Titania-catalyzed radiofluorination of tosylated precursors in highly aqueous medium

Article abstract of DOI:10.1021/jacs.5b02659

Nucleophilic radiofluorination is an efficient synthetic route to many positron-emission tomography (PET) probes, but removal of water to activate the cyclotron-produced [¹⁸F]fluoride has to be performed prior to reaction, which significantly increases overall radiolabeling time and causes radioactivity loss. In this report, we demonstrate the possibility of ¹⁸F-radiofluorination in highly aqueous medium. The method utilizes titania nanoparticles, 1:1 (v/v) acetonitrile-thexyl alcohol solvent mixture, and tetra-n-butylammonium bicarbonate as a phase-transfer agent. Efficient radiolabeling is directly performed with aqueous [¹⁸F]fluoride without the need for a drying/azeotroping step to significantly reduce radiosynthesis time. High radiochemical purity of the target compound is also achieved. The substrate scope of the synthetic strategy is demonstrated with a range of aromatic, aliphatic, and cycloaliphatic tosylated precursors.

Full text of DOI:10.1021/jacs.5b02659

Article
pubs.acs.org/JACS
Titania-Catalyzed Radiofluorination of Tosylated Precursors in Highly
Aqueous Medium
Maxim E. Sergeev,* Federica Morgia, Mark Lazari, Christopher Wang, Jr., and R. Michael van Dam*
Department of Molecular and Medical Pharmacology and Crump Institute for Molecular Imaging, David Geffen School of Medicine
at University of California, Los Angeles, 570 Westwood Plaza, Los Angeles, California 90095, United States
S
* Supporting Information
ABSTRACT: Nucleophilic radiofluorination is an efficient synthetic
route to many positron-emission tomography (PET) probes, but removal
of water to activate the cyclotron-produced [18F]fluoride has to be
performed prior to reaction, which significantly increases overall
radiolabeling time and causes radioactivity loss. In this report, we
demonstrate the possibility of 18F-radiofluorination in highly aqueous
medium. The method utilizes titania nanoparticles, 1:1 (v/v) acetonitrile−
thexyl alcohol solvent mixture, and tetra-n-butylammonium bicarbonate as
a phase-transfer agent. Efficient radiolabeling is directly performed with
aqueous [18F]fluoride without the need for a drying/azeotroping step to
significantly reduce radiosynthesis time. High radiochemical purity of the
target compound is also achieved. The substrate scope of the synthetic
strategy is demonstrated with a range of aromatic, aliphatic, and
cycloaliphatic tosylated precursors.
such as enzymatic9−12 and transition-metal mediated/cata-
1. INTRODUCTION
lyzed13−18 reactions, and the development of specific substrates
Positron emission tomography (PET) is an extremely effective
imaging tool in clinical care, preclinical and clinical research, and
drug discovery. PET enables visualization of physiological states
and fluorinating reagents.19,20 The majority of these methods
directly use cyclotron-delivered [18F]fluoride/[18O]H2O, and
reactions are conducted in organic−aqueous media with water
content of 0.5−20%. However, most of these methods are suited
only for a narrow compound class (e.g., aromatic, benzylic
nucleophilic fluorination), and the reported fluorination
efficiency was typically low to moderate (4−48% of the desired
fluorinated product). For enzymatic catalysis, the substrate scope
is limited to only a few specific precursors.
We report herein a conceptually new method based on titania
(titanium dioxide, TiO2) nanoparticles (crystalline composition
45% rutile, 55% anatase; <200 nm size) as a catalyst. The
precursor solution is first incubated with the particles, and then a
mixture of phase-transfer agent and [18F]fluoride/[18O]H2O
(taken directly from cyclotron-delivered vial or trapped and
eluted from QMA cartridge) is added and reacted, followed by
removal of the catalyst particles by filtration and then purification
and reformulation steps. The use of TiO2 obviates the laborious
synthesis and purification of a metal−precursor complex prior to
radiolabeling, and the method is compatible with many
commercially available or easily synthesized precursors. We
have demonstrated that this route for radiofluorination is suitable
for aromatic, aliphatic, and cycloaliphatic precursors in organic−
aqueous media with a tolerated water content up to 25 vol %.
