From spherical to osmotically shrunken paramagnetic liposomes: an improved generation of LIPOCEST MRI agents with highly shifted water protons.
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Contrast Media↗
Biomedical subjects
Silvio Aime
Publications and source records attributed to Silvio Aime.
Magnetic resonance imaging of gadolinium-labeled pancreatic islets for experimental transplantation.
New imaging techniques that couple anatomical resolution to sensitivity may greatly contribute to improving islet transplantation. In the present work, a report is given of the direct detection of islets by magnetic resonance imaging (MRI) after ex vivo cell labeling with the MRI T(1) contrast agent GdHPDO3A. Experiments on mouse and human islets demonstrated well-tolerated uptake of GdHPDO3A, based on morphology, viability, glucose-dependent insulin response and apoptosis/toxicity gene array profile. GdHPDO3A loading was sufficient for in vitro MRI cell detection. In vivo isotransplanted mouse islets into the kidney capsule and xenotransplanted human islets within the mouse liver were detected. Imaging specificity was supported by the absence of signal in unlabeled islet transplants, its persistence upon using fat-suppression MRI protocols and the colocalization with the transplanted islets. In conclusion, direct islet imaging with high spatial and contrast resolution after labeling with GdHPDO3A is demonstrated, allowing visualization of kidney subcapsular mouse islet grafts and intrahepatic human islet xenografts.
Animals↗
High-relaxivity supramolecular aggregates containing peptides and Gd complexes as contrast agents in MRI.
Mixed supramolecular aggregates, obtained by assembling together two amphiphilic monomers (C(18)H(37))(2)NCO(CH(2))(2)CO(AdOO)(5)-G-CCK8 (AdOO is 8-amino-3,6-dioxaoctanoic acid, CCK8 is C-terminal octapeptide of cholecystokinin) and (C(18)H(37))(2)NCO(CH(2))(2)COLys(DTPAGlu)CONH(2) (DTPAGlu is N,N-bis[2-[bis(carboxyethyl)amino]ethyl]-L-glutamic acid), are characterized for their structural parameters by dynamic light scattering and for their relaxometric properties, in the absence and in the presence of 0.9 wt% NaCl. Two different aggregates (micelles and bilayer structures) are present in the absence of NaCl, while only bilayer structures are observed at physiological ionic strength. The presence of NaCl increases the ionic strength, promoting a decrease in the repulsions between the polar heads and among the aggregates in solution, thus supporting the formation of large-curvature aggregates such as bilayer structures like vesicles. In these conditions the closed, vesicular shape and the large size (hydrodynamic radius of about 300 A) of the aggregates allow a high number of paramagnetic gadolinium complexes and bioactive peptides to be accommodated on the inner and external surfaces . The presence of the salt causes a variation in the structural arrangement of the molecules and a partial rigidification of the assembled Gd(III) complexes on the surface vesicles, reducing their internal motions and giving an approximately 15% higher relaxivity value (r (1p) = 21.0 and 18.6 Mm(-1) s(-1) in the presence and in the absence of NaCl, respectively). The vesicles obtained, for the high relaxivity of each gadolidium complex and for the presence of a surface-exposed bioactive peptide, are very promising candidates as target-selective MRI contrast agents.
Chelating Agents↗
Substituent effects on Gd(III)-based MRI contrast agents: optimizing the stability and selectivity of the complex and the number of coordinated water molecules.
