Design, Synthesis, and validation of
Imaging Probes
MRI CEST agents: basic principles,
mechanism of action and classification
Enzo Terreno
Torino, September 19 to 30, 2011
Lesson outline

CEST agents mechanism of action

Sensitivity issue
 CEST agents vs conventional MRI agents
 CEST agents classification
 LIPOCEST
CEST (Chemical Exchange Saturation Transfer) agents
Dw
(rad.Hz)
= wWAT - wCEST
If Dw > kex then….
wWAT
CEST
Agent
wCEST
40
water
molecule
CEST agent
mobile protons
30
20
10
Dw
0
-10 -20 -30 -40
KHz
The MR-CEST experiment
OFF resonance
irradiation
Aqueous solution of
a CEST agent
IS
IS = 7.8 a.u.
Bulk
water
40 30 20 10
IWAT = 16.9 a.u.
rf field
rf field
0 -10 -20 -30 -40
kHz
I0
CEST agent
40 30 20 10
Dw
0 -10 -20 -30 -40
Dw
kHz
OFF* resonance
irradiation
40 30 20 10
I0 = 14.5 a.u.
0 -10 -20 -30 -40
kHz
The decrease of IWAT is the source of the MR contrast
The net ST % effect is calculated as ( 1 - IS/I0 )
.
100
CEST agents: the sensitivity issue
Sherry D. et al.Chem. Soc. Rev., 2006, 35, 500–511
 tsat  R1w  kex fCEST 
IS
kex fCEST
ST  1   w
(1  e
)
I 0 R1  kex fCEST
Where:
fCEST 
n CA
2[ BulkW ]
Exchange rate of the mobile protons belonging to the contrast agent(CA)
Relaxation rate of the bulk water protons
Concentration of the contrast agent
Number of magnetically equivalent mobile protons
Experimental parameter: irradiation time
 tsat  R1w  kex fCEST 
IS
kex fCEST
ST  1   w
(1  e
)
I 0 R1  kex fCEST
Increase of
R1 bulk
water pool
Simulation parameters*
fCEST= 5×10-4, R2bw=
2×R1bw, R1CEST = 2 s-1,
R2CEST = 50 s-1, kCEST =
1600 s-1, B2 = 6 mT
Woessner et al, MRM
2005
The steady-state condition is reached when
e
CEST
 R1bw  kex
f CEST
In addition to R1bw, also an increase of the exchange rate
accelerates the achievement of the steady state condition
 0
Sensitivity of CEST agents: playing with kex
In principle, the CEST efficiency is proportional to kex, but the exchange rate
cannot be increased at will, because:
i) the condition Dw > kex has to be satisfied
ii)
increasing kex
Max. CEST
kex
 2
B2
Fast exchange requires high-intensity saturation fields for achieving full saturation
 overcoming SAR* limitations
 direct saturation of bulk water
unsafe saturation
less efficient CEST contrast
*SAR = Specific Absorption Rate. It is defined as the RF power absorbed per unit of mass of
an object, and is measured in watts per kilogram (W/kg).SAR of 1 Wkg-1 applied for an
hour would result in a temperature rise of about 1 °C.
Sensitivity of CEST agents: increasing the number of saturated protons
The CEST efficiency is proportional to the number of mobile protons
A CEST contrast of 10% requires
few millimolar of mobile protons
The sensitivity can be improved by increasing
the number of mobile protons (with equal or
similar Dw values) per CEST molecule
Mobile protons
Sensitivity
Chemical system
< 10
mM
Low MW molecules
103
mM
Macromolecules
106
nM
Nanoparticles
Optimizing the saturation scheme
The overall saturation time can be covered by:
 a single cw rectangular pulse
 a train of shaped pulses
Single cw saturation pulses (2-10 s long) have been observed to be more efficient in
case of probes with relatively slow exchange: DIACEST (B2 1-3 mT), PARACEST
(typically amide protons or slowly exchanging Ln-bound water protons, B2 12-24 mT),
and LipoCEST (B2 6-12 mT)
BUT
Pulsed shaped pulses are definitely better in terms of deposited energy
and clinical translation
CEST agents: the sensitivity issue
CEST contrast is quite sensitive because few millimolar of saturated mobile
protons are sufficient to affect the MRI signal (> 100 molar)
However, the development of highly sensitive agents is an hot issue,
especially for molecular imaging purposes.
H2 O
Saturation
rf pulse
Variables
affecting the
CEST contrast
