Synthesis and evaluation of [18F] labeled pyrimidine nucleosides for positron emission tomography imaging of herpes simplex virus 1 thymidine kinase gene expression

Article abstract of DOI:10.1021/jm0512847

Synthesis of three novel 2′-deoxy-2′-[¹⁸F]fluoro-1- β-D-arabinofuranosyluracil derivatives [¹⁸F]FPAU, [ ¹⁸F]-FBrVAU, and [¹⁸F]FTMAU is reported. The compounds were synthesized by coupling of 1-bromo-2-deoxy-2-fluoro sugars with corresponding silylated uracil derivatives. In vitro cell uptake indicated that all three compounds are taken up selectively in RG2TK+ cells with negligible uptake in RG2 cells. The results indicate that [¹⁸F]FBrVAU and [ ¹⁸F]FTMAU have better uptake profiles in comparison to [ ¹⁸F]FPAU and have potential as PET probes for imaging HSV1-tk gene expression.

Full text of DOI:10.1021/jm0512847

J. Med. Chem. 2006, 49, 5377-5381
5377
18
Synthesis and Evaluation of [ F] Labeled Pyrimidine Nucleosides for Positron Emission
Tomography Imaging of Herpes Simplex Virus 1 Thymidine Kinase Gene Expression
Nagavarakishore Pillarsetty, Shangde Cai, Lyudmila Ageyeva, Ronald D. Finn, and Ronald G. Blasberg*,‡,§
†
†
‡
†,§
Radiochemistry and Cyclotron Core Facility, Department of Neurology, and Department of Radiology, Memorial Sloan Kettering Cancer
Center, 1275 York AVenue, New York, New York 10021
ReceiVed December 23, 2005
1
8
18
18
Synthesis of three novel 2′-deoxy-2′-[ F]fluoro-1-â-D-arabinofuranosyluracil derivatives [ F]FPAU, [ F]-
18
FBrVAU, and [ F]FTMAU is reported. The compounds were synthesized by coupling of 1-bromo-2-deoxy-
-fluoro sugars with corresponding silylated uracil derivatives. In vitro cell uptake indicated that all three
2
compounds are taken up selectively in RG2TK+ cells with negligible uptake in RG2 cells. The results
18
18
18
indicate that [ F]FBrVAU and [ F]FTMAU have better uptake profiles in comparison to [ F]FPAU and
have potential as PET probes for imaging HSV1-tk gene expression.
1
8
18
Introduction
arabinofuranosyluracil derivatives, such as [ F]FMAU, [ F]-
1
8
FBAU, [ F]FAU, etc., and are currently evaluating their
potential as PET markers for cellular proliferation. The 5-sub-
stituted thymidine derivatives have also been shown to be good
substrates for HSV1-TK. Our aim was to synthesize
-substituted derivatives of 2′-fluoro-2′-deoxythymidine, with
a range of different hydrophobicities, and then to evaluate them
as potential radiolabeled PET reporter probes for imaging HSV1-
tk gene expression. Hydrophobic derivatives are of interest
because the currently available probes are hydrophilic and do
not readily cross the intact blood-brain barrier.
The herpes simplex virus 1 thymidine kinase (HSV1-tk) gene
is one of the most widely used reporter genes for imaging gene
regulation and expression using positron emission tomography
3
,18-24
(PET). We initially demonstrated the feasibility of using HSV1-
5
1
tk as a reporter gene, with a radiolabeled probe and by γ
2
3
camera and PET imaging. This was subsequently confirmed
by reports from other labs.4 Thymidine kinases are enzymes
that phosphorylate thymidine to thymidine 5′-monophosphate.
HSV-tk gene encodes the HSV1-TK enzyme. Unlike mam-
malian thymidine kinase 1 (TK-1), which is highly specific for
thymidine, HSV1-TK has a fairly broad range of substrate
specificity.6 HSV1-TK can phosphorylate thymidine nucleoside
analogues, which have modifications on the heterocyclic ring
or on the sugar moiety, and various acyclic analogues of
,5
A variety of 5-substituted thymidine derivatives were previ-
ously synthesized and evaluated for their potential as antiherpes
,7
1
8-24
agents.
The primary mechanism of action was based on
the ability of herpes viral thymidine kinase to phosphorylate
the thymidine derivatives, which in turn would be incorporated
into viral DNA causing multistrand breaks leading to destruction
of virus. We hypothesized that antiviral efficacy of a particular
molecule is a good indicator of its rate of phosphorylation.
