click labeling strategy for M(CO)3 (M = Re, 99mTc) prostate cancer targeted Flutamide agents

Article abstract of DOI:10.1039/b921413e

Two distinct click chemistry labeling approaches were investigated with dipyridylamine alkyne derivatives and M(CO)₃ ⁺ (M = Re, ^99mTc). The triazole ring was found uncoordinated and was incorporated into the preparation of a crossover androgen receptor targeting inhibitor for prostate cancer.

Full text of DOI:10.1039/b921413e

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“Click” labeling strategy for M(CO)3 (M = Re, 99mTc) prostate cancer
targeted Flutamide agents†
Adam L. Moore,a Dejan-Kresˇimir Bucˇar,b Leonard R. MacGillivrayb and Paul D. Benny*a
Received 14th October 2009, Accepted 15th December 2009
First published as an Advance Article on the web 6th January 2010
DOI: 10.1039/b921413e
Two distinct “click” chemistry labeling approaches were
investigated with dipyridylamine alkyne derivatives and
dinates to 99mTcI(CO)3.16,17 However, this approach still utilizes
strong radiolabeling conditions and does not take advantage of
the “click” chemistry for coupling of radiometals to biotargeting
agents, particularly temperature sensitive molecules.
Interest in adapting “click” chemistry for prostate cancer diag-
nostic imaging, has led us to investigate functionalized versions
of non-steroidal antagonists (i.e., Flutamide, Bicalutamide) for
targeting the androgen receptor (AR), overexpressed in early stage
prostate cancer.18 Previously, we reported 99mTcI(CO)3 Flutamide
derivatives that showed AR affinity.19 To improve AR activity,
we proposed to utilize “click” chemistry to generate a crossover
M(CO)3 (M = Re, 99mTc). The triazole ring was found
+
uncoordinated and was incorporated into the preparation of
a crossover androgen receptor targeting inhibitor for prostate
cancer.
Used in over 90% of clinical diagnostic imaging scans, technetium-
99m (g = 140 KeV, t1/2 = 6.0 h) has a long history as a Single
Photon Emission Computed Tomography (SPECT) radionuclide.
A number of labeling strategies (ligands and complexes) have
been proposed over the years that have primarily consisted of
mid-valent coordination complexes.1 A unique organometallic
alternative, [99mTcI(OH2)3(CO)3]+ has provided a smaller molecular
volume and weight species to improve biological activity.2-5 Several
[
99mTcI(OH2)3(CO)3]+ labeling strategies (tridentate, 2+1, mono-
dentate) have been identified.6-10 However, thermodynamic li◦mi-
tations of complexation still require high temperatures (~90 C)
and large ligand concentrations (≥10-6 M) for effective yields, even
with the most promising tridentate ligand systems (i.e., cysteine,
dipyridylamine, histidine, iminodiacetic acid).6,11,12
To improve radiolabeling efficiency, new labeling strategies for
[
99mTcI(OH2)3(CO)3]+ utilizing fast and efficient chemical reactions
are being examined to develop an alternative to current labeling
conditions. The catalytic, Huisgen [3+2] cyclolization, or “click”
reaction of an azide and an alkyne, has provided an excellent
platform for coupling two molecules for a variety of applica-
tions (nanoparticles, polymers, biomolecules).13 Sutcliffe et al.
pioneered the use of “click” chemistry in radiopharmaceutical
applications with 18F that has led to an explosion of applications
with this isotope in the literature.14,15 Whereas, investigations of
“click” chemistry with radiometals have been limited; most likely
due to competition of the copper catalyst for complexation.
Recently, “click to chelate” used the “click” reaction to generate
a combinatorial series of ligands, where the triazole ring coor-
aDepartment of Chemistry, Washington State University, PO Box 644630,
Pullman, WA, 99164, USA. E-mail: bennyp@wsu.edu; Fax: 509-335-8867;
Tel: 509-335-3858
bDepartment of Chemistry, University of Iowa, Iowa City, IA, 52242-1294,
USA. E-mail: len-macgillivray@uiowa.edu; Fax: 319-335-1270; Tel: 319-
335-3504
† Electronic supplementary information (ESI) available: Full characteri-
zation. CCDC reference numbers 751744–751746. For ESI and crystallo-
graphic data in CIF or other electronic format see DOI: 10.1039/b921413e
Scheme 1
1926 | Dalton Trans., 2010, 39, 1926–1928
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molecule by incorporating a second aromatic group into the
targeting molecule mimicing Bicalutamide and increasing the
distance between the inhibitor core and the 99mTcI(CO)3 complex,
while developing a facile method to couple the two molecules
under mild conditions.
Two general “click” chemistry strategies were employed to
determine the most effective route of complexation (Scheme 1).
Approach #1 involves the complexation of the ligand with
+
M(CO)3 followed by a second “click” reaction. Approach #2
involves “clicking” the two molecules followed by complexation
+
with M(CO)3 . Several distinct advantages can be predicted with
approach #1 over #2: separate optimization of radiolabeling can
be applied without degradation of the biomolecule, the geometry
about the M(CO)3 core can be limited to a single coordination
