U.S. patent application number 14/803860 was filed with the patent office on 2015-11-12 for radioiodinated fatty acids.
This patent application is currently assigned to GE HEALTHCARE LIMITED. The applicant listed for this patent is GE HEALTHCARE LIMITED. Invention is credited to MICHELLE E. AVORY, ROBERT JAMES DOMETT NAIRNE, HARRY JOHN WADSWORTH.
Application Number | 20150320891 14/803860 |
Document ID | / |
Family ID | 43567327 |
Filed Date | 2015-11-12 |
United States Patent
Application |
20150320891 |
Kind Code |
A1 |
AVORY; MICHELLE E. ; et
al. |
November 12, 2015 |
Radioiodinated Fatty Acids
Abstract
The present invention provides novel radioiodinated fatty acids.
Also provided are methods of preparation of said radioiodinated
fatty acids from non-radioactive precursors, as well as
radiopharmaceutical compositions comprising such radioiodinated
fatty acids. The invention also provides in vivo imaging methods
using the radioiodinated fatty acids.
Inventors: |
AVORY; MICHELLE E.;
(WENDOVER, GB) ; WADSWORTH; HARRY JOHN;
(STORTFORD, GB) ; NAIRNE; ROBERT JAMES DOMETT;
(AMERSHAM, GB) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
GE HEALTHCARE LIMITED |
Buckinghamshire |
|
GB |
|
|
Assignee: |
GE HEALTHCARE LIMITED
BUCKINGHAMSHIRE
GG
|
Family ID: |
43567327 |
Appl. No.: |
14/803860 |
Filed: |
July 20, 2015 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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13993473 |
Jun 12, 2013 |
9138493 |
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PCT/EP2011/072993 |
Dec 15, 2011 |
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14803860 |
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Current U.S.
Class: |
424/1.85 ;
548/247 |
Current CPC
Class: |
C07D 261/10 20130101;
A61K 51/0497 20130101; A61K 51/0402 20130101; A61P 9/10 20180101;
A61K 51/0453 20130101; C07B 59/002 20130101 |
International
Class: |
A61K 51/04 20060101
A61K051/04; C07B 59/00 20060101 C07B059/00; C07D 261/10 20060101
C07D261/10 |
Foreign Application Data
Date |
Code |
Application Number |
Dec 16, 2010 |
GB |
1021369.2 |
Claims
1-18. (canceled)
19. A radioiodinated fatty acid of Formula (I): ##STR00031## where:
R.sup.1 and R.sup.2 are independently H or C.sub.1-2 alkyl; Y is a
Y.sup.2 group: ##STR00032## p and q are each independently integers
of value 0 to 10 which are chosen such that [p+q] is in the range
10 to 16; L.sup.1 is a linker group of formula -(A).sub.n- where n
is an integer of value 0 to 3, and each A group is independently
chosen from --CH.sub.2--, --O--, --S-- and --C.sub.6H.sub.4-- with
the proviso that L.sup.1 does not comprise --O--O--, --S--S-- or
--O--S-- linkages; and I* is a radioisotope of iodine.
20. The radioiodinated fatty acid of claim 19, wherein I* is chosen
from .sup.123I, .sup.124I or .sup.131I.
21. The radioiodinated fatty acid of claim 19, where R.sup.1 is
CH.sub.3 and R.sup.2 is H.
22. The radioiodinated fatty acid of claim 19, where L.sup.1 is
chosen from --CH.sub.2--, --O-- and --S--.
23. The radioiodinated fatty acid of claim 19, where n=0, and [p+q]
is in the range 11 to 13.
24. A radiopharmaceutical composition comprising an effective
amount of the radioiodinated fatty acid of Formula (I) as defined
in claim 19, together with a biocompatible carrier medium.
Description
FIELD OF THE INVENTION
[0001] The present invention provides novel radioiodinated fatty
acids. Also provided are methods of preparation of said
radioiodinated fatty acids from non-radioactive precursors, as well
as radiopharmaceutical compositions comprising such radioiodinated
fatty acids. The invention also provides in vivo imaging methods
using the radioiodinated fatty acids.
BACKGROUND TO THE INVENTION
[0002] Under normal conditions, the human heart derives more than
60% of its energy from the oxidative metabolism of long chain fatty
acids. In the ischaemic myocardium, however, oxidative metabolism
of free fatty acids is suppressed, and anaerobic glucose metabolism
predominates. Metabolic imaging can therefore provide useful
information in the diagnosis and monitoring of various forms of
heart disease.
[0003] Fatty acids have been radiolabelled with .sup.11C and
.sup.18F for PET imaging, and .sup.123I and .sup.99mTc for SPECT
radiopharmaceutical imaging [Eckelman et al, J.Nucl. Cardiol., 14,
S100-S109 (2007)]. Eckelman et at stress that radiolabelling with
an isotope other than .sup.11C is in fact labelling a fatty acid
analogue, and that care is needed that the substituent does not
affect the ability of the analogue to trace important steps of the
metabolic pathway.
[0004] Taki et at [Eur. J. Nucl. Med. Mol. Imaging, 34, S34-S48
(2007)] point out that early radioiodinated fatty acid analogues
based on iodo-alkyl substituents were found to suffer significant
in vivo metabolic deioidination. Radioiodinated fatty acid
analogues incorporating iodo-phenyl moieties such as
.sup.123I-BMIPP and .sup.123I-IPPA have, however, become
established agents for such metabolic imaging (Taki et al, cited
above):
##STR00001## [0005] where I*=.sup.123I.
[0006] The applications of "click chemistry" in biomedical
research, including radiochemistry, have been reviewed by Nwe et at
[Cancer Biother. Radiopharm., 24(3), 289-302 (2009)]. As noted
therein, the main interest has been in the PET radioisotope
.sup.18F (and to a lesser extent .sup.11C), plus "click to chelate"
approaches for radiometals suitable for SPECT imaging such as
.sup.99mTc or .sup.111In. .sup.18F click-labelling of targeting
peptides, giving products incorporating an
.sup.18F-fluoroalkyl-substituted triazole have been reported by Li
et at [Bioconj. Chem., 18(6), 1987-1994 (2007)] and Hausner et at
[J. Med. Chem., 51(19), 5901-5904 (2008)].
[0007] WO 2006/067376 discloses a method for labelling a vector
comprising reaction of a compound of formula (I) with a compound of
formula (II):
##STR00002##
or, a compound of formula (III) with a compound of formula (IV)
##STR00003##
in the presence of a Cu(I) catalyst, to give a conjugate of formula
(V) or (VI) respectively:
##STR00004##
wherein L1, L2, L3, and L4 are each Linker groups; [0008] R* is a
reporter moiety which comprises a radionuclide.
