Gd complexes of macrocyclic diethylenetriaminepentaacetic acid (DTPA) biphenyl-2,2′-bisamides as strong blood-pool magnetic resonance imaging contrast agents

Article abstract of DOI:10.1021/jm2002052

We report the synthesis of macrocyclic DTPA conjugates of 2,2′-diaminobiphenyl and their Gd complexes of the type [Gd(L)(H ₂O)]?xH₂O (2a,b; L = 1a,b) for use as new MRI blood-pool contrast agents (MRI BPCAs). Pharmacokinetic inertnes

Full text of DOI:10.1021/jm2002052

ARTICLE
pubs.acs.org/jmc
Gd Complexes of Macrocyclic Diethylenetriaminepentaacetic Acid
(DTPA) Biphenyl-2,2 -bisamides as Strong Blood-Pool Magnetic
0
Resonance Imaging Contrast Agents
†
‡
§
||
^
^
Ki-Hye Jung, Hee-Kyung Kim, Gang Ho Lee, Duk-Sik Kang, Ji-Ae Park, Kyeong Min Kim,
,‡,||
,†
Yongmin Chang,* and Tae-Jeong Kim*
†
‡
§
Department of Applied Chemistry, Department of Medical and Biological Engineering, and Department of Chemistry,
Kyungpook National University, University Road 80, Daegu, 702-701, Republic of Korea
Department of Diagnostic Radiology and Molecular Medicine, Kyungpook National University, Dongin-dong 2-ga, Daegu,
00-422, Republic of Korea
7
^
Laboratory of Nuclear Medicine Research, Molecular Imaging Research Center, Korea Institute of Radiological Medical Science,
Nowon-gil 75, Seoul, 139-706, Republic of Korea
S Supporting Information
b
0
ABSTRACT: We report the synthesis of macrocyclic DTPA conjugates of 2,2 -diaminobi-
phenyl and their Gd complexes of the type [Gd(L)(H O)] xH O (2a,b; L = 1a,b) for use as
2
3
2
new MRI blood-pool contrast agents (MRI BPCAs). Pharmacokinetic inertness of 2
compares well with those of analogous Gd-DTPA MRI CAs currently in use. The present
ꢀ
1
system also shows very high stability in human serum. The R relaxivity reaches 10.9 mM
1
ꢀ
1
s
, which is approximately 3 times as high as that of structurally related Gd-DOTA (R =
1
ꢀ
1 ꢀ1
ꢀ1 ꢀ1
3
.7 mM
s
). The R relaxivity in HSA goes up to 37.2 mM
s , which is almost twice as
1
high as that of MS-325, a leading BPCA, demonstrating a strong blood pool effect. The
in vivo MR images of mice obtained with 2b are coherent, showing strong signal
enhancement in heart, abdominal aorta, and small vessels. Even the brain tumor is vividly
enhanced for an extended period of time. The structural uniqueness of 2 is that it is neutral in
charge and thus makes no resort to electrostatic interaction, supposedly one of the essential
factors for the blood-pool effect.
’
INTRODUCTION
strong and prolonged vascular enhancement. The BPCAs thus
far developed may be divided into three classes according to their
mechanism of action: (i) the noncovalent binding of low-molecular
Gd chelates (GdL) to HSA to prevent immediate leakage into the
Magnetic resonance imaging (MRI) is a powerful, noninvasive
diagnostic technique of the human anatomy, physiology, and
pathophysiology on the basis of superior spatial resolution and
contrast useful in providing anatomical and functional images of
6
ꢀ8
interstitial space;
liposomes based on an increase in the size of CAs, which slows
(ii) systems incorporating polymers or
1
the human body. At present, a large number of MRI techniques
9
ꢀ12
leakage through endothelial pores;
(iii) systems based on
are performed employing Gd(III) complexes to enhance the
image contrast by increasing the water proton relaxation rate in
1
3ꢀ15
nanoparticles, involving a change in the route of elimination.
2
The most successful approach so far is the class i represented by
MS-325, a GdꢀDTPA derivative containing a cholic acid moiety.
These complexes bind strongly and reversibly to HSA, leading to
not only high relaxivity at clinical field strengths but also much
longer residence time in the blood compared to ECF agents. The
lipophilic component represented by aromatic moieties in the
chelate is responsible for the generation of noncovalent interac-
the body. Some representative advantages of employing the
Gd(III) ion come from their unique properties such as (i) seven
unpaired electrons, (ii) nine coordination sites, and (iii) suitable
electronic relaxation time. Despite their wide and successful
applications in clinics, however, conventional Gd(III)-based low-
molecular-weight CAs are mostly extracellular fluid (ECF) agents
exhibiting rapid extravasation from the vascular space. As a result,
the time window for imaging is considerably reduced, thus
1
6,17
tions between the corresponding Gd chelate and HSA.
The
3
ꢀ5
anionic phosphodiester moiety in MS-325 is also known to
contribute in a synergistic manner to the noncovalent binding
interaction with HSA. Yet, since anionic Gd complexes lead to
limiting acquisition of high-resolution images.
In order to overcome such limitations inherent to ECF CAs,
the necessity for the development of BPCAs has risen in the
expectation that they reside in the blood vessel for an extended
period of time and thus are eliminated much more slowly from
the circulation than their ECF counterparts. The BPCAs provide
Received: February 23, 2011
Published: June 27, 2011
r 2011 American Chemical Society
5385
dx.doi.org/10.1021/jm2002052
|J. Med. Chem. 2011, 54, 5385–5394

Journal of Medicinal Chemistry
ARTICLE
Scheme 1. Synthesis of 1and 2
high osmolality under physiological conditions giving rise to
some adverse effects, efforts have focused on the preparation of
neutral analogues based on the Gd macrocycle with the con-
comitant aim to achieve thermodynamic stability as well as
crude product thus obtained was further purified by chromatography
(SiO
2
, hexane/ethyl acetate 75:25) to give a yellow crystalline solid after
1
usual workups. Yield: 3.2 g (77%). H NMR (CDCl
): δ = 8.90 (s, 2H,
3
0
0
0
3,3 -H), 8.35 (d, 2H, 5,5 -H), 7.40ꢀ7.38 (d, 2H, 6,6 -H), 4.02 (s, 6H,
2
,9ꢀ12
13
ꢀ
COOCH
3
). C NMR (CDCl
3
): δ = 164.5 (ꢀCOOCH
3
), 146.8
kinetic inertness.
