“In-loop” carbonylation—A simplified method for carbon-11 labelling of drugs and radioligands

Article abstract of DOI:10.1002/jlcr.3805

Transition-metal mediated carbonylation with ¹¹C-labelled carbon monoxide ([¹¹C]CO) is a versatile method for introducing ¹¹C (t_1/2 = 20.3 min) into drugs and radioligands for subsequent use in positron emission tomography (PET). The aim of the current study was to perform the ¹¹C-carbonylation reaction on the interior surface of a stainless-steel loop used for high performance liquid chromatography (HPLC). In the experimental setup, cyclotron produced ¹¹C-labelled carbon dioxide ([¹¹C]CO₂) was converted to [¹¹C]CO by reduction over heated Molybdenum and swept into an HPLC loop pre-charged with the appropriate reaction mixture. Following a 5 min reaction, the radiochemical purity (RCP) and the trapping efficiency (TE) of the reaction mixture was determined. After optimization, [¹¹C]N-Benzylbenzamide was obtained in quantitative radiochemical yield (RCY) following a 5 min reaction at room temperature. The methodology was further applied to label [¹¹C]benzoic acid (RCP≥99%, TE>91%), [¹¹C]methyl benzoate (RCP≥99%, TE>93%) and [¹¹C]phthalide (RCP≥99%, TE>88%). A set of pharmaceuticals was finally radiolabelled using non-optimized conditions. Excellent yields were obtained for the histamine-3 receptor radioligand [¹¹C]AZ13198083, the oncology drug [¹¹C]olaparib and the dopamine D2 receptor radioligand [¹¹C]raclopride, whereas a moderate yield was observed for the high-affinity dopamine D2 receptor radioligand [¹¹C]FLB457. The presented “in-loop” process proved efficient for diverse ¹¹C-carbonylations, providing [¹¹C]amides, [¹¹C]esters and [¹¹C]carboxylic acids in moderate to excellent RCYs. Based on the advantages associated with performing the radiolabelling step as an integrated part of the purification system, this methodology may become a valuable addition to the toolbox of methodologies used for ¹¹C-carbonylation of drugs and radioligands for PET.

Full text of DOI:10.1002/jlcr.3805

Ferrat Mélodie (Orcid ID: 0000-0001-7004-3565)
Dahl Kenneth (Orcid ID: 0000-0003-2948-2042)
“In-loop” carbonylation – a simplified method for carbon-11 labeling of drugs and
radioligands
Mélodie Ferrata*, Kenneth Dahlb, Christer Halldina and Magnus Schoua,c
aDepartment of Clinical Neuroscience, Center for Psychiatry Research, Karolinska Institutet
and Stockholm County Council, SE-171 76 Stockholm, Sweden
b CAMH & University of Toronto, Toronto, ON M5T-1R8, Canada
c
AstraZeneca PET Science Centre, Department of Clinical Neuroscience, Karolinska
Institutet, S-171 76 Stockholm, Sweden
*Author to whom correspondence should be addressed: Email: melodie.ferrat@ki.se
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been through the copyediting, typesetting, pagination and proofreading process which may
lead to differences between this version and the Version of Record. Please cite this article as
doi: 10.1002/jlcr.3805
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Abstract:
Transition-metal mediated carbonylation with 11C-labelled carbon monoxide ([11C]CO) is a
versatile method for introducing 11C (t1/2 = 20.3 min) into drugs and radioligands for
subsequent use in positron emission tomography (PET). The aim of the current study was to
11
perform the C-carbonylation reaction on the interior surface of a stainless-steel loop used
for high performance liquid chromatography (HPLC). In the experimental setup, cyclotron
produced 11C-labelled carbon dioxide ([11C]CO2) was converted to [11C]CO by reduction over
heated Molybdenum and swept into an HPLC loop pre-charged with the appropriate reaction
mixture. Following a 5 min reaction, the radiochemical purity (RCP) and the trapping
efficiency (TE) of the reaction mixture was determined. After optimization, [11C]N-
Benzylbenzamide was obtained in quantitative radiochemical yield (RCY) following a 5 min
reaction at room temperature. The methodology was further applied to label [11C]benzoic
acid (RCP≥99%, TE>91%), [11C]methyl benzoate (RCP≥99%, TE>93%) and [11C]phtalide
(RCP≥99%, TE>88%). A set of pharmaceuticals was finally radiolabelled using non-
optimized conditions. Excellent yields were obtained for the histamine-3 receptor radioligand
[11C]AZ13198083, the oncology drug [11C]olaparib and the dopamine D2 receptor
radioligand [11C]raclopride, whereas a moderate yield was observed for the high-affinity
dopamine D2 receptor radioligand [11C]FLB457. The presented “in-loop” process proved
efficient for diverse 11C-carbonylations, providing [11C]amides, [11C]esters and
[11C]carboxylic acids in moderate to excellent RCYs. Based on the advantages associated
with performing the radiolabelling step as an integrated part of the purification system, this
11
methodology may become a valuable addition to the toolbox of methodologies used for C-
carbonylation of drugs and radioligands for PET.