[0009] R* of WO 2006/067376 is a reporter moiety which comprises a
radionuclide for example a positron-emitting radionuclide. Suitable
positron-emitting radionuclides for this purpose are said to
include .sup.11C, .sup.18F, .sup.75Br, .sup.76Br, .sup.124I,
.sup.82Rb, .sup.68Ga, .sup.64Cu and .sup.62Cu, of which .sup.11C
and .sup.18F are preferred. Other useful radionuclides are stated
to include .sup.123I, .sup.125I, .sup.131I, .sup.211At, .sup.99mTc,
and .sup.111In.
[0010] WO 2007/148089 discloses a method for radiolabelling a
vector comprising reaction of a compound of formula (I) with a
compound of formula (II):
##STR00005##
or, a compound of formula (III) with a compound of formula
(IV):
##STR00006##
in the presence of a Cu(I) catalyst to give a conjugate of formula
(V) or (VI) respectively:
##STR00007##
wherein: [0011] L1, L2, L3, and L4 are each Linker groups; [0012]
R* is a reporter moiety which comprises a radionuclide.
[0013] In both WO 2006/067376 and WO 2007/148089, metallic
radionuclides are stated to be suitably incorporated into a
chelating agent, for example by direct incorporation by methods
known to the person skilled in the art.
[0014] WO 2006/116629 (Siemens Medical Solutions USA, Inc.)
discloses a method of preparation of a radiolabelled ligand or
substrate having affinity for a target biomacromolecule, the method
comprising: [0015] (a) reacting a first compound comprising [0016]
(i) a first molecular structure; [0017] (ii) a leaving group;
[0018] (iii) a first functional group capable of participating in a
click chemistry reaction; and optionally, [0019] (iv) a linker
between the first functional group and the molecular structure,
with a radioactive reagent under conditions sufficient to displace
the leaving group with a radioactive component of the radioactive
reagent to form a first radioactive compound; [0020] (b) providing
a second compound comprising [0021] (i) a second molecular
structure; [0022] (ii) a second complementary functional group
capable of participating in a click chemistry reaction with the
first functional group, wherein the second compound optionally
comprises a linker between the second compound and the second
functional group; [0023] (c) reacting the first functional group of
the first radioactive compound with the complementary functional
group of the second compound via a click chemistry reaction to form
the radioactive ligand or substrate; and [0024] (d) isolating the
radioactive ligand or substrate.
[0025] WO 2006/116629 teaches that the method therein is suitable
for use with the radioisotopes: .sup.124I, .sup.18F, .sup.13N and
.sup.15O with preferred radioisotopes being: .sup.18F, .sup.11C,
.sup.123I, .sup.124I, .sup.127I, .sup.131I, .sup.76Br, .sup.64 Cu,
.sup.99mTc, .sup.90Y, .sup.67Ga, .sup.51Cr, .sup.192Ir, .sup.99Mo,
.sup.153SM and .sup.201Tl. WO 2006/116629 teaches that other
radioisotopes that may be employed include: .sup.72As, .sup.74As,
.sup.75Br, .sup.55Co, .sup.61Cu, .sup.67Cu, .sup.68Ga, .sup.68Ge,
.sup.125I, .sup.132I, .sup.111In, .sup.52Mn, .sup.203Pb and
.sup.97Ru. WO 2006/116629 does not, however, provide any specific
teaching on how to apply the method to the radioiodination of
biological molecules.
[0026] WO 2010/129572 describes PET radiotracers for imaging fatty
acid metabolism and storage having one of the following
formulae:
##STR00008## [0027] where: n is 10-24, m is 1-10 and X is a
halogen.
[0028] WO 2010/129572 teaches that at least one atom of the above
chemical structures can be a radionuclide, preferably a
positron-emitting radioisotope. .sup.18F is the main radioisotope
described. The structures shown would not be expected to be
suitable for labelling with radioiodine, since if X were to be
iodine that requires an iodoalkyl group, and such groups are known
to be unstable with respect to deiodination in vivo.
[0029] Kim et at [Bioconj. Chem., 20(6), 1139-1145 (2009) disclose
.sup.18F-labelled fatty acid analogues for PET imaging of
myocardial metabolism:
##STR00009##
[0030] The .sup.18F-fatty acids were prepared via click
cycloaddition, wherein an .sup.18F-alkyne was coupled to an
azido-fatty acid, to generate the triazole ring.
[0031] PET imaging with .sup.18F typically requires the
availability of a cyclotron facility on the same site as the
hospital, since .sup.18F has a short half-life (110 minutes) and
the desired radiotracer needs to be synthesised. The availability
of cameras suitable for PET imaging is consequently much less
widespread than SPECT cameras. There is therefore still a need for
alternative radioiodinated fatty acids suitable for more routine
clinical imaging, especially using SPECT radiopharmaceutical
imaging.
[0032] The longer half-life of .sup.123I compared to .sup.18F
enables the cyclotron for its production to be up to one day's
transport time from the end user. This makes it possible for a
single cyclotron to be able to supply a continent rather than a
city, as is the case with .sup.18F fluorine production.
THE PRESENT INVENTION
[0033] The present invention provides radioiodinated fatty acid
analogues comprising triazole or isoxazole rings. The triazole and
isoxazole rings do not hydrolyse and are highly stable to oxidation
and reduction, meaning that the labelled fatty acid has high in
vivo stability. The triazole ring is also comparable to an amide in
size and polarity. The triazole and isoxazole rings of the fatty
acids of Formula (I) of the present invention are not expected to
be recognized by thyroid deiodination enzymes known to metabolise
iodo-tyrosine more rapidly than iodobenzene, and are thus expected
to be sufficiently stable in vivo for radiopharmaceutical imaging
and/or radiotherapy.
[0034] When the iodine isotope is .sup.123I or .sup.131I, the fatty
acids of the present invention have the advantage that they are
suitable for SPECT imaging, and hence have a wider clinical
potential than PET agents, due to the wider availability of gamma
cameras. The radioiodinated fatty acid analogues can be synthesised
readily using either click chemistry, or organometallic precursors.
Mild reaction conditions are required for the synthesis of the
carbon iodine bond and this enables sensitive molecules to be
radioiodinated. In general radiofluorination requires much more
forcing conditions rendering it unsuitable for very sensitive
molecules.