Hence, the target molecules were chosen on the basis of the
ability of the nonradioactive analogues to inhibit HSV1 viral
replication. Here, we report the synthesis and in vitro evaluation
6
,7
guanosine. This property has been utilized in gene therapy
protocols involving HSV1-TK enzyme activated prodrug therapy,
wherein the target tissue is transfected or transduced with HSV1-
tk gene. Cells expressing HSV-tk activate prodrugs such as
gancyclovir or acyclovir by phosphorylation, and their ac-
cumulation in the cell leads to apoptosis, whereas normal cells
remain unaffected by these drugs. This type of gene therapy
1
8
of three novel radiolabeled probes 2′-[ F]fluoro-2′-deoxy-5-
8
-11
has been used for selective destruction of malignant tissue.
A number of different PET reporter probes for imaging HSV-
tk gene expression are currently available. These include C,
1
8
bromovinyl-1â-D-arabinofuranosyluracil ([ F]FBrVAU), 2′-
18
18
[
F]fluoro-2′-deoxy-5-propyl-1â-D-arabinofuranosyluracil ([ F]-
FPAU), and 2′-[ F]fluoro-2′-deoxy-5-trifluoromethyl-1â-D-
arabinofuranosyluracil ([ F]FTMAU) for PET imaging of HSV-
1
1
18
1
8
124
11
F, or I radiolabeled nucleoside analogues such as [ C]-
18
FMAU,1
2
[
124
I]FIAU, [ F]FHBG,
3
18
13,14 18
[ F]FHPG,
13,14 18
[ F]-
tk gene expression.
FMAU,1 etc. Although each probe offers certain advantages
over others, there are also certain limitations. For example, our
group demonstrated that 2′-fluoro-2′-deoxy-1â-D-arabinofura-
3,15
Materials and Methods
Chemicals. All reagents and solvents were purchased from
Aldrich Chemical Co. (St. Louis, MO) and used without further
purification, unless stated otherwise. The triflate sugar (2-O-(tri-
fluoromethylsulfonyl)-1,3,5-tri-O-benzoyl-R-D-arabinofuranose) 1
was synthesized according to literature procedure18 and used in the
reactions. The silyl ethers 5-alkyl-2,4-bis-trimethylsilanyloxypyri-
midine were synthesized in situ according to published procedure
and used in the reaction. Sep-Pak extraction cartridges were
purchased from Waters Associates (Milford, MA). Radioactively
1
24
124
nosyl-5-[ I]iodouracil ([ I]FIAU) has excellent imaging
properties for studying HSV-tk gene expression in transduced
animals. However, the long half-life of I (t1/2 ) 4.28 days)
may be a disadvantage where sequential and repetitive imaging
is necessary.
1-3
124
19
16
Alauddin and co workers and more recently Shields and
17
co workers have developed a general procedure for the
synthesis of 18F labeled, 5-substituted-2′-deoxy-2′-[ F]fluoro-
18
3
14
labeled ( H and C) nucleosides were purchased from Moravek
Biochemicals (Brea, CA).
Cell Lines. RG2 rat glioma cell line was kindly provided by
Dr. Darell Bigner (Duke University Medical Center, Durham, NC).
RG2 cells were transduced with the recombinant replication-
deficient STK retrovirus containing the NeoR gene and HSV1-tk
*
To whom correspondence should be addressed. Phone: +1-212-639-
7
337. Fax: +1-212-717-3063. E-mail: blasberg@neuro1.mskcc.org.
†
Radiochemistry and Cyclotron Core Facility.
Department of Neurology.
Department of Radiology.
‡
§
1
gene as previously described (see Supporting Information). The
1
0.1021/jm0512847 CCC: $33.50 © 2006 American Chemical Society
Published on Web 08/01/2006

5
378 Journal of Medicinal Chemistry, 2006, Vol. 49, No. 17
Brief Articles
18
18
Scheme 1. Synthesis of [ F]FPAU, [ F]FBrVAU, and
18
[
F]FTMAU
1
8
Table 1. Decay-Corrected Radiochemical Yields of [ F]FBrVAU,
18
18
18
[
F]FPAU, and [ F]FTMAU
Figure 1. HPLC chromatogram of purified [ F]FBrVAU co-injected
with its nonradioactive analogue.
compd
decay-corrected yield, %
1
1
1
8
8
8
[
[
[
F]FBrVAU
F]FPAU
F]FTMAU
12 ( 4
20 ( 3
16 ( 3
using NMR and mass spectrometry (see Supporting Informa-
20,21,24
tion).
The purified radioactive samples were co-injected with
pure nonradioactive analogues and had very similar retention times,
ensuring the chemical identity and purity of the compounds.
Octanol/Water Partition Coefficient Study. Octanol/water
transduced cell line, RG2TK+, has remained stable and has
expressed constant levels of HSV1TK+ since 1994; this cell line
has been characterized previously.