Keywords: Loop, 11C-carbonylation, Carbon-11, Radioligands, Radiopharmaceutical
chemistry
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1. INTRODUCTION
Positron emission tomography (PET) is a nuclear medicine imaging technique frequently
used in translational research and drug development. After injection of a radiolabelled
compound, PET produces a series of three-dimensional images of the tracer tissue
distribution inside the living organism. A fundamental pre-requisite for PET is that the tracer
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molecule is labelled with a positron emitting radionuclide, such as C (t1/2 = 20.3 min). An
11
12
important advantage with C is that it can be introduced in the place of C in bio-organic
compounds without any structural modification of the target compound.
Due to the short half-life of carbon-11, its radiochemistry needs to be rapid, efficient and
preferably automated to minimize radiation exposure to the radiochemist. The radionuclide is
commonly produced by a medical cyclotron in the form of [11C]methane or [11C]carbon
dioxide. Because of the convenient conversion of these species into [11C]methyl iodide,1,2 the
radionuclide is most commonly introduced into tracer molecules via 11C-methylation.
Although methyl groups are common in bioactive molecules, the majority lack this structural
motif, which makes other labelling methods necessary to avoid structural modification of the
tracer molecule.
Carbonyl groups are present in most biologically active molecules and are featured in a large
number of drug molecules. Significant efforts have thus been put into the development of
methods that use [11C]carbon monoxide ([11C]CO), available in one step from [11C]CO2, to
11
introduce the C-carbonyl motif into tracer molecules. Radiochemistry with [11C]CO is far
from trivial, however, because of its low reactivity, poor solubility in organic solvents, high
dilution in inert gas and the low concentration of isotopically labelled [11C]CO.3 Many of
these challenges have been successfully addressed via recirculating [11C]CO through the
11
reaction media,4 by performing the C-carbonylation in a micro-autoclave (at 350 bar)3,5 or
inside a microtube reactor.6 Other fruitful methods for improving the yield for these reactions
include chemical complexation of [11C]CO with diborane (BH3-[11C]CO),7 copper
scorpionates (Cu-Sc-[11C]CO),8 or introducing [11C]CO into the reaction media in a stream of
Xenon carrier gas.9 These developments have resulted in numerous PET studies with tracer
molecules labelled using palladium mediated 11C-carbonylation chemistry.10,11,12,13,14,15
In an effort to simplify 11C-carbonylation reactions from a technical perspective, we were
inspired by previous work that employed captive solvents and reagents confined within
plastic or stainless-steel tubes for the radiochemical synthesis.16,17 It was considered that such
an approach, which maximizes the surface area between the gas and liquid phase, may be
ideally suited for [11C]CO radiochemistry because of the increased opportunity for the
radioactive gas to equilibrate with solution. In addition, an approach in which the HPLC loop
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is the reaction vessel for the C-carbonylation was considered ideal, since transfer losses in
such case are minimized and process automation simplified according to Wilson et al.17 Thus,
the aim of this study was to develop a novel “in-loop” method for PET tracer synthesis via
11C-carbonylation using [11C]CO (Fig. 1).
