Patent Application
20130338361
The present invention relates to a method for radiosynthesis and more specifically a novel method for the synthesis of 18F-labelled compounds. The invention also relates to a novel synthon for use in the inventive method of synthesis.
In order to expand the range of applications for positron emission tomography (PET) there is an interest in developing synthetic methods for new PET tracers, i.e. biologically useful compounds labelled with 11C, 18F or 76Br. Currently, the most widely-used of these radiotracers for PET imaging is 18F.
Typically, the synthesis of a PET tracer including its purification should be completed within three half-lives of the radiotracer. 18F has a relatively short half-life of 109.7 minutes and as such methods for its incorporation into a PET tracer demands fast and high-yielding reactions that can be performed on a small scale and under mild conditions.
Direct labelling is desirable as it introduces 18F at the last possible step. However, direct labelling tends only to be possible using [18F]fluoride in a nucleophilic substitution reaction can require the presence of activating groups, proton-free conditions and typically high temperatures of above 100° C. The reader is referred to Coenen (“PET Chemistry: The Driving Force in Molecular Imaging”, Ernst Schering Research Foundation Workshop 62, Schubiger et al, Eds; Springer 2007 pp 15-50) for more detail on typical direct labelling reaction conditions.
Alternatively, 18F can be introduced as part of a synthon. With this approach, an activated precursor is radiofluorinated, and used in subsequent reactions to prepare the desired radiofluorinated product. Many classes of synthons are known for the introduction of an 18F-labelled aromatic group, e.g. [18F]fluorobenzaldehydes, [18F]fluoroarylketones, [18F]fluorobenzoic acid, [18F]fluoronitrobenzene, [18F]fluorobenzonitrile, [18F]fluorosulfonyl arenes, and [18F]fluorohalobenzenes. These classes of synthons, methods to obtain them, and how they can be converted into PET tracers are described in a review by Ermert and Coenen (2010 Current Radiopharmaceuticals; 3: 127-160).
18F-labelled fluoropyridines have found increasing application in PET imaging, and strategies to obtain these compounds are gaining increasing attention. A review by Dolle (2005 Curr Pharm Des; 11: 3221-3235) describes how a variety of [18F]fluoropyridyl-containing compounds can be obtained by nucleophilic heteroaromatic substitution at the ortho position with [18F]fluoride. A specific example of this labelling strategy is reported by Roger et al (2006 J Label Comp Radiopharm; 49: 489-504), who describe the synthesis of 2-exo-(2′-[18F]fluoro-3′-(4-fluorophenyl)-pyridin-5′-yl)-7-azabicyclo[2.2.1]heptane by nucleophilic aromatic substitution of a precursor compound as follows:
wherein X in the scheme represents Cl or Br, with overall radiochemical yields reported as 8-9%. 4-[18F]fluoropyridyl derivatives can also be obtained using such an approach, but not feasibly for 3-[18F]fluoropyridyl derivatives where very strongly electron-withdrawing groups would need to be present, and even then the reaction would likely be low-yielding.
Abrahim et al (2006 J Label Comp Radiopharm; 49: 345-356) report the synthesis of 5-[18F]fluoro-2-pyridinamine and 6-[18F]fluoro-2-pyridinamine. In this approach a carbonyl was used para to a bromine leaving group to obtain the para radiofluorinated intermediate in 20-30% radiochemical yields as follows:
In the initial attempts to obtain the 5-[18F]fluoro-2-pyridinamine synthon using a nitro starting compound, Abrahim reported obtaining 5-bromo-2-[18F]fluoropyridine as an unwanted side-product and consequently abandoned this approach.
LaBeaume et al (2010 Tet Letts; 51: 1906-1909) describe microwave-assisted methods for direct fluorination of nitro intermediates to obtain fluorinated compounds. A variety of nitro substrates were fluorinated using the methods described, including 2-bromo-6-nitropyridine, which was fluorinated with an excess of tetrabutylammonium fluoride (TBAF), yielding >95% 2-bromo-6-fluoropyridine. LaBeaume highlights that as the method gives good to excellent yields in less than 10 minutes, it is practical for use in the preparation of 18F-labelled ligands for PET imaging. LaBeaume notes that where conventional heating was tried in place of microwave heating the conversion to fluorinated product took up to −4 hours, which would clearly be unsuitable for the successful production of an 18F-labelled compound.
As a further alternative, Carroll et al (2007 J Label Comp Radiopharm; 50: 452-454) suggested diaryliodonium salts as a more generic route to obtain 3-fluoropyridines as this approach has been shown to place little or no restriction on the aromatic substituents, allowing it be used much later in the synthetic sequence as compared with the earlier-reported techniques. Radiochemical yields of 55-63% for 3-[18F]fluoropyridine were reported in this paper.