Hydroxypyridinone (HOPO)-based Gd(III) complexes have previously been shown to exhibit high relaxivity, especially at the high magnetic fields that are clinically relevant for present and future clinical use. This is due to more than one coordinated water molecule exchanging rapidly with bulk solvent. These complexes, however, present poor water solubility. Heteropodal complexes which include a terephthalamide (TAM) moiety maintain the high relaxivity characteristics of the HOPO family and have been functionalized with solubilizing substituents of various charges. The charge of the substituent significantly affects the stability of the Gd(III) complex, with the most stable complex presenting a neutral charge. The solubilizing substituent also moderately affects the affinity of the complex for physiological anions, with the highest affinity observed for the positively charged complex. In any case, only two anions, phosphate and oxalate, measureably bind the Gd(III) complex with weak affinities that are comparable to other q = 1 complexes and much weaker than DO3A, q = 2 based complexes. Furthermore, unlike poly(amino-carboxylate)-based complexes, HOPO-based Gd(III) complexes do not show any noticeable interaction with carbonates. The nature of the substituent can also favorably stabilize the coordination of a third water molecule on the Gd(III) center and lead to a nine-coordinate ground state. Such complexes that attain q = 3 incorporate a substituent beta to the terminal amide of the TAM podand that is a hydrogen-bond acceptor, suggesting that the third water molecule is coordinated to the metal center through a hydrogen-bond network. These substituents include alcohols, primary amines, and acids. Moreover, the coordination of a third water molecule has been achieved without destabilizing the complex.
Contrast Media↗
Magnetic resonance visualization of tumor angiogenesis by targeting neural cell adhesion molecules with the highly sensitive gadolinium-loaded apoferritin probe.
Tumor vessel imaging could be useful in identifying angiogenic blood vessels as well as being a potential predictive marker of antiangiogenic treatment response. We recently reported the expression of the neural cell adhesion molecule (NCAM) in the immature and tumor endothelial cell (TEC) lining vessels of human carcinomas. Exploiting an in vivo model of human tumor angiogenesis obtained by implantation of TEC in Matrigel in severe combined immunodeficiency mice, we aimed to image angiogenesis by detecting the expression of NCAM with magnetic resonance imaging. The imaging procedure consisted of (a) targeting NCAMs with a biotinylated derivative of C3d peptide that is known to have high affinity for these epitopes and (b) delivery of a streptavidin/gadolinium (Gd)-loaded apoferritin 1:1 adduct at the biotinylated target sites. The remarkable relaxation enhancement ability of the Gd-loaded apoferritin system allowed the visualization of TEC both in vitro and in vivo when organized in microvessels connected to the mouse vasculature. Gd-loaded apoferritin displayed good in vivo stability and tolerability. The procedure reported herein may be easily extended to the magnetic resonance visualization of other epitopes suitably targeted by proper biotinylated vectors.
Animals↗
A R2/R1 ratiometric procedure for a concentration-independent, pH-responsive, Gd(III)-based MRI agent.
A novel procedure for making the pH responsiveness of a Gd-based probe independent of its concentration has been developed. The method is based on a ratiometric approach and requires the measurement, at magnetic fields higher than 1 T, of the paramagnetic contribution to the longitudinal and the transverse NMR relaxation rates of water protons in the presence of a Gd-based probe with a rotational mobility in the nanosecond time scale. The method has been tested and validated in vitro by using a macromolecular adduct based on the polyornithine backbone, whose rotational mobility is pH dependent. The results reported in this communication improve substantially the diagnostic potential of MRI responsive agents.
Journal Article↗
On the role of the counter-ion in defining water structure and dynamics: order, structure and dynamics in hydrophilic and hydrophobic gadolinium salt complexes.
The crystal structures of the hydrated salts of [Gd.DOTAM]3+ and its more hydrophobic derivative [Gd.]3+, bearing 4 alpha-phenylethyl groups, (both Gd and Yb salts) are reported and compared. The nature of the anion determines the degree of ordering in the lattice and the extent of hydration. These effects are correlated with the results of 17O and 1H NMR measurements of water exchange dynamics in solution. With [Gd.DOTAM]3+, structural ordering or the extent of hydration in the hydrated lattice follows the sequence Cl->Br->I- and this order also defines the water exchange rate in solution: 7.3, 19.5, 33.3x10(4) s-1 (298 K), respectively. For [Gd.]3+ salts, the measured relaxivity is determined purely by the outer sphere term and the water exchange rate at 298 K is very similar (typically 1x10(4) s-1) for chloride, bromide, iodide, acetate, triflate and nitrate salts, notwithstanding the different nature and extent of hydration found in the crystalline lattice.