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RESULTS AND DISCUSSION
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An initial requirement for most “in-loop” C-labeling processes is that the product must be
formed in a one-pot, one-step procedure. We thus started our investigation on the “in-loop”
11C-aminocarbonylation of iodobenzene with benzylamine, forming the product [11C]N-
benzylbenzamide, a reaction that has been well characterized previously.18 DMF was first
tested as a solvent for the reaction because of its captive properties and compatibility with
“in-loop” 11C-methylation.17 Likewise, the Pd-xantphos catalytic system was used as a
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starting point for the optimization, since it has been shown to be efficient in promoting C-
aminocarbonylation reactions.18,19 To our delight, a 5 min reaction at room temperature with
Pd-xantphos in DMF furnished [11C]N-benzylbenzamide at high trapping efficiency (TE),
albeit at a moderate radiochemical purity (RCP) (Table 1, entry 1). Other solvents, such as
DMSO and toluene, also provided good TEs in the “in-loop” 11C-carbonylation (Table 1,
entries 2 and 3), but the best yields were obtained in 1,4-dioxane and THF with near
quantitative radiochemical yields (RCYs) (Table 1, entries 4 and 5). It is worth noting that an
autoradiography experiment revealed that the majority of the radioactivity was trapped in the
first quarter (~0.5 mL) of the stainless steel loop (Fig. 2). In comparison, a control reaction
with Pd(PPh3)4 provided [11C]N-benzylbenzamide in 70% radiochemical purity, but with a
poor TE (0.2%) and thus low RCY (0.14%).
With the improved conditions in hand, the methodology was next applied to radiolabel
functional groups other than amides. Thus, a lactone, an ester and a carboxylic acid, were
each radiolabelled with only minor modifications to those previously used in the
radiosynthesis of [11C]N-benzylbenzamide (Scheme 1). Modifications to the original
procedure include an increase in aryl halide concentration to improve trapping efficiency for
[11C]phtalide, and an increased temperature to improve the RCP of [11C]methyl benzoate. The
observed yields from these reactions are on par with those observed for reactions performed
in either vial or in micro-reactor setups, thus demonstrating that the utility of the developed
“in-loop” 11C-carbonylation method extends beyond aminocarbonylation.18,20
The “in-loop” carbonylation was finally tested on a set of druglike molecules (Scheme 2).
Thus, carbon-11 radiolabelling of the histamine type-3 receptor radioligand AZ13198083, the
oncology drug olaparib and the dopamine D2 receptor radioligands raclopride14 and FLB457
were performed. Again, some modifications to the developed procedure were important to
achieve good RCYs for these compounds. Most importantly, heating was required for all
substrates to obtain the target tracers at high RCP. In addition, as was previously
demonstrated by Skrydstrup et al., [11C]olaparib cannot be produced with the Pd-xantphos
system because of a phenyl scrambling from xantphos onto the aryl palladium complex.21
Instead, the Pd-tBu3 system was used to produce the target compound, thus demonstrating
that other catalytic systems than the Pd-xantphos system are also compatible with the “in-
loop” 11C-carbonylation methodology. It is noteworthy that [11C]FLB457 could only be
produced at a moderate TE under the employed conditions, but this may well be improved
following further optimization. Finally, after integrating the loop system into a commercially
available radiochemistry system (GE tracerlab FX-C, Uppsala, Sweden), [11C]AZ13198083
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was obtained in a 38% decay-corrected RCY. The isolated radiochemical yield was lower
than expected because of losses during HPLC isolation, formulation and sterile filtration.
However, the RCP was >99% and the molar activity was 110 GBq/µmol.
CONCLUSION
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The presented “in-loop” process proved to be useful for diverse C-carbonylation, providing
both simple reference compounds (e.g. [11C]amides, [11C]esters and [11C]carboxylic acids) as
well as PET tracer molecules (e.g. [11C]raclopride, [11C]olaparib) in moderate to excellent
RCYs. Given its simplicity and efficiency, the methodology has the potential to be widely
implemented for carbonylations in PET tracer synthesis.