DETAILED DESCRIPTION OF THE INVENTION
[0035] In a first aspect, the present invention provides a
radioiodinated fatty acid of Formula (I):
##STR00010## [0036] where: [0037] R.sup.1 and R.sup.2 are
independently H or C.sub.1-2 alkyl; [0038] Y is a Y.sup.1 or
Y.sup.2 group:
[0038] ##STR00011## [0039] p and q are each independently integers
of value 0 to 10 which are chosen such that [p+q] is in the range
10 to 16; [0040] L.sup.1 is a linker group of formula -(A).sub.n-
where n is an integer of value 0 to 3, and each A group is
independently chosen from --CH.sub.2--, --O--, --S-- and
--C.sub.6H.sub.4-- with the proviso that L.sup.1 does not comprise
--O--O--, --S--S-- or --O--S-- linkages; [0041] I* is a
radioisotope of iodine.
[0042] The term "radioiodinated" has its conventional meaning, i.e.
a radiolabelled compound wherein the radioisotope used for the
radiolabelling is a radioisotope of iodine. The term "radioisotope
of iodine" has its conventional meaning, i.e. an isotope of the
element iodine that is radioactive. Suitable such radioisotopes
include: .sup.123I, .sup.124I, .sup.125I and .sup.131I.
[0043] The term "fatty acid" has its conventional meaning, i.e. a
monobasic aliphatic carboxylic acid, typically having at least a
10-carbon chain.
Preferred Aspects.
[0044] Preferred radioisotopes of iodine for use in the present
invention are those suitable for medical imaging in vivo using PET
or SPECT, preferably .sup.123I, .sup.124I or .sup.131I, more
preferably .sup.123I or .sup.124I, most preferably .sup.123I.
[0045] A preferred radioiodinated fatty acid of the first aspect is
where Y is Y.sup.1, i.e. the radioiodine isotope is attached to a
triazole ring.
[0046] In Formula (I), preferably at least one of R.sup.1 and
R.sup.2 is C.sub.1-2 alkyl, more preferably methyl. Most
preferably, le is CH.sub.3 and R.sup.2 is H.
[0047] In Formula (I), L.sup.1 is preferably chosen from
--CH.sub.2--, --O-- and --S--. In Formula (I), n is preferably 0.
In Formula (I), [p+q] is preferably in the range 10 to 14. When
n=0, [p+q] is preferably in the range 11 to 14, more preferably 11
to 13.
[0048] The radioiodinated fatty acids of Formula (I) may be
obtained as described in the second or third aspects (below). The
preparation method of the second aspect (via Precursor IA) is
preferred, since that comprises only a single step in which
radioactive manipulations are involved (a single step
iododemetallation reaction)--thus minimising the radiation dose to
the operator.
[0049] Included within the scope of the first aspect is an imaging
agent which comprises the radioiodinated fatty acid of Formula (I).
By the term "imaging agent" is meant a compound suitable for
imaging the mammalian body. Preferably, the mammal is an intact
mammalian body in vivo, and is more preferably a human subject.
Preferably, the imaging agent can be administered to the mammalian
body in a minimally invasive manner, i.e. without a substantial
health risk to the mammalian subject when carried out under
professional medical expertise. Such minimally invasive
administration is preferably intravenous administration into a
peripheral vein of said subject, without the need for local or
general anaesthetic. The imaging agents of the first aspect are
preferably used as radiopharmaceutical compositions, as described
in the fourth aspect (below).
[0050] In a second aspect, the present invention a method of
preparation of the radioiodinated fatty acid of Formula (I) of the
first aspect, where said method comprises: [0051] (i) provision of
a precursor of Formula (IA)
[0051] ##STR00012## [0052] where: [0053] R.sup.1, R.sup.2, L.sup.1,
p and q are as defined in any one of claims 1 to 6; [0054] Visa
Y.sup.1a or Y.sup.2a group:
[0054] ##STR00013## [0055] wherein Q is R.sup.a.sub.3Sn-- or
KF.sub.3B--, where each R.sup.a is independently C.sub.1-4 alkyl;
[0056] (ii) reaction of said precursor with radioactive iodide ion
in the presence of an oxidising agent to give the radioiodinated
fatty acid of Formula (I).
[0057] Preferred embodiments of R.sup.1, R.sup.2, L.sup.1, p and q
and I* in the second aspect are as defined in the first aspect.
[0058] In Y.sup.a, when Q is KF.sub.3B--, that corresponds to a
potassium trifluoroborate derivative as described below.
[0059] By the term "oxidising agent" is meant an oxidant capable of
oxidising iodide ion to form the electrophilic species (HOT,
H.sub.2OI), wherein the active iodinating agent is I.sup.+.
Suitable oxidising agents are described by Bolton [J. Lab. Comp.
Radiopharm., 45, 485-528 (2002)], and Eersels et al [J. Lab. Comp.
Radiopharm., 48, 241-257 (2005)] and include peracetic acid and
N-chloro compounds, such as chloramine-T, iodogen, iodogen tubes
and succinimides. Preferred oxidising agents are peracetic acid
(which is commercially available) at pH ca. 4, and hydrogen
peroxide/aqueous HCl at pH ca. 1. Iodogen tubes are commercially
available from Thermo Scientific Pierce Protein Research
Products.
[0060] By the term "radioactive iodide ion" is meant a radioisotope
of iodine (as defined above), in the chemical form of iodide ion
(I.sup.-).
[0061] When Q is R.sup.a.sub.3Sn--, the radioiodination method of
the third aspect is carried out as described by Bolton [J. Lab.
Comp. Radiopharm., 45, 485-528 (2002)] and Eersels et at [J. Lab.
Comp. Radiopharm., 48, 241-257 (2005)]. The organotin precursors
are prepared as described by Ali et at [Synthesis, 423-445
(1996)].
[0062] When Q is KF.sub.3B--, the radioiodination reaction method
of the third aspect can be carried out as described by Kabalka et
al [J. Lab. Comp. Radiopharm., 48, 359-362 (2005)], who use
peracetic acid as the oxidising agent. Precursors where Q is
KF.sub.3B-- can be obtained from the corresponding alkyne as
described by Kabalka et at [J. Lab. Comp. Radiopharm., 48, 359-362
(2005) and, J. Lab. Comp. Radiopharm., 49, 11-15 (2006)]. The
potassium trifluoroborate precursors are stated to be crystalline
solids, which are stable to both air and water.