Synthesis. [ F]FBrVAU, [ F]FPAU, and [ F]FTMAU were
synthesized by coupling the radiolabeled fluoro sugar with the
corresponding silylated pyrimidine derivatives following a proce-
dure reported by Alauddin and co-workers, with minor modifica-
tions as shown in Scheme 1. Briefly, 18F in the form of [ F]HF,
trapped on an anion-exchange resin, was eluted with 0.5 mL of
1
8
18
partition coefficients were determined for [ F]FBrVAU, [ F]-
1,2
18
FPAU, and [ F]FTMAU at four different pH values of 6.4, 7.0,
18
18
18
7.4, and 8.4 by measuring the distribution of radiolabeled compound
in 1-octanol and phosphate buffered saline (PBS). A 20 µL sample
18
18
18
of radiolabeled compound ([ F]FBrVAU, [ F]FPAU, or [ F]-
FTMAU) in saline was added to a vial containing 0.5 mL each of
1-octanol and PBS. After vortexing for 1 min, the vial was
centrifuged for 5 min to ensure complete separation of layers. Then
16
18
1
8
0
.8% tetrabutylammonium bicarbonate into a flask containing 1
20 µL of each layer was taken in a preweighed vial and F counts
(400-550 keV energy range) were measured using an AutoGamma
5550 spectrometer (Packard Instruments, Meriden, CT). Counts per
unit weight of sample were calculated, and log P values were
calculated using the formula
mL of acetonitrile. The water acetonitrile azeotrope was completely
removed by heating the flask to 80 °C under reduced pressure while
maintaining a slow stream of argon gas. To the flask, 1 mL of
anhydrous acetonitrile was added, and the flask was dried as
described above. The flask was cooled using dry ice-2-propanol
slush, and 20 mg of 2-O-(trifluoromethylsulfonyl)-1,3,5-tri-O-
benzoyl-R-D-arabinofuranose in 0.7 mL of anhydrous acetonitrile
was added. The mixture was heated at 80 °C for 30 min. The
mixture was cooled to room temperature and passed through a silica
Sep-Pak plus column (preconditioned with 5 mL of hexane) under
an argon atmosphere and eluted with ethyl acetate (2 × 1.5 mL).
The ethyl acetate solution was dried as described earlier, and 0.4
mL of 1,2-dichloroethane and 0.1 mL 30% HBr in acetic acid were
added. The reaction mixture was heated at 80 °C for 10 min, and
0% HBr in acetic acid was removed by azeotropic distillation with
mL of toluene. After ensuring complete removal of excess
reagents and solvent, 50 mg of silyl ether derivative in 0.7 mL of
,2-dichloroethane was added under an argon atmosphere, and the
reaction mixture was heated to 110 °C for 1 h. Then reaction
mixture was cooled and passed through a silica Sep-Pak plus
column (preconditioned with 5 mL of hexane) under an argon
counts in 1g of octanol
log10 P ) log10
( counts in 1 g of water
)
In Vitro Studies. RG2TK+ and RG2 cells in culture were used
1
4
3
to compare the accumulation of [ C]FIAU, [ H]thymidine, and each
18
18
18
of the thymidine analogues [ F]FBrVAU, [ F]FPAU, and [ F]-
FTMAU, as previously described. Paired time-course radiotracer
1
accumulation experiments were run over 120 min, and the data
3
were normalized to [ H]TdR accumulation. The incubation medium
1
4
18
18
3
1
contained [ C]FIAU (0.37 kBq/mL), [ F]FBrVAU, [ F]FPAU (10
18
3
kBq/mL), or [ F]FTMAU and [ H]TdR (3.7 kBq/mL). The
radioactivity assay for the in vitro tissue culture studies involved
F γ-counting (AutoGamma 5550 spectrometer; Packard Instru-
ments, Meriden, CT) immediately after the experiment, followed
18
1
3
14
24-36 h later by H and C â-isotope counting (Tri-Carb liquid
scintillation analyzer, model 1600TR; Packard Instruments) using
external standard quench correction and standard dual-counting
techniques. All data were expressed as disintegrations per minute
(dpm)/g cells (or dpm/mL medium). The net accumulation rate was
calculated from the slope of the probe accumulation versus time
plot ((dpm/g cells)/(dpm/mL medium) vs time; units ) (mL
medium/min)/g cells - a medium clearance constant).
atmosphere and eluted with 10% MeOH in CH
Cl
2 2
(2 × 1.25 mL).
The solvent was removed, and 0.4 mL of 0.5 M sodium methoxide
in MeOH was added. The reaction mixture was heated at 80 °C
for 10 min and then cooled to room temperature. The reaction
mixture was neutralized with 0.1 mL of 2 N HCl in MeOH, and
the solvent was removed under reduced pressure. The products,
including the R and â anomers, were purified using reverse-phase
HPLC using a C-18 column (250 × 10 mm) with an appropriate
Results
18
HPLC solvent (12% ethanol in 0.01 M saline solution for [ F]-
18
Synthesis. The synthesis of [ F]FBrVAU was carried out
1
8
FBrVAU, 8% ethanol in 0.01 M saline solution for [ F]FPAU,
with minor modifications to the procedure previously reported.16
Although the reaction time was almost the same as that reported
18
and 5% ethanol in 0.01 M saline solution for [ F]FTMAU) at a
flow rate of 7 mL/min. The radiochemical purity of the product
was >98% with decay-corrected yields of 12-20% (see Table 1).