MATERIALS AND METHODS
General experimental information
All chemical and solvents were obtained from Sigma-Aldrich (Sweden) and used without
further purification, unless specified otherwise.
High performance liquid chromatography analysis (HPLC) was performed using a Hitachi L-
6200 gradient pump and a Hitachi L-4000 variable wavelength UV-detector in a series with a
Bioscan β+-flow detector. Analytical HPLC analysis was performed using a reverse phase
column (XBridge, C18, 5 μm, 4.6 х 150 mm):
a- For [11C]N-benzylbenzamide, eluted with a gradient between acetonitrile (A) and
0.1% TFA (B). The gradient was linear between 10 - 90% over 5 minutes and
isocratic in between 5-8 min (ACN: 0.1% TFA, 90:10), at a flow rate of 3 ml/min.
b- [11C]phtalide, eluted with a gradient between acetonitrile (A) and 0.1 M NH4HCO2
(B). The gradient was linear between 10 - 90% over 9 minutes and isocratic in
between 9-10 min (ACN: 0.1 M NH4HCO2, 90:10), at a flow rate of 2 ml/min.
c- [11C]methyl benzoate, eluted with a gradient between acetonitrile (A) and 0.1% TFA
(B). The gradient was linear between 10 - 90% over 5 minutes and isocratic in
between 5-10 min (ACN: 0.1% TFA, 90:10), at a flow rate of 3 ml/min.
d- [11C]benzoic acid, eluted with a gradient between acetonitrile (A) and 0.1% TFA (B).
The gradient was linear between 10 - 90% over 7 minutes and isocratic in between 7-
10 min (ACN: 0.1% TFA, 90:10), at a flow rate of 3 ml/min.
e- For [11C]AZ13198083, eluted with a gradient between acetonitrile (A) and 0.1 M
NH4HCO2 (B). The gradient was linear between 10 - 90% over 9 minutes and
isocratic in between 9-10 min (ACN: 0.1 M NH4HCO2, 90:10), at a flow rate of 2
ml/min.
f- For [11C]olaparib, eluted with a gradient between acetonitrile (A) and 0.01 M
NH4HCO2 (B). The gradient was linear between 30 - 70% over 5 minutes and
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isocratic in between 5-6 min (ACN: 0.1 M NH4HCO2, 90:10), at a flow rate of 3
ml/min.
g- For [11C]raclopride, eluted with a gradient between acetonitrile (A) and 0.1% TFA
(B). The gradient was linear between 10 - 90% over 9 minutes and isocratic in
between 9-10 min (ACN: 0.1% TFA, 90:10), at a flow rate of 2 ml/min.
h- For [11C]FLB457, eluted with a gradient between acetonitrile (A) and 0.1 M
NH4HCO2 (B). The gradient was linear between 30 - 70% over 9 minutes and
isocratic in between 9-10 min (ACN: 0.1 M NH4HCO2, 90:10), at a flow rate of 2
ml/min.
The identity of all radioactive products was confirmed by co-elution with the corresponding
non-radioactive compound.
Radiochemistry
All experiments were performed in accordance with local and national rules and laws that
govern the work with open sources of radiation. The study was carried out at the PET Centre,
Karolinska Institutet, (Stockholm, Sweden).
[11C]CO2 was produced by a GEMS PETtrace cyclotron (General Electrics Medical Systems,
Uppsala, Sweden). The 14N(p,α)11C nuclear reaction, performed by proton bombardment of a
pressurized gas target containing nitrogen (AGA, Nitrogen 6.0) and 1% oxygen (AGA,
Oxygen 4.8), was used to produce [11C]CO2. At the end of bombardment (EOB), [11C]CO2
was transferred from the target to a [11C]CO-synthesizer prototype (General Electrics
Medical Systems, Uppsala, Sweden). [11C]CO2 was trapped on a molecular sieve column (0.6
mg packed in a ¼” tube, mesh 80/100, GRACE, USA) at room temperature. The accumulated
[11C]CO2 was released into a controlled stream of helium (10 ml/min) while heating the
molecular sieve trap to 360°C. The purified [11C]CO2 was pre-concentrated by a stainless
steel loop trap immersed in liquid nitrogen (-196˚C) prior to on-line reduction over a heated
Molybdenum column (850°C). The effluent gas was purified through an ascarite® (sodium
hydroxide-coated silica, 20-30 mesh, Sigma-Aldrich, Sweden) column prior to concentrating
the [11C]CO on a silica gel trap immersed in liquid nitrogen. Following completed
entrapment, [11C]CO was released in a stream of Helium into a stainless steel HPLC loop for
11
the C-carbonylation reaction. Untrapped [11C]CO was collected in a gas-tight bag (Tedlar®
gas sampling bag push lock valve 1L, Sigma-Aldrich, Sweden).
