hexafluorophosphate (HATU),
O-(benzotriazol-1-yl)-N,N,N'N'-tetramethyluronium tetrafluoroborate
(TBTU), N,N,N',N'-tetramethyl-O-(1H-benzotriazol-1-yl)uranium
hexafluorophosphate (HBTU),
O-[(ethoxycarbonyl)cyanomethylenamino]-N,N,N',N'-tetramethyluronium
tetrafluoroborate (TOTU).
(1-cyano-2-ethoxy-2-oxoethylidenaminooxy)dimethylamino-morpholino-carbeni-
um hexafluorophosphate (COMU) or a combination thereof. Suitable
reaction solvents include, but are not limited to aprotic solvents.
Suitable examples of aprotic solvents useful in the reaction shown
in Scheme 3, can include: acetonitrile, diethyl ether, diisopropyl
ether, 2-methoxyethyl ether, 1,2-dimethoxyethane, tert-butyl methyl
ether, tetrahydrofuran (THF), 1,4-dioxane, benzene, toluene,
.alpha.,.alpha.,.alpha.-trifluorotolune, cyclohexane,
methylcyclohexane carbon tetrachloride, methylene chloride (DCM),
N,N-dimethylformamide (DMF), dimethyl sulfoxide (DMSO),
N-methyl-2-pyrrolidone (NMP) or a combination thereof. The times
necessary for the exemplified coupling reactions comprising a
compound of Formula 5 and a compound of Formula 6, may vary with
the identities of the reactants, the solvent system and the chosen
temperature. Exemplary reaction times can range from about 2 hours
to about 10 hours. In some embodiments, the reaction time is about
5 hours. The reaction can be performed at temperatures ranging from
about 20.degree. C. to about 30.degree. C.
##STR00035##
[0095] In illustrative Scheme 4, the ester compound of Formula 7
wherein R.sup.1 and R.sup.2 are as defined above, is saponified in
the presence of an alkaline or an alkaline earth metal hydroxide to
generate an acid compound of Formula 8. Examples of alkaline or
alkaline metal hydroxides can include: sodium hydroxide, lithium
hydroxide, caesium hydroxide, and potassium hydroxide. Suitable
solvents can include methanol, ethanol, isopropanol, isopropanol,
water or a combination thereof. The times necessary for the
exemplified saponification reaction comprising a compound of
Formula 7 may vary with the identities of the reactants, the
solvent system and the chosen temperature. Exemplary reaction times
can range from about 5 hours to about 32 hours. In some
embodiments, the reaction time is about 24 hours. The reaction can
be performed at temperatures ranging from about 20.degree. C. to
about 45.degree. C.
##STR00036##
[0096] In illustrative Scheme 5, the coupling of the acid compound
of Formula 8 wherein R.sup.1 and R.sup.2 are as defined above, with
the aniline compound of Formula 9, wherein R.sup.4 is F, .sup.18F,
I, Cl, or Br, can occur in the presence of a coupling reagent to
generate the amide compound of Formula I. Examples of suitable
coupling reagents include: N,N'-dicyclohexylcarbodiimide (DCC),
N,N'-diisopropylcarbodiimide (DIC),
1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDCI),
hydroxybenzotriazole (HOBt), 1-hydroxy-7-azabenzotriazole (HOAt),
benzotriazole-1-yl-oxytripyrrolidinophosphonium hexafluorophosphate
(BOP reagent) benzotriazol-1-yl-oxytripyrrolidinophosphonium
hexafluorophosphate (PyBOP),
1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium
3-oxid hexafluorophosphate (HATU),
O-(benzotriazol-1-yl)-N,N,N'N'-tetramethyluronium tetrafluoroborate
(TBTU), N,N,N',N'-tetramethyl-O-(1H-benzotriazol-1-yl)uranium
hexafluorophosphate (HBTU),
O-[(ethoxycarbonyl)cyanomethylenamino]-N,N,N',N'-tetramethyluronium
tetrafluoroborate (TOTU),
(1-cyano-2-ethoxy-2-oxoethylidenaminooxy)dimethylamino-morpholino-carbeni-
um hexafluorophosphate (COMU) or a combination thereof.
[0097] Suitable solvents for the use in the coupling reaction of
Scheme 5, can include, but are not limited to aprotic solvents.
Examples f aprotic solvents include: acetonitrile, diethyl ether,
diisopropyl ether, 2-methoxyethyl ether, 1,2-dimethoxyethane,
tert-butyl methyl ether, tetrahydrofuran (THF), 1,4-dioxane,
benzene, toluene, .alpha.,.alpha.,.alpha.-trifluorotolune,
cyclohexane, methylcyclohexane carbon tetrachloride, methylene
chloride (DCM), N,N-dimethylformamide (DMF), dimethyl sulfoxide
(DMSO), N-methyl-2-pyrrolidone (NMP) or a combination thereof.
[0098] The coupling reaction of Scheme 5 can be assisted by heating
the reactants using microwave radiation ranging from about 10 watts
to about 50 watts. In some embodiments, the coupling reaction of
Scheme 5 can proceed in the presence of a base, for example:
diisopropylethyl amine (DIPEA), triethyl amine (TEA), N-methyl
imidazole, pyridine, N,N-dimethylamino-4-pyridine (DMAP),
34-lutidine, 4-methoxypyridine (NMO),
1,4-diazabicycle[2.2.2]octane(DABCO), 1,8-diazabicyloundec-7-ene
(DSU) or a combination thereof.
[0099] The coupling reaction of Scheme 5 can be heated to a desired
temperature varying from about 25.degree. C. to about 100.degree.
C. In some embodiments, the coupling reaction of Scheme 5 can be
heated to the desired temperature using microwave radiation. In
some embodiments, the coupling reaction when heated to a
temperature ranging from about 25.degree. C. to about 100.degree.