The specific activity of the compounds was >58 GBq/µmol. The
total time of synthesis was about 180 min. The nonradioactive
analogues were prepared using a similar procedure and characterized
1
6
previously, the application of a vacuum resulted in complete
drying of the products during the intermediate steps, and this
ensured consistent yields. Both R and â anomers were obtained
during the synthesis, and the biologically useful â anomer was

Brief Articles
Journal of Medicinal Chemistry, 2006, Vol. 49, No. 17 5379
14
18
Figure 3. Typical in vitro uptake profile of [ C]FIAU and [ F]FPAU
in RG2TK+ and RG2 cells.
1
4
18
Figure 2. Typical in vitro uptake profile of [ C]FIAU and [ F]-
FBrVAU in RG2TK+ and RG2 cells.
1
8
obtained as the major product. The ratio of anomers obtained
Figures 2-4. It is apparent from the plots that [ F]FBrVAU,
18
18
varied slightly between 1:4 and 1:5 (R/â). This method was
[ F]FPAU, and [ F]FTMAU are accumulated only in RG2TK+
18
18
18
easily extended for the synthesis of [ F]FPAU and [ F]FTMAU
and did not result in any significant difference in the ratio of
the anomers. The yields varied slightly, but the method has not
been optimized for each individual compound. The nonradioac-
cells and not in RG2 cells. The uptake rates for [ F]FBrVAU,
18
18
[ F]FPAU, and [ F]FTMAU in RG2TK+ and RG2 cells were
calculated from the slopes of the plots and normalized to that
of TdR (Table 2). The rate of uptake follows the order [ F]-
FTMAU > [ F]FBrVAU > [ F]FPAU. The corresponding
values for FIAU, which is considered as a reference standard,
are also included in Table 2.
1
18
19
18
18
tive ( F) analogues were synthesized using a similar procedure,
and they were characterized by NMR and mass spectrometry
(
see Supporting Information). The results corresponded to the
2
0,21
reported values.
The purified compounds were used as
Discussion
reference standards for HPLC analysis. Figure 1 shows the
18
HPLC chromatogram of purified [ F]FBrVAU co-injected with
its nonradioactive analogue.
Medical imaging (radionuclide, magnetic resonance, and
optical) has undergone extraordinary changes in the past decade.
This has largely been due to improved imaging technology and
to convergence with complementary advances in molecular
biology and cell biology. Noninvasive reporter gene imaging
Octanol/Water Partition Coefficient Studies. The log of
18
the octanol/water partition coefficients (log P) of [ F]FBrVAU,
1
8
18
[
F]FPAU, and [ F]FTMAU are 1.02 ( 0.06, 0.50 ( 0.03,
1
-3,25-27
and 0.03 ( 0.03 at pH 7.4, respectively. The corresponding
value for FIAU is -0.10 ( 0.01 at pH 7.4. The higher log P
is now widely applied in preclinical studies,
and initial
2
8,29
clinical applications are now being reported.
Reporter gene
1
8
18
values for [ F]FBrVAU and [ F]FPAU compared to FIAU is
in accordance with the observed trend of increasing alkyl chain
length corresponding with increasing lipophilicity. The low log P
constructs driven by constitutive promoter elements can be used
to monitor transduced cells in adoptive therapies and gene
therapy vectors. Inducible promoters function as sensor elements
and can be used to provide information about endogenous gene
18
value of [ F]FTMAU probably reflects the high polarizing
effect of the trifluoromethyl group, which can impart a slight
ionic character to the molecule.
3
0,31
regulation,
mRNA stabilization, and specific protein-protein
3
2-34
interactions.
In addition, the promoter can also be cell-
1
8
In Vitro Uptake Studies. In vitro uptake studies of [ F]-
specific, restricting expression of the transgene to certain tissue
and organs. These “molecular images” can provide spatial,
temporal, and functional information about various biological
processes at the cellular level, averaged over the resolution
element of the acquired image.
1
8
18
FBrVAU, [ F]FPAU, and [ F]FTMAU were performed con-
currently in HSV1-TK expressing RG2TK+ cells and in wild-
3
type RG2 cells (negative control). Two reference tracers, [ H]-
14
TdR (for cell viability) and [ C]FIAU (for HSV1-TK expression),
were also included in the incubation medium for paired
comparisons. The results of these uptake studies are shows in
Radiotracer-based reporter imaging (γ camera or PET)
involves a complementary “reporter gene” and “reporter probe”.

