Tris(pyrone) chelates of Gd(III) as high solubility MRI-CA

Article abstract of DOI:10.1021/ja057954f

Two tripodal, hexadentate pyrone-based chelators have been prepared. These ligands form stable, soluble complexes with gadolinium(III). The complexes show aqueous stability comparable to that of [Gd(DTPA)]^2- at pH 7.4. Evaluation by relaxometry shows that these complexes have two inner-sphere water molecules and a very fast water exchange rate. The solution behavior of these complexes suggests that they may be attractive as high relaxivity, next-generation magnetic resonance imaging contrast agents. Copyright

Full text of DOI:10.1021/ja057954f

Published on Web 02/01/2006
Tris(pyrone) Chelates of Gd(III) as High Solubility MRI-CA
David T. Puerta, Mauro Botta,* Christoph J. Jocher, Eric J. Werner, Stefano Avedano,
Kenneth N. Raymond, and Seth M. Cohen*
Department of Chemistry and Biochemistry, UniVersity of California, San Diego, La Jolla, California 92093,
Department of Chemistry, UniVersity of California, Berkeley, Berkeley, California 94720, and Department of
EnVironmental and Life Sciences, UniVersity of Eastern Piedmont “A. AVogadro”, Via Bellini 25/G,
15100 Alessandria, Italy
Received December 7, 2005; E-mail: scohen@ucsd.edu
Scheme 1 a
This report describes the synthesis, characterization, and evalu-
ation of a new type of magnetic resonance imaging contrast agent
based on pyrone chelators that demonstrate superb aqueous stability,
relaxivity, and solubility. The success of magnetic resonance
imaging (MRI) as a noninvasive medical imaging technique has
been significantly advanced by the development of contrast agents
(MRI-CA). These paramagnetic metal chelates or superparamagnetic
metal clusters can be used to greatly increase the image quality of
MRI by affecting the relaxation of water protons in the immediate
vicinity of the MRI-CA.1
In 1995, Raymond and co-workers introduced a new class of
MRI-CA inspired by the biological coordination chemistry of
bacterial siderophores. The ligands used in these MRI-CA are
generally comprised of hydroxypyridinones (HOPOs), a class of
monoanionic nitrogen heterocycles, which are linked through a
common backbone to form hexadentate, tripodal chelators. The
archetypical member of this class of compounds, [Gd(TREN-Me-
3,2-HOPO)(H2O)2] (TREN-Me-3,2-HOPO ) tris[(3-hydroxy-1-
methyl-2-oxo-1,2-didehydropyridine-4-carboxamido)ethyl] amine),2
shows good stability and selectivity for Gd(III) and high relaxivity
(10.5 mM-1 s-1 at 20 MHz and 37 °C). The high relaxivity of the
eight-coordinate [Gd(TREN-Me-3,2-HOPO)(H2O)2] and its deriva-
tives is attributed to the presence of two inner-sphere water mole-
cules (q ) 2) and a fast rate of water exchange (kex ) 1/τM > 25
× 106 s-1) as a consequence of the associative nature of the ex-
change mechanism.3,4 The major liability of [Gd(TREN-Me-3,2-
HOPO)(H2O)2] is poor aqueous solubility.5 A variety of strategies
to solubilize this compound have been investigated.4,6,7 Derivatives
have been prepared with improved solubility, although these gener-
ally rely on the introduction of one non-HOPO chelator (resulting
in a charged complex)5 derivatized with a large substituent, such
as poly(ethylene)glycols (PEGs)8 or solubilizing dendrimers.9
Pyrones, such as the food additive maltol (3-hydroxy-2-methyl-
4-pyrone), are a class of oxygen heterocycles with many parallels