In addition various reports have discussed 18F-labelled synthons for use in obtaining 18F-labelled macromolecules. These are illustrated below:
Olberg et al (2010 J Med Chem; 53: 1732) report the use of F-Py-TFP (6-[18F]fluoronicotinic acid 2,3,5,6-tetrafluorophenyl ester) for peptide coupling reactions. Dolle et al (2003 J Label Comp Radiopharm; 46: S15) report the use of FPyME ([18F]fluoropyridine maleimide) for linking to thiol groups, in particular on peptides. Kuhnast et al (2008 J Label Comp Radiopharm; 51: 336) describe FPyKYNE (2-[18F]Fluoro-3-pent-4-ynyloxy-pyridine) for use in click reactions with macromolecules. Kuhnast et al (2004 Bioconj Chem; 15: 617) describe the design and use of FPyBrA (2-bromo-N-[3-(2-[18F]fluoropyridin-3-yloxy)propyl]acetamide), a [18F]fluoropyridine based halo-acetamide reagent for the labelling of oligonucleotides. Each of these synthons is useful for obtaining 18F-labelled macromolecules, but due to their relative complexity may change the physicochemical properties of a small molecule if used to add 18F.
Alternative means to obtain synthons useful in the synthesis of a broader range of 18F-labelled pyridine-containing compounds would be desirable.
Provided by the present invention is a novel method for obtaining an 18F-labelled compound wherein said compound comprises an 18F-labelled pyridyl ring. The method of the invention is advantageous over the prior art methods as it provides these compounds in higher radiochemical yields than have been possible with previous methods. Also provided by the present invention is an 18F-labelled synthon useful in the method of the invention.
In one aspect, the present invention provides a method for [18F] labelling synthesis comprising reacting a radiolabelling precursor of Formula X:
The term “synthon” refers to a constituent part of a molecule to be synthesised which is regarded as the basis of a synthetic procedure.
[18F]Fluoride used in providing the 18F-labelled synthon of Formula Y is typically obtained as an aqueous solution which is a product of the irradiation of an [18O]-water target. Various steps are carried out on the aqueous solution to convert [18F]fluoride into a reactive nucleophilic reagent, such that it is suitable for use in nucleophilic radiolabelling reactions. These steps include the elimination of water and the provision of a suitable counterion (Handbook of Radiopharmaceuticals 2003 Welch & Redvanly eds. ch. 6 pp 195-227).
Suitable counterions include large but soft metal ions such as rubidium or caesium, potassium complexed with a cryptand such as Kryptofix™, or tetraalkylammonium salts.
In a most preferred embodiment, the synthon of Formula Y is either of the following:
The relevant dibromo-substituted pyridines are commercially-available. For example, 2-Bromo-6[18F]-fluoropyridine, can be readily prepared from 2,6-dibromopyridine. The present inventors have done so in 10 minutes at an end of synthesis (EOS) non-decay corrected yield of 53%.
In a preferred embodiment, the method of the present invention further comprises the step:
The term “cross-coupling partner” refers to a compound that can react with the synthon of Formula Y with the elimination of the synthon bromo leaving group to result in a desired 18F-labelled product. The cross-coupling partner therefore suitably comprises a chemical group that effects nucleophilic displacement of the bromo of the synthon. Non-limiting examples of such chemical groups include terminal alkene, amino, terminal alkyne, boronic acid, and organotin.
By the term “terminal alkene” is meant a double bond at the terminal end of a substituent. A preferred cross-coupling partner comprising a terminal alkene is a compound of Formula Ia as defined below.
The term “amino” refers to the group NR2 wherein each R is hydrogen or a monovalent aliphatic or aromatic hydrocarbon substituent, as defined below. Preferably at least one R is hydrogen. A preferred cross-coupling partner comprising an amine is a compound of Formula Ie as defined below.
The term “terminal alkyne” refers to a triple bond at the terminal end of a substituent. A preferred cross-coupling partner comprising a terminal alkyne is a compound of Formula Ic as defined below.
The term “boronic acid” refers to the group —B(OH2). A preferred cross-coupling partner comprising boronic acid is a compound of Formula Id as defined below.
The term “organotin” refers to a chemical group comprising tin and hydrocarbon substituents. Organotin compounds are also referred to as stannanes. A preferred cross-coupling partner comprising an organotin is a compound of Formula Ib as defined below.