or changes in the living body, investigation of the mechanism of
disease, and quantification of biological processes such as
receptor occupancy, cell proliferation, metabolic activity,
apoptosis, and gene expression. Since PET allows for highly
sensitive measurement of phenotypic changes associated with a
malignant condition, the onset of a particular disease such as
cancer can be detected, diagnosed, and treated at an early stage
prior to the development of metastasis.1 Typically, 18F-labeled
PET tracers are synthesized by nucleophilic fluorination of
activated precursors with [18F]fluoride/[18O]H2O obtained
from a cyclotron.2−5 Due to high 18F ion solvation energy in
water, aqueous fluoride is relatively unreactive unless it is
released from its aqueous surroundings. This is usually achieved
by mixing aqueous fluoride with a phase-transfer agent [e.g.,
K2CO3/Kryptofix-222 (K222), tetra-n-butylammonium bicarbon-
ate (TBAB), or Cs2CO3] and azeotropically drying the resulting
solution with acetonitrile. The dried active complex (e.g.,
[18F]KF/K222) is then used for radiolabeling in an anhydrous
organic solvent. Depending on the technical method of fluoride
activation (e.g., cartridge-based solvent exchange followed by
evaporation), this procedure may take up to 20−30 min, and
some loss of initial radioactivity is observed.6−8
To reduce the need for a lengthy drying process, there has
been recent interest in the development of radiofluorination
methods that allow direct use of aqueous [18F]fluoride without a
separate activation step. Several novel approaches were reported,
Received: August 21, 2014
© XXXX American Chemical Society
A
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Titania is widely used in a variety of chemical processes,
including electro- and photochemistry.21,22 Along with broad
catalytic activity, it possesses a strong ability to adsorb water from
surrounding medium.23−25 Numerous reports have demonstra-
ted the mechanism of water dissociation at titania surface,26,27
and we hypothesize that this feature of TiO2 might be used for in
situ desolvation of [18F]fluoride from water to catalyze
nucleophilic radiofluorination reactions in organic−aqueous
solutions without preliminary drying of the [18F]fluoride. We
present an investigation of the influence of various reaction
parameters on the radiofluorination of a model PET probe
precursor and also discuss studies performed to further elucidate
the mechanism of TiO2-catalyzed radiofluorination.
reaction, methanol was added to the reaction mixture, and solid
catalyst particles were removed by filtration through a 20 nm
filter (Whatman Anotop 10 Plus). In general, we found that some
[18F]fluoride trapped onto the catalyst and could not be
extracted. By use of a dose calibrator, radioactivity measurement
of the filtered extract was compared to the initial radioactivity to
determine the radioactivity extraction efficiency (REE): REE =
[decay-corrected radioactivity recovered in the organic extract]/
[initial radioactivity]. The fraction of initial radioactivity trapped
onto the catalyst is calculated as 1 − REE. Analysis with radio-
TLC (thin-layer chromatography) and radio-HPLC (high-
performance liquid chromatography) were used to determine
fluorination efficiency. This in turn enabled calculation of the
radiochemical conversion (RCC): RCC = REE × fluorination
efficiency. All results are averaged over at least n = 3 experiments.
Identity of the radiofluorinated product was confirmed by
coinjection with standard [19F]Fallypride (ABX GmbH,
Germany).
2. RESULTS AND DISCUSSION
2.1. Preliminary Observations. As a model system for our
primary studies, we have chosen the radiofluorination of tosyl-
Fallypride 1a to [18F]Fallypride 2a (Scheme 1), a highly specific
We performed a series of experiments (Table 1) to confirm the
importance of each species in the reaction. Entries 1 and 2 show
the conventional synthesis conditions where [18F]fluoride is
predried and reaction takes place in anhydrous organic medium.
The improvement in RCC due to addition of thexyl alcohol is
apparent. Entry 5 shows 68% RCC resulting from catalytic
synthesis conditions with all species included as described above.
Comparative runs without the phase-transfer catalyst resulted in
only 18% RCC. Thus, the phase-transfer agent appears to be
important as well, possibly due to better solubilization of in situ
generated tetra-n-butylammonium fluoride ([18F]TBAF), com-
pared to [18F]fluoride, in organic−aqueous medium. As
expected, experiments in the absence of catalyst did not lead to
formation of desired product 2a in organic−aqueous medium
(entry 3), indicating that TiO2 is essential when there is water in
the reaction mixture. If the [18F]fluoride was dried (as in
conventional synthesis) prior to catalytic reaction, no conversion
was observed (entry 6). In fact, the filtrate contained neither
fluorinated product nor parent [18F]fluoride; all the radioactivity
was found bound to the catalyst. This means that, in nonaqueous
medium, TiO2 addition results in total [18F]fluoride trapping
(REE ∼0%). On the other hand, REE was found to be
remarkably constant (∼80%) for all of the TiO2-catalyzed
conditions containing water. Even if the reaction was performed
without the precursor, the REE was unchanged (80% 2%).