In-loop 11C-carbonylation setup
A commercial HPLC injection valve (VICI, #C52-12061, Switzerland) was equipped with a
stainless-steel loop (2 mL, VICI, #SL2KCUW, Switzerland) and an injection port (for 1/16’’
fill port, VICI #VISF-2, Switzerland). Prior to start of synthesis, the injection valve was set to
LOAD position (Fig. 1) and a solution of aryl halide, palladium-source, supporting ligand and
nucleophile in anhydrous solvent, was injected using a 1 mL syringe capped with a filter
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(Acrodisc 13 mm minispike with 0,45 μm GHP Membrane, Sigma Aldrich, Sweden) through
the injection port of the loop. The isolated and concentrated [11C]CO was then swept into the
loop at 5 mL/min, and after a 5 min reaction, the contents of the loop were transferred to a
sealed test tube by purging the loop with THF (~2 mL). After transfer to the test tube, the
radioactivity in the test tube was measured before and after it was purged with N2 (to assess
TE), followed by a measurement of the gas waste bag that was connected on the outlet of the
HPLC loop. The radioactivity in the test tube was also analyzed using radio-HPLC to assess
the RCP that gives information about reaction selectivity. Moreover, as [11C]CO is a
radioactive gas, the parameter TE needs to be taken into account. TE is the total amount of
[11C]CO trapped in the reaction solution that is converted to non-volatile entities at the end of
the reaction. RCY was calculated from the RCP and TE (i.e. RCY = RCP x TE).
Application to GE Tracerlab FX-C
A Tracerlab FX-C system (GE PETtrace, Uppsala, Sweden) was used for semi-preparative
HPLC purification, in parallel with the [11C]CO-synthesizer prototype (General Electrics
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Medical Systems, Uppsala, Sweden) to produce [11C]7. In short, C-carbon monoxide was
transferred from the [11C]CO-system to the stainless-steel loop in the Tracerlab FX-C system
where the reaction was taking place. The crude product was obtained in the HPLC loop and
then purified using a semi-preparative HPLC column (ACE C-18, 5µm, 10 × 250 mm;
Advanced Chromatography Technologies Ltd; Aberdeen, UK) eluted with an isocratic
mobile phase consisting of 30% MeCN in HCO2NH4 (0.1M). The eluate was monitored for
absorbance (
= 254 nm) and for radioactivity. [11C]7 (tR = 8-10 min) was collected and
diluted with water (~50 mL) and then concentrated on a C-18 SepPak (Waters, Sweden). The
SepPak was then washed with water (~8 mL) prior to elution with ethanol (99%, 1.5 mL) into
saline (10 mL). The formulation was finally filtered through a yellow sterile filter (0.22 µm,
Millipore, Millex®GV) to yield the final product in a sterile injection vial.
Synthesis of [11C]N,N-benzylbenzamide
An oven-dried disposable 4 mL vial was equipped with a screw thread cap, a rubber septum
and cooled under Nitrogen (N2) atmosphere. Pd2[π-cinnamyl]Cl2 (2.3 mg; 1eq), xantphos (5,0
mg; 2 eq), iodobenzene (1.4 μL, 1.42 eq) were dissolved in anhydrous THF (700 μL) and the
resulting mixture was flushed with N2 until the THF was fully evaporated (20 min). 1,4-
Dioxane (150 μL) and Benzylamine (10 μL) were added and the reaction mixture was
introduced into the HPLC loop approximately 2 min prior to start of synthesis. The resulting
mixture was allowed to react for 5 min at r.t.
