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A Gadolinium Complex Confined in Silica Nanoparticles as a Highly Efficient T1/T2 MRI Contrast Agent Article in Chemistry - A European Journal · May 2013 DOI: 10.1002/chem.201300635 · Source: PubMed
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DOI: 10.1002/chem.201300635
A Gadolinium Complex Confined in Silica Nanoparticles as a Highly Efficient T1/T2 MRI Contrast Agent Nicolas Wartenberg,[b] Pascal Fries,*[a] Olivier Raccurt,[b] Armel Guillermo,[c] Daniel Imbert,[a] and Marinella Mazzanti*[a]
MRI is a powerful, noninvasive diagnostic tool that provides high-resolution three-dimensional anatomical images of soft tissues consisting of 3D maps of longitudinal and transverse local proton relaxation rates R1 = 1/T1 and R2 = 1/ T2.[1] However, due to the relatively low sensitivity of these rates to natural variation between the tissues, paramagnetic compounds are administered in up to 40 % of clinical MRI examinations to improve the image contrast. Such compounds, named contrast agents (CAs), act as enhancers of the R1 and R2 values in the tissues in which they distribute. The efficacy of a CA is measured by its relaxivities r1 and r2 defined as ra = (RaRa0)/[CA] (a = 1, 2), in which Ra0 is the measured relaxation rate without the CA. The most common clinically approved CAs are monoaqua GdIII complexes that have r1 values of about 4 s1 mm1 at the imaging field of 1.5 T. While obviously successful for basic anatomical applications, these relaxivities are not sufficient to enable many applications that might be envisaged, notably for molecular or functional imaging. Much effort has been invested in the past years to obtain contrast agents with higher relaxivity to image biological targets.[2] The standard relaxivity theory[1a, b, 3] provides a reasonable guide for optimization at imaging fields above 1 T.[4] It was recognized early on that high relaxivity in the range of 0.5–1.5 T can be achieved by optimizing the inner-sphere (IS) relaxivity mechanism, that is, by slowing down the tumbling of Gd
[a] Dr. P. H. Fries, Dr. D. Imbert, Dr. M. Mazzanti Laboratoire de Reconnaissance Ionique et Chimie de Coordination, Service de Chimie Inorganique et Biologique (UMR E-3 CEA/UJF), CEA-Grenoble, INAC 17 rue des Martyrs, 38054 Grenoble cedex 9 (France) Fax: (+ 33) 4-38-78-50-90 E-mail:
[email protected] [email protected] [b] Dr. N. Wartenberg, Dr. O. Raccurt Laboratoire de Chimie et de Scurit des NanoMatriaux (DTNM/LITEN/DRT), CEA-Grenoble 17 rue des Martyrs, 38054 Grenoble cedex 9 (France) [c] Dr. A. Guillermo Groupe Polymres Conducteurs Ioniques Service Structure et Proprits d’Architectures Molculaires (UMR 5819, CEA-CNRS-UJF), CEA-Grenoble, INAC 17 rue des Martyrs, 38054 Grenoble cedex 9 (France) Supporting information for this article is available on the WWW under http://dx.doi.org/10.1002/chem.201300635.
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chelates endowed with a fast exchange of the coordinated water molecule. Thus, enhancement of the relaxivity on a per-Gd basis has been achieved by increasing the size of the ligand coordinating the metal ion, assembling the complexes together, or linking them to macromolecules, proteins, or nanoparticles.[2, 5] Nanoparticles (NPs) are increasingly studied because they provide versatile platforms for the delivery of a large number of paramagnetic complexes at the site of interest.[2b] More recently, a few examples of enhanced per-Gd relaxivities r1 were obtained through the noncovalent confinement of Gd complexes in nanosystems permeable to water, such as zeolites, apoferritine, silicon microparticles, and hydrogels.[2b, 6] The mechanisms leading to high relaxivity in these fascinating systems remain unclear, but theoretical analysis suggests that water permeable nanoparticles with confined complexes may provide a new route to high relaxivities. If water molecules can freely move in and out of the porous framework, while the nanopore boundaries restrict the motion of the complex and water molecules inside the NPs, this would lead to an increase of the local viscosity. In such a case both the outer-sphere (OS) and IS water molecules are locally slowed down and can contribute to the relaxivity.[7] From the previous theoretical discussion, silica NPs with confined Gd chelates are particularly attractive because they are nontoxic, with sizes smaller than 100 nm, meeting the requirements of in vivo applications.[13] Such NPs can be used to store and deliver drugs and their surface can be easily derivatized with biomolecules to yield targeting contrast agents.[5e, f, 8] However, to date nano- or microparticles of amorphous and mesoporous silica have not been used for the noncovalent confinement of Gd chelates. Here, we present monoaqua Gd chelates embedded in silica nanoparticles which show exceptionally high relaxivities r1 (up to 84 s1 mm1 at 0.8 T) and r2 (larger than 227 s1 mm1 above 11.5 T) at 298 K. These NPs containing water-accessible GdIII complexes were prepared by the simple noncovalent incorporation of the monoaqua complex [GdACHTUNGRE(ebpatcn)ACHTUNGRE(H2O)][9] into silica through a sol-gel method. The [GdACHTUNGRE(ebpatcn)ACHTUNGRE(H2O)] complex which displays a relaxivity of r1 = 4.7 s1 mm1 at 45 MHz and 298 K was selected in this study because the exchange rate kex298 = 86 106 s1 of the coordinated water molecule is one of the fastest reported in the literature for a neutral complex[10] with only one
