Investigation of Zr(IV) and 89Zr(IV) complexation with hydroxamates: progress towards designing a better chelator than desferrioxamine B for immuno-PET imaging.
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ABSTRACT:
SUBMITTER: Guerard F
PROVIDER: S-EPMC3538910 | biostudies-literature | 2013 Feb
REPOSITORIES: biostudies-literature
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Investigation of Zr(IV) and 89Zr(IV) complexation with hydroxamates: progress towards designing a better chelator than desferrioxamine B for immuno-PET imaging.
Guérard François F Lee Yong-Sok YS Tripier Raphaël R Szajek Lawrence P LP Deschamps Jeffrey R JR Brechbiel Martin W MW
Chemical communications (Cambridge, England) 20121219 10
Single crystal X-ray diffraction shows that Zr(iv) forms an octa-coordinated complex with 4 bidentate hydroxamates whose solution structures were investigated by utilizing density functional theory at the level of B3LYP/DGDZVP. Stability constants obtained by potentiometry were in accordance with the tendency observed when radiolabeling with (89)Zr. ...[more]
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Project description:Three hydroxypyridinone (HOPO) positional isomers - 1,2-HOPO (L1H) and its water soluble analogue (L1'H), 3,2-HOPO (L2H) and 3,4-HOPO (L3H) have been investigated for the complexation of Zr(iv). Potentiometric and UV-Vis spectrometric studies show a higher thermodynamic stability for the formation of Zr(L1')4 in comparison with Zr(L2)4 and Zr(L3)4 as well as a higher kinetic inertness in competition studies with EDTA or Fe3+ at a radiotracer concentration with 89Zr. Besides the low pKa of L1H or L1'H (pKa = 5.01) in comparison with L2H and L3H (pKa = 8.83 and 9.55, respectively), the higher stability of Zr(L1')4 can be attributed in part to the presence of the amide group next to the chelating oxygen that induces intramolecular H-bond and amide/? interactions that were observed by X-ray crystallography and confirmed by quantum chemical calculations. The data presented here indicate that the 1,2-HOPO L1' exhibits the best characteristics for Zr(iv) complexation. However, 3,2-HOPO and 3,4-HOPO patterns, if appropriately tuned, for instance with the addition of an amide group as in the 1,2-HOPO ligand, may also become interesting alternatives for the design of Zr(iv) chelators with improved characteristics for applications in nuclear imaging with 89Zr.
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Project description:PurposeAll clinical 89Zr-immuno-PET studies are currently performed with the chelator desferrioxamine (DFO). This chelator provides hexadentate coordination to zirconium, leaving two coordination sites available for coordination with, e.g., water molecules, which are relatively labile ligands. The unsaturated coordination of DFO to zirconium has been suggested to result in impaired stability of the complex in vivo and consequently in unwanted bone uptake of 89Zr. Aiming at clinical improvements, we report here on a bifunctional isothiocyanate variant of the octadentate chelator DFO* and the in vitro and in vivo comparison of its 89Zr-DFO*-mAb complex with 89Zr-DFO-mAb.MethodsThe bifunctional chelator DFO*-pPhe-NCS was prepared from previously reported DFO* and p-phenylenediisothiocyanate. Subsequently, trastuzumab was conjugated with either DFO*-pPhe-NCS or commercial DFO-pPhe-NCS and radiolabeled with Zr-89 according to published procedures. In vitro stability experiments were carried out in saline, a histidine/sucrose buffer, and blood serum. The in vivo performance of the chelators was compared in N87 tumor-bearing mice by biodistribution studies and PET imaging.ResultsIn 0.9 % NaCl 89Zr-DFO*-trastuzumab was more stable than 89Zr-DFO-trastuzumab; after 72 h incubation at 2-8 °C 95 % and 58 % intact tracer were left, respectively, while in a histidine-sucrose buffer no difference was observed, both products were???92 % intact. In vivo uptake at 144 h post injection (p.i.) in tumors, blood, and most normal organs was similar for both conjugates, except for skin, liver, spleen, ileum, and bone. Tumor uptake was 32.59?±?11.95 and 29.06?±?8.66 % ID/g for 89Zr-DFO*-trastuzumab and 89Zr-DFO-trastuzumab, respectively. The bone uptake was significantly lower for 89Zr-DFO*-trastuzumab compared to 89Zr-DFO-trastuzumab. At 144 h p.i. for 89Zr-DFO*-trastuzumab and 89Zr-DFO-trastuzumab, the uptake in sternum was 0.92?±?0.16 and 3.33?±?0.32 % ID/g, in femur 0.78?±?0.11 and 3.85,?±?0.80 and in knee 1.38?±?0.23 and 8.20?±?2.94 % ID/g, respectively. The uptake in bone decreased from 24 h to 144 h p.i. about two fold for the DFO* conjugate, while it increased about two fold for the DFO conjugate.ConclusionsZr-DFO*-trastuzumab showed superior in vitro stability and in vivo performance when compared to 89Zr-DFO-trastuzumab. This makes the new octadentate DFO* chelator a candidate successor of DFO for future clinical 89Zr-immuno-PET.