[0063] The radioiodination reaction of the second aspect may be
effected in a suitable solvent, for example acetonitrile, a
C.sub.1-4 alkylalcohol, dimethylformamide, tetrahydrofuran (THF),
or dimethylsulfoxide, or mixtures thereof, or aqueous mixtures
thereof, or in water. Aqueous buffers can also be used. The pH will
depend on the oxidant used, and will typically be pH 0 to 1 when
eg. hydrogen peroxide/aqueous acid is used, or in the range pH 6-8
when iodogen or iodogen tubes are used. The radioiodination
reaction temperature is preferably 10 to 60.degree. C., more
preferably at 15 to 50.degree. C., most preferably at ambient
temperature (typically 15-37.degree. C.). Organic solvents such as
acetonitrile or THF and/or the use of more elevated temperature may
conveniently be used to solubilise any precursors of Formula (TB)
which are poorly soluble in water.
[0064] The precursor of Formula (IA) is suitably non-radioactive,
so can be prepared and purified by conventional means without the
need for radiation handling safety precautions.
[0065] In a third aspect, the present invention provides a method
of preparation of the radioiodinated fatty acid of Formula (I) as
defined in the first aspect, where said method comprises: [0066]
(i) provision of a precursor of Formula (IB)
[0066] ##STR00014## [0067] where: [0068] R.sup.1, R.sup.2, L.sup.1,
p and q are as defined in any one of claims 1 to 6; [0069] Y.sup.b
is a Y.sup.th or Y.sup.2b group:
[0069] ##STR00015## [0070] (ii) reaction of said precursor with a
compound of Formula (II):
[0070] ##STR00016## [0071] in the presence of a click cycloaddition
catalyst, to give the radioiodinated fatty acid of Formula (I) via
click cycloaddition, [0072] wherein I* is a radioisotope of iodine,
as defined in claim 1 or claim 2.
[0073] In Formula (IB), Y can be either an azide substituent
(Y=Y.sup.1a), or an isonitrile oxide substituent (Y=Y.sup.2a).
[0074] Preferred embodiments of R.sup.1, R.sup.2, L.sup.1, p and q
and I* in the third aspect are as defined in the first aspect.
[0075] By the term "click cycloaddition catalyst" is meant a
catalyst known to catalyse the click (alkyne plus azide) or click
(alkyne plus isonitrile oxide) cycloaddition reaction. Suitable
such catalysts are known in the art for use in click cycloaddition
reactions. Preferred such catalysts include Cu(I), and are
described below. Further details of suitable catalysts are
described by Wu and Fokin [Aldrichim. Acta, 40(1), 7-17 (2007)] and
Meldal and Tornoe [Chem. Rev., 108, 2952-3015 (2008)]. The
applications of "click chemistry" in biomedical research, including
radiochemistry, have been reviewed by Nwe et al [Cancer Biother.
Radiopharm., 24(3), 289-302 (2009)].
[0076] A preferred click cycloaddition catalyst comprises Cu(I).
The Cu(I) catalyst is present in an amount sufficient for the
reaction to progress, typically either in a catalytic amount or in
excess, such as 0.02 to 1.5 molar equivalents relative to the
compound of Formula (Ia) or (Ib). Suitable Cu(I) catalysts include
Cu(I) salts such as Cul or [Cu(NCCH.sub.3).sub.4][PF.sub.6], but
advantageously Cu(II) salts such as copper (II) sulphate may be
used in the presence of a reducing agent to generate Cu(I) in situ.
Suitable reducing agents include: ascorbic acid or a salt thereof
for example sodium ascorbate, hydroquinone, metallic copper,
glutathione, cysteine, Fe.sup.2+, or Co.sup.2+. Cu(I) is also
intrinsically present on the surface of elemental copper particles,
thus elemental copper, for example in the form of powder or
granules may also be used as catalyst. Elemental copper, with a
controlled particle size is a preferred source of the Cu(I)
catalyst. A more preferred such catalyst is elemental copper as
copper powder, having a particle size in the range 0.001 to 1 mm,
preferably 0.1 mm to 0.7 mm, more preferably around 0.4 mm.
Alternatively, coiled copper wire can be used with a diameter in
the range of 0.01 to 1.0 mm, preferably 0.05 to 0.5 mm, and more
preferably with a diameter of 0.1 mm. The Cu(I) catalyst may
optionally be used in the presence of bathophenanthroline, which is
used to stabilise Cu(I) in click chemistry.
[0077] In the method of the third aspect, the compound of Formula
(II) may optionally be generated in situ by deprotection of a
compound of Formula (IIa):
##STR00017## [0078] wherein M.sup.1 is an alkyne-protecting group,
and I* is as defined for Formula (II). Preferred aspects of I* in
Formula (IIa), are as described for Formula (II).
[0079] By the term "protecting group" is meant a group which
inhibits or suppresses undesirable chemical reactions, but which is
designed to be sufficiently reactive that it may be cleaved from
the functional group in question under mild enough conditions that
do not modify the rest of the molecule. After deprotection the
desired product is obtained. Suitable alkyne protecting groups are
described in Protective Groups in Organic Synthesis, Theodora W.
Greene and Peter G. M. Wuts, Chapter 8, pages 927-933, 4.sup.th
edition (John Wiley & Sons, 2007), and include: an
trialkylsilyl group where each alkyl group is independently
C.sub.1-4 alkyl; an aryldialkylsilyl group where the aryl group is
preferably benzyl or biphenyl and the alkyl groups are each
independently C.sub.1-4 alkyl; hydroxymethyl or
2-(2-hydroxypropyl). A preferred such alkyne protecting group is
trimethylsilyl. The protected iodoalkynes of Formula (IIa) have the
advantages that the volatility of the radioactive iodoalkyne can be
controlled, and that the desired alkyne of Formula (II) can be
generated in a controlled manner in situ so that the efficiency of
the reaction with the precursor of Formula (IA) is maximised.
[0080] The click cycloaddition method of the second aspect may be
effected in a suitable solvent, for example acetonitrile, a
C.sub.1-4 alkylalcohol, dimethylformamide, tetrahydrofuran, or
dimethylsulfoxide, or aqueous mixtures of any thereof, or in water.
Aqueous buffers can be used in the pH range of 4-8, more preferably
5-7. The reaction temperature is preferably 5 to 100.degree. C.,
more preferably at 75 to 85.degree. C., most preferably at ambient
temperature (typically 15-37.degree. C.). The click cycloaddition
may optionally be carried out in the presence of an organic base,
as is described by Meldal and Tornoe [Chem. Rev. 108, 2952, Table 1
(2008)].
[0081] A preferred precursor of Formula (IB) has Y.sup.b=Y.sup.ib.
One reason is that the isonitrile oxides are typically less stable
than azides. Consequently, whilst the azide of Formula (IB,
Y.sup.b=Y.sup.1b) can be isolated and purified, the isonitrile
oxide of Formula (IB, Y.sup.b=Y.sup.2b) will typically need to be
generated in situ.