Scheme 1. Catalytic Formation of [18F]Fallypride
dopamine D2/D3 receptor radioligand used in PET imaging of
the brain to study receptor occupancy and density and which has
potential clinical application in relation to neuropsychiatric
conditions and aging.28,29
The radiofluorination reaction was performed in a 1:1 (v/v)
mixture of acetonitrile (MeCN) and 2,3-dimethyl-2-butanol
(thexyl alcohol). Thexyl alcohol was included due to a report by
Lee and co-workers,30 showing that addition of alcohol to the
reaction medium facilitates the radiofluorination SN2 process,
and a report by Javed et al.,31 showing a substantial increase in
fluorination efficiency of [18F]Fallypride when this solvent
mixture was used compared to pure MeCN. Reactions were
performed in a sealed vial, heated to 110 °C for 7 min by use of a
Peltier heater. The reaction mixture was mixed by refluxing
solvent; magnetic stirring was not used. After completion of the
a
Table 1. Influence of Reaction Components on Formation of 2a
b
c
entry
catalyst
phase-transfer agent
MeCN (μL)
thexyl alcohol (μL)
water (μL)
REE (%)
RCC (%)
d
1
2
none
none
none
TiO2
TiO2
TiO2
TiO2
none
MgSO4
CaCl2
TBAB
40
20
15
15
15
20
15
20
15
15
0
20
15
15
15
20
15
20
15
15
0
0
100
31
64
0
2
4
d
TBAB
100
d
3
none
10
10
10
0
100
4
none
79
80
0
4
3
18
68
0
3
2
5
TBAB
6
TBAB
7
K2CO3/K222
K2CO3/K222
TBAB
10
0
78
100
99
99
3
39
31
0
6
4
d
8
9
10
10
1
1
10
TBAB
0
a
Reactions were performed with 2.3 μmol of 1a and 140 μmol of TiO2 in 40 μL reaction volume at 110 °C for 7 min without magnetic stirring.
Amounts of phase-transfer agent used, when applicable: 0.36 μmol of TBAB; 0.36 μmol of K2CO3, and 0.36 μmol of K222. For cases where water is
b
c
0 μL, [18F]fluoride was added as dry complex with phase-transfer agent (1.5−4 mCi) reconstituted in 10 μL of MeCN−thexyl alcohol (1:1 v/v); for
d
other cases, radioactivity was introduced as a solution of aqueous [18F]fluoride (1.5−4 mCi) containing phase-transfer agent; In the case of catalyst
absence, no extraction was performed.
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The influence of the type of phase-transfer agent was evaluated
by carrying out the reaction with K2CO3/K222 mixture (entry 7).
This resulted in a substantial decrease in RCC, suggesting that
TBAB is a superior phase-transfer agent for these reactions.
Finally, to assess whether the role of TiO2 was simply to
sequester water to facilitate fluorination in mixed aqueous−
organic medium, comparative runs with particles of other
common non-oxide drying agents, MgSO4 and CaCl2 (140 μmol
each), were also performed (entries 9 and 10). These showed
zero RCC, suggesting an effect of TiO2 beyond simple water
adsorption but most probably its ability of water splitting.
2.3. Hypothesized Mechanism of TiO2-Catalyzed
Radiofluorination. Previous reports show that oxo- and oxy-
containing species readily coordinate on a TiO2 surface through
hydrogen bonding.32−35 Thus, we hypothesize that in addition to
an interaction with water, the TiO2 catalyst could also serve to
coordinate tosyl-Fallypride 1a via oxygen atoms of the sulfonyl
group, facilitating reaction with desolvated [18F]fluoride in close
proximity. Based on these ideas, the mechanism of catalyzed
fluorination shown in Figure 1 is suggested.
heating at 110 °C for 12 min in an oil bath. The reaction mixture
was mixed by refluxing solvent; no magnetic stirring was used.
We observed a modest enhancement in RCC starting after 20−
30 min of incubation and reaching a maximum improvement
after ∼1 h (Figure 2).
Figure 2. Effect of substrate−catalyst preincubation time on radio-
fluorination efficiency of 1a.
Analytical samples of organic solution after incubation with
catalyst contained significant amount of precursor 1a; thus, it is
likely that the observed saturation is due to occupation of all
accessible active sites of TiO2 capable of binding the oxygen
atoms of the leaving group. Attempts to perform preincubation at
elevated temperatures (30, 45, and 60 °C) showed similar
results; that is, the time scale was not shortened. It should be
noted that the preincubation step was performed before the
introduction of [18F]fluoride; therefore, incorporation of
preincubation into the synthesis protocol does not introduce
any delays that would affect the yield due to decay.
