Synthesis of a technetium-99m labeled tricyclic ganciclovir analog for non-invasive reporter gene expression imaging

Article abstract of DOI:10.1016/j.bmcl.2006.10.077

A potential radiopharmaceutical and HSV1-TK substrate, 3-((1,3-dihydroxypropan-2-yloxy)-methyl)-6-(4-(3-((2-mercaptoethyl)(2-(2-mercaptoethyl-amino)-ethyl)amino)propoxy)phenyl)-3H-imidazopurin-9(5H)-one-oxo-technetium(V), was synthesized via a converging

Full text of DOI:10.1016/j.bmcl.2006.10.077

Bioorganic & Medicinal Chemistry Letters 17 (2007) 741–744
Synthesis of a technetium-99m labeled tricyclic ganciclovir analog
for non-invasive reporter gene expression imaging
Yi Zhang, Jing Lin and Dongfeng Pan*
The Department of Radiology, University of Virginia, Charlottesville, VA 22908, USA
Received 19 September 2006; revised 24 October 2006; accepted 25 October 2006
Available online 28 October 2006
Abstract—A potential radiopharmaceutical and HSV1-TK substrate, 3-((1,3-dihydroxypropan-2-yloxy)-methyl)-6-(4-(3-((2-merca-
ptoethyl)(2-(2-mercaptoethyl-amino)-ethyl)amino)propoxy)phenyl)-3H-imidazopurin-9(5H)-one-oxo-technetium(V), was synthe-
sized via a converging approach and its chemical structure was comparatively characterized with a non-radioactive analog. The
final radiochemical purity and yield were 97 and 73%, respectively.
ꢀ 2006 Elsevier Ltd. All rights reserved.
Non-invasive nuclear imaging of the Herpes simplex
virus type-1 thymidine kinase (HSV1-TK) expression
has received considerable attention due to its potential
clinical application in gene therapy.1–4 Current nuclear
reporter probes of HSV1-TK include fluorine and iodine
isotope labeled pyrimidine nucleoside derivatives5–9 and
acycloguanosine derivatives.10,11 However, their use in
routine procedures is hampered by either the limited
supply of the cyclotron-produced isotopes, F-18 and
I-123/124, or the suboptimal imaging characteristics of
I-131, which yields high and multiple emission energies.
There are currently interests for novel tracers suitable
for routine application.
the 4-position of the phenyl ring are permitted
(Fig. 1). Radiometal labeling was accomplished through
a neutral bisaminoethanethiol (N2S2) based chelating
moiety.
In this communication, we report the synthesis and
characterization of 3-((1,3-dihydroxypropan-2-yloxy)
methyl)-6-(4-(3-((2-mercaptoethyl)(2-(2-mercaptoethyl-
amino)ethyl)amino)-propoxy)phenyl)-3H-imidazo[1,2-f]-
purin-9(5H)-one-oxo-[Tc-99m] 1. To characterize the
chemical structure of the Tc-99m labeled probe, a non-
radioactive mimic compound, 3-((1,3-dihydroxypropan-
2-yloxy)methyl)-6-(4-(3-((2-mercaptoethyl)(2-(2-merca-
ptoethylamino)ethyl)amino)propoxy)-phenyl)-3H-imi-
dazo-[1,2-f]purin-9(5H)-one-oxo-[Re] 215 is synthesized
for NMR and MS analysis and HPLC identification.
Miscellaneous potential HSV-TK subjects have been
studied as either imaging or antiherpes therapeutic
agent. For example, a series of tricyclic analogs of
ACV and GCV have been synthesized and evaluated
as potential fluorescent cellular imaging probes for
herpes virus assay and some of them demonstrated the
similar activity and selectivity to herpes simplex virus
type I as ACV or GCV.12–14
At first, the tricyclic fragment 316 was synthesized
(Scheme 1) as follows. Coupling GCV 4 and phenacyl
bromide 5 yielded the tricyclic TGCV 6,12 followed by
O
In our exploration of the Tc-99m labeled TK substrates
for nuclear gene imaging, tricyclic ganciclovir (TGCV)
was selected as a template because it seems to be suffi-
cient for HSV-TK interaction and the modifications at
N
N
O
N
S
O
S
N
99mTc
N
N
H
N
O
HO
Keywords: Nuclear imaging tracer; HSV1-TK substrate; Technetium-
99m.
HO
1
*
Corresponding author. Tel.: +1 434 243 2893; fax: +1 434 924
9435; e-mail: dp3r@virginia.edu
Figure 1. Technetium-99m complex of tricyclic ganciclovir (TGCV).
0960-894X/$ - see front matter ꢀ 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2006.10.077