5
380 Journal of Medicinal Chemistry, 2006, Vol. 49, No. 17
Brief Articles
Table 3. Comparison of Normalized Accumulation Rates in RG2TK+
and RG2 Cell Lines
RG2TK+,
RG2,
RG2TK+/
RG2
-
1
-1
mL mg-1 min-1
compd
mL mg min
1
1
1
1
4
8
8
8
[
[
[
[
C]FIAU
F]FFEAU
F]FHPG
F]FHBG
0.31 ( 0.07
0.29 ( 0.05
0.0077 ( 0.0003
0.022 ( 0.001
0.036 ( 0.012
∼8.6
∼550
∼19
0.00053 ( 0.00058
0.0004 ( 0.0003
0.0032 ( 0.0026
∼6.9
All three compounds demonstrate rapid equilibration across
cell membranes and selective accumulation in HSV1-tk trans-
duced RG2 cells compared to wild-type RG2 cells. Although
the net accumulation rates were not as high as that reported for
1
35
FIAU and FFEAU, they are substantially higher or compa-
rable to that obtained with FHBG and FHPG in the same cell
1
4
lines (Table 3). Importantly, all three compounds have very
low uptake in wild-type RG2 cells in comparison to that
obtained with FIAU. The uptake ratio of FTMAU (RG2TK+/
RG2) is comparable to that obtained with FIAU and with FHBG
and FHPG (Tables 2 and 3). The uptake ratio (transduced to
nontransduced cells) of FBrVAU and FPAU is likely to be even
higher (more comparable to that of FFEAU) because of the very
low uptake in wild-type RG2 cells. Thus, background activity
due to phosphorylation by endogenous mammalian thymidine
kinase in rapidly proliferating tissue would be expected to be
low in comparison to that with FIAU, and this will contribute
to higher specificity for HSV1 thymidine kinase activity.
Conclusion
These results demonstrate that the synthetic procedure
developed by Alauddin et al. for the synthesis of 2′-deoxy-2′-
18
[
F]fluoro-1â-D-arabinofuranosyl-5-methyluracil can easily be
extended to other 5-alkyl substituted analogues without a
significant loss in yields or changes in the anomeric ratio. The
in vitro experiments reveal that [ F]FBrVAU and [ F]FTMAU
1
4
18
Figure 4. Typical in vitro uptake profile of [ C]FIAU and [ F]-
FTMAU in RG2TK+ and RG2 cells.
18
18
Table 2. Comparison of Normalized Accumulation Rates in RG2TK+
are good substrates for the HSV1-tk expressing RG2TK+ cells.
and RG2 Cell Lines
18
The higher hydrophobicity (log P ) 1.02 at pH 7.4) of [ F]-
RG2TK+,
RG2,
RG2TK+/
RG2
n
FBrVAU indicates that it has a better chance of passively
diffusing across cell membranes and crossing an intact blood-
brain barrier than FIAU or the other radiolabeled HSV1-TK
1
-1
mL mg min-1
-1
compd
mL mg- min
(no. expts)
[1
4
8
8
8
C]FIAU
0.274 ( 0.095
0.048 ( 0.036
∼5.7
10
4
3
1
[
F]FBrVAU 0.035 ( 0.017 -0.001 ( 0.001
F]FPAU
F]FTMAU
18
probes. The hydrophilicity and pH dependency of [ F]FTMAU
1
[
0.014 ( 0.003 -0.001 ( 0.001
0.051 ( 0.005 0.004 ( 0.001
1
may limit its use as a PET probe for imaging HSV1-tk gene
expression in non-CNS tissue.
[
∼13
3
The reporter gene product is frequently an enzyme that converts
a radiolabeled substrate (the “reporter probe”) to a metabolite
that is selectively trapped within transduced cells. Alternatively,
the reporter gene product can be a receptor or transporter that
Acknowledgment. This work was supported in part by NIH
Grant P50-CA86438, U.S. Department of Energy Grant DE-
FG02-86ER60407, Center Support Core Grant NCI-P30-CA-
0
8748, and National Center for Research Resources (NCRR)
“
irreversibly” traps the radiolabeled probe in transduced cells
Grant C06 RR11192.
during the period of image acquisition.
Three novel pyrimidine analogues were developed with the
goal to attain higher lipophilicity than FIAU to facilitate passive
diffusion across cell membranes and across the intact blood-
brain barrier while retaining selective phosphorylation by HSV1-
Supporting Information Available: Spectroscopic character-
ization data of nonradioactive analogues of 5a-c and details on
cell cultures, transduction, and in vitro uptake study. This material
is available free of charge via the Internet at http://pubs.acs.org.
1
8
18
18
tk. [ F]FBrVAU, [ F]FPAU, and [ F]FTMAU were synthe-
sized in good yields following a procedure developed by
Alauddin et al. The yields obtained for each of the compounds
were consistent (Table 1). The R/â anomeric ratio of the
compounds varied between 1:4 to 1:5. The total time for
synthesis was about 3 h from EOB.