C. using microwave radiation results in shorter reaction times
compared to the coupling reaction without microwave heating. The
reaction times necessary for the exemplified coupling reaction
comprising a compound of Formula 8 and a compound of Formula 9 may
vary with the identities of the reactants, the solvent system and
the chosen temperature. In some embodiments, the reactants of the
coupling reaction of Scheme 5 can be heated using about 10 watts to
about 20 watts of microwave radiation to reach the desired
temperature of from about 25.degree. C. to about 100.degree. C.,
which provides a higher yield of the desired product.
##STR00037##
[0100] In illustrative Scheme 6, the acid compound of Formula 8,
wherein R.sup.1 and R.sup.2 are as defined above, is reacted with a
halogenating agent to generate the corresponding acid halide
compound 8a, wherein X is chloro or bromo. Examples of suitable
halogenating agents include: thionyl chloride, thionyl bromide,
oxalyl chloride, phosphorus pentachloride or phosphorus
trichloride. Suitable solvents for the reaction include diethyl
ether, diisopropyl ether, 2-methoxyethyl ether,
1,2-dimethoxyethane, tert-butyl methyl ether, tetrahydrofuran
(THF), 1,4-dioxane, benzene, toluene,
.alpha.,.alpha.,.alpha.-trifluorotolune, cyclohexane,
methylcyclohexane or a combination thereof. The reaction times
necessary for the exemplified halogenation reaction of Scheme 6
comprising a compound of Formula 8 may vary with the identities of
the reactants, the solvent system and the chosen temperature. In
some embodiments, the reaction of Scheme 6 may proceed at a desired
temperature ranging from about 20.degree. C. to about 25.degree.
C.
##STR00038##
[0101] In illustrative Scheme 7, the compound of Formula 8a,
wherein R.sup.1 and R.sup.2 are as defined above and X is chloro or
bromo, is coupled with an aniline compound 9 in the presence of a
base to yield a compound of Formula I. Suitable bases for use in
the reaction of Scheme 7 can include potassium carbonate, sodium
carbonate, sodium bicarbonate, triethyl amine (TEA), diisopropyl
ethyl amine (DIPEA), pyridine, N,N-dimethylamino-4-pyridine (DMAP)
or N-methylmorpholine (NMO). Suitable solvents include water,
ethanol, isopropanol, dimethyl sulfoxide or a combination thereof.
The reaction times necessary for the exemplified coupling reaction
of Scheme 7 comprising a compound of Formula 8a and a compound of
Formula 9 may vary with the identities of the reactants, the
solvent system and the chosen temperature. In some embodiments, the
reaction of Scheme 7 may proceed at a desired temperature ranging
from about 20.degree. C. to about 25.degree. C.
EXAMPLES
[0102] Additional embodiments are disclosed in further detail in
the following examples, which are not in any way intended to limit
the scope of the claims.
Materials and General Methods
[0103] [.sup.18F]-Fluoride was purchased from P.E.T. Net.RTM.
Pharmaceuticals. Inc. All reagents were obtained from Aldrich
Chemical Company or Lancaster.RTM. and were either ACS grade or of
the highest quality material commercially available. Zorbax C18
analytical, semi-preparative HPLC columns, 25 mm (0.45 .mu.m) nylon
syringe filters (Pall P.N. 4438T), Merck LiChrolut.RTM. SCX (P.N.
48219-242), and Merck LiChrolut.RTM. EN cartridges (P.N. 48219-232
200 mg) were obtained from VWR Inc. Basic alumina light (Waters
P.N. WAT023555, 280 mg), C18-plus (Waters P.N. WAT 020515, 360 mg),
and QMA light (Waters P.N. WAT023525, 130 mg) were obtained from
Waters. Microvials (5 ml) were obtained from Kontes. The microwave
heating model RI 520A was obtained from Resonance Instruments Inc.
(Skokie, Ill.). Mass spectra were obtained with a Finnigan TSQ or a
Finnigan LCQ mass spectrometer. Proton NMR spectra were recorded on
a Jeol EC+500 MHz NMR.
[0104] All reported radiochemical yields were decay corrected to
the start of the radiochemical synthesis. HPLC purification and
analysis was performed on a Varian Prostar HPLC system consisting
of two pumps, a Varian UV detector and a Lab Logic y-RAM
radioactive flow through detector. Radiochemical purity was
determined by analytical HPLC. System A, in this system analytical
samples were loaded into a Zorbax SB C18 column (4.6.times.250 mm)
with a mobile phase of 50% MeCN and 50% 25 mM potassium phosphate
dibasic solution (pH 9.0) at a flow rate of 1 ml/min. The UV
detector was set at 254 nm. System B, In this system, analytical
samples were loaded into a Luna C-8(2) column (4.6.times.150 mm)
with a gradient program using a mobile phase that went from 5% MeCN
and 95% 0.1% TFA at a flow rate of 1 ml/min at time 0 min to 95%
MeCN and 5% 0.1% TFA at a flow rate of 1 ml/min at time 30 min. The
UV detector was set at 254 nm. RadioTLC was completed on a Bioscan
AR2000 using uniplate--Silica Gel GHLF, scored 10.times.20 cm, 250
microns TLC plates and an 8% methanol in dichloromethane as the
solvent.
Example 1
Synthesis of 4-chloro-6,7-dimethoxyquinolone (11) from
6,7-Dimethoxy-quinolin-4-ol (10)
##STR00039##
[0106] A reactor was charged sequentially with
6,7-dimethoxy-quinoline-4-ol (47.0 kg) and acetonitrile (318.8 kg).
The resulting mixture was heated to approximately 60.degree. C.,
and phosphorus oxychloride (POCl.sub.3, 130.6 kg) was added. After
the addition of POCl.sub.3, the temperature of the reaction mixture
was raised to approximately 77.degree. C. The reaction was deemed
complete (approximately 13 hours) when less than 3% of the starting
material remained (in-process high-performance liquid
chromatography [HPLC] analysis). The reaction mixture was cooled to
approximately 2-7.degree. C. and then quenched into a chilled
solution of dichloromethane (DCM, 482.8 kg), 26% NH.sub.4OH (251.3
kg), and water (900 L). The resulting mixture was warmed to
approximately 20-25.degree. C., and phases were separated. The
organic phase was filtered through a bed of AW hyflo super-cel NF
(Celite; 5.4 kg) and the filter bed was washed with DCM (118.9 kg).