742
Y. Zhang et al. / Bioorg. Med. Chem. Lett. 17 (2007) 741–744
O
N
O
N
O
O
N
N
N
N
N
HN
H2N
O
O
N
H
i
+
O
O
Br
OH
OH
O
OH
OH
6
4
5
O
O
N
O
N
N
N
N
N
N
HO
O
N
H
N
N
iii
H
ii
O
O
O
O
O
O
7
8
O
N
N
N
N
HO
iv
N
O
O
O
O
O
3
Scheme 1. Reagents and condition: (i) NaH/DMF, 86%; (ii) DMF, 2-methoxypropene, TsOHÆH2O, 74%; (iii) NH3–MeOH, 52%; (iv) Boc2O,
1,4-dioxane, aq Na2CO3, 78%.
protecting the free hydroxyls with 2-methoxypropene in
presence of TsOH to give isopropylidene ketal 7.17
Removal of the isobutanoyl group of 7 with saturated
methanolic ammonia gave phenol 8.18 The secondary
amino group at position 5 was protected with t-Boc by
mixing compound 8 with di-tert-butyl dicarbonate in
1,4-dioxane and aqueous Na2CO3.
the presence of Sn-glucoheptonate was added 2.1 mCi
of 99mTc pertechnetate in PBS buffer, and 1.5 mCi of
the target radiochemical 1 was obtained after HPLC
purification with a C18 reversed-phase Econosphere
column (250 · 10 mm, 30% CH3CN in H2O, 3 mL/
min, 254 nm). The radiotracer was collected with reten-
tion time at 10.38 min and radiochemical yield of 73%.
To characterize the chemical structure of the radiochem-
ical 1, a rhenium conjugated analog 2 was synthesized
by adding tetrabutylammonium tetrachlorooxorhe-
nate(V) into compound 14 in methanol and stirring for
12 h. The rhenium conjugated 2 was purified by flash
The synthesis of radiometal-chelating moiety 919 started
from N,N0-bis-[2-(4-methoxy-benzylsulfanyl)-ethyl]-eth-
ylenediamine 10, which was synthesized as described
previously (Scheme 2).20 After blocking one of the sec-
ondary amino groups of 10 with t-Boc to produce
11,21 the monoalkylated 1222 was obtained by coupling
of 11 with 3-bromo-1-propanol. The subsequent bromi-
nation of 12 with NBS/triphenylphosphine yielded 9 (see
Scheme 3).
1
chromatography and characterized with H NMR and
mass spectrum. The characterization of Tc-99m tracer
1 was carried out by co-injection with rhenium analog
2 using reverse phase HPLC.
In conclusion, we have successfully synthesized a Tc-
99m labeled acycloguanosine derivative 1, a potential
HSV1-TK substrate for nuclear gene imaging. In vitro
cell uptake assays of the compound are currently in
progress.
Coupling of TGCV derivative 3 and chelator fragment 9
produced 13,23 the precursor of the target radiotracer 1.
After deprotection of 13 with Hg(OAc)2 in TFA, to the
crude intermediate 14 (50 lg) in aqueous methanol in