Studies were performed by running the radiofluorination
reaction in the presence of various amounts of a nonreactive SO-
containing compound (dimethyl sulfoxide, DMSO) to assess
whether the presence of another cocoordinating species could
block the binding of the precursor and lower the RCC (Figure 3).
Figure 1. Proposed mechanism for TiO2-catalyzed radiofluorination.
SN2 substitution is improved with alcohol as a cosolvent, perhaps by
inclusion in intermediate complex formation as determined by Oh et
al.30 (not shown here).
Hydrogen bonding occurs between oxygen atoms of the
sulfonyl moiety and TiO2, which coordinates tosylated precursor
to the surface of the catalyst; when aqueous [18F]fluoride/TBAB
solution is added, solvated [18F]fluoride is adsorbed at active sites
of TiO2 where the aqueous shell is split, resulting in [18F]fluoride
release (i.e., activation); TBAB phase-transfer catalyst then
serves as a [18F]fluoride-trapping agent and intercepts activated
[18F]fluoride, subsequently conducting the phase transfer of
[18F]fluoride to the surface-coordinated precursor to faciliate the
SN2-type reaction. Because the coordination with the precursor is
at the leaving group, the resulting radiofluorinated product is
released upon formation from TiO2.
2.4. Evidence for Coordination. To explore the role of
coordination in the reaction mechanism, we performed radio-
labeling after first incubating the tosylated substrate with catalyst.
If coordination indeed occurs, prolonged exposure of the
precursor to the catalyst should increase the amount of precursor
bound to the surface of the catalyst and thus promote increased
SN2-reactions (i.e., radiolabeling efficiency). Thus, a solution of
tosyl-Fallypride 1a in 1:1 (v/v) MeCN−thexyl alcohol was
added to the catalyst and incubated at room temperature for
various durations. After incubation, aqueous [18F]fluoride/
TBAB mixture was added, and radiolabeling was performed by
Figure 3. Influence of DMSO on TiO2-catalyzed radiochemical
conversion of 1a.
It was revealed that even slight addition of DMSO dramatically
affected the RCC; a 2-fold decrease in RCC was registered at 5
vol % DMSO, with nearly complete inhibition at 37.5 vol %
DMSO, suggesting that DMSO molecules may indeed bind to
active sites of TiO2, thus reducing sites available for precursor
coordination. Surprisingly, the REE was also significantly
affected, and the amount of [18F]fluoride trapped onto the
catalyst increased with increasing DMSO content. Almost total
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[18F]fluoride trapping (90%) was registered at 75 vol % DMSO.
This suggests that instead of merely affecting precursor
coordination, DMSO may interact with the catalyst in additional
ways leading to trapping of fluoride, such as DMSO−water
cluster formation,36−39 intercalation of DMSO−water clathrate
inside the oxide structure,40 or DMSO-induced creation of
positively charged sites.41
To investigate further the need for leaving-group coordination
at the TiO2 surface, experiments were performed comparing
tosylated compounds 1b, 1c, and 1s with their brominated
versions 3−5 (not suspected to coordinate with surface) in TiO2-
catalyzed [18F]fluorination (Table 2). No product was observed
amount is increased, one of the reaction components becomes
depleted, reducing the interactions necessary for fluorination.
On the other hand, looking at extraction efficiency, we
observed that REE decreases (i.e., trapping increases) with
increasing amount of catalyst. Due to this linear relationship, we
suspect there are specific trapping sites for fluoride on the TiO2,
the number of which depends on the amount of catalyst present.
Perhaps trapping occurs via exchange of fluoride with terminal
hydroxyl groups.42−44 The consistent ∼20% trapping for a fixed
amount of catalyst may represent the equilibrium exchange
between the surface and the reaction solution containing both
fluoride and hydroxide ions. The case of 100% trapping under
dry conditions (Table 1, entry 6), where the fluoride
concentration dominates, could represent a shift in this
equilibrium.45
Table 2. TiO2-Catalyzed Radiofluorination of Tosylated
a
versus Brominated Substrates
To determine if the optimal amount of catalyst is universal or
should be adjusted for every precursor, similar experiments were
performed with substrates 1i, 1q, and 1t. The loading of 140
μmol of TiO2 remained optimal for these substrates as well
(Figure 4).
a
Preincubation time 1 h, 2.3 μmol of precursor, 140 μmol of TiO2,
130 °C for 5 min, 40 μL total reaction volume, no magnetic stirring;
radioactivity was introduced as 10 μL solution of aqueous [18F]fluoride
(1.5−4 mCi) containing 0.36 μmol of TBAB. Radioactivity extraction
efficiency (REE) ∼80% was observed for every entry.
in the case of bromo derivatives, while high RCCs of ∼80% were
determined for tosylated precursors, which agrees with the
necessity for the proposed oxygen-coordinating mechanism at
the leaving group. The REE in the case of bromo-substituted
substrates remained close to 80%, that is., similar to that for
tosylated reactants.