Journal Article↗
In vitro and in vivo magnetic resonance detection of tumor cells by targeting glutamine transporters with Gd-based probes.
The glutamine transporting system is up-regulated in tumor cells because cell proliferation requires the uptake of large quantities of glutamine. It has been found that the paramagnetic magnetic resonance imaging (MRI) reporter Gd-DOTAMA-C6-Gln, where the glutamine residue is covalently bound to the Gd chelate through a C6 spacer, accumulates in tumor cells both "in vitro" and "in vivo" experiments. The observation that the relaxivity of cellular pellets does not increase with the increase in the amounts of entrapped Gd chelate is taken as an indication that the internalization has occurred through receptor mediated endocytosis. The iv administration of Gd-DOTAMA-C6-Gln allowed the MRI visualization of tumor masses in A/J mice grafted with the murine neuroblastoma cell line Neuro-2a and in Her-2/neu transgenic mice developing multiple mammary carcinoma, respectively.
Animals↗
Fe(III)-templated Gd(III) self-assemblies-a new route toward macromolecular MRI contrast agents.
The self-assembly of supramolcular clusters of Gd(III) hydroxypyridinone complexes, templated by an Fe(III) terephthalamide center, is presented. The peripheral Gd(III) ions are each coordinated by two water molecules which exchange rapidly with the bulk solvent. These properties, along with the high rigidity of the supramolecules, efficiently increase the rotational correlation times of the cluster, resulting in high relaxivities at high magnetic fields and hence making these complexes good candidates for MRI contrast agents.
Contrast Media↗
Magnetic resonance imaging of tumor cells by targeting the amino acid transport system.
An early diagnosis of cancer is crucial in the battle against this disease and the in vivo visualization of tumors at cellular level is still the most challenging goal. In order to target tumor cells, we took into account their increased metabolism and amino acid nutrients or pseudo-nutrients, which are actively transported through the cell membrane, have been chosen as vectors for new MRI contrast agents. For this reason new gadolinium complexes conjugated to agmatine, arginine, and glutamine have been synthesized and studied.
Amino Acid Transport Systems↗
Tuning the coordination number of hydroxypyridonate-based gadolinium complexes: implications for MRI contrast agents.
Eight-coordinate hydroxypyridinone/terephthalamide GdIII complexes display high relaxivities due to their two inner sphere water molecules. This relaxivity can be further increased by functionalizing the terephthalamide moiety with an amine. A significant hydrogen bonding interaction between the amine and another water molecule close to the GdIII apparently facilitates its coordination on the open site of the metal. The resulting nine-coordinate complex has three inner sphere water molecules, while maintaining high stability and fast ligand exchange rates.
Contrast Media↗
Relaxometric and solution NMR structural studies on ditopic lanthanide(III) complexes of a phosphinate analogue of DOTA with a fast rate of water exchange.