to the hydroxypyridinones. They are also monoanionic, hard oxygen
donor ligands that form five-membered chelate rings upon metal
binding. Notably, chelators, such as maltol, have been found to
form metal complexes with good aqueous solubility and biocom-
patibility; for example, [VdO(maltolato)2] has been widely studied
as a soluble therapeutic vanadate source for treating type II
diabetes.10 With this in mind, we turned our efforts to preparing a
pyrone analogue of TREN-Me-3,2-HOPO, which we anticipated
would form Gd3+ complexes with similarly favorable features as
an MRI-CA, but with notably improved aqueous solubility. The
results of these efforts were two new tripodal chelators, which
were used to prepare the MRI-CA, [Gd(TRENMAM)(H2O)2] and
[Gd(TREN-Me-MAM)(H2O)2] (Scheme 1).
a Key: (a) BnBr, NaOH(aq), MeOH, 75 °C, 83%; (b) SeO2, bromoben-
zene, 155 °C; (c) NaClO2, NH2SO3H, H2O/acetone, 70% (combined yield
for steps b and c); (d) NHS, DCC, dry THF; (e) TREN, dry THF, 88%
(combined yield for steps d and e); (f) 1:1 HCl:CH3COOH, 83%; (g)
Gd(NO3)3‚5H2O, pyridine, H2O, 94%.
3-hydroxyl group of maltol is benzyl protected, followed by
oxidation of the 2-methyl group in two steps to give 2-carboxy-
3-benzyloxypyran-4(1H)-one (3).11 Acid 3 is then converted to the
NHS activated ester (4), coupled to tris(2-aminoethyl)amine
(TREN), and deprotected to give the TRENMAM ligand. The facile
and inexpensive synthesis (∼$25/g) of the TRENMAM ligand is
another attractive feature of this new class of MRI-CA, as these
agents generally need to be administered in gram quantities during
clinical use. The related ligand TREN-Me-MAM was prepared by
a similar route starting from 2-carboxy-3-benzyloxy-6-methylpyran-
4(1H)-one (see Supporting Information).12 Combination of either
ligand with Gd(NO3)3‚(H2O)5 gives the desired metal complex in
∼90% isolated yield. Despite numerous attempts, we were unable
to crystallize these Gd3+ complexes; however, we did obtain a
crystal structure of [Fe(TREN-Me-MAM)] (Figure S1), which
illustrated the expected ligand composition, metal coordination, and
internal hydrogen bonding typical of these tripodal complexes.2,5,13
To determine whether TRENMAM and TREN-Me-MAM formed
sufficiently stable Gd3+ complexes for use as MRI-CA, the solution
thermodynamics of these compounds was evaluated. The ligands
and Gd3+ complexes were sufficiently soluble for the protonation
constants to be measured by potentiometry (Table 1).5 The pGd
values ([Gd] ) 10-6 M, [L] ) 10-5 M) were independently
determined by competition titrations with DTPA.14
The ligands TRENMAM and TREN-Me-MAM are generally
more acidic when compared to TREN-Me-3,2-HOPO, with the
average of the first three protonation constants (which are attributed
to the hydroxyl groups) being 6.0, 6.5, and 7.0, respectively. As
expected, the electron-donating methyl substituent on TREN-Me-
MAM makes this ligand more basic when compared with TREN-
MAM. A species distribution plot shows that at physiological pH
(7.4) both ligands are essentially fully deprotonated and thereby
The synthesis of [Gd(TRENMAM)(H2O)2], beginning with
commercially available maltol, is shown in Scheme 1. The
9
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J. AM. CHEM. SOC. 2006, 128, 2222-2223
10.1021/ja057954f CCC: $33.50 © 2006 American Chemical Society