In line with these efforts and as part of our continued search
0
0
0
0
0
(
2,2 -C), 137.7 (1,1 -C), 134.3 (5,5 -C), 131.8 (4,4 -C), 131.0 (6,6 -C),
18
0
1
26.0 (3,3 -C), 53.0 (ꢀCOOCH ). Anal. Calcd for C H N O : C,
for Gd-based MRI BPCAs, we have designed and synthesized
3
16 12 2 8
0
53.34; H, 3.36; N, 7.78. Found: C, 53.47; H, 3.34; N, 7.77.
macrocyclic DTPA-(biphenyl-2,2 -bisamide) (1a,b, Scheme 1)
0
0
Dimethyl 2,2 -Diaminobiphenyl-4,4 -dicarboxylate. To a
for use as a ligand leading to the formation of MRI BPCA
belonging to class i. Our justification for their synthesis may
come from the expectation that 1 would form a neutral, macro-
cyclic Gd(III) complex [Gd(1)(H O)] xH O (Scheme 1) car-
0
0
solution of dimethyl 2,2 -dinitrobiphenyl-4,4 -dicarboxylate (3.0 g, 8.3
mmol) in acetic acid (60 mL) in an autoclave was added Pd/C (10%, 1.5
g). The reaction vessel was flushed several times and pressurized with
dihydrogen to 700 kPa. The vessel was sealed and the mixture stirred at
2
3
2
rying a highly liphophilic biphenyl on the chelate backbone.
3
0 ꢀC overnight, after which the pressure was released and the catalyst
separated by filtration with acetic acid (3 ꢁ 5 mL). The solvent was
removed under reduced pressure and the residue purified by chroma-
’
EXPERIMENTAL SECTION
tography (SiO
solid. Yield: 2.3 g (91%). H NMR (CDCl
2
, hexane/ethyl acetate 6:4) to give a yellow crystalline
1
0
General Remarks. All reactions were carried out under an atmo-
3
): δ = 7.51ꢀ7.50 (d, 2H, 5,5 -
0
0
sphere of dinitrogen using the standard Schlenk techniques. Solvents
were purified and dried using standard procedures. DTPA was obtained
from TCI and used without further purification. The N,N-bis(anhydride) of
H), 7.47 (d, 2H, 6,6 -H), 7.19ꢀ7.17 (s, 2H, 3,3 -H), 3.92 (s, 6H,
1
3
ꢀCOOCH
3
), 3.83 (br, 4H, NH
2
ꢀ). C NMR (CDCl
3
): δ = 167.0
0
0
0
(ꢀCOOCH
3
), 146.0 (2,2 -C), 131.2 (4,4 -C), 130.0 (6,6 -C), 128.0
19
0
0
0
DTPA was prepared according to the literature methods. All other
commercial reagents were purchased from Aldrich and used as received
unless otherwise stated. Deionized water was used for all experiments.
(1,1 -C), 117.6 (5,5 -C), 116.2 (3,3 -C), 52.3 (ꢀCOOCH
3
). Anal. Calcd
for C16
H
16
N
2
O
4
: C, 63.99; H, 5.37; N, 9.33. Found: C, 63.85; H, 5.42;
N, 9.24.
1
0
The H NMR experiment was performed on a Bruker Advance 400
1a. To a stirred solution of 2,2 -diaminobiphenyl (0.8 g, 4.4 mmol) in
DMF (10 mL) was added DTPA-bis(anhydride) (1.6 g, 4.4 mmol) in
DMF (5 mL) in one portion. The mixture was stirred at 70 ꢀC for 16 h,
after which the solvent was removed under reduced pressure. The solid
residue was taken up in methanol (5 mL) and filtered through a short
silica gel column to remove any solid impurities and ether added to
precipitate out a white solid. This was isolated by filtration and dried
under vacuum to give the desired product as a white powder. Yield: 1.9 g
spectrometer by Center for Instrumental Analysis, Kyungpook National
University (KNU). Chemical shifts are given as δ values with reference
to tetramethylsilane (TMS) as the internal standard. Coupling constants
are in Hz. Elemental analyses were performed by Center for Instru-
mental Analysis, KNU, and Catholic University in Daegu. FAB mass
spectra were obtained by using a JMS-700 model (Jeol, Japan) mass
spectrophotometer by Korea Basic Science Institute (KBSI). The purity
of all products was determined to be above 95% by elemental analysis.
1
0
(81%). H NMR (CDCl
3
): δ = 7.52ꢀ7.50 (d, 2H, 3,3 -H), 7.46ꢀ7.45
0
0
0
0
0
Synthesis. 2,2 -Diaminobiphenyl. To a solution of 2,2 -dini-
trobiphenyl (5.0 g, 20.5 mmol) in ethyl acetate (90 mL) in an autoclave
was added Pd/C (10%, 2.0 g). The reaction vessel was flushed several
times and pressurized with dihydrogen to 700 kPa. The vessel was sealed
and the mixture stirred at room temperature for 18 h, after which the
pressure was released and the catalyst separated by filtration with ethyl
acetate (3 ꢁ 10 mL). The solvent was removed under reduced pressure
(d, 2H, 6,6 -H), 7.37ꢀ7.35 (t, 2H, 4,4 -H), 7.22 (t, 2H, 5,5 -H),
3.43ꢀ3.38 (m, 10H, ꢀNHCOCH
(m, 8H, ꢀNCH CH N). Anal. Calcd for C26
55.87; H, 5.94; N, 12.52. Found: C, 55.27; H, 5.96; N, 12.53. FAB-
2
and ꢀNCH
2
COO), 3.28ꢀ3.09
2
2
H
31
N
5
O
8
1H O: C,
2
3
+
MS (m/z): calcd for C26H N O , 542.22 ([MH] ). Found: 542.41.
32 5 8
1b. The title compound was prepared as described above for the
0
preparation of 1a by replacing 2,2 -diaminobiphenyl with dimethyl 2,
0
0
and the residue recrystallized from hexane/chloroform to give pale violet
crystals. Yield: 3.5 g (94%). H NMR (CDCl
2 -diaminobiphenyl-4,4 -dicarboxylate (0.8 g, 2.7 mmol) in DMF (10 mL).
The product was obtained as an off-white powder. Yield: 1.3 g (76%).