50 40 30 20 10
0 -10 -20 -30 -40 -50
ppm
 Exchange rate of the CEST protons
 Duration of saturation time
 Resonance frequency of the CEST protons
 Saturation scheme (cw, pulsed)
 Total concentration of saturated spins
 Saturation pulse power
 T1 and T2 of both the exchanging sites
 Magnetic field strength (affects Dw)
CEST agents: mechanism of action
H2 O
Saturation
rf pulse
50
ST-spectrum
45
40
0 -10 -20 -30 -40 -50
ppm
30
ST %
50 40 30 20 10
ST % = 100  (1- IS/I0)
35
25
20
15
Dw
100
10
Dw
5
0
I
water
%
80
I0
60
40
20
IS
Z-spectrum
0
50 40 30 20 10
0 -10 -20 -30 -40 -50
ppm
0
10
20
ppm
30
40
50
Field homogeneity
B0 field homogeneity plays a key-role for the detection of CEST contrast,
especially in biological specimen
Dw
100
Dw
50
40
I
I0
60
IS
ST % = 100  (1- IS/I0)
35
30
ST %
water
%
80
40
ST-spectrum
45
25
20
15
20
10
Z-spectrum
5
0
50 40 30 20 10
0 -10 -20 -30 -40 -50
ppm
0
0
10
20
ppm
30
40
50
An accurate contrast assessment requires that the two MR signal intensities
are measured at frequency offsets symmetrically distributed with respect to
the resonance frequency of the bulk water
Field homogeneity
The pixel-by-pixel evaluation of the spatial distribution of the frequency offset
of the bulk water is necessary to avoid CEST artifacts…
Human astrocytoma
Zhou et al., MRM 2008, 60, 842
Field homogeneity
…but, of course, acquiring B0 maps takes time
Several methods have been proposed so far:
 B0 (and also B2) compensation algorithm (Sun et al., MRM 2007)
 relatively fast (in addition to the couple of CEST scans, it requires few images for
generating the B0/B1 maps)
 Not suitable for large inhomogeneities
 Z-spectrum interpolation (Zhou et al., Nat. Med. 2003 – Stancanello et al., CMMI 2008)
 broad applicability, good accuracy
 Relatively time consuming (depending on the frequency sampling)
 WAter Saturation Shift Referencing (WASSR) (Kim et al., MRM 2009)
 excellent accuracy; optimal for detecting CEST contrast from very little shifted agents
 Time consuming (an additional Z spectrum is required)
CEST agents vs conventional MRI contrast agents
Gd-based fibrin-targeting agent
(visualisation of non-occlusive thrombi)
3D-Gd-MR Angiography
Do we actually need a
new class of MRI
agent ?
Gd-enhanced MR images
MRI cell tracking experiment
(Glioma)
(Limph node targeting by tumor specific
SPIO-labeled dendritic cells)
in vivo pH mapping of tumors
Multiple visualization of MRI agents
In vivo multiple visualization of MRI probes would considerably improve
the potential of MRI in many molecular imaging experiments (e.g. multi-
detection
of
epitopes,
simultaneous
tracking
of
different
populations, dynamic measurements,...)
Example: vivo Fluorescence image of a lymph node
R .N. Germain et al, Nature Rev. Imm., 2006, 6, 497
Can we get similar results using MRI agents?
cell
NMR provides a parameter that can characterise any molecule:
the resonance frequency of its spins
1H-NMR
spectrum
bulk
water
Multiple detection can be
achieved by generating a
“frequency encoded” contrast
Agent A
10
8
6
4
Agent B
2
0
ppm
For imaging purposes, this information must be transferred to the intensity
of bulk water protons
CEST agents vs conventional MRI agents
MULTIPLE DETECTION
1,8
1,5
B
D
A
1,2
50
A
40
B
30
C
20
ppm
10
C
D
0
-10
Concentration dependence of the MRI contrast
R2
10
60
-1
8
R1water s
CEST %
80
40
6
4
R1
20
2
0
0
20
40
60
[CEST] - mM
80
100
0
0,0
0,2
0,4
0,6
0,8
1,0
[agent] mM
Responsive contrast requires the MRI observable to depend only on the
parameter of interest
A ratiometric analysis of the intensity of the water signal after the irradiation of two
different mobile pools allows to get rid of the concentration of the MRI probes
CEST agents are very suitable Responsive probes
Ratiometric analysis
ST  1 
IS
I0
(1);
IS
1