Fig. 2 Molecular structure of fac-[Re(CO)3(4)]+, 6. Thermal ellipsoids
are shown at the 30% probability level. Hydrogen atoms are omitted for
clarity.
+
mode, and deactivation of the copper catalyst through ligand
complexation, can be eliminated. In both approaches, dipyridy-
lamine (dpa) was utilized to achieve high labeling efficiency and
In approach #2, the assembled ligand was prepared by “click-
ing” ligand 1 with an azide under standard Sharpless conditions
prior to complexation. The “clicked” benzyl azide ligand, 4,
was reacted with [ReI(CO)3(OH2)3]+. Interestingly, the reaction
of 4 with [ReI(CO)3(OH2)3]+ at 70 ◦C for 2 hours produced a
colorless solid (74%) that corresponded to the dpa coordinated
product fac-[ReI(CO)3(4)]+, 6, prepared in #1. HPLC analysis
of the reaction mixture indicated a single peak (tR 20.2 min.).
Reaction progression followed by 1H NMR showed the presence
of the free ligand and the dpa coordinated complex. Attempts to
generate the triazole coordinated complex or shift the coordination
geometry of the system by extended heating or initial cooling of
the sample did not perturb the ReI(CO)3 dpa coordination in 6.
Approaches #1 & #2 were also investigated with the flutamide
analog, either the azide, 3, or the dpa “clicked” ligand, 5.
In both approaches, the reactions yielded the same product,
fac-[ReI(CO)3(5)]+, 7. 1H NMR and X-ray structure analysis
confirmed ReI(CO)3 dpa coordination and the uncoordinated
triazole ligand (Fig. 3).
+
specificity for M(CO)3 . The alkyne functionalized dpa ligand,
1, was prepared by alkylation of the secondary amine with
propargylbromide.20
In approach #1, the dpa alkyne, 1, was reacted with
[ReI(CO)3(OH2)3]+ in equal concentrations and heated at 70 ◦C
for 2 hours. The product, fac-[Re(CO)3(1)]+, 2, precipitated out
of solution as a colorless solid (37%) and was characterized by
normal analytical methods and HPLC (tR 19.0 min).20 The X-
ray structure confirmed the dpa coordination of Re(CO)3 core
in 2 and had similar bond angles and distances to other dpa
complexes (Fig. 1). The “click” reaction of 2 was carried out with
standard Sharpless conditions21 first using the model, benzyl azide
to probe the conditions and complex speciation. The reaction of
2 with benzylazide proceeded smoothly and in a reasonable time
frame (90 min. r.t.) to produce fac-[Re(CO)3(4)]+, 6, in good yield
(84%). As expected in approach #1, the triazole ligand was found
uncoordinated to the Re(CO)3 core in both the X-ray structure
1
and the H NMR (Fig. 2). No rearrangement of the Re(CO)3
coordination environment from the dpa to triazole was observed
during the course of the reaction. Several biologically relevant
temperatures (25, 37, 50, 70 ◦C) and time points (15, 45, 90 min)
were also examined to probe efficiency of the triazole formation
in 6. At 70 ◦C, triazole formation was completed in 15 min, while
at room temperature it required 90 min to reach completion.
Fig. 1 Molecular structure of fac-[Re(CO)3(1)]+, 2. Thermal ellipsoids
are shown at the 30% probability level. Hydrogen atoms are omitted for
clarity.
Fig. 3 Molecular structure of fac-[Re(CO)3(5)]+, 7. Thermal ellipsoids
are shown at the 30% probability level. Hydrogen atoms are omitted for
clarity.
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Table 1 “Clicked” triazole formation yields of 99mTc-4, (6A), from 99mTc-
1, (2A), with benzyl azide (10-4 to 10-6 M) and temperatures (◦C) in a
15 min reaction time
and the Office of Science (BER), U.S. Department of Energy
(DE-FG02-08-ER64672).
10-4
10-5
10-6 (M)
Notes and references
70 ◦
50 ◦
37 ◦
25 ◦
C
C
C
C
100
100
100
100
100
100
100
93
75
61
23
23
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examined at 10-4–10-5 M benzylazide in 15 min. However, at 10-6
M
benzylazide, incomplete reaction yields were observed at 70 ◦C and
yields declined as the reaction temperature decreased. The reaction
of 2A with the Flutamide azide yielded similar radiolabeling yields.
In conclusion, we have successfully compared two approaches
using “click” chemistry with a dpa alkyne ligand, azides, and
[M(CO)3]+. In both cases, the dpa ligand exclusively favored
coordination to the metal over the triazole demonstrating the
power of click chemistry to provide a specific coordination mode
and permit the incorporation of the triazole into the structural
design of targeting molecules. Furthermore, the “chelate then
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21 Standard Sharpless Conditions: A 25 cm3 scintillation vial was charged
with [ReI(CO)3(1)]+, 2, (0.050 g, 0.076 mmol), benzylazide (0.010 g,
0.076 mmol) and dissolved in tert-butyl alcohol (4 cm3). To the mixture
was added sodium ascorbate (0.003 g, 0.0015 mmol) in water (2 cm3)
followed by CuII(OAc)2 (0.002 g, 0.0076 mmol) in water (2 cm3). This
mixture was stirred at room temperature for 90 minutes.
Acknowledgements
This work was funded in part by the Department of Defense
Prostate Cancer New Investigator Award (W81XWH0510556)
1928 | Dalton Trans., 2010, 39, 1926–1928
This journal is
The Royal Society of Chemistry 2010
©