The coupling reaction of step (ii) of the preferred embodiment of the invention is preferably site-specific and may consequently require the presence of one or more protecting groups on the cross-coupling partner. 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 to obtain the desired product under mild enough conditions that do not modify the rest of the molecule. Protecting groups are well known to those skilled in the art and are described in ‘Protective Groups in Organic Synthesis’, Theorodora W. Greene and Peter G. M. Wuts, (Fourth Edition, John Wiley & Sons, 2007).
The term “transition metal” includes palladium, platinum, gold, ruthenium, rhodium, and iridium. The most typically used transition metal for the coupling reactions encompassed by step (ii) of the method of the invention is palladium. Typical forms of palladium for use as a catalyst include palladium acetate, tetrakis(triphenylphosphine)palladium(0), bis(triphenylphosphine)palladium(II) dichloride, [1,1′-bis(diphenylphosphino)ferrocene]palladium(II) dichloride, and palladium on carbon (Pd/C).
Preferably, the 18F-labelled product obtained in step (ii) is a tracer suitable for PET imaging and preferably has a molecular weight <1500 Daltons; preferably <1000 Daltons. Where the 18F-labelled product is intended to be a PET tracer for imaging the central nervous system, the molecular weight is preferably <500, which is optimal for blood-brain barrier penetration.
As reported by Ermert and Coenen (2010 Current Radiopharmaceuticals; 3: 127-160), [18F]fluorohalobenzenes can be converted into a range of different target 18F-labelled molecules by means of transition metal-mediated coupling reactions, as illustrated in Scheme 1 below:
In Scheme 1, X represents bromo, chloro or iodo, and A-E represent:
Each of these transition-metal catalysed reactions are well-known to the skilled person and are described e.g. in “March's Advanced Organic Chemistry: Reactions, Mechanisms, and Structures (6th Edition Wiley 2007, Smith and March, Eds.); see page 792 for the Stille reaction, page 875 for Hartwig-Buchwald N-arylation, page 904 for the Sonogashira reaction and page 899 for Suzuki coupling. The [18F]-fluorobromopyridine synthon provided in step (i) of the method of the present invention can therefore be converted into a range of 18F-labelled products using these same reactions. The method of the present invention therefore allows for the synthesis of a wide range of 18F-labelled heteroaromatic PET tracers.
In one preferred embodiment of the method of the invention, said transition metal coupling reaction comprises reaction of the 18F-labelled synthon of Formula Y with a compound of Formula Ia:
The term “monovalent aliphatic hydrocarbon group” used here and elsewhere in the specification is intended to encompass substituted or unsubstituted linear, branched or cyclic alkyl, alkenyl, or alkynyl radicals, wherein one or more of the carbons in the chain is optionally a heteroatom selected from O, S and N. The term “alkyl” refers to monovalent radical having the general formula CnH2n+1, the term “alkenyl” refers to an alkyl comprising one or more double bonds, and the term “alkynyl” refers to an alkyl comprising one or more triple bonds. The term “aliphatic” relates to those parts of the radical arranged in straight or branched chains, and not containing aromatic rings.
The term “monovalent aromatic hydrocarbon group” used here and elsewhere in the specification is intended to encompass substituted or unsubstituted radicals containing one or more six-carbon rings characteristic of the benzene series and related organic groups, wherein one or more of the carbons is optionally a heteroatom selected from O, S and N.
The term also includes radicals comprising aliphatic elements in addition, wherein the aliphatic elements can be monovalent aliphatic hydrocarbon groups as defined above, or divalent derivatives thereof, provided that the designated atom's normal valency under the existing circumstances is not exceeded.
The term “substituted” as used throughout the specification means that one or more hydrogens on a designated atom is replaced with a substituent, provided that the designated atom's normal valency under the existing circumstances is not exceeded, and that the substitution results in a stable compound. Combinations of substituents and/or variables are permissible only if such combinations result in stable compounds. The term “stable compound” is meant a compound that is sufficiently robust to survive isolation to a useful degree of purity from a reaction mixture.
Non-limiting examples of “substituents” include, halo groups, hydroxy groups, oxo groups, mercapto groups, amino groups, carbamoyl groups, carboxyl groups, cyano groups, nitro groups, acyl groups, phosphate groups, sulfamyl groups, sulfonyl groups, sulfinyl groups, and combinations thereof. A substituent can also be a substituted or unsubstituted monovalent aliphatic or aromatic hydrocarbon group as defined above.
As used herein, the term “halo” or “halogen” means refers to chlorine, bromine, fluorine or iodine.
The term “oxo” refers to the group ═O.
The term “mercapto” refers to the group —SH, which is also known as thiol or sulfhydryl.