I 0 1  kbwT1w
(2);
IS

I0
by substitution of eq.3 into eq.2
kex n CAT1w
I0
 1
IS
2[ BulkW ]
kbw 
(5);
kex n CA
2[ BulkW ]
1
kex n CA
1
T1w
2[ BulkW ]
(3);
(4);
kex n CAT1w
I0
1 
IS
2[ BulkW ]
If two exchanging pools (A and B) are present in a known ratio (R) then...
poolA
A
A A
 I0

k
n
CA
T1w


ex

1
A


A A
A
k
n
CA
I


k
ex
 S

2[ BulkW ]
ex



R
B
poolB
B
B
B B
B B
kex
kex n CA T1w kex n CA
 I0

  1
2[ BulkW ]
 IS

Classification of CEST agents
Diamagnetic
CEST agents
Paramagnetic
CEST agents
Diamagnetic - vs. Paramagnetic -CEST agents
Dw > kex
Dwpara
R
HN
O
Dwdia
OH2
NH R
N
O
R
N
O
Ln3+
N
N
HN
O
NH R
DIACEST
PARACEST
30
25
20
15
10
5
0
ppm
The extension of the Dw range facilitates multiple visualization and allow
to exploit larger exchange rate before coalescence takes place, but the
associated line broadening may introduce SAR issues and the T1 and T2
shortening may be detrimental for the CEST eficacy
Field homogeneity and highly shifted agents
The detrimental effect of B0 inhomogeneity progressively vanishes moving
away from the resonance of the bulk water
mouse bearing a
B16 melanoma xenograft
(B2 6 mT)
ST@ 3.5 ppm
Uncorrected CEST maps
Morphological T2w- image
Water shift map
Highly shifted CEST agents can be accurately
detected by a simple two scans experiments
ST@ 20 ppm
Classification of CEST agents
Small-sized
(sugars,
aminoacids)
DIACEST
Macromolecular
/Polymeric
(poliaminoacids,
RNA-like,…)
Diamagnetic
CEST agents
EndoCEST
peptides/proteins,
glycogen...
Paramagnetic
CEST agents
Endogenous CEST agents (2): APT imaging
Human patient with a meningioma (3 T)
APT imaging may help to discriminate between tumor and edema
Endogenous CEST agents (4): gagCEST
In vitro
cartilage depletion by tripsine
In vivo CEST-MR images of
a human patella
Classification of CEST agents
Small-sized
(sugars,
aminoacids)
DIACEST
Macromolecular
/Polymeric
(poliaminoacids,
RNA-like,…)
Diamagnetic
CEST agents
EndoCEST
peptides/proteins,
glycogen...
Small-sized
ParaCEST
Paramagnetic
CEST agents
SupraCEST
LipoCEST
Eu
CEST agents for cell-labeling experiments (1)
CEST agents can be used to label different cell populations
1,4
Eu-H2O
1,2
Tb-H2O
1,0
200
0
-200
-400
-600
-800
ppm
Angew. Chem. Int. Ed. 2005, 44, 1813
Classification of CEST agents
Small-sized
(sugars,
aminoacids)
DIACEST
Macromolecular
/Polymeric
(poliaminoacids,
RNA-like,…)
Diamagnetic
CEST agents
EndoCEST
peptides/proteins,
glycogen...
Small-sized
ParaCEST
Paramagnetic
CEST agents
SupraCEST
LipoCEST
Macromolecular
/Polymeric
(dendrimers...)
CEST agents for assessing tumor vascular permeability
Simultaneous injection of two agents with different size
Yb-G2
Eu-G5
MCF-7 xenograft tumor on mice
Meser Ali et al., Mol. Pharm. 2009, 6, 1409
Classification of CEST agents
Small-sized
(sugars,
aminoacids)
DIACEST
Macromolecular
/Polymeric
(poliaminoacids,
RNA-like,…)
Diamagnetic
CEST agents
EndoCEST
peptides/proteins,
glycogen...
Small-sized
ParaCEST
Paramagnetic
CEST agents
SupraCEST
LipoCEST
Macromolecular
/Polymeric
(dendrimers...)
Nano-sized,
(perfluorocarbon...)
The route to high sensitivity: exploiting nanotechnology
Usually, the typical approach consists of loading a large number of CEST agents to
the external surface of the nanosystem:
Perfluorocarbon nanoemulsions
P.M. Winter et al. Mag. Reson. Med., 2006, 56, 1384
R
HN
O
OH2
NH R
N
O
N
Ln3+
N
N
O
Adenovirus
R HN
O
NH
O. Vasalatiy et al. Bioconjugate Chem., 2008, 19, 598
The sensitivity of such nanoprobes is primarily dependent on the maximum payload
that can be achieved (generally 103-105 PARACEST units per nanosystem)
Macromolecular Paramagnetic CEST agents
To exploit the reversible interaction
between a paramagnetic
Shift Reagent and a substrate rich of
mobile protons
SUPRACEST
Example:
Interaction between
[TmDOTP]52-
O3P
N
N
PO32-
3+
Tm
2-
N
O3P
N
PO32-
and polyArginine
Sensitivity threshold (referred to the paramagnetic complex) of tens of mmolar
S. Aime et al., Angew. Chemie Int. Ed., 2003, 42, 4527
1) Small-sized molecules
small number of mobile protons (<10) per molecule
Sensitivity
a)Diamagnetic agents
(sugars, aminoacids,…)
B1 field intensity
tens of mM