Y. Zhang et al. / Bioorg. Med. Chem. Lett. 17 (2007) 741–744
743
HO
S
S
PMB
PMB
S
S
PMB
PMB
NH
NH
NH
N
ii
S
S
PMB
PMB
i
N
N
Boc
Boc
11
12
10
Br
S
PMB
PMB
N
N
iii
S
Boc
9
Scheme 2. Reagents and condition: (i) Boc2O, 1,4-dioxane, aq Na2CO3, 74%; (ii) acetonitrile, 3-bromo-1-propanol, DIEA, 71%; (iii) NBS, PPh3,
DMF, 64%.
O
N
N
O
N
N
S
S
PMB
PMB
ii
N
N
N
i
3 + 9
O
O
Boc
O
O
O
13
O
N
O
N
N
N
N
N
N
N
O
N
N
S
S
O
iii
N
SH
SH
Re
N
O
N
H
H
NH
O
O
HO
14
HO
HO
HO
2
iv
1
Scheme 3. Reagents and condition: (i) CHCl3, aq Na2CO3, Bu4N+Brꢀ, 81%; (ii) Hg(OAc)2/TFA, H2S; (iii) (Bu4N)+(ReOCl4)ꢀ, 58%; (iv)
99mTc]NaTcO4, Sn-gluceptate, 73%.
[
3. Shah, K.; Jacobs, A.; Breakefield, X. O.; Weissleder, R.
Gene Ther. 2004, 11, 1175.
Acknowledgments
4. Lyer, M.; Gambhir, S. S. Lung Biol. Health Dis. 2005, 203,
171.
5. Mercer, J. R.; Xu, L. H.; Knaus, E. E.; Wiebe, L. I.
J. Med. Chem. 1989, 32, 1289.
This work was supported in part by Grants from DOD
PC010112, the Nihon Medi-Physics Co. Ltd (NMP),
and the Department of Radiology, University of Virginia.
6. Tjuvajev, J. G.; Avril, N.; Oku, T.; Sasajima, T.; Miya-
gawa, 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. Cancer
Res. 1998, 58, 4333.
References and notes
1. Yaghoubi, S. S.; Barrio, J. R.; Namavari, M.; Satyamur-
thy, N.; Phelps, M. E.; Herschman, H. R.; Gambhir, S. S.
Cancer Gene Ther. 2005, 12, 329.
7. Morin, K. W.; Atrazheva, E. D.; Knaus, E. E.; Wiebe, L.
I. J. Med. Chem. 1997, 40, 2184.
2. Min, J. J.; Lyer, M.; Gambhir, S. S. Eur. J. Nucl. Med.
Mol. Imaging 2003, 30, 1547.
8. Wiebe, L. I.; Knaus, E. E.; Morin, K. W. Nucleosides
Nucleotides 1999, 18, 1065.