1
3
): δ = 7.19ꢀ7.15 (t, 2H,
13
0
0
0
1
0
4
6
,4 -H), 7.12ꢀ7.10 (d, 2H, 6,6 -H), 6.84ꢀ6.80 (t, 2H, 5,5 -H),
H NMR (DMSO): δ = 9.83 (br, 3H, ꢀCOOH), 8.14 (s, 2H, 3,3 -H),
0
0
0
.78ꢀ6.75 (d, 2H, 3,3 -H), 3.68 (s, 4H, ꢀNH ). C NMR (CDCl ):
7.89ꢀ7.86 (d, 2H, 5,5 -H), 7.41ꢀ7.39 (d, 2H, 6,6 -H), 3.87 (s, 6H,
2
3
0
0
0
0
δ = 144.5 (2,2 -C), 131.5 (4,4 -C), 129.2(6,6 -C), 125.1 (5,5 -C),
ꢀCOOCH
2.91ꢀ2.87 (m, 8H, ꢀNCH
C, 54.79; H, 5.36; N, 10.65. Found: C, 54.06; H, 5.43; N, 10.64. FAB-
3
), 3.24ꢀ3.07 (m, 10H, ꢀNHCOCH
2
and ꢀNCH
2
COO),
H N O :
35 5 12
0
0
1
19.3(1,1 -C), 116.1(3,3 -C). Anal. Calcd for C12
H
12
N
2
: C, 78.23; H,
2
CH N). Anal. Calcd for C30
2
6
.57; N, 15.21. Found: C, 77.92; H, 6.53; N, 15.12.
Dimethyl 2,2 -Dinitrobiphenyl-4,4 -dicarboxylate. The title
0
0
+
MS (m/z): calcd for C30
([MH] ).
H
36
N
5
O
12, 658.24 ([MH] ). Found: 658.23
2
0
+
compound was prepared according to the literature methods. The
5
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Journal of Medicinal Chemistry
ARTICLE
Typical Procedure for the Preparation of 2. To a solution of 1
1.5 mmol) in pyridine (7 mL) was added an equivalent amount of
were sealed in glass spheres, fitting into 10 nm NMR tubes in order to
eliminate susceptibility correction to the chemical shift. The hydration
numbers in 3 were fitted using eq 5
(
2
2
Gd(OAc) (1.5 mmol). The suspension was stirred overnight at 70 ꢀC
3
during which time the solution became homogeneous. The solvent was
removed and the residue dissolved in a minimum amount of ethanol to
be passed through Celite to remove any solid impurities. Ether was
added to the filtrate to precipitate an off-white powder which was dried
Δδ ¼ qΔ½DyðligandÞ ðH2OÞ ꢂ=½H2Oꢂ
ð5Þ
n
q
The slope of a plot of the Dy(III)-induced shift versus the Dy(III)
concentration is proportional to the hydration number in the complex.
The hydration number is obtained by linear fitting of the dysprosium-
induced shift data using OriginPro 8.
in vacuo. 2a. Yield: 0.9 g (91%). Anal. Calcd for C26
C, 40.67; H, 4.73; N, 9.12. Found: C, 40.79; H, 4.95; N, 9.29. FAB-MS
5 8 2
H28GdN O 4H O:
3
+
(
2
4
m/z): calcd for C26
H
28GdN
b. Yield: 1.1 g (88%). Anal. Calcd for C30
0.76; H, 4.56; N, 7.92. Found: C, 40.01; H, 4.37; N, 7.97. FAB-MS
5
NaO
8
, 719.11 ([MNa] ). Found; 719.54.
1
Relaxivity. T measurements were carried out using an inversion
H
32GdN O: C,
5 2
O12 4H
3
recovery method with a variable inversion time (TI) at 1.5 T (64 MHz,
GE Healthcare, Milwaukee, WI, U.S.). The magnetic resonance (MR)
images were acquired at 35 different TI values ranging from 50 to
+
(
(
m/z): calcd for C H GdN O , 812.13 ([MH] ). Found: 812.12
30 33 5 12
+
[MH] ).
Dy(1)(H O)] (3). The title compound was prepared as described
1
1
750 ms. T relaxation times were obtained from the nonlinear least-
[
2
squares fit of the signal intensity measured at each TI value. For T2
measurements the CPMG (CarrꢀPurcellꢀMeiboonꢀGill) pulse se-
quence was adapted for multiple spinꢀecho measurements. Thirty-four
images were acquired with 34 different echo time (TE) values ranging
above for the preparation of 2 by replacing Gd(OAc) with DyCl . The
3
3
product was obtained as an off-white powder. 3a. Yield: 1.4 g (80%).
+
FAB-MS (m/z): calcd for C26
H
28DyN
5 8 2
NaO H O, 721.24 ([MH] ).
3
+
Found; 721.16 ([MH] ). 3b. Yield: 1.7 g (89%). FAB-MS (m/z): calcd
from 10 to 1900 ms. T relaxation times were obtained from the
+
+
2
5 12 2
for C30H33DyN O H O, 837.15 ([MH] ). Found: 837.28 ([MH] ).
3
nonlinear least-squares fit of the mean pixel values for the multiple
Transmetalation Kinetics. This experiment was performed ac-
spinꢀecho measurements at each echo time. Relaxivities (R
were then calculated as an inverse of the relaxation time per mM. The
determined relaxation times (T and T ) and relaxivities (R and R ) are
1 2
and R )
cording to the literature method, where the water proton relaxation rates
P
(
R
1
) of buffered solutions (phosphate buffer, pH 7.4) containing
2
1
1
2
1
2
2
equimolar amounts of 2 (2.5 mM) and ZnCl (2.5 mM) are measured.
finally image-processed to give the relaxation time map and relaxivity
map, respectively.
Brain Tumor Model. The C6 glioma cells were grown in a
monolayer with Dulbecco’s modified Eagle medium (DMEM), supple-
mented with fetal calf serum (10%), L-glutamine (2%), penicillin/
Namely, to a buffered solution of 2 (1 mL) was added a buffered solution
of ZnCl (10 μL). The mixture was vigorously stirred and 300 μL of the
2
solution taken up for the relaxometric study. Control studies have also
been carried out with Gd-DTPA-BMA, Gd-DOTA, Gd-EOB-DTPA,
P
and Gd-BOPTA for comparative purposes. The R1 relaxation rate was
streptomycin (0.2%) and incubated at 37 ꢀC in a mixture of air/CO
2
obtained after subtraction of the diamagnetic contribution of the proton
(
95%/5%). Cells were suspended in an agar solution (1%) prepared with
water relaxation from the observed relaxation rate R
1
= (1/T
1
). The
6
ꢀ1
supplemented DMEM (10 cell mL ) shortly before injection and
injected with 10 μL of the mixture. Agarose was used to prevent the cells
from spreading out of the injection site. The rats (40ꢀ60 g of body
weight) were anesthetized (1.5% isoflurane in oxygen) and placed on a
stereotactic head holder. A middle scalp incision was made. The cell
suspension (10 μL) was slowly injected for 10 min using a Hamilton
syringe (1.0 mm diameter). The syringe was slowly removed 2ꢀ5 min
after cell injection, and the scalp was sutured. The MR experiments were
measurements were performed on a 3 T whole body system (Magnetom
Tim Trio, Siemens, Germany) at room temperature.