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inner-sphere water molecule and, therefore, is optimized for the design of high relaxivity macromolecular MRI contrast agents. Moreover [GdACHTUNGRE(ebpatcn)ACHTUNGRE(H2O)] has a reasonably high stability (pGd = 13.1) with respect to ligand dissociation providing an additional barrier capable of preventing the release of the toxic aqua ion from the NPs in the absence of covalent binding to the silica matrix. The high stability of the Gd NPs in H2O over a pH range of 7–8 with respect to metal leakage over a month period has been confirmed by inductively coupled plasma mass spectrometry (ICPMS) and by optical and radiotracing studies performed on the analogous EuIII and TbIII ebpatcn complexes.[11] The value of the luminescence emission lifetime measured for the Tb NPs in H2O (1.86(3) ms) is slightly longer than the value measured for the monoaqua [TbACHTUNGRE(ebpatcn)ACHTUNGRE(H2O)] complex in water (1.60(2) ms, which indicates the presence of only one coordinated water molecule in the encapsulated complex. Gd–ebpatcn embedded silica NPs of different sizes, NPGd25 (diameter = 25 5 nm), NPGd50 (diameter = 50 5 nm), and NPGd60 (diameter = 60 5 nm), were prepared by a reverse microemulsion procedure,[12] such that the ammonia-catalyzed hydrolysis of tetraethyl orthosilicate (TEOS) takes place in water-in-oil droplets in the presence of the [GdACHTUNGRE(ebpatcn)ACHTUNGRE(H2O)] complex. TEM measurements (Figure 1) confirmed that the obtained NPs are spherical and uniform in size.
Figure 1. TEM image of NPGd60.
The used procedure leads to a nanospherical structure with tunable narrow pore channels.[13] Moreover, it results in a random distribution of the GdIII complexes in the silica matrix through electrostatic and hydrogen-bonding interactions. Then, the complexes do not mutually hinder the accessibility of the water molecules to the Gd3 + ions and their mobility is restricted by the confinement in the silica matrix.
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Moreover, the water molecules inside the NPs can be slowed down in the pores because they bind to the slowly moving complexes or undergo the geometrical restraints of the proximate pore walls. These structural and dynamic properties result in remarkably high values of the water proton relaxivities r1 and r2 due to the incorporated complex [GdACHTUNGRE(ebpatcn)ACHTUNGRE(H2O)]. These values are easily about 20 and 50 times larger than those of the free complex for r1 and r2, respectively. The water proton relaxivities due to NPGd25 and NPGd50 were measured at 25 8C over a very large range of resonance frequencies nI (from 5 to 800 MHz for r1). They are shown on a per mm of GdIII basis in Figure 2 and addi-
Figure 2. Water proton relaxivities of r1 (left) and r2 (right) for NPGd25 and NPGd50 at different frequencies.
tional information is reported in the Supporting Information. The relaxivity values measured for NPGd25 are significantly higher than those measured for the larger NPGd50. Near 35 MHz, the r1 profiles for the large and small NPs show high local maxima of 33 and 84 s1 mm1, respectively. The r1 values at 50 MHz are still 19 and 56 s1 mm1, that is, 4 and 12 times larger than the value 4.7 s1 mm1 of the free complex. The relaxivity r2, measured above 20 MHz, rises monotonously. The rate of increase is first slow, so that r2 r1 and the NPs can be used as T1 CAs,[1a, b] and then more rapid. Above 500 MHz, r2 exceeds 150 s1 mm1 in the case of the large NPs and even 227 s1 mm1 in the case of the small ones. Here, using the GdIII concentration in the solution measured either from the NMR paramagnetic susceptibility shift[14] or by ICPMS,[11] the NPGd50 were shown to contain about 1,000 [GdACHTUNGRE(ebpatcn)ACHTUNGRE(H2O)] complexes (see the Supporting Information) so that their relaxivities on a per NP basis are r1ffi1,9 104 s1·ACHTUNGRE(mm of NP)1 at 50 MHz and r2 > 1,5 105 s1·ACHTUNGRE(mm of NP)1 for above 500 MHz. Larger NPs are obtained when a larger number of complexes are incorporated in the NP (see the Supporting Information).