A novel "ditopic" ligand containing two monophosphinate triacetate DOTA-like units linked by a thiourea bridge has been synthesized and its complexes with Ln3+ ions (Ln = Y, Eu, Gd, Dy) investigated by NMR spectroscopy and relaxometry. The presence of one water molecule in the first coordination sphere has been determined by the measurement of the dysprosium(III)-induced 17O NMR shifts. The 1H and 31P NMR spectra of the Eu(III) derivative indicate a higher abundance of the fast-exchanging twisted square antiprismatic (m) isomer than the isomeric square antiprismatic (M; m/M = 3:2) complex. The analysis of the 89Y and 13C T1 NMR relaxation times in the Gd(III)/Y(III) mixed complex have provided useful structural information. Values of ca. 6.3 and 8.2 A for the Gd...Y and Gd...C distances, respectively, have been estimated which indicate a rather compact solution structure. This result finds support in the value of the relaxivity whose increase (at 20 MHz and 298 K) on passing from the monomeric (5.7 s(-1) mM(-1)) to the ditopic complex (8.2 s(-1) mM(-1)) can be attributed to the doubling of the inner-sphere term following the doubling of the molecular size. The structural and dynamic relaxivity-controlling parameters were assessed by a simultaneous fitting of the variable temperature 17O NMR and 1H NMRD relaxometric data. The mean water residence lifetime (298tauM) has been found to be 53 ns, one of the shortest values reported for ditopic complexes. The reorientational correlation time is two times longer (298tauR = 183 ps) than the corresponding value of the parent monomeric Gd(III) complex, thus supporting the view of a limited degree of internal rotation. The possible influence of magnetic Gd-Gd coupling has been excluded by a comparison of the 1H NMRD profiles of the homodinuclear Gd(III)/Gd(III) and the heterodinuclear Gd(III)/Y(III) complexes.
Benzene Derivatives↗
Glycoconjugates of gadolinium complexes for MRI applications.
Examples of C-4 symmetric, medium MW conjugates incorporating 12 glucose or galactose groups linked via four dendritic wedges to a central Gd complex have been characterised; their enhanced relaxivity is interpreted in terms of effective motional coupling and large contributions from second sphere water molecules.
Gadolinium↗
Improved syntheses of bis(beta-cyclodextrin) derivatives, new carriers for gadolinium complexes.
In the last decade a number of reports have been published on the synthesis and characterization of bridged cyclodextrin dimers (bis-CDs) connected with linkers of different lengths and structures. These dimers, having two hydrophobic cavities in close proximity, display much higher binding affinities and molecular selectivities than parent CDs, forming stable supramolecular adducts. We describe new synthetic protocols for the preparation of bis(beta-CDs) bearing 2-2', 3-3' and 6-6' bridges. Some of the critical steps were carried out either under high-intensity ultrasound (US) or microwave (MW) irradiation. Bis(beta-CDs) containing 6-6' ureido- and thioureido-bridges were prepared in high yields by a MW-promoted aza-Wittig reaction using polymer-bound triphenylphosphine, while those containing 2,2' and 3,3' bridges were prepared from mono-alkenyl beta-CDs by the cross-metathesis reaction (homodimerization) in the presence of 2(nd)-generation Grubbs catalyst under sonochemical conditions. By these improved protocols CD dimers could be obtained in gram amounts to prepare stable adducts of bis-CDs with contrast agents (CAs) containing gadolinium(iii) chelates. In the case of Gd(iii) chleate "G-1" the inclusion complexes were found to be 2 to 3 orders of magnitude more stable than that formed by beta-CD (K(ass) = 4.3 x 10(4) M(-1)vs 8.0 x 10(2) M(-1)). Relaxivity increased as well by factors of 3 and 4, viz. from 9.1 mM(-1) s(-1) (beta-CD) to 27.7 and 35 mM(-1) s(-1).
Carbohydrate Conformation↗
New paramagnetic supramolecular adducts for MRI applications based on non-covalent interactions between Gd(III)-complexes and beta- or gamma-cyclodextrin units anchored to chitosan.
Gd(III) complexes are used as magnetic resonance imaging (MRI) contrast agents because they greatly enhance the relaxation rate of water protons of tissues in which they distribute, an effect that is much more marked if the paramagnetic complex is part of a macromolecular system. Furthermore applications in molecular imaging, require that as many units of contrast agent as possible be directed to the site of interest. To this end we synthesised a polymer made of chitosan functionalized with beta- and gamma-cyclodextrins (CDs) that is able to form high-affinity adducts with suitably functionalized Gd(III) complexes. beta- and gamma-CDs were first treated with maleic anhydride to afford 6-monosubstituted derivatives that reacted regioselectively with the amino groups of chitosan. Reaction times and yields were markedly improved by carrying out these reactions under high-intensity ultrasound or microwave irradiation. Compared to the CD monomers, beta- and gamma-CD-chitosan adducts show large increases both in terms of their binding affinity towards Gd(III) complexes and in relaxivity values and they appear promising carriers for the in vivo vehiculation of Gd(III) complexes.