The term “carbamoyl” refers to the group —C(═O)NH2.
The term “carboxyl” refers to the group —C(═O)OH.
The term “cyano” refers to the group —C≡N.
The term “nitro” refers to the group —NO2.
The term “acyl” refers to the group —C(═O)-alkyl wherein alkyl is as defined above.
The term “phosphate” refers to the group —O—P(OH)3.
The term “sulfamyl” refers to the group —S(═O)2-amino wherein amino is as defined above.
The term “sulfonyl” refers to the group —S(═O)2-alkyl wherein alkyl is as defined above.
The term “sulfinyl” refers to the group —S(═O)-alkyl wherein alkyl is as defined above.
In another preferred embodiment, in the method of the invention, said transition metal coupling reaction comprises reaction of the 18F-labelled synthon of Formula Y with a compound of Formula Ib:
Bu3SnR2 (Ib)
In a further preferred embodiment, in the method the invention said transition metal coupling reaction comprises reaction of the 18F-labelled synthon of Formula Y with a compound of Formula Ic:
In another further preferred embodiment, in the method the invention said transition metal coupling reaction comprises reaction of the 18F-labelled synthon of Formula Y with a compound of Formula Id:
Example 4 describes such a reaction.
In a yet further preferred embodiment, in the method the invention said transition metal coupling reaction comprises reaction of the 18F-labelled synthon of Formula Y with a compound of Formula Ie:
Example 2 describes such a reaction.
In the case of each of the 18F-labelled products of Formulas IIa-IIe, the suitable and preferred positions for 18F and for each substituent are as defined respectively for 18F and 8r in the synthon of Formula Y.
The term “nitrogen-containing aliphatic or aromatic ring” refers to any substituted or unsubstituted cyclic substituent that comprises at least one nitrogen heteroatom, preferably having between 4-7 carbon atoms, most preferably between 4-5 carbon atoms. It is preferred that such rings have between 1-3, and most preferably between 1-2 nitrogen heteroatoms.
In an even further preferred embodiment, in the method the invention said transition metal coupling reaction comprises reaction of the 18F-labelled synthon of Formula Y with the above-defined compound of Formula Ie in the presence of a source of carbon monoxide to obtain an 18F-labelled product of Formula IIf:
Example 3 relates to such a reaction.
The alternative known synthetic routes to form the above specific classes of compounds are relatively low-yielding as compared with the method of the present invention. For example, to obtain the compound of Formula IIf, one known method is via direct labelling, although this can be prohibitively low yielding in unactivated substrates. An alternative known method is a multi-stage activated ester strategy which is not straightforward to implement.
In a preferred embodiment, the method of the invention is automated, preferably on an automated synthesiser. [18F]-radiotracers are now often conveniently prepared on an automated radiosynthesis apparatus. There are several commercially-available examples of such apparatus, including Tracerlab™ and Fastlab™ (both from GE Healthcare Ltd). Such apparatus commonly comprises a “cassette”, often disposable, in which the radiochemistry is performed, which is fitted to the apparatus in order to perform a radiosynthesis. The cassette normally includes fluid pathways, a reaction vessel, and ports for receiving reagent vials as well as any solid-phase extraction cartridges used in post-radiosynthetic clean up steps. The present invention therefore provides in another aspect a cassette for carrying out the automated method of the invention wherein said cassette comprises:
The cassette preferably also comprises:
Where the cassette comprises the vessel comprising a compound of any one of Formula Ia-e, this vessel is eluted with the purified product of the reaction between the precursor compound of Formula X and [18F]fluoride, i.e. the synthon of Formula Y as defined herein. Purification is typically carried out by solid phase extraction on the cassette.
The cassette may also additionally comprise:
Example 1 describes the Preparation of 2-Bromo-6[18F]-fluoropyridine.
Example 2 describes the preparation of 1-benzyl-4-(6-[18F]fluoropyridin-2-yl)piperazine.
Example 3 describes the preparation of N-benzyl-6-[18F]fluoropicolinamide.
Example 4 describes the preparation of 2-[18F]fluoro-6-(p-tolyl)pyridine.
Example 5 describes the preparation of 3-bromo-5-[18F]fluoropyridine.
Example 6 describes the preparation of 3-[18F]fluoro-5-(p-tolyl)pyridine.