few mM

hundreds of mM

b)Paramagnetic agents
PARACEST
Amide, hydroxilic
protons
metal bound
water protons
2) Macromolecular agents
a)Diamagnetic agents
(poliaminoacids, RNA-like,…)
large number of mobile protons (~103) per molecule
mM

few mM

b)Paramagnetic agents
SUPRACEST agents
3) Nanoparticles
extremely high number of mobile protons (>106) per molecule
LIPOCEST agents
tens of pM

Liposomes
Biocompatibles, extremely versatiles, successfully used in pharmaceutical field
Cryo-TEM image
DPPC: DiPalmitoyl-PhosphatidylCholine
 The external surface may be easily functionalized with a
wide variety of chemicals including targeting vectors, or
PEG chains for prolonging the blood half lifetime (Stealth®
liposomes).
 Liposomes can be passively accumulated in
pathological body regions (tumors, atherosclerotic
plaques,…)
The ST efficiency  to kex
and
number of mobile protons
WATER
MOLECULE
kex
The number of mobile protons for Large Unilamellar Vescicles (LUV) range from
2,4x106 (50 nm) to 2,1x109 (500 nm)

Liposomes can be very efficient CEST Probes
kex
can be modulated by:
 varying the liposome membrane permeability (P)
(kex= P  S/V)
Saturated phospholipids
Membrane tightly packed
Slow exchange
Unsaturated phospholipids
Less tightly packed
Fast exchange
Cholesterol insert himself
in the hole
Reduce the exchange rate
 Varying the liposome size (kex= P  S/V=P x 3/radius)
kex
n° of mobile protons
How the resonance frequencies of inner and outer water
protons can be separated ?
Encapsulating a paramagnetic shift reagent (SR) in the liposome
Bulk water
intraliposomal water
Bulk+ water
O
kex
O
OH2
N
O
O
N
Ln3+
N
N
O
O
O
O
intraliposomal water
10
O
O
O
O
N
Ln3+
N
N
O
4
2
0
-2
-4
-6
-8 -10
O
O
O
6
ppm
OH2
N
8
Lanthanide-based SR
Angew. Chem. Int. Ed., 2005, 44, 5513.
The chemical shift of the water protons () in the presence of
a paramagnetic SR is the sum of three contributions:
   DIA   HYP   BMS
DIA often negligible
HYP requires a “chemical” interaction between the paramagnetic
center (the Ln(III) ion) and the water molecule
(through bond: contact shift; through space: pseudocontact shift)
BMS does not require a “chemical” interaction and it is dependent on
the bulk magnetic susceptibility of the compartment containing the SR
In the case of spherical compartment BMS=0
Conventional liposomes
Shift Reagent for intraliposomal water protons
When kex  Dw then:
[ H 2O ]bound to SR
 int ralipo =
  bound
[ H 2O ]total
water
water
H
H
O
Ln
bound   HYP   pseudo  D  G
water
contact
 D is the magnetic anisotropy of the lanthanide complex Ln