The combined organic phase was washed with brine (282.9 kg) and
mixed with water (120 L). The phases were separated and the organic
phase was concentrated by vacuum distillation with the removal of
solvent (approximately 95 L residual volume). DCM (686.5 kg) was
charged to the reactor containing organic phase and concentrated by
vacuum distillation with the removal f solvent (approximately 90 L
residual volume). Methyl t-butyl ether (MTBE, 226.0 kg) was then
charged and the temperature of the mixture was adjusted to -20 to
-25.degree. C. and held for 2.5 hours resulting in solid
precipitate which was then filtered and washed with n-heptane (92.0
kg), and dried on a filter at approximately 25.degree. C. under
nitrogen to afford the title compound. (35.6 kg).
Example 2
Synthesis of 4-(6,7-dimethoxyquinolin-3-yloxy)aniline (12)
##STR00040##
[0108] 4-Aminophenol (24.4 kg) dissolved in N,N-dimethylacetamide
(DMA, 184.3 kg) was charged to a reactor containing
4-chloro-6,7-dimethoxyquinoline (35.3 kg), sodium t-butoxide (21.4
kg) and DMA (167.2 kg) at 20-25.degree. C. This mixture was then
heated to 100-105.degree. C. for approximately 13 hours. After the
reaction was deemed complete as determined using in-process HPLC
analysis (<2% starting material remaining), the reactor contents
were cooled at 15 to 20.degree. C. and water (pre-cooled, 2 to
7.degree. C., 587 L) charged at a rate to maintain 15 to 30.degree.
C. temperature. The resulting solid precipitate was filtered,
washed with a mixture of water (47 L) and DMA (89.1 kg) and finally
with water (214 L). The filter cake was then dried at approximately
25.degree. C. on filter to yield crude
4-(6,7-dimethoxy-quinoline-4-yloxy)-phenylamine (59.4 kg wet, 41.6
kg dry calculated based on LOD). Crude
4-(6,7-dimethoxy-quinoline-4-yloxy)-phenylamine was refluxed
(approximately 75.degree. C.) in a mixture of tetrahydrofuran (THF,
211.4 kg) and DMA (108.8 kg) for approximately 1 hour and then
cooled to 0-5.degree. C. and aged for approximately 1 h after which
time the solid was filtered, washed with THF (147.6 kg) and dried
on a filter under vacuum at approximately 25.degree. C. to yield
4-(6,7-dimethoxyquinolin-3-yloxy)aniline (34.0 kg).
Example 3
Synthesis of methyl
1-(4-(6,7-dimethoxyquinolin-3-yloxy)phenylcarbamoyl)cyclopropanecarboxyla-
te (14)
##STR00041##
[0110] 1-(Methoxycarbonyl)cyclopropanecarboxylic acid (13, 0.2 g,
1.2 mmol), 1,3-diisopropylcarbodiimide (0.2 mL, 1.2 mmol), and
1-hydroxybenzotriazole monohydrate (0.2 g, 1.2 mmol) were added to
a 25 mL round bottom flask and dissolved in DMF (3.2 mL). This
reaction mixture was allowed to stir at ambient temperature for 10
minutes. To it 4-(6,7-dimethoxyquinolin-3-yloxy)aniline (12, 0.3 g,
1.0 mmol) was added to reaction mixture, and allowed to stir at
ambient temperature for 4.5 hours. After this time period, 100 mL
of DI water was added and the reaction mixture was extracted with
3.times.50 mL of ethyl acetate. The organic layer was dried over
sodium sulfate and concentrated to afford a yellow/white solid. The
crude reaction mixture was purified via flash silica gel column
chromatography using ethyl acetate as the mobile phase. Methyl
1-(4-(6,7-dimethoxyquinolin-3-yloxy)phenylcarbamoyl)cyclopropaneca-
rboxylate (14, 0.4 g, 0.9 mmol, 89% yield) was obtained as white
solid. .sup.1H NMR (CDCl.sub.3, 500 MHZ) .delta. 10.95 (s, 1H);
8.45 (d, 1H, J=5.5 Hz); 7.67 (d, 2H, J=6.6 Hz); 7.54 (s, 1H); 7.41
(s, 1H); 7.15 (d, 2H, J=6.6 Hz); 6.43 (d, 1H, J=5.5 Hz); 4.04 (s,
6H); 3.75 (s, 3H); 1.85-1.82 (m, 2H), 1.71-1.69 (m, 2H); LCMS (m/z)
C.sub.23H.sub.23N.sub.2O.sub.6 (M+H) Calcd: 423.16; Found: 423.1;
.sup.13C NMR (CDCl.sub.3, 125 mHz) .delta. 174.4, 166.9, 160.9,
152.9, 150.4, 149.5, 148.8, 146.8, 135.6, 121.9, 121.6, 116.1,
107.8, 103.3, 99.5, 56.1, 52.5, 26.5, 20.9.