Chitosan↗
Neural-cell adhesion molecule (NCAM) expression by immature and tumor-derived endothelial cells favors cell organization into capillary-like structures.
The neural cell adhesion molecule (NCAM) is widely expressed during embryogenesis, down-regulated in the course of differentiation to be re-expressed during progression of some tumors. We here found that renal tumor-derived endothelial cells (TEC) but not normal endothelial cells (HMEC) expressed NCAM. In TEC, NCAM expression was regulated by the renal embryonic transcription factor PAX2, as transfection with PAX2 antisense abrogated NCAM expression. NCAM stimulation with an agonistic synthetic NCAM peptide enhanced apoptosis resistance and increased ability of TEC to organize in vessel-like structures. The angiogenic effect of NCAM peptide was, at least in part, mediated by the association of NCAM and FGFR1. HMEC transiently acquired NCAM when organized in vessel-like structures after VEGF stimulation or when transfected with PAX2 gene. During the process of VEGF-induced endothelial differentiation of renal stem cells and of circulating endothelial progenitors, NCAM was transiently expressed to disappear at complete endothelial maturation. Targeting NCAM with a saporin-conjugated peptide induced a cytotoxic effect on TEC but not on HMEC. In conclusion, we identified a new role of NCAM in tumor neo-angiogenesis relevant for endothelial cell organization into capillary-like structures. In addition, we found that NCAM expression was associated with an immature phenotype of endothelial cells.
Angiogenesis Inducing Agents↗
Effect of the intracellular localization of a Gd-based imaging probe on the relaxation enhancement of water protons.
Gd-HPDO3A has been internalized into rat hepatocarcinoma cells in the cytoplasm (by electroporation) or in intracellular vesicles (by pinocytosis), respectively. In the former case, the observed relaxation rates are likely dependent upon the amount of internalized paramagnetic complex, whereas in the latter case the relaxation enhancement is "quenched" to a plateau value (about 3 s(-1)) when the entrapped amount of Gd-chelate is higher than 1 x 10(10) Gd/cell. The observed behavior has been accounted in terms of a theoretical treatment based on equations formally derived by Labadie et al. (J Magn Reson B 1994;105:99-102). On this basis, entrapment into intracellular vesicles has been treated as a three-site water exchange (extracellular/cytoplasm/vesicle compartments), whereas the cell pellets containing the paramagnetic agent spread out in the cytoplasm can be analyzed by a two-site exchange system.
Animals↗
Quantification of the expression level of integrin receptor alpha(v)beta3 in cell lines and MR imaging with antibody-coated iron oxide particles.
Targeted imaging requires site-specific accumulation of a contrast agent (CA), and the properties of that agent must be selected according to the abundance of the target to obtain a signal above the detection limit of the instrument. However, numerical estimates of receptors per cell are rarely found in the literature. Integrin receptors would be particularly promising targets because of their accessibility from the blood stream and expression on activated neovascular endothelial cells. We systematically estimated the number of integrin receptors of cell lines and primary cells by flow cytometry analysis. Since integrin receptors are heterodimeric molecules, and alpha(v) forms complexes with various beta subunits, the numbers of alpha(v) and beta(3) subunits are therefore dissimilar. The observed values are 3 . 10(3)-1.4 . 10(4)/cell for alpha(v), and 5.3 . 10(2)-1.1 . 10(4)/cell for beta(3). Despite the low number of exposed receptors, we show that up to single-cell MR visualization can be achieved with the use of iron oxide beads complexed with antibodies as CAs.
Antibodies, Monoclonal↗