Figure 4. Determination of optimal catalyst amount for radiolabeling of
1a, 1i, 1q, and 1t.
2.5. Optimization of Catalyst Loading. It was found that
the amount of catalyst added had a substantial effect on RCC.
Studies with different catalyst amounts (see Supporting
Information) revealed that the optimal loading of TiO2 is 140
μmol for 40 μL reaction volume with 25 vol % water content for
the catalyst particle size used (<200 nm). This translates to
∼60:1 ratio of catalyst to precursor 1a and enables production of
target compound 2a with the highest RCC (78%). The RCC
drops for both increasing and decreasing amounts of catalyst.
Looking at fluorination efficiency, we observed catalyst
amounts lower than the optimum to result in decreasing
fluorination efficiency, perhaps due to reduced desolvation of
fluoride. Surprisingly, increasing the amount of catalyst also
decreased the fluorination efficiency. Perhaps as the catalyst
2.6. Evaluation of Optimal Reaction Conditions. To
maximize fluorination efficiency of the precursor 1a, further
evaluation of reaction conditions has been performed to
determine optimal reaction parameters.
The range of water content that provided maximum RCC was
found to be up to 25 vol % (see Supporting Information). In this
aqueous range, the RCC and fluoride trapping remained
remarkably constant. With higher water content, the trapping
remained constant, but a strong decrease in [18F]fluorination was
observed. We hypothesize that higher water content exceeds the
capacity of the catalyst to adsorb and split water, and thus the
[18F]fluoride is not as effectively desolvated, reducing the
fluorination efficiency. If true, this suggests that increased catalyst
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a
Table 3. Titania-Catalyzed Production of [18F]Fallypride
parameter
run 1
run 2
run 3
average
initial activity (mCi)
REE (%)
5.20
79.1
79.1
79.1
75.7
71.2
28.8
71.2
5.19
81.2
81.2
81.2
76.5
68.6
31.4
68.6
5.23
81.9
81.5
81.9
77.6
73.2
26.8
73.2
5.20 0.02
80.7 1.5
80.6 1.3
80.7 1.5
76.6 1.0
71.0 2.3
28.9 2.3
71.0 2.3
RCC (%) (nonisolated)
b
activity remaining after synthesis and extraction (%)
b
activity remaining after HPLC purification (%)
b
activity remaining after reformulation (%)
total loss (%)
c
isolated RCY (%)
a
Optimized reaction conditions: 1 h preincubation time, 2.3 μmol of precursor, 140 μmol of TiO2, 40 μL total reaction volume, no magnetic stirring;
radioactivity introduced as 10 μL solution of aqueous [18F]fluoride (∼2.6 mCi) containing 0.36 μmol of TBAB; heated to 130° for 5 min in an oil
bath. For each run, two vials are pooled after extraction to improve accuracy of measurements. Fractions of remaining radioactivity determined by
measuring the radioactivity after the relevant step, correcting for decay, and dividing by the initial radioactivity. RCY = radiochemical yield.
b
c
Table 4. Comparison of Catalytic Production of 2a to Known Procedures
a
ref
reactor type
radiolabeling conditions
mean RCY (%)
total time (min)
Moon et al.48
Pike et al.46
Javed et al.7
Lazari et al.49
this report
macroscale automated. TracerLab FX
microscale automated, Nanotek Advion
microscale automated, EWOD chip
macroscale automated, ELIXYS
100 °C, 30 min
150−190 °C, 4−23 min
105 °C, 7 min
105 °C, 7 min
130 °C, 5 min
68
74
16−88
83
50−218
70
b
66
56
small-volume vial, manual
71
50
a
b
RCY = radiochemical yield. Total time reported without reformulation step.
