Iron-based T1 MRI contrast agent for MR-guided drug delivery from temperature sensitive liposomes
Esther Kneepkens1, Adriana Fernandes2, Klaas Nicolay3, and Holger Grüll3,4

1Biomedical NMR, Biomedical Engineering, Eindhoven University of Technology, Eindhoven, Netherlands, 2Universidade de Lisboa, Lisbon, Portugal, 3Eindhoven University of Technology, Eindhoven, Netherlands, 4Philips Research, Eindhoven, Netherlands

Synopsis

The aim of this study was to investigate the potential of Fe(III) N-succinyl deferoxamine (Fe-SDFO) as a safe T1 contrast agent for encapsulation in temperature sensitive liposomes (TSLs) in order to visualize drug release from TSLs when using Magnetic Resonance-guided High Intensity Focused Ultrasound (MR-HIFU). Two TSLs were developed that contained either Fe-SDFO or doxorubicin. Both TSLs showed suitable release and stability characteristics in vitro. An in vivo proof-of-concept study was carried out in tumor-bearing rats treated with MR-HIFU. Treated tumors showed an increase in R1 and future work aims to correlate the R1 change with tumor drug concentrations.

Introduction

For malignant lesions, the combination of a local temperature increase inflicted by High Intensity Focused Ultrasound (HIFU) and temperature sensitive liposomes (TSL) enables a triggered drug release confined to the disease site, thereby limiting side effects. Co-release of an MR contrast agent (CA) allows for indirect imaging of the drug release with MR, providing an indirect assessment of the HIFU treatment.1 However, liposome encapsulation of commonly used Gd-based MR-CA leads to prolonged retention times in the liver and spleen, increasing the risk on nephrogenic systemic fibrosis.2 In this study an Fe-deferoxamine derivative is proposed as a safe alternative T1- CA for TSL encapsulation.

Materials and Methods

In vitro: TSLs were prepared using DPPC, DSPC, cholesterol, DPPE-PEG2000 (61:14:15:3 molar ratio). They were either actively loaded with doxorubicin (dox) or passively loaded with Fe(III) N-succinyl Deferoxamine (Fe-SDFO, synthetized by SyMO-Chem, the Netherlands)3. TSLs were characterized by Dynamic Light Scattering, Differential Scanning Calorimetry and Inductive Coupled Plasma-Optical Emission Spectroscopy. Release assays were carried out in Fetal Bovine Serum (FBS) at 37, 40 and 42˚C either fluorometrically (dox) or by measuring the longitudinal relaxivity r1 at 1.41T (Fe-SDFO).

In vivo: 9L glioma tumors were inoculated subcutaneously on the hind limb of Fisher 344 rats. The tumor-bearing animals were placed into a rat MR-HIFU setup in a clinical Philips 3T Sonalleve® MR-HIFU system4. Animals were divided into a MR-HIFU treatment group (n = 4) and a non-treated control group (n = 3). All animals were injected i.v. with a mixture of TSLs encapsulating dox and Fe-SDFO (5 mg/kg and 79 μg/kg respectively) just prior to MR-HIFU or control treatment. The MR-HIFU treatment consisted of two sonication periods of 10-15 minutes each (acoustic power 9 – 15 W, acoustic frequency 1.44 MHz) to reach and maintain a tumor temperature of 42˚C. Tumor temperature changes were monitored by proton resonance frequency shift MR thermometry. T1 maps were acquired before and after the two hyperthermia treatments or at corresponding time points for the control animals, using a single slice Look Locker sequence (Figure 1).

Results

The characteristics of both dox TSLs and Fe-SDFO TSLs are summarized in Figure 2 and were considered comparable and suitable for in vivo application. Both TSLs showed a fast release at 42˚C (Figure 3). 90 ± 4 % of the dox was released within 2 min, compared to a release of 80 ± 4% of the Fe-SDFO. At 37 ˚C, the dox TSLs showed no release over 1.5 h, while the Fe-SDFO TSLs showed a slight release of 15 ± 11%. At 1.41T, unheated Fe-SDFO TSLs displayed an longitudinal relaxation rate r1 of 0.80 ± 0.01 mM-1s-1. After 60 min at 42 ˚C, r1 increased to 1.35 ± 0.02 mM-1s-1.

T1 maps obtained before and at various time points after TSL injection are shown in Figure 4. All treated tumors showed a contrast change upon the heat treatment (Figure 4, ΔR1 = 0.18 ± 0.10 s-1). In the non-treated tumors, the average change in R1 change was much smaller (Figure 4, ΔR1 = 0.028 ± 0.007 s-1). Strikingly, the R1 change across the treated tumors was homogeneous in some (animal 2, animal 4, Figure 4), while very inhomogeneous in others (animals 1 and 3, Figure 4), suggesting untreated tumor areas in the latter.

Discussion and Conclusion

Dox-loaded TSLs and Fe-SDFO loaded TSLs were characterized in vitro, showing the potential of this combined system for MR-guided triggered drug release, even though the r1 of the proposed CA is around a factor 3 lower than that of more commonly used Gd-based CA.1 An in vivo proof-of-concept study was conducted to assess the feasibility of monitoring drug release using the newly designed drug and CA loaded TSL systems. Treated tumors showed an increase in R1 while the R1 of the untreated control tumors remained essentially unchanged. Moreover, the pattern of R1 change could elucidate the pattern of drug release across the tumor. Future work aims to correlate the dox delivery to the tumors with the observed R1 changes.

Acknowledgements

This work is supported by NanoNextNL, a micro and nanotechnology consortium of the Government of the Netherlands and 130 partners.

References

1.Smet, M. De, et al. J. Control. Release 150, 102–110 (2011), 2.MacNeil, S. et al. Invest. Radiol. 46, 711–7 (2011), 3. Muetterties, K. a. et al. Magn. Reson. Med. 22, 88–100 (1991). 4.Hijnen, N. M. et al. Int. J. Hyperthermia 28, 141–55 (2012).

Figures

Schematic overview of the study timeline for both of the animal groups.

Overview of relevant liposomal characteristics for both Fe-SDFO TSL as well as Dox TSL.

Release curves of the TSLs at several temperatures in FBS. A) Fe-SDFO release as deduced from the increase in longitudinal relaxation rate R1 upon release of Fe-SDFO from the TSLs. B) Dox release derived from its change in fluorescence upon release from the TSLs.

T1 maps obtained at various time points throughout the treatment for HIFU-treated animals (left panel) and non-treated control animals at corresponding time points (TP, right panel), overlaid onto an anatomical fast field echo image.



Proc. Intl. Soc. Mag. Reson. Med. 24 (2016)
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