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Synthesis of [11C]Phtalide
Pd2[π-cinnamyl]Cl2 (6.7 mg; 3 eq), xantphos (7.5 mg; 3 eq) and iodobenzylalcohol (3.0 μL, 3
eq) were dissolved in anhydrous THF (700 μL) kept at r.t. for 15 min and then heated at 75˚C
during 5 min. THF (150 μL) was added and the resulting mixture was introduced into the
HPLC loop approximately 2 min prior to start of synthesis. The resulting mixture was
allowed to react for 5 min at 75˚C.
Synthesis of [11C]Methyl benzoate
Pd2[π-cinnamyl]Cl2 (2.3 mg; 1eq), xantphos (5.0 mg; 2eq) and iodobenzene (2.2 μL, 2.9 eq)
were dissolved in anhydrous THF (700 μL) and the resulting mixture was flushed with N2
until the THF was fully evaporated (20 min). THF (150 μL) and a solution of lithium
methoxide in MeOH (1M, 40 μL) were added and the reaction mixture was introduced into
the HPLC loop approximately 2 min prior to start of synthesis. The resulting mixture was
allowed to react for 5 min at 120˚C.
Synthesis of [11C]Benzoic acid
Pd2[π-cinnamyl]Cl2 (2.2 mg), xantphos (5.0 mg) and iodobenzene (1.4 μL) were dissolved in
anhydrous THF (700 μL) and the resulting mixture was flushed with N2 until the THF was
fully evaporated (20 min). 1,4-Dioxane (150 μL) and a solution of NaOH (0.35 M, 3.5 μL)
were added and the reaction mixture was introduced into the HPLC loop approximately 2 min
prior to start of synthesis. [11C]CO was swept at r.t to the loop in which the coupling reagents
were already added. The resulting mixture was allowed to react for 5 min at r.t.
Synthesis of [11C]AZ13198083
Pd2[π-cinnamyl]Cl2 (2.2 mg;
1
eq), xantphos (5.0 mg;
2
eq), ((2S)-2-(4-
bromophenyl)cyclopropyl)(4-cyclobutylpiperazin-1-yl)methanone (3.3 μL) were dissolved in
anhydrous THF (700 μL) and the resulting mixture was flushed with N2 until the THF was
fully evaporated (20 min). A solution of ethylamine in THF (2M, 600 μL) was added and the
reaction mixture was introduced into the HPLC loop approximately 2 min prior to start of
synthesis. The resulting mixture was allowed to react for 5 min at 100˚C.
Synthesis of [11C]Olaparib
Pd(dba)2 (11.5 mg), PtBu3 (4.9 μL) and 4-(4-fluoro-3-iodobenzyl)phthalazin-1(2H)-one (7.6
mg) were dissolved in anhydrous THF (700 μL) and the resulting mixture was flushed with
N2 until the THF was fully evaporated (20 min). THF (150 μL) and cyclopropyl(piperazin-1-
yl)methanone (10.1 μL) were added and the reaction mixture was introduced into the HPLC
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loop approximately 2 min prior to start of synthesis. The resulting mixture was allowed to
react for 5 min at 100˚C.
Synthesis of [11C]Raclopride
Pd2[π-cinnamyl]Cl2 (6.4 mg; 1 eq), xantphos (7.1 mg; 1eq) and 4,6-dichloro-2-iodo-3-
methoxyphenol (5.9 mg, 1.5 eq) were dissolved in anhydrous THF (1 mL) and the resulting
mixture was flushed with N2 until the THF was fully evaporated (20 min). THF (150 μL) and
(S)-(-)-2-Aminomethyl-1-ethylpyrrolidine (30 μL) were added and the reaction mixture was
introduced into the HPLC loop approximately 2 min prior to start of synthesis. The resulting
mixture was allowed to react for 5 min at 75˚C.