Example 4
Synthesis of
1-(4(6,7-dimethoxyquinolin-3-yloxy)phenylcarbamoyl)cyclopropanecarboxylic
acid (15)
##STR00042##
[0112] Methyl
1-(4-(6,7-dimethoxyquinolin-3-yloxy)phenylcarbamoyl)cyclopropanecarboxyla-
te (14, 0.3 mg, 0.6 mmol) was dissolved in methanol (5.0 mL) and
heated at 35-45.degree. C., sodium hydroxide (1.0 N, 1.0 mL, 1.0
mmol) was then added to the reaction mixture and was allowed to
stir at 35-45.degree. C. over a 24 hour period. The reaction was
then concentrated to afford the
1-(4-(6,7-dimethoxyquinolin-3-yloxy)phenylcarbamoyl)
cyclopropanecarboxylic acid (15, 0.2 g, 0.5 mmol, 88% yield) as a
light yellow solid. The crude product was taken up in Dl water (5.0
mL) and the pH of the solution was adjusted to 3 with concentrated
sulfuric acid (0.5 ml, 1.0 mmol), yielding a white precipitate. The
white precipitate was filtered, washed with 3 portions of DI water
(5.0 mL) and then lyophilized over a 24 hour period and finally
dried in a vacuum oven at 70.degree. C. for 24 hours to afford
1-(4-(6,7-dimethoxyquinolin-3-yloxy)phenylcarbamoyl)cyclopropanecarboxyli-
c acid (15, 0.2 mg, 0.5 mmol, 88% yield). HRMS (m/z)
C.sub.22H.sub.21N.sub.2O.sub.6 (M+H) Calcd 409.13992; found
409.14007; .sup.1H NMR (d.sub.6-DMSO, 500 MHZ) .delta. 8.64 (d, 1H,
J=6.6 Hz); 7.83 (s, 1H); 7.81 (d, 2H, J=8.8 Hz); 7.44 (s, 1H); 7.33
(d, 2H, J=8.8 Hz); 6.91 (d, 1H, 6.6 Hz); 4.13 (s, 3H); 4.08 (s,
3H); 1.83-1.68 (m, 4H); .sup.13C NMR (d.sub.6-DMSO, 125 mHz)
.delta. 168.3, 163.7, 158.5, 154.3, 153.5, 150.1, 143.4, 138.8,
135.9, 123.6, 122.7, 117.5, 108.2, 101.6, 100.2, 57.4, 57.1, 20.9,
20.2.
Example 5
Preparation of
1-[4-(6,7-dimethoxy-quinolin-4-yloxy)-phenylcarbamoyl]-cyclopropanecarbox-
ylic Acid (15)
##STR00043##
[0114] Preparation of
1-[4-(6,7-dimethoxy-quinolin-4-yloxy)-phenylcarbamoyl]-cyclopropanecarbox-
ylic acid 15. To the cyclopropyl di-carboxylic acid 12a (449 mg,
3.45 mmol) in THF (3.5 mL) was added TEA (485 .mu.L, 3.45 mmol).
The resulting solution was stirred at room temperature under a
nitrogen atmosphere for 40 minutes before adding thionyl chloride
(250 .mu.L, 3.44 mmol). The reaction was monitored by LCMS for the
formation of mono acid chloride 12 (quenched the sample with MeOH
and looked for corresponding mono methyl ester). After 3 hours
stirring at room temperature,
4-(6,7-dimethoxy-quinolin-4-yloxy)-phenylamine 16 (1.02 g, 3.44
mmol) was added as a solid, followed by more THF (1.5 mL).
Continued to stir at room temperature for 16 hours. The resulting
thick slurry was diluted with EtOAc (10 mL) and extracted with IN
NaOH. The biphasic slurry was filtered and the aqueous phase was
acidified with conc. HCl to pH=6 and filtered. Both solids were
combined and washed with EtOAc, then dried under vacuum. The
desired product,
1-[4-(6,7-dimethoxy-quinolin-4-yloxy)-phenylcarbamoyl]-cyclopropanecarbox-
ylic acid, 15 was obtained (962 mg, 68.7% yield, 97% pure) as a
white solid. .sup.1H NMR (D.sub.2O/NaOH): 7.97 (d, 1H), 7.18 (d,
2H), 6.76 (m, 4H), 6.08 (d, 1H), 3.73 (s, 3H), 3.56 (s, 3H), 1.15
(d, 4H).
Example 6
Synthesis of Cabozantinib (1)
##STR00044##
[0116] Method A: Cabozantinib (1) was obtained via a
microwave-assisted reaction with 4-fluoroaniline, HATU, DIPEA, and
1-(4-(6,7-dimethoxyquinolin-4-yloxy)phenylcarbamoyl)cyclopropanecarboxyli-
c acid (15) in 80% isolated yields. Using a capped microvial in the
microwave apparatus, microwave powers of greater than 50 W led to
vigorous refluxing of these solutions; therefore, powers of 10-50 W
were investigated. It was determined that using lower wattages of
10-20 W, allowed the reaction temperature to reach 85.degree. C.
over a 20 minute period.
[0117] Method B: To a solution of
1-(4-(6,7-dimethoxyquinolin-4-yloxy)phenylcarbamoyl)cyclopropanecarboxyli-
c acid (15, 123 mg, 0.30 mmol), 4-fluoroaniline (40 mg, 0.36 mmol),
DIPEA (234 mg, 1.81 mmol) and DMF (3 mL) was added
(benzotriazol-1-yloxy)tripyrrolidinophosphonium hexafluorophosphate
(PyBOP, 470 mg, 0.90 mmol) with stirring at 25.degree. C. The
resulting solution was stirred at 25.degree. C. for 1 h. The
mixture was diluted with EtOAc, washed with 0.2 N NaOH, and brine,
dried with MgSO.sub.4. Removal of EtOAc and column chromatography
gave the desired product 1 (133 mg, 88% yield). .sup.1H NMR
(DMSO-d.sub.6): 10.20 (s, 1H), 10.07 (s, 1H), 8.47 (d, 1H), 7.77
(d, 2H), 7.65 (m, 2H), 7.50 (s, 1H), 7.39 (s, 1H), 7.24 (dd, 2H),
7.16 (m, 2), 6.42 (d, 1H), 3.94 (s, 3H), 3.93 (s, 3H), 1.47 (s, 4H)
ppm. LC/MS: Calcd for [M+H].sup.+502.2, found 502.2. Anal. HPLC (10
min gradient): 97.5% purity, 6.62 min.