D  CJ  A02 r 2
- CJ is a constant of the metal
CJ > 0 for Eu, Er, Tm e Yb
CJ < 0 for Ce, Pr, Nd, Sm, Tb, Dy and Ho
CJ = 0 for Gd
- (A02 <r2>) depends on the crystal field
Magnetic axis
of the complex
H
H
O
Ln
G
3 cos 2   1

r3
O
DyDOTMA
O
DOTA
O
DyDOTA
N
N
N
N
O
DyHPDO3A
O
DyDTPA
O
TmDOTMA
O
O
O
TmDOTA
O
O
DOTMA
TmDTPA
35 30 25 20 15 10 5
N
N
TmHPDO3A
N
N
O
O
0 -5 -10-15-20-25-30-35
O
HYP - ppm/M
O
O
O
O
HOOC
O
COOH
N
N
N
N
O
HOOC
N
N
OH
COOH
HPDO3A
N
DTPA
O
O
COOH
-Shift differences are due to the geometric differences among the complexes
(parameter G)
LIPOCEST agents
SR
Hydration
Vortexing
MLV
Extrusion
55 °C
LUV
Dialisis
55 °C
Lipidic film
Bulk water
SR unit
Water protons
inside liposomes
1H-NMR
spectrum (7 T, 298 K)
Tm
[TmDOTMA]0.12 M inside liposomes
DPPC/DPPG 95/5 (w/w) liposomes
LipoCEST agents: sensitivity
LipoCEST formulation: POPC/DPPG/Chol (55/5/40 in moles) size:250 nm
Experimental condition: 7 T – 37°C – pH 7.4 - B2 field 6 mT
O
O
N
O
O
N
OH2
N
Ln3+
N
O
O
O
O
Normalized Intensity Values, a.u.
Z-spectra GLOBAL ROI
NMR
spectrum
1
0.8
Z-spectrum
sample 1
0.6
0.4
0.2
0
-15
-10
-5
0
5
Sat. offset, ppm
10
15
Seleziona le ROI manualmente - sulla prima viene calcolato l'ST globale
LipoCEST conc.
1.
2.
3.
4.
5.
6.
1.5 nM
750 pM
320 pM
160 pM
80 pM
40 pM
1
2
6
5
3 4
T2W Image
CEST map @ 3.8 ppm
LipoCEST agents: factors affecting sensitivity
- Water permeability of the liposome bilayer (Pw)
Can be modulated by changing the packaging
properties of the phospholipids
Phospholipids with saturated aliphatic chains (e.g. dipalmitoyl) displays lower Pw than unsaturated ones (e.g.
di-oleyl)
Cholesterol intercalates in the bilayer and reduces Pw
1000
-5
-1
10 x Pw (cm s )
800
600
400
200
9
6
3
0
DPPC
POPC/CHOL (20) POPC/CHOL (40)
DSPE-PEG2000 DSPE-PEG2000 DSPE-PEG2000
J. Inorg. Biochem., 2008, 102, 1112.
First generation LIPOCEST: spherical liposomes
Pro: Highly sensitive (pM range)
Con: Little frequency range
Tm
4ppm
-4ppm
Dy
How to increase the shift?
Exploiting the BMS shift
LISBulk = BMS  Dip
water
BMS depends on the concentration of the shift reagent and its sign
depends on the shape and orientation (wrt B0) of the compartment
in which the shift reagent is confined
Second generation of LIPOCEST: non-spherical liposomes
Osmotic
shrinkage
-H2O
Cryo-TEM images of osmotically
shrunken LIPOCEST agents
in collaboration with E. Sanders and N.
Sommerdijk from University of
Eindhoven (NL)
Cj Gd=0
D  0
 bound   dia
water
Gd
Before dialysis
Gd(III)-complexes has Dip = 0
BMS   SR    meff