Determination of Kinetic Constants. These constants were
fitted according to eqs 1ꢀ4.
kf
GdL þ Zn þ
2
s
Gd þ ½ZnLꢂ1ꢀ
3þ
ð1Þ
f
s
r
kr
2
3
where k and k are the rate constants for the forward and reverse
carried out 6 or 7 weeks after tumor cell implantation.
f
r
transmetalation reactions, respectively. The rate of transmetalation can
Serum Stability Assay. The serum stability assay was performed
in two ways. The first method involves traditional HPLC monitoring of
the signal intensity of the Gd complex in human serum from the UV
be written as in eq 2. Here [ZnL] = [GdL]
the initial concentration of the gadolinium complex.
0
ꢀ [GdL], with [GdL]
0
being
2
4,25
absorbance detector at 240 nm for the present studies.
The HPLC
d½GdLꢂ
0
0
r
system (high pressure liquid chromatography, KNAUER, Berlin,
¼
ꢀ k ½GdLꢂ þ k ½ZnLꢂ
ð2Þ
f
dt
Germany) equipped with a Develosil ODS-HG-5 column (6.0 mm ꢁ
250 mm; Nomura Chemical Co., Ltd., Aichi, Japan) was used for the
The concentration of GdL at time t can be written as in eq 3.
measurement. Elution conditions were as follows: A mixture of an
aqueous solution of TFA (0.1%, v/v) and an acetonitrile solution of TFA
k1t
½
GdLꢂ ¼ ½GdLꢂ∞ þ ð½GdLꢂ ꢀ ½GdLꢂ Þe
ð3Þ
0
∞
(0.1%, v/v) with a 30 min linear gradient (from 5% to 80%) at a flow rate
0
0
k
k
1
f
= k
f
+ k
r
, and [GdL]
∞
is the equilibrium concentration of GdL. Thus,
of 1.7 mL/min was used. Samples were prepared as follows: 2b (0.2 mg)
was dissolved initially in DMSO (20 μL) to be diluted with water (80
μL). The solution was divided into two vials: the first solution (60 μL)
was added to human serum (940 μL) in a 2 mL vial for the purpose of
incubation at 37 ꢀC, and the second one (30 μL) was diluted with water
(470 μL) in a 1 mL vial as a standard. Aliquots (60 μL each) of the first
solution were analyzed without dilution at different time points. In
addition, a standard serum solution was prepared by mixing DMSO (12
μL), water (48 μL), and human serum (940 μL). The second method
can be calculated as
½
GdLꢂ ꢀ ½GdLꢂ
0
0
∞
k ¼ k1
ð4Þ
f
½
GdLꢂ0
0
The k
f
is obtainable by nonlinear curve fitting of transmetalation data
using MATLAB R2008a in eq 4.
1
7
Dy(III)-Induced Water O Shifts. 3 was dissolved in water
17
2
containing 20% D O and 5% O-enriched water (Isotech, U.S.). The
3
+
pH of the solution was adjusted to the physiological value (pH 7.0). The
dysprosium-induced shift measurements were performed at a Dy(III)/
ligand molar ratio corresponding to the desired complex stoichiometry
was developed by us and involves the determination of Gd concentra-
p
p
tion by measuring the ratio of relaxivities, R (t)/R (0), as a function of
1
1
1
8b
incubation time for the serum solution of 2. This method, although
qualitative, is a handy and simple replacement of the traditional HPLC
method. The typical procedure is as follows: To a solution of aqueous
human serum (Innovative Research) (180 μL) was added an aqueous
ꢀ
3
by varying the concentration of 3 over the range 20ꢀ80 mmol dm
.
1
7
O NMR experiments were run at 500 MHz using a Varian Unity
INOVA instrument (11.8T, 67.814 MHz) at room temperature. Samples
5
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dx.doi.org/10.1021/jm2002052 |J. Med. Chem. 2011, 54, 5385–5394

Journal of Medicinal Chemistry
ARTICLE
solution of 2 (20 μL, 5 mM) at 4 ꢀC. The mixture was incubated at 37 ꢀC
at different intervals (0 min, 30 min, 1 h, 2 h, 4 h, 48 h, and 96 h), after
which time aqueous TCA (20%, 100 μL) was added to precipitate free
human serum. The supernatant solution, after removal of the white
precipitates of human serum by centrifugation at 4 ꢀC for 10 min, was
taken for relaxivity measurement at a 3 T whole body system at room
temperature. In this way the serum stability assay can be qualitatively
accomplished, since any changes in R relaxivities (an increase, in fact) of
1
the solutions prepared before and after incubation would indicate the
release of free Gd(III) ion from 2.
Table 1. Kinetic Constants of 2, Gd Ferromide, Gd-DOTA,
Gd-EOB-DTPA, Gd-BOPTA, and Gd-DTPA-BMA
ꢀ1
0
ꢀ1
complex
K
1
(min
)
[Lig Gd]
∞
(mol/L)
K (min )
f
ꢀ3
ꢀ3
ꢀ3
ꢀ4
ꢀ3
ꢀ4
ꢀ4
ꢀ4
ꢀ4
ꢀ4
ꢀ4
ꢀ4
2
a
1.11 ꢁ 10
1.04 ꢁ 10
1.19 ꢁ 10
1.81 ꢁ 10
1.39 ꢁ 10
1.65 ꢁ 10
1.41 ꢁ 10
1.19 ꢁ 10
1.26 ꢁ 10
1.35 ꢁ 10
1.53 ꢁ 10
1.37 ꢁ 10
1.24 ꢁ 10
1.09 ꢁ 10
5.79 ꢁ 10
5.15 ꢁ 10
5.46 ꢁ 10
0.69 ꢁ 10
6.25 ꢁ 10
8.32 ꢁ 10
7.96 ꢁ 10
ꢀ3
ꢀ3
ꢀ3
ꢀ3
ꢀ3
ꢀ3
2
b
a
Gd ferromide
a
Gd-DOTA
ꢀ3
Gd-EOB-DPTA
a
ꢀ3
ꢀ3
In Vitro Cell Toxicity. The CT-26 mouse colon carcinoma cell line
was purchased from the American Type Culture Collection (ATCC,
Rockville, MD). Cells were maintained in DMEM (Welgene), supple-
mented with 10% FBS and 1% antibiotic (all purchased from Gibco).
The medium was replaced every 2 days, and CT-26 cells were placed into
Gd-BOPTA
a
Gd-DTPA-BMA
a
Data obtained from ref 18b.