[0082] The non-radioactive precursor compound of Formula (IB),
where Y.sup.b is Y.sup.1b (azido derivatives) may be prepared by
either: [0083] (i) reaction of the corresponding bromo-fatty acid
with sodium azide; [0084] (ii) conversion of the corresponding
hydroxy-fatty acid to a tosylate or mesylate derivative, and
subsequent reaction with sodium azide.
[0085] Further details are provided by Kim et al [Bioconj. Chem.,
20(6), 1139-1145 (2009)], and Kostiuk et al [Meth. Enzymol., 457,
149-165 (2009)]. Many functionalised fatty acids are commercially
available.
[0086] The non-radioactive precursor compound of Formula (IB),
where Y.sup.b is Y.sup.2b (isonitrile oxide derivatives) may be
prepared by the methods described by Ku et al [Org. Lett., 3(26),
4185-4187 (2001)], and references therein. Thus, they are typically
generated in situ by treatment of an alpha-halo aldoxime with an
organic base such as triethylamine. A preferred method of
generation, as well as conditions for the subsequent click
cyclisation to the desired isoxazole are described by Hansen et al
[J. Org. Chem., 70(19), 7761-7764 (2005)]. Hansen et al generate
the desired alpha-halo aldoxime in situ by reaction of the
corresponding aldoxime with chloramine-T trihydrate, and then
dechlorinating this with sodium hydroxide. The corresponding
aldoxime is prepared by reacting the corresponding aldehyde with
hydroxylamine hydrochloride at pH 9-10. See also K. B. G. Torsell
Nitrile Oxides, Nitrones and Nitronates in Organic Synthesis [VCH,
New York (1988)]. .OMEGA.-aldehyde functionalised fatty acids are
readily accessible by the oxidation of the corresponding alcohol in
a Swern oxidation. The alcohols are generally commercially
available but are also accessible by ozone oxidation to the ozonide
of the corresponding unsatuturated fatty acid followed by
borohydride reduction to the alcohol. A very wide range of
unsaturated fatty acids are available from natural product
sources.
[0087] Included within the scope of this third aspect, is the
option of using an aldoxime precursor, wherein instead of Y.sup.2b,
Y.sup.b is chosen to be (HO)N.dbd.CH--, so that the isonitrile
oxide (Y.sup.b=Y.sup.2b) is generated in situ. The steps involved
can be carried out sequentially without workup. The resulting
nitrile oxide is not particularly stable and is best used
immediately, preferably in situ without workup.
[0088] The preparation methods of the second and third aspects are
preferably carried out in an aseptic manner, such that the product
of Formula (I) is obtained as a radiopharmaceutical composition.
Thus, the method is carried out under aseptic manufacture
conditions to give the desired sterile, non-pyrogenic
radiopharmaceutical product. It is preferred therefore that the key
components, especially any parts of the apparatus which come into
contact with the product of Formula (I) (e.g. vials and transfer
tubing) are sterile. The components and reagents can be sterilised
by methods known in the art, including: sterile filtration,
terminal sterilisation using e.g. gamma-irradiation, autoclaving,
dry heat or chemical treatment (e.g. with ethylene oxide). It is
preferred to sterilise the non-radioactive components in advance,
so that the minimum number of manipulations need to be carried out
on the radioiodinated radiopharmaceutical product. As a precaution,
however, it is preferred to include at least a final sterile
filtration step.
[0089] The precursors of Formula (IA) or (IB), and other reactants,
reagents and solvents are each supplied in suitable vials or
vessels which comprise a sealed container which permits maintenance
of sterile integrity and/or radioactive safety, plus optionally an
inert headspace gas (eg. nitrogen or argon), whilst permitting
addition and withdrawal of solutions by syringe or cannula. A
preferred such container is a septum-sealed vial, wherein the
gas-tight closure is crimped on with an overseal (typically of
aluminium). The closure is suitable for single or multiple
puncturing with a hypodermic needle (e.g. a crimped-on septum seal
closure) whilst maintaining sterile integrity. Such containers have
the additional advantage that the closure can withstand vacuum if
desired (eg. to change the headspace gas or degas solutions), and
withstand pressure changes such as reductions in pressure without
permitting ingress of external atmospheric gases, such as oxygen or
water vapour. The reaction vessel is suitably chosen from such
containers, and preferred embodiments thereof. The reaction vessel
is preferably made of a biocompatible plastic (eg. PEEK).
[0090] When the radioiodinated fatty acid is used as a
pharmaceutical composition, the method of the second or third
aspects is preferably carried out using an automated synthesizer
apparatus. By the term "automated synthesizer" is meant an
automated module based on the principle of unit operations as
described by Satyamurthy et at [Clin. Positr. Imag., 2(5), 233-253
(1999)]. The term `unit operations` means that complex processes
are reduced to a series of simple operations or reactions, which
can be applied to a range of materials. Such automated synthesizers
are preferred for the method of the present invention especially
when a radiopharmaceutical product is desired. They are
commercially available from a range of suppliers [Satyamurthy et
al, above], including: GE Healthcare; CTI Inc; Ion Beam
Applications S.A. (Chemin du Cyclotron 3, B-1348 Louvain-La-Neuve,
Belgium); Raytest (Germany) and Bioscan (USA).
[0091] Commercial automated synthesizers also provide suitable
containers for the liquid radioactive waste generated as a result
of the radiopharmaceutical preparation. Automated synthesizers are
not typically provided with radiation shielding, since they are
designed to be employed in a suitably configured radioactive work
cell. The radioactive work cell provides suitable radiation
shielding to protect the operator from potential radiation dose, as
well as ventilation to remove chemical and/or radioactive vapours.
The automated synthesizer preferably comprises a cassette.
[0092] By the term "cassette" is meant a piece of apparatus
designed to fit removably and interchangeably onto an automated
synthesizer apparatus (as defined below), in such a way that
mechanical movement of moving parts of the synthesizer controls the
operation of the cassette from outside the cassette, i.e.
externally. Suitable cassettes comprise a linear array of valves,
each linked to a port where reagents or vials can be attached, by
either needle puncture of an inverted septum-sealed vial, or by
gas-tight, marrying joints. Each valve has a male-female joint
which interfaces with a corresponding moving arm of the automated
synthesizer. External rotation of the arm thus controls the opening
or closing of the valve when the cassette is attached to the
automated synthesizer. Additional moving parts of the automated
synthesizer are designed to clip onto syringe plunger tips, and
thus raise or depress syringe barrels.