Example 7
Synthesis of Cabozantinib
##STR00045##
[0119] To a mixture of the carboxylic acid (15, 123 mg, 0.30 mmol),
THF (1.0 mL), and DMF (5 .mu.L) was added oxalyl chloride (38 mg;
0.30 mmol) dropwise at room temperature. After 15 min, the acid
chloride (15a) slurry was added to another flask containing a
stirred suspension of 4-fluoroaniline (37 mg, 0.33 mmol),
K.sub.2CO.sub.3 (104 mg, 035 mmol), in THF (1.0 mL) and water (0.5
mL) over approximately 2 min. After 45 min, the lower aqueous layer
was removed. The upper organic layer was concentrated in vacuo and
purified by column chromatography to give the desired product 1
(108 mg, 72% yield). LC/MS: Calcd for [M+H].sup.+ 502.2, found
502.2. Anal. HPLC (10 min gradient): 99% purity, 6.62 min.
Example 8
Synthesis of 4-nitro-N,N,N-trimethylanilinium
trifluoromethanesulfonate (18)
##STR00046##
[0121] N,N-dimethyl-4-nitroaniline (17, 0.7 g, 4.0 mmol) and
benzene (10 mL) were added to a nitrogen-purged flask equipped with
a stir bar. To this stirring solution was added methyl
trifluoromethanesulfonate (0.7 mL, 6.1 mmol) at ambient
temperature. The reaction was then heated to 40.degree. C. for a
24-hour period. At the conclusion of the reaction the
N,N,N-trimethyl-4-nitrobenzenaminium trifluoromethanesulfonate (18,
1.0 g, 3.0 mmol, 75% yield) precipitated from solution as an orange
solid. The product was filtered, washed with ether (3.times.20 mL)
and placed in a high vacuum drying tube at 100.degree. C. for 1
hour. LCMS C.sub.9H.sub.13N.sub.2O.sub.2 (M.sup.+) Calcd 181.10;
Found 181.1; .sup.1H NMR (d.sub.6-DMSO, 500 mHz) .delta. 8.45 (d,
2H, J=9.4 Hz); 8.26 (d, 2H, J=9.3 Hz); 3.66 (s, 12H); .sup.13C NMR
(d.sub.b-DMSO, 125 mHz) .delta. 151.0, 147.6, 124.9, 122.5,
56.24.
Radiochemical Synthesis
Example 9
Synthesis of K.2.2.2/K[.sup.18F]F Complex (19)
[0122] The aqueous [.sup.18F]-Fluoride solution (1.0 mL, 18.5
GBq/500 mCi) was purchased from P.E.T. Net.RTM. Pharmaceuticals in
West Point Pa. and shipped to the site. The average delivery time
at the end of bombardment was 80 minutes, and once delivered, was
transferred to our remote controlled synthesis apparatus and
delivered to a Sep-Pak light QMA [The Sep-Pak light QMA cartridge
was pre-conditioned sequentially with 5 mL of 0.5 M potassium
bicarbonate, 5 mL of deionized water, and 5 mL of MeCN before use].
Upon completion of this transfer, the aqueous [.sup.18F]-fluoride
was released from the QMA Sep-Pak by the sequential addition of
potassium carbonate (15 mg/mL; 0.1 mL) followed by a mixture of
potassium carbonate (30 mg/ml, 0.1 mL),
4,7,13,16,21,24-hexaoxa-1,10-diazabicyclo[8.8.8]hexacosane
(Kryptofix.RTM. 2.2.2., 15 mg, 0.04 mmol) and 1.2 mL of MeCN. The
solvent was evaporated under a gentle stream of nitrogen at
90.degree. C. and vacuum. Azeotropic drying was repeated twice with
1 mL portions of acetonitrile to generate the anhydrous Kryptofix
2.2.2/K[.sup.18F]F complex.
Example 10
Radiosynthesis of [.sup.18F]-fluoroaniline (9a) Using
4-nitro-N,N,N-trimethylanilinium trifluoromethanesulfonate (18)
##STR00047##
[0124] 4-Nitro-N,N,N-trimethylanilinium trifluoromethanesulfonate
(18) (5.0 mg, 0.02 mmol) was dissolved in 0.3 mL MSO and added to
the dried Kryptofix 2.2.2/K[.sup.18F]-F salt (19). The resultant
solution was heated at 120.degree. C. for 3 minutes. After heating,
the contents of this reaction vial were diluted with 10 mL of DI
water. This solution was passed through three Sep-Pak cartridges
that were connected in series. In the first step, the crude
reaction mixture is passed through a basic alumina light cartridge
followed by a Merck LiChrolut.RTM. SCX cartridge to remove
unreacted K[.sup.18F]F. In the second step, the reaction mixture is
passed through a SCX Sep-Pak cartridge to remove unreacted
trimethylanilinium salt (18). In the final step, the compound is
passed through C-18 Sep-Pak cartridge (Waters P.N. WAT 020515, 360
mg) that was pre-conditioned with 10 mL ethanol & 10 mL DI
water to remove the reaction solvent (for example, dimethyl
sulfoxide). The [.sup.18F]-1,4-fluoronitrobenzene (22a) was
retained onto C-18 Plus Sep-Pak. Residual Kryptofix 2.2.2 was
rinsed from the C-18 Plus Sep-Pak by an addition of 10 mL of 0.1 N
HCl. [.sup.18F]-1,4-fluoronitrobenzene was eluted from this Sep-Pak
via the addition of 2 mL of anhydrous methanol. The schematic
diagram showing the various steps of purification of
[.sup.18F]-1,4-fluoronitrobenzene is shown in FIG. 1. The eluted
methanol solution containing [.sup.18F]-1,4-fluoronitrobenzene was
transferred into a sealed 5 mL microvial that contained palladium
black (11.0 mg, 0.1 mmol), phosphorous acid (0.10 g, 1.3 mmol) and
conical stir bar. The resulting solution was then heated at
60.degree. C. for 15 minutes and then this reaction mixture was
filtered through a 25 mm, 0.45 .mu.m nylon membrane syringe filter
to remove any palladium black. The remaining solution was diluted
with 25 mL of 1 N NaOH and transferred to the Merck EN Sep-Pak
(this Sep-Pak was pre-activated with 5 mL of EtOH followed by 10 mL
1 N NaOH). 2 mL of DCM was added to the EN Sep-Pak to elute the
[.sup.18F]-4-fluoroaniline (9a) into the 5 mL microvial microwave
station. The volume of DCM was reduced with a gentle stream of
nitrogen and heated at 40.degree. C. until approximately 0.2 mL of
this solution remained. To this reaction mixture was added 0.5 mL
DMF and this mixture was azeotropic dried twice with 1 mL portions
of DCM to generate the anhydrous [.sup.18F]-fluoroaniline (9a) in
DMF. Attempts to reduce the volume of (9a) below 0.2 mL resulted in
significant volitization of the product and were subsequently
avoided.