Synthesis of [11C]FLB457
Pd2[π-cinnamyl]Cl2 (2.2 mg; 1 eq), Xantphos (5.0 mg; 2 eq), 5-bromo-1-iodo-2,3-
dimethoxybenzene (5.1 mg) were dissolved in anhydrous THF (700 μL) and the resulting
mixture was flushed with N2 until the THF was fully evaporated (20 min). 1,4-Dioxane (150
μL) and (S)-(-)-2-Aminomethyl-1-ethylpyrrolidine (27 μL) were added and the reaction
mixture was introduced into the HPLC loop approximately 2 min prior to start of synthesis.
The resulting mixture was allowed to react for 5 min at 75˚C.
ACKNOWLEDGMENTS
This project has received funding from the European Union’s Horizon 2020 research and
innovation programme under the Marie Sklodowska-Curie grant agreement N°675071.
The authors would like to thank members of PET group at Karolinska Institutet.
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REFERENCES
1. Långström B, Lundqvist H. The preparation of 11C-methyl iodide and its use in the
synthesis of 11C-methyl-L-methionine. Int J Appl Radiat Isot 1976;27(7):357-363.
2. Larsen P, Ulin J, Dahlström K, Jensen M. Synthesis of [11C]Iodomethane by Iodination
of [11C]Methane. Appl Radiat Isot 1997;48:153-157.
3. Långström B, Itsenko O, Rahman O. [11C]Carbon monoxide, a versatile and useful
precursor in labelling chemistry for PET-ligand development. J Label Compd
Radiopharm 2007;50(9-10):794-810.
4. Lidström P, Kihlberg T, Långström B. [11C]Carbon monoxide in the palladium-mediated
synthesis of 11C-labelled ketones. J Chem Soc Perkin Trans 1 1997;(18):2701-2706.
5. Kihlberg T, Långströms B. Biologically active C-11labeled amides using palladium-
mediated reactions with aryl halides and [C-11]carbon monoxide. J Org Chem
1999;64(25):9201-9205.
6. Miller PW, Long NJ, De Mello AJ, Vilar R, Audrain H, Bender D, Passchier J, Gee A.
Rapid multiphase carbonylation reactions by using a microtube reactor: applications in
positron emission tomography 11 C‐radiolabeling. Angew Chem Int Ed
2007;46(16):2875-2878.
7. Audrain H, Martarello L, Gee A, Bender D. Utilisation of 11C-labelled boron carbonyl
complexes in palladium carbonylation reaction. Chem Commun 2004;(5):558-559.
8. Kealey S, Miller PW, Long NJ, Plisson C, Martarello L, Gee AD. Copper(I) scorpionate
complexes and their application in palladium-mediated [11C]carbonylation reactions.
Chem Commun 2009;(25):3696-3698.
9. Eriksson J, Hoek JVD, Windhorst AD. Transition metal mediated synthesis using
[11C]CO at low pressure – a simplified method for 11C‐carbonylation. J Label Compd
Radiopharm 2012;55(6):223-228.
10. Lindhe Ö, Almqvist P, Kågedal M, Gustafsson S-Å, Bergström M, Nilsson D, Antoni G,
Lindhe Ö, Almqvist P, Kågedal M, Gustafsson S-Å, Bergström M, Nilsson D, Antoni G.
Autoradiographic mapping of 5-HT(1B/1D) binding sites in the rhesus monkey brain
using [carbonyl-C]zolmitriptan. Int J Mol Imaging 2011;2011(2011):6.
11. Wall A, Kågedal M, Bergström M, Jacobsson E, Nilsson D, Antoni G, Frändberg P,
Gustavsson SA, Långström B, Yates R. Distribution of Zolmitriptan into the CNS in
Healthy Volunteers. Drugs in R&D 2005;6(3):139-47.
12. Lu S, Hong J, Itoh T, Fujita M, Inoue O, Innis RB, Pike VW. [carbonyl‐11C]Benzyl
acetate: Automated radiosynthesis via Pd‐mediated [11C]carbon monoxide chemistry and
PET measurement of brain uptake in monkey. J Label Compd Radiopharm
2010;53(8):548-551.