[0093] The cassette is versatile, typically having several
positions where reagents can be attached, and several suitable for
attachment of syringe vials of reagents or chromatography
cartridges (eg. solid phase extraction, SPE). The cassette always
comprises a reaction vessel. Such reaction vessels are preferably 1
to 10 cm.sup.3, most preferably 2 to 5 cm.sup.3 in volume and are
configured such that 3 or more ports of the cassette are connected
thereto, to permit transfer of reagents or solvents from various
ports on the cassette. Preferably the cassette has 15 to 40 valves
in a linear array, most preferably 20 to 30, with 25 being
especially preferred. The valves of the cassette are preferably
each identical, and most preferably are 3-way valves. The cassettes
of the present invention are designed to be suitable for
radiopharmaceutical manufacture and are therefore manufactured from
materials which are of pharmaceutical grade and ideally also are
resistant to radiolysis.
[0094] Preferred automated synthesizers of the present invention
are those comprising a disposable or single use cassette which
comprises all the reagents, reaction vessels and apparatus
necessary to carry out the preparation of a given batch of
radioiodinated radiopharmaceutical. The cassette means that the
automated synthesizer has the flexibility to be capable of making a
variety of different radioiodine-labelled radiopharmaceuticals with
minimal risk of cross-contamination, by simply changing the
cassette. The cassette approach also has the advantages of:
simplified set-up hence reduced risk of operator error; improved
GMP (Good Manufacturing Practice) compliance; multi-tracer
capability; rapid change between production runs; pre-run automated
diagnostic checking of the cassette and reagents; automated barcode
cross-check of chemical reagents vs the synthesis to be carried
out; reagent traceability; single-use and hence no risk of
cross-contamination, tamper and abuse resistance.
[0095] In a fourth aspect, the present invention provides a
radiopharmaceutical composition comprising an effective amount of
the radioiodinated fatty acid of Formula (I) as defined in the
first aspect, together with a biocompatible carrier medium.
[0096] Preferred embodiments of the radioiodinated fatty acid of
Formula (I) in the fourth aspect are as defined in the first
aspect.
[0097] The "biocompatible carrier medium" comprises one or more
pharmaceutically acceptable adjuvants, excipients or diluents. It
is preferably a fluid, especially a liquid, in which the
radioiodinated fatty acid of Formula (I) is suspended or dissolved,
such that the composition is physiologically tolerable, i.e. can be
administered to the mammalian body without toxicity or undue
discomfort. The biocompatible carrier medium is suitably an
injectable carrier liquid such as sterile, pyrogen-free water for
injection; an aqueous solution such as saline (which may
advantageously be balanced so that the final product for injection
is either isotonic or not hypotonic); an aqueous solution of one or
more tonicity-adjusting substances (eg. salts of plasma cations
with biocompatible counterions), sugars (e.g. glucose or sucrose),
sugar alcohols (eg. sorbitol or mannitol), glycols (eg. glycerol),
or other non-ionic polyol materials (eg. polyethyleneglycols,
propylene glycols and the like). The biocompatible carrier medium
may also comprise biocompatible organic solvents such as ethanol.
Such organic solvents are useful to solubilise more lipophilic
compounds or formulations. Preferably the biocompatible carrier
medium is pyrogen-free water for injection, isotonic saline or an
aqueous ethanol solution. The pH of the biocompatible carrier
medium for intravenous injection is suitably in the range 4.0 to
10.5.
[0098] In a fifth aspect, the present invention provides the use of
the precursor of Formula (IA) as defined in the second aspect, or
the precursor of Formula (IB) as defined in the third aspect in the
manufacture of the radioiodinated fatty acid of Formula (I) as
defined in the first aspect, or for the manufacture of the
radiopharmaceutical composition of the fourth aspect.
[0099] Preferred embodiments of the radioiodinated fatty acid of
Formula (I), precursor of Formula (IA) or of Formula (IB) in the
use of the fifth aspect, are as defined in the first, second and
third aspects respectively.
[0100] In a sixth aspect, the present invention provides the use of
an automated synthesizer apparatus to carry out the method of
preparation of the second or third aspects.
[0101] Preferred embodiments of the precursors, methods and
automated synthesizer in the use of the sixth aspect are as
described in the second and third aspects.
[0102] In a seventh aspect, the present invention provides a method
of generating an image of a human or animal body comprising
administering the radioiodinated fatty acid of Formula (I) of the
first aspect, or the radiopharmaceutical composition of the fourth
aspect, and generating an image of at least a part of said body to
which said compound or composition has distributed using PET or
SPECT.
[0103] Preferred aspects of the radioiodinated fatty acid and
radiopharmaceutical composition in the seventh aspect are as
described in the first and fourth aspects respectively.
[0104] The radioiodinated fatty acids of the invention are useful
for imaging myocardial metabolism, in particular for patients with
coronary artery disease. Such imaging includes the imaging of:
acute myocardial infarction; unstable angina; myocardial viability
assessment and prediction of recovery of function in chronic
coronary artery disease; risk stratification and prognosis. The
agent may also be useful, in conjunction with myocardial perfusion
assessment for patients with cardiomyopathy. Further details are
provided by Taki et al [Eur. J. Nucl. Med. Mol. Imaging, 34,
S34-S48 (2007)].
[0105] In a further aspect, the present invention provides a method
of monitoring the effect of treatment of a human or animal body
with a drug, said method comprising administering to said body the
radioiodinated fatty acid of Formula (I) as defined in the first
aspect, or the radiopharmaceutical composition of the fourth
aspect, and detecting the uptake of said fatty acid or composition
in at least a part of said body to which said fatty acid or
composition has distributed using PET or SPECT, said administration
and detection optionally but preferably being effected before,
during and after treatment with said drug. The administration and
detection of this final aspect are preferably effected before and
after treatment with said drug, so that the effect of the drug
treatment on the human or animal patient can be determined. Where
the drug treatment involves a course of therapy, the imaging can
also be carried out during the treatment. The diseases or
conditions being treated in the further aspect are as described in
the seventh aspect (above)
[0106] The invention is illustrated by the following Examples.
Example 1 provides the synthesis of .sup.123I-iodoacetylene.
Example 2 provides the click cycloaddition of
.sup.123I-iodoacetylene to an azide derivative, to form a
radioiodinated triazole ring. Example 3 provides the click
cycloaddition of .sup.123I-iodoacetylene to an isonitrile oxide
derivative, to form a radioiodinated isoxazole ring. Example 4
provides a click cycloaddition of a tributyltin-alkyne to an azide
derivative, to form a triazole radioiodination precursor having a
triazole-tributyltin bond. Example 5 provides the conditions for
converting the precursor of Example 4, to the radioiodinated
product. Example 6 provides a synthesis of an isoxazole
radioiodination precursor having an isoxazole-tributyltin bond via
click cycloaddition from an isonitrile oxide derivative. Example 7
provides the radioiodination of the precursor of Example 6. Example
8 provides the synthesis of an iodotriazole-substituted fatty acid.