C O M M U N I C A T I O N S
Table 1. Protonation Constants for Pyrone Ligands as Measured
by Potentiometric Titrations
Table 2. Parameters Obtained from the Simultaneous Fitting of
1H NMRD and 17O NMR Data
constant
TRENMAM
TREN-Me-MAM
[Gd(TRENMAM)]
[Gd(TREN-Me-MAM)]
log K1
log K2
log K3
log K4
7.33(1)
5.76(1)
4.97(2)
3.84(2)
7.91(1)
6.30(2)
5.48(2)
4.46(2)
∆2/1019 s-2
298τv/ps
5.6 ( 0.3
19.0 ( 0.8
145 ( 6
1.1 ( 0.3
27.6 ( 0.9
1
10.8 ( 0.2
15.2 ( 1.2
120 ( 9
1.0 ( 0.4
22.4 ( 1.8
1
298τR/ps
298τM/ns
∆HM/kJ mol-1
Ev/kJa
r
r
GdH/Å
3.09 ( 0.2
3.04 ( 0.3
GdO/Åa
2.48
2.48
A/p/106 rad‚s-1
qa
-3.6 ( 0.2
2
4.0
2.27 ( 0.3
22
-3.8 ( 0.1
2
4.0
2.30 ( 0.2
22
a/Åa
D/10-5 cm2 s-1
ED/kJ mol-1a
a Values were fixed in the fitting procedure.
rate of the complexes versus concentration in the range of
0.5-100 mM (at 0.1 MHz and 298 K). From this relaxometric
approach, 0.1 M represents the lower limit of the solubility of the
complexes, a value that is almost an order of magnitude higher
than that for similar tripodal chelates.4
In conclusion, we have described a new class of pyrone-derived
tripodal chelators that form stable and soluble Gd3+ complexes with
high relaxivity for use as MRI-CA. The compounds are among the
most soluble with a q ) 2 value, and one of them can be readily
and economically synthesized from the food additive maltol.
Continuing studies with derivatives of TRENMAM and TREN-
Me-MAM are ongoing and will be reported in due course.
Figure 1. 1/T1 NMRD profiles of [Gd(TRENMAM)(H2O)2] (b) and
[Gd(TREN-Me-MAM)(H2O)2] (O), at 298 K and pH 7.2.
competent to bind metal ions without competition from protons
under these conditions (Figure S2). The pGd values of 19.27 (
0.08 and 19.03 ( 0.04 were determined for TRENMAM and
TREN-Me-MAM, respectively, indicating these pyrone ligands form
Gd3+ complexes with high stability (Figure S3). On the basis of
these experiments, the stability of these complexes rivals those of
the currently used MRI-CA1,4 and therefore should be quite adequate
for in vivo applications.
Acknowledgment. S.M.C. is supported by the University of
California, San Diego, a Hellman Faculty Scholar award, the
American Heart Association (0430009N), and the Research Cor-
poration. D.T.P. is supported by the NIH (GM-72129-01). K.N.R.
is supported by the NIH (HL69832). M.B. acknowledges support
from MIUR.
Having demonstrated that pyrone-based complexes were suf-
ficiently stable and soluble for use as MRI-CA, we sought to
evaluate their physical properties as contrast agents. Several
measurements were performed, including nuclear magnetic reso-
nance dispersion (NMRD) profiles, pH-dependent relaxometry, and
variable temperature 17O NMR relaxometry. The relaxivity, r1p, of
the complexes, as measured at 20 MHz and 298 K, is 9.3 and 8.2
mM-1 s-1 for [Gd(TRENMAM)(H2O)2] and [Gd(TREN-Me-
MAM)(H2O)2], respectively, and these values are constant in the
pH range of 4-9. These values are very similar to those found for
[Gd(TREN-Me-3,2-HOPO)(H2O)2] and related chelates, suggesting
that in the pyrone complexes the Gd3+ ion retains two water
molecules in its inner coordination sphere. This was confirmed by
a detailed NMRD (at 298 and 310 K, Figure S4) and variable
temperature 17O NMR (at 2.12 T, Figure S5) study. Figure 1 shows
the 1/T1 NMRD profiles of the two complexes recorded at 298 K
over the frequency range of 0.01-70 MHz. A simultaneous fitting
of both the NMRD and 17O NMR data provided the parameters
listed in Table 2.
The complex [Gd(TRENMAM)(H2O)2] shows a slightly higher
relaxivity than that of [Gd(TREN-Me-MAM)(H2O)2] over the entire
range of magnetic field strength that is explained by a shorter value
of ∆2 and a slightly longer rotational correlation time τR. On the
other hand, both complexes are endowed with a fast rate of water
exchange (298kex ≈ 8 × 108 s-1), similar to that measured for the
Gd3+ aqua ion.4 The very rapid water exchange kinetics15 may
represent an advantage for MRI-CA applications at high fields
(80-100 MHz), where the optimal τM values for achieving high
relaxivities are close to 1 ns.8 Finally, we measured the relaxation
Supporting Information Available: Details for syntheses, struc-
tures, titration experiments, and complete ref 15. X-ray crystallographic
files in CIF format are available at http://www.ccdc.cam.ac.uk (CCDC
ref 289563). This material is available free of charge via the Internet
at http://pubs.acs.org.
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