3
a 96-well plate (2 ꢁ 10 cells/well per 50 μL). Magnevist Gd complexes
were added to each well (1 nM to 1 mM per 50 μL). After incubation for
5
5
days at 37 ꢀC in a 5% CO
2
atmosphere, 3-(4,5-dimethylthiazol-2-yl)-
-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner
salt (MTS, Promega, Madison, WI) was added to each well (20 μL/
well). After an additional 1 h of incubation, proliferation of the treated
cells was quantified by measuring the absorbance of the culture medium
at 492 nm with a 96-well plate reader (GENios, TECAN Co., Boston,
MA). The cytotoxicities were compared. IMR-90 human embryonic
lung fibroblasts were also employed. Cells were maintained in DMEM
(
Gibco) supplemented with heat-inactivated FCS (10%), penicillin (100
IU/mL), streptomycin (100 mg/mL), and gentamicin (200 mg/mL)
all purchased from Gibco). The medium was replaced every 2 days, and
(
4
cells placed into a 96-well plate (1 ꢁ 10 cells/well per 200 μL). Various
Gd(III) concentrations (IMR-90: 1 nM to 0.1 mM) of the contrast agent
were added into the culture serum free medium and incubated for 24 h.
CCK-8 (10 μL) was then added to each well to evaluate cell viability.
The solution was removed after 4 h at 37 ꢀC. The OD (optical density)
was read at 450 nm using a microplate reader (Molecular Device, USA
Bio-Rad 550 reader) to determine the cell viability/toxicity.
In Vivo MR Imaging. All animal experiments were performed in
accordance with the rules of the animal research committee of Kyungpook
National University. Six-week-old male ICR (Institute of Cancer Research)
mice with weights of 29ꢀ31 g were used for the MRI. Eight-week-old male
SD (SpragueꢀDawley) rats with weights of 100ꢀ120 g were used for the
brain tumor MR imaging. Ten-week-old male SD rats with weights of
p
p
Figure 1. Evolution of R
MRI CAs.
1
(t)/R
1
(0) as a function of time for various
slice thickness; 192 ꢁ 65 matrix size; number of acquisitions (NEX) = 1.
Each image was taken at an interval of 11 s.
Image Analysis. The anatomical locations with enhanced contrast
were identified with respect to heart, kidney, and liver on postcontrast
MR images. For quantitative measurement, signal intensities in specific
regions of interest (ROI) were measured using Advantage Window
software (GE Medical, U.S.). The CNR (contrast to noise ratio) was
calculated using eq 6, where SNR is the signal-to-noise ratio.
350ꢀ400 g were used for the CE-MRA. The mice (n = 4) and rats (n = 8)
were anesthetized with 1.5% isoflurane in oxygen. Measurements were made
before and after injection of 2b via tail vein. The amount of CA per each
injection is 0.1 mmol [Gd]/kg for MR and CE-MRA images. After each
measurement the mouse was revived from anesthesia and placed in the cage
with free access to food and water. During these measurements, the animals
were maintained at approximately 37 ꢀC using a warm water blanket. Whole
body MR images were taken with a 1.5 T MR unit (GE Healthcare,
Milwaukee, WI, U.S.) equipped with a homemade small animal rf coil. The
coil was of the receiver type with its inner diameter being 50 mm. The imaging
parameters for 3D fast SPGR (spoiled GRASS images) are as follows: repeti-
tion time (TR) = 9.2 ms; echo time (TE) = 2.1 ms; 12 mm field of view
CNR ¼ SNRpost ꢀ SNRpre
ð6Þ
’
RESULTS AND DISCUSSION
Synthesis and Characterization. Scheme 1 shows the pre-
paration of 1 and 2. The scheme initially requires the preparation
0
of 2,2 -diaminobiphenyls by catalytic hydrogenation of the
0
corresponding 2,2 -dinitrobiphenyls. Subsequent condensation
with DTPA-bis(anhydride) resulted in the formation of 1 as
macrocyclic DTPA-bis(amide) chelates anchored by biphenyl. It
is noted here that the condensation proceeded to form a macrocyclic
chelate rather than linear, oligomeric analogues. The macrocy-
(
FOV); 256 ꢁ 192 matrix size; 0.8 mm slice thickness; number of
acquisitions (NEX) = 8. Images were obtained for 110 min after injection.
Brain tumor MR images were taken with a 3 T MR unit (GE Healthcare,
Milwaukee, WI, U.S.) equipped with a homemade small animal rf coil. The
imaging parameters for SE (spin echo) are as follows: repetition time (TR)
0
clization such as that observed here has a precedent with 1,1 -
ferrocenediyldiamines which on reaction with DTPA-bis(anhydride)
led to the formation of analogous macrocyclic DTPA-bis-
18b
(
amides). These ligands form Gd(III) complexes of the type
[Gd(L)(H O)] xH O (2a,b; L = 1a,b) by simple complexation
with an equimolar amount of Gd(OAc) in pyridine. All com-
=
550 ms; echo time (TE) = 11 ms; 4 mm field of view (FOV); 128 ꢁ 128
matrix size; 0.8 mm slice thickness; number of acquisitions (NEX) = 4. CE-
MRA was carried out with a 3.0 T MR unit (Magnetom Tim Trio, Siemens
Medical Solution, Erlangen, Germany) equipped with 8HR wrist coil. The
imaging parameters for CE-MRA vibe are as follows: repetition time (TR) =
2
2
3
3
plexes were isolated as off-white, hygroscopic solids by repeated
precipitation with cold diethyl ether from the reaction mixture.
The formation of 1 and 2 was confirmed by microanalysis and
3.3 ms; echo time (TE) = 1.3 ms; 17 mm field of view (FOV); 0.9 mm
5
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Journal of Medicinal Chemistry
ARTICLE
1
13
spectroscopic techniques such as H and C NMR, HRFAB
endogenous ions. In such a process, the paramagnetic Gd(III)
+
mass spectrometry. For instance, 2a and 2b exhibit [MNa] and
MH] signals at m/z of 719.54 and 812.12, respectively.
ion is expelled from the complex. Endogenous ions likely to
+
26ꢀ28
[
compete with Gd(III) are Cu(II), Ca(II), and Zn(II).
The
Although ligands are insoluble in water, their Gd complexes
exhibit fairly good water solubility (up to 0.25 M).