Ac acetyl
BINAP 2,2′-bis(diphenylphosphino)-1,1′-binaphthalene
dba dibenzylideneacetone
DBU 1,8-diazabicyclo[5.4.0]undec-7-ene
DMF dimethylformamide
DMSO dimethylsulfoxide
HPLC high performance liquid chromatography
MeCN acetonitrile
NEt3 triethylamine
Ph phenyl
THF tetrahydrofuran
UV ultraviolet
Experiments were undertaken to explore the optimum reaction conditions for preparing 2-bromo-6-[18F]fluoropyridine from 2,6-dibromo pyridine.
All reactions were performed by conventional heating for ten minutes and the resulting 2-bromo-6-[18F]fluoropyridine was purified by semi-preparative HPLC using the following method:
MOBILE phase A=H2O
Flow rate 3 mL/min
Gradient 0-15 min, 5-95% B
In the table below, yields (from fluoride) are of the isolated product after HPLC purification, with yields in brackets being decay corrected.
2.0
DMF
100
53 (79)
64
The reaction highlighted in bold in the above table resulted in the highest yield.
A Buchwald-Hartwig coupling reaction was tested with 1-benzylpiperazine, using tris(dibenzylideneacetone) dipalladium(0) and (±)BINAP with sodium t-butoxide in MeCN. After 25 min heating at 100° C., 49% of the activity was the desired product. The analytical HPLC using the following method:
Mobile phase A=0.8% NEt3 in H2O, corrected to pH ˜7.5 with H3PO4
Mobile phase B=MeCN
Flow rate=1 mL/min
The traces are displayed in
A reaction using molybdenum(0) hexacarbonyl as CO source was performed. In a procedure based on that described by Wannberg et al (2003 J Org Chem; 68: 5750), palladium acetate, molybdenum hexacarbonyl, benzylamine and 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU) were added and the reaction was heated at 100° C.
Analytical HPLC was carried out using the following method:
Mobile phase A=0.8% NEt3 in H2O, corrected to pH ˜7.5 with H3PO4
Mobile phase B=MeCN
Flow rate=1 mL/min
Traces of the aminocarbonylation reaction after 5, 15, and 30 min heating are displayed in
Analytical HPLC data a. Radio-HPLC 5 min, b. Radio-HPLC 15 min c. Radio-HPLC 30 min d. UV-HPLC of cold standard.
The absence of any [18F]-Buchwald-Hartwig product in the radiolabelling reaction (formed without insertion of CO) can be noted, demonstrating the efficiency of the aminocarbonylation reaction.
A Suzuki coupling was performed with p-tolylboronic acid, using tetrakis (triphenylphosphino) palladium and sodium carbonate in H2O-acetonitrile mixture. After 5 min heating at 100° C., 98% of the activity was the desired product. Analytical HPLC was carried out using the following method:
Mobile phase A=0.8% NEt3 in H2O, corrected to pH ˜7.5 with H3PO4
Mobile phase B=MeCN
Flow rate=1 mL/min
HPLC traces of the unpurified reaction is shown in the
Experiments to assess the [18F]fluorine labelling in the 3-position were undertaken.
Several experiments to explore various reaction conditions were performed. Yields given are after HPLC purification of the synthon (with the exception of entry 1). The HPLC method was as follows:
Mobile phase A=H2O
Flow rate 3 mL/min
The highest yielding reaction was performed as follows:
3,5-dibromopyridine (3.0 mg) was added to dried [18F]fluoride/kryptofix/potassium carbonate in DMSO and subjected to microwave heating (50 W) for 1 min. After purification by semi-preparative HPLC, the isolated non-decay corrected yield from fluoride was 16%.
Suzuki coupling of 3-bromo-5[18F]fluoropyridine was performed with p-tolylboronic acid, using tetrakis(triphenylphosphino) palladium and sodium carbonate in an acetonitrile-H2O mixture. After 5 min heating at 100° C., 82% of the activity was the desired product.
Semi-preparative radio-HPLC was carried out as follows:
Flow rate 3 mL/min
Analytical HPLC was carried out as follows:
Mobile phase A=0.8% NEt3 in H2O, corrected to pH ˜7.5 with H3PO4
Mobile phase B=MeCN
Flow rate=1 mL/min
a shows the semi-preparative radio-HPLC trace of the Suzuki coupling reaction of p-tolylboronic acid and 3-bromo-5-[18F]fluoropyridine after 5 min at 100° C. Rt desired product=14.1 min.
| Number | Date | Country | Kind |
|---|---|---|---|
| 1103455.0 | Mar 2011 | GB | national |
| Filing Document | Filing Date | Country | Kind | 371c Date |
|---|---|---|---|---|
| PCT/EP2012/053547 | 3/1/2012 | WO | 00 | 8/28/2013 |
| Number | Date | Country | |
|---|---|---|---|
| 61447724 | Mar 2011 | US |