Example 9 provides the synthesis of an iodoisoxazole-substituted
fatty acid.
Abbreviations Used in the Examples
[0107] HPLC: high performance liquid chromatography, PAA: peracetic
acid, RCP: radiochemical purity, THF: tetrahydrofuran.
Example 1
Preparation and Distillation of [.sup.123I]-Iodoacetylene Using
Peracetic Acid Oxidant
##STR00018##
[0109] To a Wheaton vial on ice was added, ammonium acetate buffer
(100 .mu.l, 0.2 M, pH 4), sodium [.sup.127I] iodide (10 .mu.l, 10
mM solution in 0.01M sodium hydroxide, 1.times.10.sup.-7 moles),
sodium [.sup.123I] iodide (20 .mu.l, 53 MBq), peracetic acid, (10
.mu.l, 10 mM solution, 1.times.10.sup.-7 moles) and a solution of
ethynyltributylstannane in THF (Sigma-Aldrich; 38 .mu.l, 1 mg/mL,
1.2.times.10.sup.-7 moles). Finally, 460 .mu.l THF was added, the
Wheaton vial sealed and the reaction mixture allowed to warm to
room temperature prior to reverse phase HPLC analysis which showed
[.sup.123I]-iodoacetylene with a radiochemical purity (RCP) of 75%
(t.sub.R 12.3 minutes, System A).
[0110] The reaction mixture was heated at 80-100.degree. C. for 30
minutes during which time, the [.sup.123I]-iodoacetylene and THF
were distilled through a short tube into a collection vial on ice.
After this time, a low flow of nitrogen was passed through the
septa of the heated vial to remove any residual liquids from the
tube. [.sup.123I]-iodoacetylene was collected in 38.6% yield (non
decay corrected) with an RCP of 94%. (t.sub.R 12.3 minutes, System
A).
[0111] HPLC System A (A=water; B=acetonitrile).
[0112] Column C18 (2) phenonenex Luna, 150.times.4.6 mm, 5
micron
TABLE-US-00001 Gradient Time (min) 0 1 20 25 25.5 30 % B 5 5 95 95
5 5
Example 2
Preparation of 1-Benzene-4-[.sup.123I]-iodo-1H-1,2,3 triazole
(Prophetic Example)
##STR00019##
[0114] To a Wheaton vial charged with copper powder (200 mg, -40
mesh), sodium phosphate buffer (200 mL, pH 6, 50 mM) and placed on
ice is added, [.sup.123I]-iodoacetylene and benzyl azide (1 mg,
7.5.times.10.sup.-6 moles). Following reagent addition, the ice
bath is removed and the reaction incubated at room temperature with
heating applied as required.
1-Benzene-4-[.sup.123I]-iodo-1H-1,2,3-triazole is purified by
reverse phase HPLC.
Example 3
Preparation of 5-[.sup.123I]-iodo-3-phenyl isoxazole (Prophetic
Example)
##STR00020##
[0116] To a Wheaton vial charged with copper powder (50 mg, -40
mesh), copper (II) sulphate (3.8 .mu.g, 1.53.times.10-8 moles, 0.5
mg/mL solution in water), sodium phosphate buffer (100 .mu.L, 50
mM, pH6) and placed on ice, is added [.sup.123I]-iodoacetylene and
benzonitrile-N-oxide (1 mg, 8.4.times.10.sup.-6 moles. Following
reagent addition, the ice bath is removed and the reaction
incubated at room temperature with heating applied as required.
5-[.sup.123I]-iodo-3-phenyl isoxazole is purified by reverse phase
HPLC.
Example 4
Preparation of 1-Phenyl-4-(tributylstannyl)-1H [1,2,3] triazole
(Prophetic Example)
##STR00021##
[0118] Phenylazide can be obtained from Sigma-Aldrich or can be
synthesized by the method described in J. Biochem., 179, 397-405
(1979). A solution of tributylethynyl stannane (Sigma Aldrich; 400
mg, 1.27 mmol) in THF (4 mL) is treated with phenylazide (169 mg,
1.27 mmol), copper (I) iodide (90 mg, 0.47 mmol), and triethylamine
(256 mg, 2.54 mmol) at room temperature over 48 h. The reaction is
then filtered through celite to remove copper (I) iodide and
chromatographed on silica in a gradient of 5-20% ethyl acetate in
petrol. The second fraction is collected and concentrated in vacuo
to give the 1-phenyl-4-(tributylstannyl)-1H [1,2,3] triazole as a
colourless oil.
Example 5
Preparation of [.sup.123I]-Phenyl-4-iodo-1H [1,2,3] triazole Using
Peracetic Acid as the Oxidant (Prophetic Example)
##STR00022##
[0120] To sodium [.sup.123I] iodide, received in 5-20 .mu.L 0.05M
sodium hydroxide is added ammonium acetate buffer (100 .mu.L pH
4.0, 0.2M), sodium [.sup.127I] iodide (10 .mu.L 1 mM solution in
0.01M sodium hydroxide, 1.times.10.sup.-8 moles), peracetic acid
(PAA) solution (10 .mu.L 1 mM solution, 1.times.10.sup.-8 moles)
and finally, 1 phenyl-4-tributylstannyl-1H [1,2,3] triazole
(Example 4; 43 .mu.g, 1.times.10.sup.-7 moles) dissolved in
acetonitrile. The reaction mixture is incubated at room temperature
for 15 minutes prior to purification by HPLC.
Example 6
Preparation of 3-Phenyl-5-(tributylstannyl) isoxazole (Prophetic
Example)
##STR00023##
[0122] (E)-benzaldehyde oxime (Sigma Aldrich; 3.3 g, 20 mmol) in
tert butanol and water (1:1) 80 mL, is treated with chloramine T
trihydrate (Sigma Aldrich; 5.9 g, 21 mmol) in small, portions over
5 min. The reaction is then treated with copper sulfate
pentahydrate (0.15 g, 0.6 mmol) and copper turnings .about.50 mg
and tributylethynylstannane (6.3 g, 20 mmol). The reaction is then
adjusted to pH 6 with sodium hydroxide solution and stirred for 6
h. The reaction mixture is treated with dilute ammonium hydroxide
solution to remove all copper salts. The product is collected by
filtration, redissolved in ethyl acetate and filtered through a
short plug of silica gel. The filtrate is concentrated in vacuo to
give 3-phenyl-5-(tributylstannyl) isoxazole.