Table 5. Quality Control Tests of Injectable [18F]Fallypride Solution
clinical QC test
optical clarity
clinical acceptance criteria
results of this study
clear and particle-free
clear and particle-free
pH
5.5−8.0
6.5
radiochemical purity (%)
radiochemical identity
18F-radionuclide half-life (min)
endotoxin level (EU/mL)
filter integrity (psig)
MeCN content (ppm)
thexyl alcohol content (ppm)
sterility
>95
>99
matches retention time of standard
matches retention time of standard
105−115
<5
111
<1
>50
>100
<410
<2
<5000
<1
no growth in 14 days
none specified
no growth in 14 days
titanium content (ng)
36
4
amount may enable improved water tolerance if desired, but in
light of results in the previous section, it would also be necessary
to increase the precursor amount in the same proportion as the
catalyst.
We also studied RCC as a function of reaction time and
temperature (see Supporting Information) to determine the
optimal reaction time (5 min) and temperature (130 °C). The
influence of the specific type of alcohol cosolvent was evaluated
as well. Several alcohols were tested in place of thexyl alcohol, but
relatively little difference in RCC was observed (see Supporting
Information). Thexyl alcohol showed the highest RCC of
alcohols tested.
2.7. Production and Quality Control of Clinically
Relevant PET Probe. To demonstrate the overall radio-
chemical yield of isolated [18F]Fallypride 2a, we performed full
production runs (radiofluorination, HPLC purification, and
formulation). The final formulated product was obtained as a
sterile, injectable solution. During production, radioactivity
measurements were recorded at key steps to assess efficiency
of each process and identify potential areas for optimization
(Table 3). The biggest loss is during extraction from the catalyst
(20% trapped), and an additional ∼10% is lost during our
purification and formulation processes. Total production times,
isolated yields, and specific activities are compared to those
previously reported in literature (Table 4). Generally, the
isolated yield of titania-catalyzed reaction tends to be higher than
the yield reported for macroscale automated production, while it
is comparable to the yields for microfluidic procedures, such as
syntheses on a digital microfluidic chip7 or use of the Advion
Nanotek capillary reactor.46 By avoiding the need for fluoride
drying/azeotroping, the synthesis process is simplified.
It is noteworthy to mention the high radiochemical purity of
target compound 2a that formed during reaction. Only two
radioactive peaks were detected by analytical HPLC, which
consisted of unreacted [18F]fluoride and the desired [18F]-
Fallypride (see Supporting Information). Regarding HPLC
analysis of the nonradioactive side products, the hydroxylated
compound, resulting from hydrolysis, is clearly observed while
no byproduct from β-elimination is apparent. With HPLC
purification, the [18F]fluoride and nonradioactive side products
were effectively removed, and the reformulated solution was then
examined by standard quality control (QC) tests47 to evaluation
its compliance with U.S. Food and Drug Administration
requirements for injectable PET tracers (Table 5).
With the introduction of TiO2 catalyst into the production
procedure, an additional QC test to assess the titanium content
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in the reformulated solution may be needed. While the 20 nm
filtration process and subsequent HPLC purification are
expected to eliminate all particles, there is still the possibility of
titanium ions in the final formulated solution. By use of
inductively coupled plasma mass spectrometry (ICP-MS), the
titanium content in representative samples was found to be 36
4 ng (n = 9) of titanium per batch. Titanium is considered very
inert and there do not appear to be established limits for titanium
in injectable solutions. According to medicinal reports,50−52
however, normal levels of titanium in human body range from 0
to ∼20 μg/L blood (∼0−100 μg per whole body of an adult) in
patients without titanium implants, while reaching 100−150 μg/
L blood (500−750 μg per whole body) in patients with artificial
titanium joints. Thus, an injection of formulated [18F]Fallypride
produced by our method would have a very miniscule impact
because it contains orders of magnitude less titanium than
normally present in the blood. This could suggest that routine
titanium testing may not be needed for every batch of injectable
PET probe.53,54
prior to scaling (Figure 5). We suspect that longer time is needed
for sufficient diffusive mixing in the larger volume. We also
Figure 5. Correlation between reaction time and fluorination efficiency
in scaled-up catalytic radiolabeling of 1a. Values are averaged from n = 9
experiments for 5 min runs and n = 6 for each of 8 and 15 min
experiments.
We have, therefore, demonstrated the practical feasibility of
the newly developed synthetic approach for synthesis of a
clinically relevant PET probe, including compliance with FDA
requirements for injectable solutions in clinical applications. By
varying the precursor, other [18F]fluorine-labeled PET probes
could be produced by this procedure and similar QC results
would be expected.
encountered some initial difficulties during the filtration step to
remove the nanoparticles after the reaction was done. We
observed clogging of the 20 nm filter due to the increased
amount of TiO2. This issue was resolved by incorporating an
initial prefiltration step with a 0.22 μm filter prior to 20 nm fine
filtering. Unfortunately, this had the effect of slightly reducing the
extraction efficiency, from 80% 2% to 71% 13%.