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Y. Zhang et al. / Bioorg. Med. Chem. Lett. 17 (2007) 741–744
1
9. Tjuvajev, J. G.; Finn, R.; Watanabe, K.; Joshi, R.; Oku,
T.; Kennedy, J.; Beattie, B.; Koutcher, J.; Larson, S.;
Blasberg, R. G. Cancer Res. 1996, 56, 4087.
10. Alauddin, M. M.; Shahinian, A.; Kundu, R. K.; Gordon,
E. M.; Conti, P. S. Nucl. Med. Biol. 1999, 26, 371.
11. Alauddin, M. M.; Conti, P. S. Nucl. Med. Biol. 1998, 25,
175.
18. Compound 8. H NMR (DMSO-d6, 300 MHz), d 1.27 (s,
6H), 3.56 (m, 1H), 3.60 (m, 2H), 3.83 (m, 2H), 5.54 (s, 2H),
6.85, 7.71 (dd, 2H, 2H), 7.97 (s, 1H), 8.06 (s, 1H), 9.82 (s,
1H). Mass spectrum (ESI) m/z 412.1615 (M + H)+
(C20H22N5O5 requires 412.1615).
19. Compound 9. 1H NMR (CDCl3, 300 MHz), d 1.44 (s, 9H),
1.88 (m, 2H), 2.54 (m, 10H), 3.17 (m, 2H), 3.32 (m, 2H),
3.45 (t, 2H), 3.67 (s, 4H), 3.79 (s, 6H), 6.85, 7.25 (dd, 2H,
12. Goslinski, T.; Golankiewicz, B.; De Clercq, E.; Balzarini,
J. J. Med. Chem. 2002, 45, 5052.
2H).
Mass
spectrum
(ESI)
m/z
641.2078
13. Golankiewicz, B.; Ostrowski, T.; Andrei, G.; Snoeck, R.;
De Clercq, E. J. Med. Chem. 1994, 37, 3187.
14. Goslinski, T.; Wenska, G.; Golankiewicz, B.; Balzarini, J.;
De Clercq, E. Nucleosides Nucleotides Nucleic Acids 2003,
22, 911.
(M+H)+(C30H46BrN2O4S2 requires 641.2077).
20. Zhang, Y.; Dai, X.; Kallmes, D. F.; Pan, D. Tetrahedron
Lett. 2004, 45, 8673.
21. Compound 11. 1H NMR (CDCl3, 300 MHz), d 1.42 (s,
9H), 2.52 (m, 6H), 2.63 (m, 2H), 2.70 (m, m, 2H), 3.21 (m,
2H), 3.62 (s, 2H), 3.65 (s, 2H), 3.73 (s, 6H), 6.80, 7.20 (dd,
4H, 4H). Mass spectrum (ESI) m/z 521.2502 (M+H)+
(C27H40N2O4S2 requires 521.2502).
15. Compound 2. 1H NMR (CDCl3, 300 MHz), d 1.59 (m,
2H), 2.21(m, 2H), 2.50 (m, 10H), 3.13 (m, 4H), 3.42 (m,
2H), 3.61 (m, 2H), 3.81 (m, 2H), 5.63(s, 2H), 6.93, 7.44
(dd, 2H, 2H), 7.69 (s, 1H), 8.06 (s, 1H). Mass spectrum
(MALDI-TOF) m/z 792.3 (M+H)+ (theorectcal m/z
791.2).
22. Compound 12. 1H NMR (CDCl3, 300 MHz), d 1.42 (s,
9H), 1.61 (m, 2H), 2.51 (m, 12H), 3.22 (m, 2H), 3.66 (s,
4H), 3.76 (s, 6H), 6.83, 7.21 (dd, 2H, 2H). Mass spectrum
(ESI) m/z 579.2920 (M+H)+ (C30H47N2O5S2 requires
579.2921).
16. Compound 3. 1H NMR (CDCl3, 300 MHz), d 1.30 (s, 3H),
1.34 (s, 3H), 1.55 (s, 9H), 3.62 (m, 1H), 3.70 (m, 2H), 3.83
(m, 2H), 5.48 (s, 2H), 7.23, 7.81 (dd, 2H, 2H), 7.85 (s, 1H),
7.93 (s, 1H). Mass spectrum (ESI) m/z 512.2142 (M+H)+
(C25H29N5O7 requires 512.2140).
23. Compound 13. 1H NMR (CDCl3, 300 MHz), d 1.35 (s,
3H), 1.38 (s, 9H), 1.41 (s, 3H), 1.58 (s, 9H), 1.79 (m, 2H),
2.48 (m, 10H), 3.06 (m, 2H), 3.21 (m, 2H), 3.62 (s, 2H),
3.65 (m, 3H), 3.71 (m, 2H), 3.75 (s, 3H), 3.76 (s, 3H), 3.86
(m, 2H), 4.12 (m, 2H), 5.59 (s, 2H), 6.81 (m, 4H), 7.19 (m,
4H), 7.34, 7.48 (dd, 2H, 2H), 7.66 (s, 1H), 7.80 (s, 1H).
Mass spectrum (ESI) m/z 1072.4879 (M+H)+
(C55H74N7O11S2 requires 1072.4882).
1
17. Compound 7. H NMR (DMSO-d6, 300 MHz), d 1.23 (s,
1H), 1.25 (s, 1H), 1.28 (s, 6H), 2.83 (m, 1H), 3.58 (m, 1H),
3.64 (m, 2H), 3.83 (m, 2H), 5.55 (s, 2H), 7.24, 7.95 (dd,
2H, 2H), 8.07 (s, 1H), 8.22 (s, 1H). Mass spectrum (ESI)
m/z 482.2035 (M+H)+ (C24H28N5O6 requires 482.2034).