References
(1) Tjuvajev, J. G.; Stockhammer, G.; Desai, R.; Uehara, H.; Watanabe,
K.; Gansbacher, B.; Blasberg, R. G. Imaging the expression of
transfected genes in vivo. Cancer Res. 1995, 55, 6126-6132.
(
2) Tjuvajev, J. G.; Finn, R.; Watanabe, K.; Joshi, R.; Oku, T.; Kennedy,
J.; Beattie, B.; Koutcher, J.; Larson, S.; Blasberg, R. G. Noninvasive
imaging of herpes virus thymidine kinase gene transfer and expres-
sion: a potential method for monitoring clinical gene therapy. Cancer
Res. 1996, 56, 4087-4095.
18
18
As expected, the log P values of [ F]FBrVAU and [ F]-
FPAU indicate that both of these compounds are significantly
more hydrophobic compared to FIAU (log P ) -0.10) and
reflects the longer carbon chain at the 5-position. On the basis
of log P, FBrVAU and FPAU would be expected to cross cell
membranes and the blood-brain barrier more readily by passive
diffusion (noncarrier mediated transport) than FIAU.
(3) Tjuvajev, J. G.; Avril, N.; Oku, T.; Sasajima, T.; Miyagawa, T.; Joshi,
R.; Safer, M.; Beattie, B.; DiResta, G.; Daghighian, F.; Augensen,
F.; Koutcher, J.; Zweit, J.; Humm, J.; Larson, S. M.; Finn, R.;
Blasberg, R. Imaging herpes virus thymidine kinase gene transfer
and expression by positron emission tomography. Cancer Res. 1998,
58, 4333-4341.

Brief Articles
Journal of Medicinal Chemistry, 2006, Vol. 49, No. 17 5381
(
4) Alauddin, M. M.; Shahinian, A.; Kundu, R. K.; Gordon, E. M.; Conti,
(21) Su, T. L.; Watanabe, K. A.; Schinazi, R. F.; Fox, J. J. Nucleosides.
136. Synthesis and antiviral effects of several 1-(2-deoxy-2-fluoro-
â-D-arabinofuranosyl)-5-alkyluracils. Some structure-activity rela-
tionships. J. Med. Chem. 1986, 29, 151-154.
(22) Watanabe, K. A.; Reichman, U.; Hirota, K.; Lopez, C.; Fox, J. J.
Nucleosides. 110. Synthesis and anti-herpes virus activity of some
2′-fluoro-2′-deoxyarabinofuranosylpyrimidine nucleosides. J. Med.
Chem. 1979, 22, 21-24.
1
8
P. S. Evaluation of 9-[(3- F-fluoro-1-hydroxy-2-propoxy)methyl]-
18
guanine ([ F]-FHPG) in vitro and in vivo as a probe for PET imaging
of gene incorporation and expression in tumors. Nucl. Med. Biol.
1999, 26, 371-376.
(
5) Gambhir, S. S.; Barrio, J. R.; Wu, L.; Iyer, M.; Namavari, M.;
Satyamurthy, N.; Bauer, E.; Parrish, C.; MacLaren, D. C.; Borghei,
A. R.; Green, L. A.; Sharfstein, S.; Berk, A. J.; Cherry, S. R.; Phelps,
M. E.; Herschman, H. R. Imaging of adenoviral-directed herpes
simplex virus type 1 thymidine kinase reporter gene expression in
mice with radiolabeled ganciclovir. J. Nucl. Med. 1998, 39, 2003-
(23) Watanabe, K. A.; Su, T. L.; Klein, R. S.; Chu, C. K.; Matsuda, A.;
Chun, M. W.; Lopez, C.; Fox, J. J. Nucleosides. 123. Synthesis of
antiviral nucleosides. 5-Substituted 1-(2-deoxy-2-halogeno-â-D-ara-
binofuranosyl)cytosines and -uracils. Some structure activity relation-
ships. J. Med. Chem. 1983, 26, 152-156.
2011.
(
6) Keller, P. M.; Fyfe, J. A.; Beauchamp, L.; Lubbers, C. M.; Furman,
P. A.; Schaeffer, H. J.; Elion, G. B. Enzymatic phosphorylation of
acyclic nucleoside analogs and correlations with antiherpetic activi-
ties. Biochem. Pharmacol. 1981, 30, 3071-3077.
(24) Watanabe, K. A.; Su, T. L.; Reichman, U.; Greenberg, N.; Lopez,
C.; Fox, J. J. Nucleosides. 129. Synthesis of antiviral nucleosides.
5
-Alkenyl-1-(2-deoxy-2-fluoro-â-D-arabinofuranosyl)uracils. J. Med.
(
(
(
7) De Clercq, E. Antivirals and antiviral strategies. Nat. ReV. Microbiol.
Chem. 1984, 27, 91-94.
2004, 2, 704-720.
