Conjugation to Biocompatible Dendrimers Increases Lanthanide T2 Relaxivity of Hydroxypyridinone (HOPO) Complexes for Magnetic Resonance Imaging (MRI)

Conjugation to Biocompatible Dendrimers Increases Lanthanide T 2 Relaxivity of Hydroxypyridinone (HOPO) Complexes for Magnetic Resonance Imaging (MRI) Piper J. Klemm, [a] William C. Floyd III, [a] Christopher M. Andolina, [b] Jean M. J. Frechet [a][b] and Kenneth N. Raymond* [a],[b] Keywords: MRI Probes / Lanthanides / Dendrimers / T 2 Contrast Agent / Conjugation Magnetic resonance imaging (MRI) contrast agents represent a worldwide billion-dollar market annually. While T 1 relaxivity enhancement contrast agents receive greater attention and a significantly larger market share, the commercial potential for T 2 relaxivity enhancing contrast agents remains a viable diagnostic option due to their increased relaxivity at high field strengths. Improving the contrast and biocompatibility of T 2 MRI probes may enable new diagnostic prospects for MRI. Paramagnetic lanthanides have the potential to decrease T 1 and T 2 proton relaxation times, but are not commercially used in MRI diagnostics as T 2 agents. In this article, oxygen donor chelates (hydroxypyridinone, HOPO, and terephthalamide, TAM) of various lanthanides are demonstrated as biocompatible macromolecular dendrimer conjugates for the development of T 2 MRI probes. These conjugates have relaxivities up to 374 m M-1 s - per dendrimer, high bioavailability, and low in vitro toxicity. class of MRI-CAs. The development of T 2 targeted imaging for MRI will require higher relaxivities than are currently attained if this approach is to be realized for diagnostic medicine. 3 High relaxivity at physiological conditions and 60 – 100 MHz, combined with adequate synthetic functionality and versatility, would allow conjugation of various targeting molecules such as antibodies, proteins or targeting ligands and enable the development of a new generation of MRI agents capable of site selective delivery. 1 Currently, T 2 commercial contrast agents are primarily superparamagnetic iron oxide nanoparticles (SPION). Current SPION’s are unsuitable for targeted imaging and bioresponsive mechanisms. In addition, SPION’s have not responded to investigations into advanced applications such as tumor targeting and the tracking of stem cells. 1 In this laboratory, Gd III complexes have been developed only for enhanced T 1 relaxivities compared to current commercial agents. 5,6 These include Gd-TREN-bis(1-methyl-3,2-HOPO)- TAM-ethylamine (1), Gd-TREN-bis(1-methyl-3,2-HOPO)-TAM- N2 (2), Gd-TREN-bis(1-methyl-3,2-HOPO)-TAM-N3 (3), and Gd- TREN-bis(1,2-HOPO)-TAM-ethylamine (4) (Figure 1). Compared to commercial agents such as Gd-DOTA (Gd-1,4,7,10- tetraazacyclododecane-1,4,7,10-tetraacetic acid) and Gd-DTPA (Gd-diethylenetriaminepentaacetic acid), the Gd-HOPO (Gd-tris- hydroxypyridinone) hexadentate oxygen donor chelators used in our studies increase relaxivity through fast water exchange via an associative water exchange mechanism, large numbers of inner- sphere water molecules (q), and the ability to conjugate to a macromolecule to decrease molecular tumbling time. 7-10 As these parameters increase both r 2 and r 1 , this research provides Ln III complexes desirable T 2 relativities as well. For r 1 , (1-Me)-3,2- HOPO chelators have outperformed the corresponding (1,2)-HOPO chelators in every comparison study. 5-10 To investigate the feasibility of using alternate lanthanides for T 2 imaging, Yb III and Dy III based agents, Dy-TREN-bis(1-methyl-3,2-HOPO)-TAM- ethylamine Yb-TREN-bis(1-methyl-3,2-HOPO)-TAM- ethylamine (6), Dy-TREN-bis(1-methyl-3,2-HOPO)-TAM-N2 (7), Yb-TREN-bis(1-methyl-3,2-HOPO)-TAM-N2 (8), Dy-TREN- bis(1-methyl-3,2-HOPO)-TAM-N3 (9), and Yb-TREN-bis(1- methyl-3,2-HOPO)-TAM-N3 (10), were also synthesized and evaluated as dendrimer conjugates. Yb III has potential as a bimodal imaging agent as a T 2 MRI contrast agent and a near infrared probe in biological systems. Dy III has been shown to be a prospective cerebral perfusion agent, superior to Gd III and therefore should be explored as a specific MRI diagnostic analysis. In addition to varying the lanthanide used in these complexes, they were also varied by the amine linking appendage used to attach the complex [a] [b] Department of Chemistry, University of California, Berkeley, Berkeley, California 94720 United States Lawrence Berkeley National Laboratory, Berkeley, California 94720 United States Fax: 510-486-5283 E-mail: raymond@socrates.berkeley.edu Supporting information for this article is available on the WWW under http://www.eurjic.org/ or from the author. Introduction Magnetic Resonance Imaging (MRI) is the leading soft tissue diagnostic imaging technique due to its unparalleled ability to generate high resolution three dimensional in vivo images. 1,2 In addition, MRI, considered a noninvasive technique, uses biologically benign 1-4 radio frequencies rather than potentially harmful X-rays such as those employed for CAT (Computerized Axial Tomography) scans. Current Gd III contrast agents (CA) based on longitudinal spin-lattice relaxation time, T 1 , dominate use in the clinic. 1,2 Recently, more emphasis is being placed on the necessity of developing CAs with greater relaxivities to reduce the quantity of Gd III -CAs administered for MRI diagnostics, resulting from concerns that Gd III is not safe for all patients, the threat of limited access to large quantities of Gd III , and environmental accumulation of Gd III . CAs based on the transverse spin-spin relaxation time, T 2 , could allow for the development of targeted imaging and alternative diagnostic agents. 1-4 All T 1 commercial contrast agents suffer a decrease in efficiency at higher field strengths, 2 which is a liability as clinical instruments increase in magnetic field strength to enhance inherent instrument contrast. Many MRI’s in the clinic have reached 60 to 125 MHz to increase resolution, but at the cost of diminishing the efficacy of the contrast agent injected. 2,3 To counteract the increase in field strength, increasing dosage is required to maintain current levels of contrast, however, gram quantities of T 1 contrast agents must already be injected to overcome low sensitivity. Therefore, alternative options need to be explored and developed, both improving T 1 relaxivity and providing an alternative such as an effective T 2 CA. 2 In contrast to T 1 imaging agents, the T 2 agents exhibit a direct correlation between increasing field strength and relaxivity, 1 and as a result they improve contrast at the larger field strengths now seen in the clinic. This relationship, combined with increasingly powerful magnets used for MRIs, suggests that over time T 2 agents may emerge as a more thoroughly studied and highly developed