2
DOsm
meff Gd = 7.94
Lanthanides showing the higher
values for meff:
Gd, Tb, Dy, Ho,
Aime S. et al., J. Am. Chem. Soc., 2007, 129, 2430
Er and Tm
O
Hydrophilic SR
O
N
O
2nd generation
O
OH2
N
Tm3+
N
O
O
O
O OH2
N
O
CH3
O
N
Amphiphilic SR
O
Tm3+
N
HO
N
O
N
O
N
3rd generation
12 ppm
22 ppm
LISint ralipo  Dip  BMS
water
Terreno E. et al., Angew. Chemie Int. Ed., 2007, 46, 966.
A further DLIPO increase can be achieved by encapsulating neutral multimeric
SRs
-
COO-
OOC
N
-
COO-
OOC
N
N
N
N
OOC
N
OH
N
-
OOC
OOC
Gd
OOC
-
N
N
N
HNOC
OOC
N
Tm3+
-
N
CONH
COO-
[Tm-dimer]
CONH
N
N
Tm
N
[Tm-HPDO3A]
-
COO-
OOC
Tm
Tm
-
-
N
HNOC
COO-
N
N
COO-
COO-
HNOC
N
N
Tm3+
Gd
Gd
CONH
COO-
N
Tm3+
-
N
OOC
N
COO-
[Tm-trimer]
Tm-HPDO3A 25 mM
Tm-dimer 25 mM
28 ppm
Tm-trimer 25 mM
DPPC/Tm-1/DSPE-PEG2000
(75/20/5 mol %)
Terreno E. et al., Chem. Commun., 2008, 600
In addition to increase the magnitude of DLIPO, the incorporation of amphiphilic
SRs may also affect he sign of the shift through the modulation of the magnetic
alignment of the vesicles.
The sign of the BMS contribution depends on the orientation of the
compartment with respect to the external B0 field
B0
For a cylinder
DBMS  0
DBMS  0
As non-spherical vesicles, also the osmotically shrunken LIPOCEST
could orient themselves in the field, thus changing the DLIPO sign
Phospholipid-based systems, e.g bicelles, are
bicelle
oriented in the field with their principal symmetry
axis perpendicular to B0
Alignment energy
The driving force of the orientation is the
interaction between B0 and the magnetic
susceptibility anisotropy (D) of the
phospholipidic membrane.
B0
Incorporation in the
membrane of a
lanthanide complex with
D 0
COOH
D  0
N
CONH
HOOC
N
22270
Ln
CONH
D  0
N
COOH
S. Prosser et al., J. Magn. Res., 1999, 141, 256.
COOH
-
CONH
HOOC
O
O
N
22270
Ln
N
COO-
OOC
N
O
O
N
N
OH2
N
Ln3+
N
O
O
N
O
CONH
N
-
OOC
N
[LnHPDO3A]
Ln
O
N
OH
COOH
( D )SR  C jLn  A02 r 2
Tm
Tm CJ > 0
(D )SR  0
(D )LIPO  0
Gd
Gd CJ = 0
(D )SR  0
(D )LIPO  0
Dy
Dy CJ < 0
(D )SR  0
(D )LIPO  0
20
10
D
LIPO
0
-10
-20
- ppm
B0
O
O
N
O
O
N
OH2
N
Ln3+
N
O
O
O
N
O
O
O
O
O
N
OH2
N
Ln3+
N
O
O
O