(
25) Yu, Y.; Annala, A. J.; Barrio, J. R.; Toyokuni, T.; Satyamurthy, N.;
Namavari, M.; Cherry, S. R.; Phelps, M. E.; Herschman, H. R.;
Gambhir, S. S. Quantification of target gene expression by imaging
reporter gene expression in living animals. Nat. Med. 2000, 6, 933-
8) Moolten, F. L. Drug sensitivity (“suicide”) genes for selective cancer
chemotherapy. Cancer Gene Ther. 1994, 1, 279-287.
9) Ram, Z.; Culver, K. W.; Oshiro, E. M.; Viola, J. J.; DeVroom, H.
L.; Otto, E.; Long, Z.; Chiang, Y.; McGarrity, G. J.; Muul, L. M.;
Katz, D.; Blaese, R. M.; Oldfield, E. H. Therapy of malignant brain
tumors by intratumoral implantation of retroviral vector-producing
cells. Nat. Med. 1997, 3, 1354-1361.
937.
(
26) Gambhir, S. S.; Bauer, E.; Black, M. E.; Liang, Q.; Kokoris, M. S.;
Barrio, J. R.; Iyer, M.; Namavari, M.; Phelps, M. E.; Herschman, H.
R. A mutant herpes simplex virus type 1 thymidine kinase reporter
gene shows improved sensitivity for imaging reporter gene expression
with positron emission tomography. Proc. Natl. Acad. Sci. U.S.A.
(
10) Sandmair, A. M.; Loimas, S.; Puranen, P.; Immonen, A.; Kossila,
M.; Puranen, M.; Hurskainen, H.; Tyynela, K.; Turunen, M.;
Vanninen, R.; Lehtolainen, P.; Paljarvi, L.; Johansson, R.; Vapalahti,
M.; Yla-Herttuala, S. Thymidine kinase gene therapy for human
malignant glioma, using replication-deficient retroviruses or aden-
oviruses. Hum. Gene Ther. 2000, 11, 2197-2205.
11) Singhal, S.; Kaiser, L. R. Cancer chemotherapy using suicide genes.
Surg. Oncol. Clin. N. Am. 1998, 7, 505-536.
12) Conti, P. S.; Alauddin, M. M.; Fissekis, J. R.; Schmall, B.; Watanabe,
2
000, 97, 2785-2790.
(
27) Gambhir, S. S.; Barrio, J. R.; Phelps, M. E.; Iyer, M.; Namavari,
M.; Satyamurthy, N.; Wu, L.; Green, L. A.; Bauer, E.; MacLaren,
D. C.; Nguyen, K.; Berk, A. J.; Cherry, S. R.; Herschman, H. R.
Imaging adenoviral-directed reporter gene expression in living
animals with positron emission tomography. Proc. Natl. Acad. Sci.
U.S.A. 1999, 96, 2333-2338.
(
(
11
K. A. Synthesis of 2′-fluoro-5-[ C]-methyl-1-beta-D-arabinofurano-
1
1
(28) Jacobs, A.; Braunlich, I.; Graf, R.; Lercher, M.; Sakaki, T.; Voges,
J.; Hesselmann, V.; Brandau, W.; Wienhard, K.; Heiss, W.-D.
Quantitative kinetics of [1 I]FIAU in cat and man. J. Nucl. Med.
syluracil ([ C]-FMAU): a potential nucleoside analog for in vivo
study of cellular proliferation with PET. Nucl. Med. Biol. 1995, 22,
24
783-789.
2
001, 42, 467-475.
(
13) Alauddin, M. M.; Shahinian, A.; Gordon, E. M.; Conti, P. S. Direct
comparison of radiolabeled probes FMAU, FHBG, and FHPG as PET
imaging agents for HSV1-tk expression in a human breast cancer
model. Mol. Imaging 2004, 3, 76-84.
14) Tjuvajev, J. G.; Doubrovin, M.; Akhurst, T.; Cai, S.; Balatoni, J.;
Alauddin, M. M.; Finn, R.; Bornmann, W.; Thaler, H.; Conti, P. S.;
Blasberg, R. G. Comparison of radiolabeled nucleoside probes (FIAU,
FHBG, and FHPG) for PET imaging of HSV1-tk gene expression.
J. Nucl. Med. 2002, 43, 1072-1083.
(
29) Yaghoubi, S.; Barrio, J. R.; Dahlbom, M.; Iyer, M.; Namavari, M.;
Satyamurthy, N.; Goldman, R.; Herschman, H. R.; Phelps, M. E.;
Gambhir, S. S. Human pharmacokinetic and dosimetry studies of
1
8
[
F]FHBG: a reporter probe for imaging herpes simplex virus type-1
thymidine kinase reporter gene expression. J. Nucl. Med. 2001, 42,
225-1234.
