New zirconium-89 and manganese-52 complexes for cell and liposome tracking with positron emission tomography

Abstract

Positron emission tomography (PET) can be used to understand the biodistribution, and potentially predict the therapeutic response, of new cellular therapies and liposomal nanomedicines. In particular, the PET tracer [ ⁸⁹Zr]Zr-oxine allows the radiolabelling of cells and liposomes without modification. However, little work has been performed looking at alternatives for this compound. Additionally, a new PET radioisotope, ⁵²Mn (t_1/2 = 5.6 days) has become available that may allow cell and liposome tracking twice as long than with ⁸⁹Zr. Hence, in this work, new methods for the direct labelling of cells and liposomes using ⁸⁹Zr and ⁵²Mn were explored. Firstly, new oxine analogues were investigated as new ionophores for liposome labelling with ⁸⁹Zr, and a potential improvement on [ ⁸⁹Zr]Zr-oxine. Secondly, we used [ ⁸⁹Zr]Zr-oxine to radiolabel and track a liposomal glucocorticoid nanomedicine (LC200) within an inflammation model. Secondly, the 52Mn analogue of [ ⁸⁹Zr]Zr-oxine ([ ⁵²Mn]Mn-oxine) was characterised and used for cell labelling and to track the liposomal nanomedicine, DOXIL, in vivo. Following that, we explored ⁵²Mn-labelled porphyrins as cell/liposome labelling agents. 
The ⁸⁹Zr-complexes of several oxine analogues were successfully synthesised and were all demonstrated to radiolabel liposomes, with varying efficiency. However, serum stability of the radiolabelled liposomes after 48 h showed that only one ligand, 5-fluoro8-hydroxyquinoline, had high stability (>95 %) – which was comparable to [ ⁸⁹Zr]Zroxine. The liposomal nanomedicine, LC200, was successfully radiolabelled without modification using [ ⁸⁹Zr]Zr-oxine (⁸⁹Zr-LC200). High uptake of ⁸⁹Zr-LC200 was seen in inflamed joints with the uptake correlating with the degree of inflammation. This correlation was not seen with unchelated ⁸⁹Zr. Additionally, a therapeutic study with LC200 in the same mouse model, showed a significant reduction in inflammation at the same time points of the PET imaging study. This indicated that PET imaging using this technique could be used to predict the therapeutic response of radiolabelled liposomes. [ ⁵²Mn]Mn-oxine was shown to have higher cell labelling than with unchelated ⁵²Mn in all cell lines tested – with similar cellular uptake to [ ⁸⁹Zr]Zr-oxine in γδT cells. However, cellular retention of ⁵²Mn was much lower after 24 h (ca. 20%), compared with [ ⁸⁹Zr]Zroxine. This was hypothesised to be due to the usage of manganese in biological processes. When tracking DOXIL labelled with [ ⁵²Mn]Mn-oxine in vivo using PET, high in vivo stability was observed up to 24 h. However, as tissue uptake of the liposomes occurred, release of ⁵²Mn was observed; with the biodistribution at 72 h matching that of unchelated ⁵²Mn. A microwave synthesis method for the ⁵²Mn radiolabelling of porphyrins was developed allowing >95 % radiochemical yield after 1 h at 165 oC. Three of the [ ⁵²Mn]Mn-porphyrins synthesised were capable of liposome labelling. Two of them were used to radiolabel MDA-MB-231 cells and showed lower labelling than with [ ⁵²Mn]Mnoxine. However, cellular retention of the [ ⁵²Mn]Mn-porphyrins was approximately double that of[ ⁵²Mn]Mn-oxine. In conclusion, alternative oxine ligands can allow liposome labelling with ⁸⁹Zr. PET imaging with [ ⁸⁹Zr]Zr-oxine can be used for the tracking of liposomal glucocorticoid nanomedicines to sites of inflammation and may allow the prediction of the therapeutic response. [ ⁵²Mn]Mn-oxine was shown to radiolabel cells and track liposomes in vivo. However, this technique is likely limited by the biological usage of manganese metal. Finally, [ ⁵²Mn]Mn-porphyrins show promise as liposome and cell labelling agents. However, further work is needed to improve these properties. This work has reported several new PET tracers which, with further development, may allow the direct labelling and imaging of cell and liposome therapies for longer time periods than currently available. Additionally, this work solidifies the utility of using [ ⁸⁹Zr]Zr-oxine for the validation of new liposomal nanomedicines, alongside its usage with cellular therapies.

Date of Award	1 Jun 2020
Original language	English
Awarding Institution	King's College London
Supervisor	Rafael T. M. de Rosales (Supervisor)