The tested scale-up factor of 3 is sufficient for production of a
human dose of [18F]Fallypride, and further increases in scale are
presumably possible, if desired; with higher volumes, stirring
during the preincubation and reaction steps may become
important, requiring additional optimization of reaction time
and extraction procedures. Another reason to perform volume
scale-up is to potentially enable automation in commercially
available radiosynthesizers, which typically require at least several
hundred microliters of solution in the reaction. Though there are
advantages in performing reactions in extremely small
volumes,7,8 it will be some time before automated and
commercialized versions of such technologies are widely
available.
As an alternative way to increase the amount of radioactivity in
the reaction without impacting reaction volume, solid-phase
extraction procedures using microscale QMA cartridges could be
used to concentrate the [18F]fluoride to obtain higher starting
radioactivity in volumes of the [18F]fluoride/TBAB solution
described here (i.e., 10 μL water content). Several reports have
shown that an entire cyclotron target volume can be trapped and
efficiently eluted in only 5−45 μL of eluent solution.61−63 This
would likely be the preferred approach to scale up the amount of
radioactivity since no increase in reaction volume would be
necessary.
2.10. Substrate Scope. In order to explore the utility of the
TiO2 catalytic approach for synthesis of other PET tracers, we
investigated the scope of applicable substrates. We first
considered the use of other leaving groups with sulfonyl moieties
(i.e., triflate and nosylate). Surprisingly, radiofluorination of
commercially available substrates 6 and 7, precursors for 2′-
deoxy-2′-[18F]fluoro-D-glucose ([18F]FDG) and 3′-deoxy-3′-
[18F]fluoro-L-thymidine ([18F]FLT) (Figure 6), resulted in
RCC = 0%.
2.8. Determination of Specific Activity. High specific
activity (SA) of PET tracers is essential to minimize the injected
quantity of the nonradioactive form of the tracer, which can
saturate rare biological targets, such as neurological receptors,
and subsequently lower the image quality and possibly cause
pharmacologic effects.55 It has been confirmed that SA plays an
important role in PET image quality,29,56,57 especially in small
animals.58,59 By use of standard methods,7 the SA for TiO2-
catalyzed synthesis of [18F]Fallypride 2a was found to be 5
2
Ci/μmol (n = 5). This is higher than typically obtained from
macroscale synthesis (0.4−3 Ci/μmol).49,60 This evidence
suggests TiO2 catalysis could be successfully used for routine
production of PET imaging probes that require high SA.
2.9. Attempts to Scale up Synthesis. Previous experi-
ments utilized operating volumes of 40 μL, 10 μL of which was
[18F]fluoride solution. While we have been able to demonstrate
the production of several millicuries of the isolated product
starting from ∼5 mCi of [18F]fluoride, it may be desirable in
some cases to produce larger amounts of product (e.g., for
clinical production), which requires a larger volume of the initial
[18F]fluoride solution.
One simple way to scale up the quantity of tracer produced is
to run several reactions in parallel and pool the results; however,
this would require multiple reaction vessels and may not be
convenient to handle or automate. Another approach is to scale
up the reaction volume, enabling a larger volume of [18F]fluoride
to be used while still keeping water content within the 25 vol %
range, and thus enabling a larger amount of starting radioactivity.
To test this possibility, we explored proportionally increasing the
amounts of all of the reaction components (i.e., precursor, TiO2,
solvent, and aqueous [18F]fluoride/TBAB solution). As an
example, the catalytic fluorination of 1a was scaled up by a factor
of 3, such that the final volume of the reaction mixture comprised
120 μL. During these experiments, we found that 5 min reaction
time was insufficient to efficiently fluorinate the precursor
(fluorination efficiency = 50%). With increased heating time, it
was possible to increase fluorination efficiency to the values seen
In the presence of catalyst, triflate and nosylate groups seemed
to become overreactive, and immediate explosive hydrolysis was
F
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Article
Table 6. Substrate Scope for TiO2-Catalyzed
a
Radiofluorination of Tosylated Substrates
Figure 6. Additional substrates with sulfonyl-containing leaving groups
and their corresponding hydrolysates.
observed upon aqueous [18F]fluoride addition. When analyzed,
only unreacted [18F]fluoride and hydroxylated compounds 6-
OH and 7-OH were detected. This phenomenon suggests that
reactivity of oxygen-containing leaving groups are increased
when incubated with TiO2. Due to this additional activation, the
ideal leaving group should initially possess lower reactivity
(tosyloxy preferred over triflyloxy or nosyloxy), otherwise
concurrent side reaction of hydrolysis prevails over [18F]-
fluorination.