Example 7
Preparation of 5-[.sup.123I]-Iodo-3-phenyl isoxazole (Prophetic
Example)
##STR00024##
[0124] To sodium [.sup.123I] iodide, received in 5-20 .mu.L 0.05M
sodium hydroxide is added ammonium acetate buffer (100 .mu.L pH
4.0, 0.2M), sodium [.sup.127I] iodide (10 .mu.L, 1 mM solution in
0.01M sodium hydroxide, 1.times.10.sup.-8 moles), peracetic acid
(PAA) solution (10 .mu.L 1 mM solution, 1.times.10.sup.-8 moles)
and finally, 3-phenyl-5-tributylstannyl-isoxazole (Example 6; 43
.mu.g, 1.times.10.sup.-7 moles) dissolved in acetonitrile. The
reaction mixture is incubated at room temperature for 15 minutes
prior to purification by HPLC.
Example 8
Preparation of 10-(4-Iodo-[1,2,3]triazol-1-yl)-decanoic acid
(Prophetic Example)
Step (a): Preparation of 10-(4-azido-[1,2,3]triazol-1-yl)-decanoic
acid
##STR00025##
[0126] 10-Bromodecanoic acid (2.51 g, 10 mmol) in acetone (50 mL)
is treated with sodium azide (0.75 g, 11 mmol) at reflux for 2 h.
The reaction is then concentrated in vacuo to a gum that is
partitioned between ethyl acetate (50 mL) and water (50 mL). The
organic phase is separated, dried over sodium sulfate and
concentrated in vacuo to give 10-azidodecanoic acid (2.02 g,
95%).
Step (b): Preparation of
10-(4-tributylstannyl-[1,2,3]triazol-1-yl)-decanoic acid
##STR00026##
[0128] 10-Azidodecanoic acid (2.0 g, 9.5 mmol) in THF (50 mL) is
treated with tributylstannyl acetylene (2.99 g, 9.5 mmol)
copper(I)iodide (90 mg 0.47 mmol) and triethylamine (256 mg, 2.54
mmol) at room temperature with constant stirring for 24 h. The
reaction is then filtered through celite and concentrated in vacuo
to a gum, and then chromatographed on silica in a gradient of
10-40% ethyl acetate in petrol. The main fraction is collected and
concentrated in vacuo to give
10-(4-tributylstannyl-[1,2,3]triazol-1-yl)-decanoic acid (3.22 g,
0.8 mmol).
Step (c): Preparation of 10-(4-iodo-[1,2,3]triazol-1-yl)-decanoic
acid
##STR00027##
[0130] To sodium [.sup.123J] iodide, received in 5-20 .mu.L 0.05M
sodium hydroxide is added ammonium acetate buffer (100 .mu.L pH
4.0, 0.2M), sodium [.sup.127J] iodide (10 .mu.L, 1 mM solution in
0.01M sodium hydroxide, 1.times.10.sup.-8 moles), peracetic acid
(PAA) solution (10 .mu.L 1 mM solution, 1.times.10.sup.-8 moles)
and finally 10-(4-tributylstannyl-[1,2,3]triazol-1-yl)-decanoic
acid solution in ethanol or acetonitrile (53 .mu.g,
1.times.10.sup.-7 moles). The reaction mixture is allowed to stand
at room temperature for 15 minutes prior to HPLC purification of
the iodinated product 10-(4-iodo-[1,2,3]triazol-1-yl)-decanoic
acid.
Example 9
Preparation of 10-(5-Iodo isoxazol-3-yl)-decanoic acid (Prophetic
Example)
Step (a): Preparation of 11[(E)-hydroximino-decanoic acid
##STR00028##
[0132] A solution of dimethyl sulfoxide (1.17 g, 15 mmol) in
dichloromethane (50 mL) is cooled to -30.degree. C. and treated
with oxalyl chloride (1.92 g, 15 mmol). The reaction mixture is
then treated with 10-hydroxydecanoic acid (2.06 g, 10 mmol) in
dichloromethane (50 mL) and allowed to warm to room temperature
over a period of 2 h. The reaction is then washed with water
(2.times.50 mL). The organic layer is dried over sodium sulfate and
then treated with hydroxylamine hydrochloride (1.03 g 15 mmol) and
sodium hydroxide (0.6 g in 10 mL water) and stirred vigorously for
1 h. The organic phase is then separated dried over sodium sulfate
and concentrated in vacuo to give 11[(E)-hydroxyimino-decanoic acid
(1.8 g, 9.0 mmol).
Step (b): Preparation of
10-(5-tributylstanyl-isoxazol-3-yl)-decanoic acid
##STR00029##
[0134] 11[(E)-Hydroxyimino-decanoic acid (1.8 g, 9.0 mmol) is
dissolved in acetonitrile (30 ml) and the solution cooled to
0.degree. C. and then treated with a solution of chloramine-T
trihydrate (2.52 g, 9.0 mmol) in water (10 mL). The reaction is
allowed to warm to room temperature over a period of 25 minutes. To
the resulting nitrile oxide solution is added
tributyl(ethynyl)stannane (2.83 g, 9.0 mmol), copper iodide (100
mg, 0.5 mmol) and triethylamine (50 mg, 0.5 mmol) and the reaction
stirred at room temperature for 24 h. The reaction is then filtered
through celite to remove the copper salts and concentrated in vacuo
to a gum. The gum is then chromatographed on silica in a gradient
of 10- 30% ethyl acetate in petrol to give
10-(5-tributylstanyl-isoxazol-3-yl)-decanoic acid.
Step (c)
Example 4
Preparation of 10-(5-iodo isoxazol-3-yl)-decanoic acid
##STR00030##
[0136] To sodium [.sup.123I] iodide, received in 5-20 .mu.L 0.05M
sodium hydroxide is added ammonium acetate buffer (100 .mu.L pH
4.0, 0.2M), sodium [.sup.127I] iodide (10 .mu.L, 1 mM solution in
0.01M sodium hydroxide, 1.times.10.sup.-8 moles), peracetic acid
(PAA) solution (10 .mu.L 1 mM solution, 1.times.10.sup.-8 moles)
and finally 10-(5-tributylstannyl-isoxazol-3-yl)-decanoic acid
solution in ethanol or acetonitrile (53 .mu.g, 1.times.10.sup.-7
moles). The reaction mixture is allowed to stand at room
temperature for 15 minutes and the iodinated product
10-(4-iodo-[1,2,3]triazol-1-yl)-decanoic acid purified by HPLC.
* * * * *