O
D LIPO 0
D LIPO 0
with the same amphiphilic ligand it is
possible to change the liposome
orientation by changing the Ln(III) ion
-
COO-
OOC
N
N
Tm
[Tm-HPDO3A] -
O
N
OOC
N
OH
O
N
O
O
N
OH2
N
Ln3+
N
O
O
O
O
( D )SR  C jLn  A02 r 2
Tm CJ > 0
Tm(III)
complexes1
-
COO-
OOC
N
N
Tm
N
-
N
B0
CON
OOC
O
O
N
O
-
O
COO
OH2
N
Ln3+
N
O
O
O
O
N
O
O
O
O
N
OH2
N
Ln3+
N
O
O
O
O
2
N
-
N
OOC
D LIPO 0
D LIPO 0
CON
N
Tm
-
OOC
N
Tm-1
COO-
3
OOC
N N
-
OOC
N
Tm N
N
-
OOC
Tm-3
N N
-
Tm-2
N
OOC
COO
-
4
N
CONH
-
OOC
N
Tm-4
Tm
CONH
Tm-5
N
COO-
COOO
N
CONH
-
OOC
O
O
Tm
N
CONH
N
COO-
O
5
30
20
10
0
D
LIPO
-10
-20
-30
-40
- ppm
O
O
Delli Castelli D. et al., Inorg. Chem., 2008, 47, 2928.
Extending the range of DLIPO values
O
N
O
O
O
O
N
OH2
N
Ln3+
N
O
O
OH2
N
O
O
O
O
O
N
N
Ln3+
N
O
N
O
O
N
O
O
N
Ln3+
N
Entrapping
monomeric
or
multimeric neutral hydrophilic
shift reagents
O
O
O
O
O
1st generation
OH2
O
O
2nd generation 3nd generation
DLIPO
80
B0
60
O
N
O
O
N
O
OH2
N
Ln3+
N
O
N
O
N
N
Ln3+
N
O
O
OH2
O
O
O
O
O
O
O
O
O OH2
N
O
O
N
N
Ln3+
N
O
O
O
O
DLIPO 0
DLIPO 0
CEST %
O
40
20
The DLIPO sign for not spherical LipoCEST agents
depends on their orientation in the B0 field
0
40
20
0
-20
Saturation frequency offset (ppm)
Terreno E. et al., Methods Enzymol. , 2009, 464, 193.
-40
Multiple detection of LipoCEST agents: buffer vs. agar
O
A
O
O
O
OH2
N
O
N
Ln3+
N
N
O
B
O
O
O
7 T – 312 K – sat. intensity 6 mT
DLIPO 6.7 ppm
O
N
O
N
O
OH2
N
Ln3+
N
O
O
O
O
DLIPO 18 ppm
Control (buffer)
A
A+B
In buffer
A
B
6.7 ppm
B
In agarose gel
Control
(agar)
A+B
@18 ppm
In vivo Multiple detection of LipoCEST agents
Intramuscolar injection
Subcutaneous injection
O
O
O
O
N
Ln3+
N
N
O
N
OH2
N
O
O
O
O
O
O
N
O
O
O
O
N
O
OH2
N
Ln3+
N
N
OH2
N
Ln3+
N
O
O
O
O
O
O
@-17 ppm
O
O
@ 7 ppm
@-17 ppm
O
O
N
O
O
N
OH2
N
Ln3+
N
O
O
O
O
@3.5 ppm
Some readings…
S. Zhang et al., Acc. Chem. Res., 36, 783, 2003
- M. Woods et al., Chem. Soc. Rev., 35, 500, 2006
- J. Zhou et al., Progr. NMR Spectr., 48, 109, 2006
- S. Viswanathan et al., Chem. Rev., 110, 2960, 2010
- E. Terreno et al., Contrast Media Mol. I., 5, 78, 2010
- Hancu I. et al., Acta Radiol., 51, 910, 2010