Cu(II) ion is present in very small concentrations in the blood
(1ꢀ10 mol/L), whereas Ca(II) has relatively low affinity for
acyclic ligands such as DTPA and DTPA-BMA (roughly 10
orders of magnitude lower than that of Gd(III)). Although the
affinity for Ca(II) by cyclic ligand such as DOTA may be high
enough, transmetalation by this ion can be ignored in the presence
of other metal ions. For instance, only Zn(II) can displace a
significant amount of Gd(III) because the concentration of the
Transmetalation Kinetics. Protonation constants and ther-
modynamic stability constants could not be obtained because of
water insolubility of ligands. Yet transmetalation kinetic data
would be sufficient enough to provide relevant information about
the stability of Gd complexes under physiological conditions.
The Gd chelates, although they are thermodynamically stable,
may be kinetically labile enough to undergo transmetalation by
29
former in blood is relatively high (55ꢀ125 mol/L). The stability
of Gd complexes in the presence of Zn(II) is thus an important issue
because transmetalation will induce a release of Gd(III) into the
body and possible depletion of the endogenous ion subsequent to its
elimination as a hydrophilic complex by the kidneys. If transmetala-
tion of a soluble, paramagnetic Gd complex by diamagnetic Zn(II)
ions were to occur in a phosphate-buffered solution, then the
released Gd(III) would react to form Gd (PO ) , the solubility of
Chart 1
2
4 3
ꢀ
22.2
2
2
which is very low (K = 10
mol /L ) and whose influence on
the longitudinal relaxation rate of water is negligible.
The relative value of R1 at any time t, R (t)/ R (0), is
sp
30
P
P
P
1
1
therefore a good estimator of the extent of transmetalation. Its
evolution over time gives relevant information about the kinetics
of the reaction, whereas the plateau value theoretically reached at
t = ∞ will reflect the thermodynamic aspects of the system.
Measurements were made of the buffered solutions (phosphate
buffer, pH 7.4) containing equimolar amounts of 2 (2.5 mM) and
21
ZnCl (2.5 mM) as described in the Experimental Section, and
2
quantitative transmetalation kinetic data are provided in Table 1.
Also provided are the data obtained with Gd-DTPA-BMA, Gd-
DOTA, Gd-EOB-DTPA, and Gd-BOPTA for comparative pur-
poses. The kinetic data are in good agreement with the semiquanti-
tative results shown in Figure 1. The fitted graph of kinetic constants
is provided in Figure S1 (ESI).
Quite obviously there must be a clear distinction in the
evolution rate between complexes incorporating macrocyclic
18c
chelates and those containing acyclic ones. As such, Gd-
DOTA reveals the highest kinetic stability (inertness), that is,
0
the smallest K , as it possesses more five-membered metalla-
f
cycles (eight in all) not alone with the macrocyclic motifs. On the
other hand, complexes incorporating an acyclic ring such as
Figure 2. Human serum stability for 2, Gd-DTPA-BMA, and Gd-DTPA.
Figure 3. HPLC spectra of (a) 2b and (b) human serum.
5
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Journal of Medicinal Chemistry
ARTICLE
Table 2. Relaxivity Data of 2, Gd-DTPA, MS-325, Gd-EOB-
DTPA, and Gd-BOPTA in Water and Blood (64 MHz, 310 K,
2
at 293 K)
water
blood (HSA, 0.67 mM)
R
1
1
,
R
2
,
R
1
,
R
2
,
ratio
/R
ꢀ
ꢀ1
ꢀ1 ꢀ1
ꢀ1 ꢀ1
ꢀ1 ꢀ1
mM
s
mM
s
mM
s
mM
s
R
1
2
2
a
8.9
10.2
13.5
3.9
31.3
37.2
4.3
47.9
0.65
0.59
0.97
0.51
0.76
0.74
2
b
10.9
3.3
62.8
4.4
a
Gd-DTPA
a
MS-325
5.2
5.9
19.0
8.52
6.91
37.0
11.18
9.33
Gd-EOB-DTPA
Gd-BOPTA
7.14
5.35
7.96
5.87
a
Data obtained from ref 33.
Figure 4. HPLC spectra of 2b incubating in human serum at 37 ꢀC at
different time intervals.
Figure 6. Plots of R
1
relaxivity of various MRI CAs in water (solid line)
and 4.5% HSA (dashed line) as a function of [Gd].
Figure 5. Quantitative results obtained by integration of area at
τ = 17 min.
Gd-EOB-DTPA, Gd-BOPTA, and Gd-DTPA-BMA exhibit much
0
lower kinetic stability with the K values being 7ꢀ12 times as
f
high as that of Gd-DOTA. The kinetic stability of 2, which also
adopts a macrocycle and possesses fewer five-membered rings, is
located between that of Gd-DOTA and Gd-DTPA as shown in
Table 1 and Figure 1. It is informative to note that Gd ferromide
(
Chart 1), an organometallic analogue of macrocyclic DTPA-
0
ꢀ4
bis(amide), has a kinetic constant (K = 5.46 ꢁ 10 ) quite close
f
to that of 2. All-in-all, a general trend, though qualitative, may
be drawn as follows, in decreasing order of kinetic stability:
Gd-DTPA ∼ Gd-DTPA-bis(amide) > macrocyclic Gd-DTPA-
bis(amide) > Gd-DOTA.
17
Figure 7. Plots of the Dy(III)-induced water O shifts as function of [Dy].
Serum Stability. In addition to the kinetic stability of the
present system, their biostability is equally essential for application
in vivo. For this purpose we have developed a qualitative method of
31
ions. The figure shows little changes in the evolution rate with 2
3+
determining the Gd concentration by measuring the ratio of
compared with clinically available Gd-DTPA analogues.
p
p
p
1 1
p
relaxivities, R (t)/R (0), as a function of incubation time for the
Although qulitative, the plot of R (t)/R (0) as a function of
1
1
1
8b
serum solution of 2.
Figure 2 shows serum stability assay
time should provide reliable information about the biostability of
performed on 2, Gd-DTPA-BMA, and Gd-DTPA. The resulting
2. We have thus performed the traditional HPLC measurement
for the purpose of consistency. The stability assay on 2b was
p
25
proton longitudinal relaxation rate enhancement (R ) thus essen-
1
tially arises from the existence of solubilized and hydrated Gd(III)
conducted by monitoring the signals from a UV absorbance
5
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Journal of Medicinal Chemistry
ARTICLE
Figure 8. In vivo MR coronal images of mice obtained with 2b (0.1 mmol/kg): H, heart; L, liver; K, kidney; B, bladder; A, abdominal aorta.