(
1
(
30) Doubrovin, M.; Ponomarev, V.; Beresten, T.; Balatoni, J.; Bornmann,
W.; Finn, R.; Humm, J.; Larson, S.; Sadelain, M.; Blasberg, R.;
Gelovani, T. J. Imaging transcriptional regulation of p53-dependent
genes with positron emission tomography in vivo. Proc. Natl. Acad.
Sci. U.S.A. 2001, 98, 9300-9305.
(
(
(
15) Alauddin, M. M.; Shahinian, A.; Gordon, E. M.; Conti, P. S.
Evaluation of 2′-deoxy-2′-flouro-5-methyl-1-beta-D-arabinofurano-
syluracil as a potential gene imaging agent for HSV-tk expression
in vivo. Mol. Imaging 2002, 1, 74-81.
(31) Serganova, I.; Doubrovin, M.; Vider, J.; Ponomarev, V.; Soghom-
onyan, S.; Beresten, T.; Ageyeva, L.; Serganov, A.; Cai, S.; Balatoni,
J.; Blasberg, R.; Gelovani, J. Molecular imaging of temporal
dynamics and spatial heterogeneity of hypoxia-inducible factor-1
signal transduction activity in tumors in living mice. Cancer Res.
2004, 64, 6101-6108.
(32) Luker, G. D.; Pica, C. M.; Song, J.; Luker, K. E.; Piwnica-Worms,
D. Imaging 26S proteasome activity and inhibition in living mice.
Nat. Med. 2003, 9, 969-973.
(33) Luker, G. D.; Sharma, V.; Pica, C. M.; Prior, J. L.; Li, W.; Piwnica-
Worms, D. Molecular imaging of protein-protein interactions:
controlled expression of p53 and large T-antigen fusion proteins in
vivo. Cancer Res. 2003, 63, 1780-1788.
16) Alauddin, M. M.; Conti, P. S.; Fissekis, J. D. A general synthesis of
2
′-deoxy-2′-[F-18]fluoro-1-beta-D-arabinofuranosyluracil and its 5-sub-
stituted nucleosides. J. Labelled Compd. Radiopharm. 2003, 46, 285-
89.
17) Mangner, T. J.; Klecker, R. W.; Anderson, L.; Shields, A. F. Synthesis
2
1
8
18
of 2′-deoxy-2′-[ F]fluoro-beta-D-arabinofuranosyl nucleosides, [ F]-
1
8
18
18
FAU, [ F]FMAU, [ F]FBAU and [ F]FIAU, as potential PET
18
agents for imaging cellular proliferation. Synthesis of [ F]labeled
FAU, FMAU, FBAU, FIAU. Nucl. Med. Biol. 2003, 30, 215-224.
18) Tann, C. H.; Brodfuehrer, P. R.; Brundidge, S. P.; Sapino, C.; Howell,
H. G. Fluorocarbohydrates in synthesis. An efficient synthesis of 1-(2-
deoxy-2-fluoro-beta-D-arabinofuranosyl)-5-iodouracil (â-FIAU) and
(
1
-(2-deoxy-2-fluoro-beta-D-arabinofuranosyl)thymine (â-FMAU). J.
(34) Luker, K. E.; Smith, M. C.; Luker, G. D.; Gammon, S. T.; Piwnica-
Worms, H.; Piwnica-Worms, D. Kinetics of regulated protein-protein
interactions revealed with firefly luciferase complementation imaging
in cells and living animals. Proc. Natl. Acad. Sci. U.S.A. 2004, 101,
12288-12293.
Org. Chem. 1985, 50, 3644-3647.
(
19) Howell, H. G.; Brodfuehrer, P. R.; Brundidge, S. P.; Benigni, D. A.;
Sapino, C. Antiviral nucleosides. A stereospecific, total synthesis of
2
′-fluoro-2′-deoxy-â-D-arabinofuranosyl nucleosides. J. Org. Chem.
1988, 53, 85-88.
(35) Balatoni, J. A.; Doubrovin, M.; Ageyeva, L.; Pillarsetty, N.; Finn,
R. D.; Gelovani, J. G.; Blasberg, R. G. Imaging herpes viral thymidine
(
20) Matulic-Adamic, J.; Takahashi, K.; Chou, T. C.; Gadler, H.; Price,
R. W.; Reddy, A. R.; Kalman, T. I.; Watanabe, K. A. Nucleosides.
18
kinase-1 reporter gene expression with a new F-labeled probe: 2′-
1
50. Synthesis and some biological properties of 5-monofluoromethyl,
18
fluoro-2′-deoxy-5-[ F]fluoroethyl-1-beta-d-arabinofuranosyl uracil.
5-difluoromethyl, and 5-trifluoromethyl derivatives of 2′-deoxyuridine
Nucl. Med. Biol. 2005, 32, 811-819.
and 2′-deoxy-2′-fluoro-â-D-arabinofuranosyluracil. J. Med. Chem.
1988, 31, 1642-1647.
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