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Project description:Herein we report a new high-denticity chelator based on the iron siderophore desferrioxamine (DFO). Our new chelator-DFO2-is acyclic and was designed and synthesized with the purpose of improving the coordination chemistry and radiolabeling performance with radioactive zirconium-89. The radionuclide zirconium-89 ([89Zr]Zr4+) has found wide use for positron emission tomography (PET) imaging when it is coupled with proteins, antibodies, and nanoparticles. DFO2 has a potential coordination number of 12, which uniquely positions this chelator for binding large, high-valent, and oxophilic metal ions. Following synthesis of the DFO2 chelator and the [natZr]Zr-(DFO2) complex we performed density functional theory calculations to study its coordination sphere, followed by zirconium-89 radiolabeling experiments for comparisons with the "gold standard" chelator DFO. DFO (CN 6) can coordinate with zirconium in a hexadentate fashion, leaving two open coordination sites where water is thought to coordinate (total CN 8). DFO2 (potential CN 12, dodecadentate) can saturate the coordination sphere of zirconium with four hydroxamate groups (CN 8), with no room left for water to directly coordinate, and only binds a single atom of zirconium per chelate. Following quantitative radiolabeling with zirconium-89, the preformed [89Zr]Zr-(DFO) and [89Zr]Zr-(DFO2) radiometal-chelate complexes were subjected to a battery of in vitro stability challenges, including human blood serum, apo-transferrin, serum albumin, iron, hydroxyapatite, and EDTA. One objective of these stability challenges was to determine if the increased denticity of DFO2 over that of DFO imparted improved complex stability, and another was to determine which of these assays is most relevant to perform with future chelators. In all of the assays DFO2 showed superior stability with zirconium-89, except for the iron challenge, where both DFO2 and DFO were identical. Substantial differences in stability were observed for human blood serum using a precipitation method of analysis, apo-transferrin, hydroxyapatite, and EDTA challenges. These results suggest that DFO2 is a promising next-generation scaffold for zirconium-89 chelators and holds promise for radiochemistry with even larger radionuclides, which we anticipate will expand the utility of DFO2 into theranostic applications.