Figure 9. In vivo MR coronal maximum intensity projection images of mice obtained by tail vein injection with 2b (0.1 mmol/kg): C, carotid artery; H,
heart; L, liver; K, kidney; B, bladder; A, abdominal aorta.
detector at 240 nm on the HPLC system for an extended period
of time. Figure 3 shows the HPLC spectra of 2b and serum as
reference standards. Figure 4 shows that 2b remains stable enough
in human serum for as long as 15 days. Figure 5 shows quantitatively
the decrease in the peak area (%) of the spectrum as a function of
time. Very little decrease (as low as 5%) is observed within the first 6
days. Even after 15 days, overall decrease was as low as 14%.
examined for comparison both in water and in blood. Even more
remarkable is the observation that not only the same relaxivity of
2 reaches almost twice as high as that of MS-325, a well-known
MRI BPCA, but also the R /R value (=0.65) for 2a is higher
1
2
than that (=0.51) for MS-325.
A slow tumbling motion (longer rotational correlation time,
τR) due to the formation of a sterically strained ring by the
biphenyl unit might have played a certain role in such high
relaxivities exhibited by 2. The relaxivity increase in the blood
further demonstrates the role of the aromatic ring that may be
Relaxivity. Table 2 summarizes R relaxivity for 2 along with
1
those for Gd-DTPA and MS-325 for comparative purpose. It is
remarkable that 2 shows the highest R relaxivities among those
1
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Journal of Medicinal Chemistry
ARTICLE
Figure 10. In vivo MR coronal images of brain tumor (C6-glyoma cell) obtained with 2b (0.1 mmol/kg).
involved in the lipophilic interaction with HSA. Figure 6 provides
solid evidence for the presence of such an interaction. In HSA
solution, the relaxation rates are significantly enhanced compared
with those in pure water. For example, the paramagnetic relaxa-
tion rate of a solution containing 2 (1 mM) and HSA (4.5%,
0
.67 mM) is 3 times as high as that in pure water. On the other
hand, little relaxivity increase is observed with either Gd-EOB-
DTPA or Gd-BOPTA, well-known BPCAs with liver specificity.
Finally, the most characteristic feature of 2 compared with MS-
3
25, EOB-DTPA, and Gd-BOPTA is that the former is neutral in
charge and therefore, unlike the latter, should exhibit no electro-
static interaction. As stated earlier, the electrostatic interaction
between the negatively charged chelate in Gd chelate and the
positively charged protein moiety is known to make an additional
Figure 11. Cell viability for 2 and Gd-DTPA.
32
contribution to the relaxivity increase. It is interesting, there-
fore, to note such high relaxivity with 2 even in the absence of
such an electrostatic interaction.
1
7
Dysprosium-Induced Water O Shifts. Figure 7 shows the
17
shift of the water O NMR signal to lower frequencies on addition
of 3compared to DyCl . As mentioned earlier, the slope of each plot
3
is linearly proportional to the hydration number (q), and hydration
numbers (q) for 3a and 3b are found to be 1.16 and 1.79,
respectively (see Experimental Section for calculation). These
observations may justify our hypothesis made above in connec-
tion with such high relaxivities exhibited by 3. In other words, a
macrocyclic ring conformation of 3, rather than the hydration
number, seems to play a more prominent role in their relaxivities.
In Vivo MR Imaging. Figure 8 shows the coronal T -weighted
images of 6-week-old male mice, ICR for 2b, and Figure 9 shows
1
Figure 12. Cell viability for 2 and Gd-DTPA-BMA.
the coronal T -weighted angiography images of 10-week-old
1
concentration in blood might have caused such enhancement in
brain tumor, and as such a noncovalent interaction of biphenyl with
HSA seems to play a certain role in the long retention time in blood.
In Vitro Cell Toxicity. Figures 11 and 12 show the cytotoxicity
tests performed on CT-26 mouse colon carcinoma cell and IMR-
male SD rats as a function of time. A strong blood-pool effect is
observed from the signal enhancement in both heart and
abdominal aorta. Signal enhancement is observed in various
arteries and vessels and lasts as long as 120 min after injection as
demonstrated in Figure 9. Renal excretion is observed 20 min
after injection as evidenced by enhancement in kidneys and
bladder. The CNR profiles shown in Figure S2 provide concur-
9
0 human embryonic lung fibroblasts with 2. Cytotoxicity of the
present series compares well with that of Gd-DTPA and Gd-
DTPA-BMA. When the comparison is made between 2a and 2b,
the former shows a little lower cytotoxicity. A related observation
is that cell viability for 2b decreases gradually with an increase in
its concentration.
34,35
ring observations made in connection with Figures 8 and 9.
Even more striking, 2b is able to enhance the brain tumor for as
long as an hour, which is a rare example of ordinary ECF MRI
CAs exhibiting such behavior (Figure 10). For instance, we have
provided supporting evidence in Figure S3 showing the CNR
profiles obtained with 2b to be much greater during the early
stage of injection than those with Gd-DOTA, a well-known
ECF CA. The long circulation time and the subsequent high
’
CONCLUSION
The reaction of 2,2 -diaminobiphenyls with DTPA-bis-
(anhydride) yielded corresponding macrocyclic DTPA-bis(amides)
0
5
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Journal of Medicinal Chemistry
2a,b) rather than acyclic, oligomeric analogues. They form on
ARTICLE
(
angiography of mice using macromolecular MR contrast agents with
polyamidoamine dendrimer core with references to their pharmacoki-
netic properties. Magn. Reson. Med. 2001, 45 (3), 454–460.
4) Ayyagari, A. L.; Zhang, X. D.; Ghaghada, K. B.; Annapragada, A.;
Hu, X. P.; Bellamkonda, R. V. Long-circulating liposomal contrast agents
reaction with Gd(OAc) corresponding Gd complexes of the
3
type [Gd(L)(H O)] xH O (2a,b; L = 1a,b). Pharmacokinetic
2
3
2
(
inertness of 2 compares well with that of analogous Gd-DTPA
MRI CAs currently in use. The R relaxivity reaches as high as
1
ꢀ
1
ꢀ1
for magnetic resonance imaging. Magn. Reson. Med. 2006, 55 (5),
1
0.9 mM
s . This value is approximately 3 times as high as
1
023–1029.
(
ꢀ
1
ꢀ1
that of Gd-DOTA (R = 3.7 mM
s
). The R relaxivity in
1
1
5) Schwickert, H. C.; Stiskal, M.; Vandijke, C. F.; Roberts, T. P.;
ꢀ1
ꢀ1
HSA is 37.2 mM
s , which is almost twice as high as that of
Mann, J. S.; Demsar, F.; Brasch, R. C. Tumor angiography using high
resolution 3D MRI: comparison of Gd-DTPA and a macromolecular
blood pool contrast agent. Acad. Radiol. 1995, 2 (10), 851–858.