Example 11
Radiosynthesis of [.sup.18F]-Cabozantinib,
Cyclopropane-1,1-dicarboxylic Acid
[4-(6,7-dimethoxy-quinolin-4-yloxy)-phenyl]-amide
(4-[.sup.18F]-fluoro-phenyl)amide) (1b)
##STR00048##
[0126] A mixture of
1-(4-(6,7-dimethoxyquinolin-3-yloxy)phenylcarbamoyl)
cyclopropanecarboxylic acid (10 mg, 0.024 mmol, 15),
2-(3H-[1,2,3]triazolo[4,5-b]pyridine-3-yl)-1,1,3,3-tetramethylisouronium
hexafluorophosphate(V) (19 mg, 0.05 mmol), and
N-ethyl-N-isopropylpropan-2-amine (20 .mu.L, 0.1 mmol) were
dissolved in 0.3 mL DMF and added to the 5 mL microvial containing
[.sup.18F]-4-fluoroaniline (9a). The reaction was then heated with
the microwave apparatus at 85.degree. C. with 10-20 W for 20
minutes. After this period of heating, 5.0 mL of 25 mM potassium
phosphate dibasic solution, 1.5 mL of acetonitrile were added to
the microvial and this solution was loaded onto a 9.4.times.250 mm,
5 micron Zorbax SB-C18 column using an isocratic mobile phase of
45% acetonitrile and 65% 25 mM potassium phosphate dibasic solution
(pH 9.0). The UV detector was set at 254 nm.
[.sup.18F]-Cabozantinib was isolated at the 30 minute mark using
these conditions. This sample was then diluted with 25 mL of 25 mM
potassium phosphate dibasic and the entire sample was loaded onto a
Waters Sep-Pak C18 Plus Short Cartridge, 360 mg Sorbent per
Cartridge, 55-105 .mu.m Particle Size (part number WAT020515) which
was preactivated with 5 ml of ethanol, followed by 10 mL of DI
water. After the entire solution was loaded onto the C18 Sep-Pak
the final product was released with 0.5 mL of ethanol to give 902.8
MBq/24.4 mCi of [.sup.18F]-Cabozantinib,
cyclopropane-1,1-dicarboxylic acid
[4-(6,7-dimethoxy-quinolin-4-yloxy)-phenyl]-amide
(4-C.sup.811-fluoro-phenyl)amide, 1b. This peak was confirmed to be
[.sup.18F]-Cabozantinib 1b via co-injection of reference standard
on analytical HPLC and radiochemical purity was >99% (FIG. 2).
In system A, [.sup.18F]-Cabozantinib 1b co-eluted with the
non-radioactive standard 1 and had a R.sub.f of 10.2-10.5 min and
in system B, [.sup.18F]-Cabozantinib 1b co-eluted with the
non-radioactive standard 1 and had a R.sub.f of 14.5-14.8 min. The
specific activity was measured using a Zorbax SB C18 HPLC column
using an isocratic mobile phase of 50% acetonitrile and 50% 25 mM
potassium phosphate dibasic in distilled water. The UV was
monitored at 254 nM and the flow rate was 1 mL/min, generating a
back pressure of 1100 PSI. The specific activity using this method
was measured against a 6 point calibration curve and determined to
be 42.2+/-10 GBq/.mu.N/.mu.mol (1.14+/-0.2 Ci/.mu.mol).
Example 12
Radiosynthesis [.sup.18F]-Cabozantinib: Variation of Microwave
Reaction Conditions
[0127] The coupling reaction as described in Example 11 was
conducted by varying microwave heating at various wattages and
solvents. The results are shown in Table 1. A significant amount of
precursor decomposition was noticed when the reactions conducted at
powers above 20 W. An increase in the isolated amount of
[.sup.18F]-Cabozantinib 1b was seen using DMF as the reaction
solvent and decreased when DMSO, THF, or a mixture of DMF/DMSO.
[.sup.18F]-Cabozantinib 1b was isolated (903 MBq+/-120 MBq; 24.4
mCi +/-3.2 mCi, n=10) in radiochemical yields (13%, decay
corrected) after HPLC purification in high radiochemical purity
(>99%). The specific activity was 42.2 GBq/.mu.mol+/-7.5
GBq/.mu.mol (1.14 Ci/.mu.mol+/-0.2 Ci/.mu.mol). The retention time
of this radioactive product was confirmed by co-injection of the
non-radioactive standard Cabozantinib as shown in FIG. 2, using two
separate analytical HPLC systems (Systems A and B).
TABLE-US-00002 TABLE 1 Investigation of [.sup.18F]-Cabozantinib
using microwave heating Radiochemical yield of Experiment Microwave
[.sup.18F]-Cabozantinib (1A) No. power Time Solvent (decay
corrected) 1 10 W 20 min. DMF:DMSO 1.1% 2 20 W 20 min DMF:DMSO 1.3%
3 20 W 20 min DMF 13.8% 4 20 W 20 min DMSO 0.0% 5 50 W* 10 min
DMF:DMSO 0.7% 6 10 W 20 min THF 0.0% *At 50 W, precursor
decomposition was observed.