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Project description:Zirconium-89 has an ideal half-life for use in antibody-based PET imaging; however, when used with the chelator DFO, there is an accumulation of radioactivity in the bone, suggesting that the (89)Zr(4+) cation is being released in vivo. Therefore, a more robust chelator for (89)Zr could reduce the in vivo release and the dose to nontarget tissues. Evaluation of the ligand 3,4,3-(LI-1,2-HOPO) demonstrated efficient binding of (89)Zr(4+) and high stability; therefore, we developed a bifunctional derivative, p-SCN-Bn-HOPO, for conjugation to an antibody. A Zr-HOPO crystal structure was obtained showing that the Zr is fully coordinated by the octadentate HOPO ligand, as expected, forming a stable complex. p-SCN-Bn-HOPO was synthesized through a novel pathway. Both p-SCN-Bn-HOPO and p-SCN-Bn-DFO were conjugated to trastuzumab and radiolabeled with (89)Zr. Both complexes labeled efficiently and achieved specific activities of approximately 2 mCi/mg. PET imaging studies in nude mice with BT474 tumors (n = 4) showed good tumor uptake for both compounds, but with a marked decrease in bone uptake for the (89)Zr-HOPO-trastuzumab images. Biodistribution data confirmed the lower bone activity, measuring 17.0%ID/g in the bone at 336 h for (89)Zr-DFO-trastuzumab while (89)Zr-HOPO-trastuzumab only had 2.4%ID/g. We successfully synthesized p-SCN-Bn-HOPO, a bifunctional derivative of 3,4,3-(LI-1,2-HOPO) as a potential chelator for (89)Zr. In vivo studies demonstrate the successful use of (89)Zr-HOPO-trastuzumab to image BT474 breast cancer with low background, good tumor to organ contrast, and, importantly, very low bone uptake. The reduced bone uptake seen with (89)Zr-HOPO-trastuzumab suggests superior stability of the (89)Zr-HOPO complex.
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Project description:Due to its long half-life (78 h) and decay properties (77% electron capture, 23% ?(+), Emax = 897 keV, Eav = 397 keV, E? = 909 keV, I? = 100%) (89)Zr is an appealing radionuclide for immunoPET imaging with whole IgG antibodies. Derivatives of the siderophore desferrioxamine-B (H3DFO) are the most widely used bifunctional chelators for coordination of (89)Zr(4+) because the radiolabeling of the resulting immunoconjugates is rapid under mild conditions. (89)Zr-DFO complexes are reportedly stable in vitro but there is evidence that (89)Zr(4+) is released in vivo, and subsequently taken up by the skeleton. We have evaluated a novel tripodal tris(hydroxypyridinone) chelator, H3CP256 and its bifunctional maleimide derivative, H3YM103, for coordination of Zr(4+) and compared the NMR spectra, and the (89)Zr(4+) radiolabeling, antibody conjugation, serum stability and in vivo distribution of radiolabelled immunoconjugates with those of H3DFO and its analogues. H3CP256 coordinates (89)Zr(4+) at carrier-free concentrations forming [(89)Zr(CP256)](+). Both H3DFO and H3CP256 were efficiently radiolabelled using [(89)Zr(C2O4)4](4-) at ambient temperature in quantitative yield at pH 6-7 at millimolar concentrations of chelator. Competition experiments demonstrate that (89)Zr(4+) dissociates from [(89)Zr(DFO)](+) in the presence of one equivalent of H3CP256 (relative to H3DFO) at pH 6-7, resulting largely in [(89)Zr(CP256)](+). To assess the stability of H3DFO and H3YM103 immunoconjugates radiolabelled with (89)Zr, maleimide derivatives of the chelators were conjugated to the monoclonal antibody trastuzumab via reduced cysteine side chains. Both immunoconjugates were labelled with (89)Zr(4+) in >98% yield at high specific activities and the labeled immunoconjugates were stable in serum with respect to dissociation of the radiometal. In vivo studies in mice indicate that (89)Zr(4+) dissociates from YM103-trastuzumab with significant amounts of activity becoming associated with bones and joints (25.88 ± 0.58% ID g(-1) 7 days post-injection). In contrast, <8% ID g(-1) of (89)Zr activity becomes associated with bone in animals administered (89)Zr-DFO-trastuzumab over the course of 7 days. The tris(hydroxypyridinone) chelator, H3CP256, coordinates (89)Zr(4+) rapidly under mild conditions, but the (89)Zr-labelled immunoconjugate, (89)Zr-YM103-trastuzumab was observed to release appreciable amounts of (89)Zr(4+)in vivo, demonstrating inferior stability when compared with (89)Zr-DFO-trastuzumab. The significantly lower in vivo stability is likely to be a result of lower kinetic stability of the Zr(4+) tris(hydroxypyridinone complex) relative to that of DFO and its derivatives.
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