(6) Lauffer, R. B.; Parmelee, D. J.; Dunham, S. U.; Ouellet, H. S.;
Dolan, R. P.; Witte, S.; McMurry, T. J.; Walovitch, R. C. MS-325:
albumin-targeted contrast agent for MR angiography. Radiology (Oak
Brook, IL, U. S.) 1998, 207 (2), 529–538.
MS-325, a leading BPCA, demonstrating a strong blood pool
effect. The in vivo MR images of mice obtained with 2b are
coherent, showing strong signal enhancement in heart, abdom-
inal aorta, and small vessels. Further, even the brain tumor is
vividly enhanced for an extended period of time. The structural
uniqueness of 2 is that it is neutral in charge and thus makes no
resort to electrostatic interaction, supposedly one of the essential
factors for the blood-pool effect.
(
7) Cavagna, F. M.; Lorusso, V.; Anelli, P. L.; Maggioni, F.; de Ha €e n,
C. Preclinical profile and clinical potential of gadocoletic acid trisodium
salt (B22956/1), a new intravascular contrast medium for MRI. Acad.
Radiol. 2002, 9, S491–S494.
’
ASSOCIATED CONTENT
(
8) Preda, A.; Novikov, V.; M €o glich, M.; Turetschek, M. K.; Shames,
D. M.; Brasch, R. C.; Cavagna, F. M.; Roberts, T. P. L. MRI monitoring
of avastin antiangiogenesis therapy using B22956/1, a new blood pool
contrast agent, in an experimental model of human cancer. J. Magn.
Reson. Imaging 2004, 20 (5), 865–873.
(9) Merbach, A. E.; T ꢀo th, E. The Chemistry of Contrast Agents in
Medial Magnetic Resonance Imaging; Wiley-VCH: Chichester, U.K.,
S
Supporting Information. Kinetic inertness fitting data,
b
in vivo CNR profiles, and osmolality data; zipped file containing a
video of MR of mouse in avi format. This material is available free
of charge via the Internet at http://pubs.acs.org.
2
001.
(
’
AUTHOR INFORMATION
10) Aime, S.; Botta, M.; Crich, S. G.; Giovenzana, G.; Palmisano,
Corresponding Author
G.; Sisti, M. Novel paramagnetic macromolecular complexes derived
from the linkage of a macrocyclic Gd(III) complex to polyamino acids
through a squaric acid moiety. Bioconjugate Chem. 1999, 10 (2),
*
For Y.C.: phone, (+)82-53-420-5471; e-mail, ychang@knu.ac.
kr. For T.-J.K.: phone, (+)82-53-950-5587; fax, (+)82-53-950-
594; e-mail, tjkim@knu.ac.kr.
192–199.
6
(11) Curtet, C.; Maton, F.; Havet, T.; Slinkin, M.; Mishra, A.;
Chatal., J. F.; Muller, R. N. Polylysine-Gd-DTPAn and polylysine-Gd-
0
DOTAn coupled to anti-CEA F(ab )2 fragments as potential immuno-
’
ACKNOWLEDGMENT
contrast agents. Relaxometry, biodistribution, and magnetic resonance
imaging in nude mice grafted with human colorectal carcinoma. Invest.
Radiol. 1998, 33 (10), 752–761.
T.-J.K. and Y.C. gratefully acknowledge NRF (Grant No.
2
010-0024143) and NRF (Grant No. 2009-0072413), respec-
tively, for financial support. KBSI, Daegu branch, is also acknowl-
edged for the mass spectral measurements.
(12) Lokling, K. E.; Fossheim, S. L.; Skurtveit, R.; Bjornerud, A.;
Klaveness, J. pH-sensitive paramagnetic liposomes as MRI contrast
agents: in vitro feasibility studies. Magn. Reson. Imaging 2001, 19 (5),
7
31–738.
’
ABBREVIATIONS USED
(13) Jung, C. W.; Jacobs, P. Physical and chemical properties of
0
00 00
DTPA, diethylentriamine-N,N,N ,N ,N -pentaacetic acid; MRI,
magnetic resonance imaging; BPCA, blood-pool contrast agent;
CA, contrast agent; DOTA, 1,4,7,10-tetraazacyclododecane-N,
superparamagnetic iron oxide MR contrast agents: ferumoxides, feru-
moxtran, ferumoxsil. Magn. Reson. Imaging 1995, 13 (5), 661–674.
(14) Taupitz, M.; Schnorr, J.; Abramjuk, C.; Wagner, S.; Pilgrimm,
0
00 000
H.; H €u nigen, H.; Hamm, B. New generation of monomer-stabilized very
small superparamagnetic iron oxide particles (VSOP) as contrast
medium for MR angiography: preclinical results in rats and rabbits.
J. Magn. Reson. Imaging 2000, 12 (6), 905–911.
N ,N ,N -1,4,7,10-tetraacetic acid; MS-325, (trisodium {(2-(R)-
[
(4,4-diphenylcyclohexyl)phosphonooxymethyl]diethylenetria-
minepentaacetato)(aquo)gadolinium(III)}; ECF, extracellular
fluid; HSA, human serum albumin; TI, inversion time; TE,
echo time; TR, repetition time; ICR, Institute of Cancer Research;
SD, SpragueꢀDawley; NEX, number of acquisitions; ROI, region
(15) Mornet, S.; Vasseur, S.; Grasset, F.; Duguet, E. Magnetic
nanoparticle design for medical diagnosis and therapy. J. Mater. Chem.
2
004, 14 (14), 2161–2175.
(16) Avedano, S.; Tei, L.; Lombardi, A.; Giovenzana, G. B.; Aime, S.;
0
0
of interest; TMS, tetramethylsilane; DTPA-BMA, N,N -bis-
(
3
4
methylamide) of DTPA; EOB-DTPA, S-[4-(4-ethoxybenzyl)-
,6,9-tris(carboxylatomethyl)-3,6,9-triazaundecanedioic acid; BOPTA,
-carboxy-5,8,11-tris(carboxylmethyl)-l-pheny1-2-oxa-5,8,11- ria-
zatridecan-13-oic acid
Longo, D.; Botta, M. Maximizing the relaxivity of HSA-bound gadoli-
nium complexes by simultaneous optimization of rotation and water
exchange. Chem. Commun. 2007, 45, 4726–4728.
(17) Henrotte, V.; Elst, L. V.; Laurent, S.; Muller, R. N. Compre-
hensive investigation of non-covalent binding of MRI contrast agents
with human serum albumin. J. Biol. Inorg. Chem. 2007, 12 (6), 929–937.
’
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