Example 13
Radiometabolite Analysis in Plasma
[0128] 7.4-6.4 MBq (0.2-0.17 mCi) of [.sup.18F]-Cabozantinib was
injected into athymic BALB/C mice (Harlan) and these animals were
maintained on 1-2% isofluorane during injection, tracer uptake, and
blood sampling. 100 .mu.L aliquots samples of blood were collected
in lithium heparin coated tubes either from the contralateral tail
vein or the retro-orbital plexus at 15 and 60 minutes after
[.sup.18F]-Cabozantinib injection. These samples were centrifuged
for 10 minutes at 10,000 rpm for plasma separation at 4.degree. C.
washed with equal amounts of methanol. 10 .mu.L of the methanol
extract of plasma was co-spotted with non-radioactive reference
compound onto and across the TLC plates (Silica Gel GHLF; Scored
2.5.times.10 cm; 250 microns), blow-dried with warm air and
developed in a TLC chamber. The TLC solution used in this study was
8% methanol in dichloromethane which gave a R.sub.f of
approximately 0.55. After the development was completed, the TLC
plate was placed onto the radio-TLC scanner (Bioscan AR2000), which
scanned the TLC plate for 15 minutes. The peak areas were
calculated to give the metabolite profile of
[.sup.18F]-Cabozantinib.
Example 14
PET Imaging in Mice
[0129] Rodent imaging experiments were performed on a dedicated
small animal microPET.RTM. F120.TM. scanner (Siemens Preclinical
Solutions, Knoxville, Tenn.). Both static and dynamic PET imaging
studies were conducted on athymic BALB/C mice (Harlan) and
stereotactically implanted U87 tumor cells. The mice were
administered about 6.4-7.4 MBq (0.17-0.2 mCi) of
[.sup.18F]-Cabozantinib via the tail vein. The mice were
anesthetized (2% isofluorane in O.sub.2 at a flow rate of 1 mL/min)
during tracer injection and throughout the imaging period. After
injection, each mouse was laid in the microPET scanner. A 10
minutes transmission scan was acquired using a .sup.57Co point
source prior to either the 2 hour dynamic scan or the 45 min static
scan starting at about 15 minutes after injection. Mice were
visually monitored for breathing, and a heating pad was used to
maintain body temperature throughout the entire procedure. PET data
were reconstructed using an OSEM 2D algorithm into a
128.times.128.times.95 image matrix (resulting in final voxel
dimensions of 0.79.times.0.79.times.0.8 mm). Dead time, decay
correction, attenuation correction, and normalization were applied
to all PET data. Data were analyzed by manually drawing
standardized volumes of interest using the Siemens ASIPro VM
software package version 6.6.
Example 15
In vivo PET Imaging of [.sup.18F]-Cabozantinib (1b)
[0130] Dynamic PET acquisitions were carried out on 3 mice with
orthotopically implanted U87 tumor cells 19 days after cells were
stereotactically implanted and 2 tumor-free animals for a 2 hour
period. As can been seen in FIG. 3, the summed PET images of the
tumor-free control mice after injection of 0.62 MBq (0.17 mCi) of
[.sup.18F]-Cabozantinib, shows little to n [.sup.18F]-Cabozantinib
(SUV mean 0.19) uptake in the brain of these control animals. A
significant increase in [.sup.18F]-Cabozantinib uptake was observed
in the PET image of the orthotopically implanted U87 mice (SUV mean
0.50 (n=3)) giving a high 2.4-2.6 tumor: brain ratio measured in
these animals. MRI confirmed the location of these tumors. In all 3
animals with orthotopically implanted U87 tumor cells,
visualization of the tumor could be observed with this tracer.
These results suggest that this tracer does not cross the blood
brain barrier in tumor-free mice and has significant uptake in U87
orthopically implanted in mice. From time activity curves, a steady
accumulation of the tracer in the tumor area can be observed over
the 2 hour period, with a maximal tumor uptake 60 minutes post
injection.
[0131] FIG. 3 depicts Positron Emission Tomography (PET) images of
mice injected with [.sup.18F]-Cabozantinib. Frame shows PET image
of control mice, i.e., non-tumor bearing. Frame B shows PET image
of mice with stereotactically implanted U87 tumor cells. Images are
summed frames over 15-45 post injection of a 6.3 MBq (0.17 mCi) of
[.sup.18F]-Cabozantinib. A tumor:brain ratio, which is the ratio of
the amount of [.sup.18F]-Cabozantinib in the tumor compared to the
amount of [.sup.18F]-Cabozantinib in the brain in non-tumor
regions, of 2.6 to 2.8 was measured for U87 stereotactically
implanted tumor cells. Frame C shows MRI image of the same
stereotatically implanted U87 tumor bearing mouse as shown in Frame
B. Frame D shows PET image in the same animal, in the same view as
MRI (Frame C) to confirm tumor location and PET tracer uptake in
the tumor region.
Example 16
Stereotactic Intracranial Surgical Procedures
[0132] All animal procedures were done in accordance with
established policy and protocols. Athymic BALB/C mice (Harlan) were
anesthetized (2% isofluorane in O.sub.2 at a flow rate of 1 L/min)
and positioned in a stereotaxic atlas prior to surgery. A midline
cranial incision was made and bregma located. From bregma, the
coordinates 2 mm lateral.times.2 mm posterior were used, and a
small bore hole was made with a dental drill. Using a Hamilton
syringe (30 g) inserted to a depth of 3 mm, 2.5.times.10.sup.5 U87
cells (ATCC) in 10 .mu.L of RPMI were injected over a 3 minute
period. Bone wax (Ethicon) was used to seal the burr hole followed
by Veterinary Tissue Adhesive (3M) to close the incision. Mice were
recovered from anesthesia, observed for any signs of pains or
distress, and if necessary, provided appropriate veterinary care.
After 7 days, mice were initiated on imaging protocols to monitor
disease progression
Example 17
Biodistribution
[0133] Groups of athymic BALB/C mice were implanted with a
subcutaneous U87 tumor cells (N=6) were anesthetized, and
administered [.sup.18F]-Cabozantinib (4-10 MBq) in 100 .mu.L 10%
ethanol in saline. After acute sedation with isofluorane (2%)
animals were sacrificed by decapitation at 15 and 60 min following
tracer injection. Blood samples were collected and portions of the
liver, kidneys, heart, lungs, muscles, stomach, large intestine,
small intestine, bladder, spleen, brain and femur were dissected
and weighed. Radioactivity of each sample was measured using a
gamma counter (1480 Wizard.TM. 3 Automatic Gamma Counter,
PerkinElmer). Decay-corrected radioactivity concentrations were
calculated as percentage of injected dose per gram tissue (%
ID/g).