We next investigated the generality of TiO2-catalyzed
radiofluorination of tosylated precursors. A library of aromatic,
aliphatic, and cycloaliphatic tosylates was tested, along with the
commercially available tosylated PET probe precursors for [18F]-
4-fluoroproline ([18F]-4-FP), [18F]fluoroazomycin arabinoside
([18F]FAZA), and [18F]fluoroerythronitroimidazole ([18F]-
FETNIM) (Table 6). The methodology was highly efficient
for low-molecular-weight precursors 1b−v (65−80% RCC) but
resulted in low to moderate yields with bulky and sterically
hindered substrates 1w−x (i.e., from the commercially available
precursors for [18F]FAZA and [18F]FETNIM, respectively).
These particular precursors also contain additional oxo moieties,
which potentially lower yields by coordinating the precursor at
the catalyst surface instead of at the OSO moiety of the
tosylate leaving group. Such coordination could significantly
reduce [18F]fluoride interaction with the tosylate reactive center
by placing the reaction center further from the catalyst surface
where fluoride desolvation occurs. We are currently looking into
methods to further understand the reaction mechanism to
predict the effectiveness of different substrates and perhaps
enable improved substrate design.
3. CONCLUSIONS
In conclusion, we have developed a novel method of TiO2-
catalyzed radiofluorination of tosylated presursors and demon-
strated its use for the preparation of 18F-labeled PET probes. The
method avoids the need for drying of [18F]fluoride/[18O]H2O
from the cyclotron before fluorination. The wet [18F]fluoride is
mixed with a phase-transfer agent and added to a solution of
precursor solution preincubated with TiO2 nanoparticles and
reacted for a short time. In this fashion, nucleophilic 18F-
fluorination is shown to proceed rapidly and efficiently in
aqueous medium with up to 25 vol % water content, which to the
best of our knowledge is the highest reported other than
a
Optimized reaction conditions: 1 h preincubation time, 2.3 μmol of
precursor, 140 μmol of TiO2, 130 °C, 5 min, 40 μL total reaction
volume, no magnetic stirring; radioactivity introduced as 10 μL
solution of aqueous [18F]fluoride (1.5−4 mCi) containing 0.36 μmol
TBAB. For all entries, REE was observed to be ∼80%.
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as well as shown compliance of the final formulated PET tracer
with QC requirements for clinical use. The product was found to
have high specific activity even with low amounts of starting
radioactivity. The applicability of the reported protocol to a range
of tosylated substrates was also demonstrated for organic
molecules containing aromatic, aliphatic, and cycloaliphatic
moieties. Although extensive additional investigations are
required to explore the substrate scope and further understand
the mechanism, we anticipate that the facile procedure and high
radiofluorination efficiency of this new method may provide a
versatile tool for practitioners in the field of PET radiochemistry.
On the basis of our hypothesized mechanism of reaction, further
studies regarding the importance of the structure of the precursor
and other effective catalysts are currently in progress in our
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ASSOCIATED CONTENT
■
S
* Supporting Information
(22) Froschl, T.; Hormann, U.; Kubiak, P.; Kuce
̌
rova,
́
G.; Pfanzelt, M.;
̈
̈
Additional text, 11 figures, and nine tables with experimental
details, as well as selected HPLC and NMR data and spectra
(PDF); files for compounds 1a−1x and 2a−2x (CDX). This
material is available free of charge via the Internet at http://pubs.
acs.org.
Weiss, C. K.; Behm, R. J.; Husing, N.; Kaiser, U.; Landfester, K.;
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AUTHOR INFORMATION
■
Corresponding Authors
*msergeev@mednet.ucla.edu.
*mvandam@mednet.ucla.edu.
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
(30) Oh, Y.-H.; Ahn, D.-S.; Chung, S.-Y.; Jeon, J.-H.; Park, S.-W.; Oh,
S. J.; Kim, D. W.; Kil, H. S.; Chi, D. Y.; Lee, S. J. Phys. Chem. A 2007, 111,
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■
We gratefully acknowledge the invaluable services of Professor
Saman Sadeghi and the staff of the UCLA Biomedical Cyclotron
for providing 18F− ion and performing QC testing of samples.
This work was supported in part by the Department of Energy
Office of Biological and Environmental Research (DE-
SC00001249) and the National Institute of Biomedical Imaging
and Bioengineering (NIBIB, Grant EB015540).
(33) Bahruji, H.; Bowker, M.; Brookes, C.; Davies, P. R.; Wawata, I.
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