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For instance, NPGd60 (diameter = 60 5 nm) contains about 3,000 Gd chelates, whereas the incorporation of 1000 chelates produces NPGd50 (diameter 50 5 nm). Accordingly, when passing from NPGd50 to NPGd60, the relaxivities r1 per NP increase from 3.3 104 to 7,2 104 s1·ACHTUNGRE(mm of NP)1 at 35 MHz, near their local maxima, and the relaxivities r2 rise from 1.4 105 to 2.7 105 s1·ACHTUNGRE(mm of NP)1 above 400 MHz. Note that the per-Gd r1 and r2 decrease in NPGd60, probably due to a lower accessibility of the water molecules to the Gd3 + ions in such overloaded systems. Finally, when the temperature increases from 25 to 37 8C, the changes of r1 are generally modest, smaller than about 10 %. As predicted by all theories, r2 is larger than r1 at any frequency. The relaxivities r1 and r2 measured for the NPGd25 nanoparticles are among the highest ones reported to date on a per-Gd basis, rendering these systems suitable for the development of dual T1/T2 MRI contrast agents. They are notably higher than the values reported for any monoaqua Gd chelates anchored to mesoporous silica nanoparticles through a covalent siloxane linkage[15] (r1 per-Gd comprised between 14 and 38 s1 mm1 at 20 MHz), for which the increase of the per-Gd relaxivity is assigned to a decrease in the tumbling rate of the Gd chelate due to the increased size of the system. This suggests the presence of a different mechanism of paramagnetic relaxation arising from the hindered motion of the outer and inner sphere water molecules associated with the water-permeable nature of the silica matrix. Very recently, high relaxivities r1 have been obtained at high field (39.26 s1 mm1 at 125 MHz and 33.57 s1 mm1 at 300 MHz)[16] by covalently anchoring Gd chelates in the core of mesoporous silica nanoparticles. As illustrated by several examples,[1] the standard relaxivity theory does not predict that r1 will significantly increase in these systems at lower field since it already has large values of similar sizes at two quite different high fields. In the present NPs, the noncovalent incorporation allows the possible free rotation of the complex in the silica matrix. Then, both the rotational IS motion of the coordinated water molecules and the translational OS motion of the free water molecules can be optimized by confinement to give optimal relaxivity values at the desired imaging fields. At imaging fields B0 1 T, the relaxivity theory[1a, b, 3, 17] stresses the importance of the correlation times tr and t of the rotational and translational dynamics of water with respect to the metal ion. This indicates that r1 has a local maximum if tr or t are long enough and that r2 increases proportionally to both correlation times. According to this theory, the steep decrease of r1 after its local maximum can be attributed to long correlation times tr and/or t of at least a few ns. By contrast, the monotonous increase of r1 as nI goes to zero is a peculiarity of some nanosystems,[6c, d] which has not yet been explained in detail, but is probably due to a restricted motion of the Gd chelate and of the water molecules combined to the water permeable structure of the silica NPs.[20] The high-frequency values of r2, which are more than twice as large as the r1 maximum, cannot either
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be explained by the standard relaxivity theory. They should likely be ascribed to the combined effects of the inhomogeneities of the magnetic field created by the magnetized NPs and the exchange of water between these NPs and the bulk.[6a] This interpretation is further supported by the relaxivities measured at nI = 50, 500, and 800 MHz for the large nanoparticles (NPDy50, diameter = 50 nm) doped with [DyACHTUNGRE(ebpatcn)ACHTUNGRE(H2O)] complexes. Indeed, the relaxivity r1 has a nearly constant, small value of about 0.5 s1 mm1, characteristic of the very short electronic relaxation time of the Dy3 + ion,[14, 18] whereas r2 rises sharply with frequency as the Dy3 + Curie spin,[18] since it is 5.4, 137, and 182 s1 mm1 at the above frequencies, respectively. The versatile incorporation of lanthanide complexes in silica NPs is particularly well suited for the development of multimodal probes. Notably luminescent complexes of visible and/or near-IR LnIII emitters[19] can be noncovalently incorporated in silica NPs.[20] To obtain the multimodal MRI/ optical nanoparticles, during the preparation of NPGd25, 5 % of [GdACHTUNGRE(ebpatcn)ACHTUNGRE(H2O)] is replaced by [Yb(thqtcn-SO3)ACHTUNGRE(H2O]3 for which a strong near-IR emission has been reported.[21] These NPs show high r1 and r2 relaxivities together with a detectable luminescence in the near-IR region even at this low YbIII concentration (see Figure S4 in the Supporting Information). In conclusion, the noncovalent incorporation of the monoaqua complex [GdACHTUNGRE(ebpatcn)ACHTUNGRE(H2O)] into silica nanoparticles by a sol-gel method is a new example of nanosized contrast agents with very high per-Gd relaxivities r1 and r2. The encapsulation of complexes is very simple and versatile, allowing the combination of LnIII ions with different imaging modalities. The easy tuning of the size of the NPs and potentially of their pore structure provides a tool to gain further insight into the underlying relaxation mechanisms giving rise to the remarkable contrast efficiency of Gd chelates confined in these water permeable nanosized systems and for the optimization of the system towards its highest possible relaxivities.
Acknowledgements Assistance from Dr. A. Favier and financial support from the TGE RMN THC and from the “French Agence Nationale de la Recherche”, grants ANR-10-P2N-001 and Labex Arcane, ANR-11-LABX-003-01 are gratefully acknowledged.
Keywords: contrast agents · gadolinium · lanthanides · luminescence · nanoparticles
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Received: February 18, 2013 Published online: April 18, 2013
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