Biodistribution Data and in vivo Metabolism of
[.sup.18F]-Cabozantinib
Example 18
In viva Metabolism of [.sup.18F]-Cabozantinib (1b)
[0134] The results of the in vivo stability of
[.sup.18F]-Cabozantinib are summarized in FIG. 4.
[.sup.18F]-Cabozantinib was extracted in the plasma samples taken
from mice with no significant differences in metabolism of this
tracer in tumored or wild type mice. 10 .mu.L of methanol extract
of mouse plasma was co-spotted with non-radioactive reference
compound onto and across the TLC plates, which were blow-dried with
warm air and developed in TLC chamber. The TLC solution used in
this study is 8% methanol in dichloromethane which gave a R.sub.f
.about.0.55. After the development was completed, the TLC plate was
placed onto the radio-TLC scanner (Bioscan AR2000), which scanned
the TLC plate for 15 minutes. The peak areas were calculated to
give the metabolite profile of [.sup.18F]-Cabozantinib. The content
of the extracted radioactivity was analyzed on TLC and showed that
over 80% of this tracer remained intact over the time course of
this study. Only one radiometabolite was seen in this analysis and
was more polar than its parent suggesting that
[.sup.18F]-Cabozantinib is stable in vivo. The biodistribution of
[.sup.18F]-Cabozantinib in mice was investigated in both through
radioactivity assay of postmortem mouse tissues harvested on
completion of the imaging studies at 15 and 60 minutes after
injection (n =6).
[0135] FIG. 5 depicts biodistribution analysis of
[.sup.18F]-Cabozantinib in mice. The bar graph represents the
biodistribution (% ID/g) of [.sup.18F]-Cabozantinib in mice (n=6)
at 15 minutes post injection (blue) and 60 minutes post injection
(red). At the 15 minute mark after injection, the highest tracer
uptake was found in the liver (14.4.+-.3.1% ID/g), followed by the
kidneys (14.2.+-.3.0% ID/g), heart (12.9.+-.3.2% ID/g), bladder
(7.1.+-.6.5% ID/g), small intestines (7.0.+-.2.4% ID/g), lungs
(6.5.+-.1.1% ID/g, stomach (5.2.+-.1.7% ID/g), spleen (4.7.+-.1.2%
ID/g), large intestines (4.7.+-.2.1% ID/g), femur (2.9.+-.1.0%
ID/g) muscles (4.4.+-.1.1% ID/g), blood (1.8.+-.0.2% ID/g), and
brain (0.9.+-.0.2% ID/g). At the 60 minutes post injection time
point, the highest tracer uptake was found in the liver
(14.0.+-.1.9% ID/g), followed by the kidneys (11.7.+-.1.5% ID/g),
small intestines (10.7.+-.1.1% ID/g), heart (9.5.+-.1.5% ID/g),
stomach (8.4.+-.1.5% ID/g), lungs (4.9.+-.0.8% ID/g, bladder
(4.3.+-.1.2% ID/g), large intestines (4.6.+-.0.9% ID/g), muscles
(4.2.+-.0.3% ID/g), spleen (3.6.+-.0.6% ID/g), blood (2.2.+-.0.3%
ID/g), femur (1.8.+-.0.8% ID/g) and brain (0.9.+-.0.1% ID/g). These
results suggest that a minimal amount of radioactivity was seen in
the femur and fluorine locating to bone was not a significant issue
with this compound and thus supporting good stability of the
radiolabel. In addition, these results show that this compound did
not appear to cross the blood brain barrier.
Example 19
MRI
[0136] All MRI analyses were performed on a Bruker 7.0T/20 cm
horizontal Biospec MRI system (Bruker BioSpin, Ettlingen, Germany),
and a Bruker 25-mm quadrature mouse head coil was used for RF
transmitting/receiving. Mice anesthesia was induced by 2%
isofluorane in 100% O.sub.2 for tail vein catheter preparation.
Gadopentetate imeglumine (Gd-DTPA, Magnevist, Bayer) contrast was
diluted from 0.5 M with distilled water into 0.0625 M for mouse
tail vein injection at 1 mL/kg body weight. Following Magnevist
injection, the mouse under anesthesia (maintained at 1-2%
isofluorane in 100% O.sub.2) was immediately moved into a
custom-made animal holder including a nose cone and bite bar for
MRI imaging. Respiration was monitored during imaging acquisition.
Scout images were collected using a RARE Tripliot of the head for
the localization f the orthotopic tumor. Subsequent axial images
were obtained using the following parameters to cover the
identified tumor: TRITE 1000/30 ms, FOV-1.8 cm.sup.2, matrix of
128.sup.2, for 6 continued slices at 1 mm slice thickness. The
mouse was kept in the same animal holder and carefully moved into a
PET scanner for .sup.18F-Cabozantinib 1b PET imaging. The post-Gd
axial MRI images were used for co-registration with PET images.
OTHER EMBODIMENTS
[0137] All publications and patents referred to in this disclosure
are incorporated herein by reference to the same extent as if each
individual publication or patent application were specifically and
individually indicated to be incorporated by reference. Should the
meaning of the terms in any of the patents r publications
incorporated by reference conflict with the meaning of the terms
used in this disclosure, the meaning of the terms in this
disclosure are intended to be controlling. Furthermore, the
foregoing discussion discloses and describes merely exemplary
embodiments of the present invention. One skilled in the art will
readily recognize from such discussion and from the accompanying
drawings and claims, that various changes, modifications and
variations can be made therein without departing from the spirit
and scope of the invention as defined in the following claims.
* * * * *