13. Takashima-Hirano M, Ishii H, Suzuki M. Synthesis of (11)CAm80 via novel Pd(0)-
mediated rapid (11)CCarbonylation using arylboronate and (11)CCarbon Monoxide. ACS
Med Chem Lett 2012; 3(10):804-807.
14. Rahman O, Takano A, Amini N, Dahl K, Kanegawa N, Långströms B, Farde L, Halldin
C. Synthesis of ([11C]carbonyl)raclopride and a comparison with ([11C]methyl)raclopride
in a monkey PET study. Nucl Med Biol 2015;42(11):893-898.
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15. Dahl K, Nakao R, Amini N, Moein MM, Finnema S, Malmquist J, Varnäs K, Schou M.
Development of [Carbonyl-11C]AZ13198083, a novel histamine type-3 receptor
radioligand with favorable kinetics. ACS Chem Neurosci 2018;9(5):906-911.
16. McCarron JA, Turton DR, Pike VW, Poole K.G. Remotely‐controlled production of the
5‐HT1A receptor radioligand, [carbonyl‐11C]WAY‐100635, via 11C‐carboxylation of an
immobilized Grignard reagent. J Label Compd Radiopharm 1996;38(10):941-953.
17. Wilson AA, Garcia A, Jin L, Houle S. Radiotracer synthesis from [C-11]-iodomethane: A
remarkably simple captive solvent method. Nucl Med Biol 2000;27(6):529-532.
18. Dahl K, Schou M, Amini N, Halldin C. Palladium‐mediated [11C]carbonylation at
atmospheric pressure: A general method using xantphos as supporting ligand. Eur J Org
Chem 2013;(7):1228-1231.
19. Martinelli JR, Watson DA, Freckmann DMM, Barder TE, Buchwald SL. Palladium-
catalyzed carbonylation reactions of aryl bromides at atmospheric pressure: A general
system based on xantphos. J Org Chem 2008;73(18):7102-7107.
11
20. Dahl K, Schou M, Ulin J, Sjöberg CO, Farde L, Halldin C. C-carbonylation reactions
using gas–liquid segmented microfluidics. RSC Adv 2015;5(108):88886-88889.
21. Andersen TL, Friis SD, Audrain H, Nordeman P, Antoni G, Skydstrup T. Efficient C-
11
carbonylation of isolated aryl palladium complexes for PET: Application to challenging
radiopharmaceutical synthesis. J Am Chem Soc 2015;137(4):1548-1555.
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Table 1. Investigation of solvent effects on the radiochemical yield of [11C]N-
benzylbenzamide.
Entry
Solventa
DMF
TE (%)
92
RCP (%)
52
RCY (%)
48
84
81
97
92
1
2
3
4
5
DMSO
95
88
Toluene
1,4-Dioxane
THF
84
96
98
99±0,5b
95±5b
97
Reaction conditions: Pd2(π-cinnamyl)Cl2 (1 eq), Xantphos (2eq), Iodobenzene(1,4 μL), THF(700 μL) in appropriate solvent (150 μL)
and Benzylamine (10 μL)
a Oxidative addition takes place during evaporation of THF (700µL)
b n = 3 (triplicate)
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Figure 1. Schematic representation of the HPLC loop applied in the 11C-carbonylation
procedure. The yellow arrows indicate the flow direction for [11C]CO.
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Figure 2. Picture (left) and autoradiographical image (right) of the HPLC loop following
entrapment of [11C]CO in the synthesis of [11C]N-benzylbenzamide. The arrows indicate the
entry and exit points of the He carrier gas through the HPLC loop.
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Scheme 1. Synthesis of [11C]phtalide, [11C]methyl benzoate and [11C]benzoic acid, via “in-
loop” 11C-carbonylation.
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Scheme 2. “In-loop” 11C-carbonylation applied to four druglike molecules.
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Graphical Abstract
A new “in-loop” methodology was developed to simplify palladium-mediated carbonylations
with [11C]carbon monoxide. The methodology proved to be useful in generating 11C-labelled
amides, esters and carboxylic acids. Given its simplicity, efficiency and compatibility with
automation, this methodology may become a valuable addition to the toolbox of
methodologies used for 11C-carbonylation of drugs and radioligands for PET.
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