Theranostics in radiology : development of targeted contrast media with treatment capability

Imaging is essential in the diagnostics and medicine of today. The development of new contrast agents is important for obtaining specific information from images and to distinguish disease. Microbubbles (MB) have previously been introduced as a contrast agent for ultrasound. By incorporating super paramagnetic iron oxide nanoparticles (SPION) to the polymer matrix of the MB or between its shell layers we obtain a contrast media for Magnetic Resonance Imaging (MRI); while functionalizing the MB by ligands for labeling with 99mTc enables imaging using Single-Photon Emission Tomography (SPECT). The use of hybrid SPECT- and Computed Tomography (CT) or MRI systems enables fusion of the images from the different modalities to obtain SPECT/CT or SPECT/MR images. In the research underlying this thesis we investigated the preclinical characteristics, biodistribution and kinetics of several types of MB in Sprague Dawley rats by injecting single- and multiple layer SPION MB as well as ligand functionalized- and SPION MB labeled with 99mTc. The results obtained from imaging was correlated and compared to the histopathology of MB findings in organs. Moreover, mice were injected with Alexa-680 Vivo Tag labeled MB for imaging using a pre-clinical In Vivo Imaging System (IVIS)/μCT. Sprague Dawley rats (300 ± 50 g) were injected with single layer SPION-, multiple layer SPION-, 99mTc-labeled ligand functionalized diethylenetriamine penta-acetic acid (DTPA)-, thiolated poly(methacrylic acid) (PMAA)-, chitosan-, 1,4,7- triazacyclononane-1,4,7-triacetic acid (NOTA)-, NOTA-SPION- or DTPA-SPION MB intravenously (i.v.) through the tail vein. The rats injected with SPION MB were scanned using MRI, while the rats injected with 99mTc-labeled DTPA-, PMAA-, chitosan- or NOTA MB were scanned using SPECT/CT. The rats injected with NOTA- SPION- or DTPA-SPION MB were co-registrated using SPECT/CT and MRI. The organs from rats injected with the nuclear medicine marker were removed post mortem and measured for radioactivity. The rats injected with SPION MB were sacrificed and their organs were removed post mortem for histopathology examination using Perls’ Prussian blue staining to show iron content and immunohistochemistry (IHC) to visualize macrophage uptake of MB. Mice (30 ± 5 g) were injected with multiple layer fluorescence Alexa-680 MB and imaged using IVIS. Their organs were removed post mortem and examined using pathology and the fluorescence of MB was visualized under the microscope. The uptake of MB was mainly seen in the lungs and liver 1-2 h post-injection, while the main distribution of MB at 24 h post-injection was seen in the liver. In conclusion the MB matrix can be functionalized by ligands, labeled by SPION, 99mTc and fluorescence Alexa-680 Vivo Tag to enable its visualization in vivo using multimodal imaging SPECT/CT, SPECT/MRI or IVIS/μCT. Furthermore we have shown that MB can be loaded with cytostatic- or inflammatory drugs for theranostics. Future studies regarding MB should address toxicity and efficiency in drug loading and delivery

Multimodality imaging using SPECT/CT and MRI and ligand functionalized 99m

BACKGROUND: In the present study, we used multimodal imaging to investigate biodistribution in rats after intravenous administration of a new 99mTc-labeled delivery system consisting of polymer-shelled microbubbles (MBs) functionalized with diethylenetriaminepentaacetic acid (DTPA), thiolated poly(methacrylic acid) (PMAA), chitosan, 1,4,7-triacyclononane-1,4,7-triacetic acid (NOTA), NOTA-super paramagnetic iron oxide nanoparticles (SPION), or DTPA-SPION. METHODS: Examinations utilizing planar dynamic scintigraphy and hybrid imaging were performed using a commercially available single-photon emission computed tomography (SPECT)/computed tomography (CT) system. For SPION containing MBs, the biodistribution pattern of 99mTc-labeled NOTA-SPION and DTPA-SPION MBs was investigated and co-registered using fusion SPECT/CT and magnetic resonance imaging (MRI). Moreover, to evaluate the biodistribution, organs were removed and radioactivity was measured and calculated as percentage of injected dose. RESULTS: SPECT/CT and MRI showed that the distribution of 99mTc-labeled ligand-functionalized MBs varied with the type of ligand as well as with the presence of SPION. The highest uptake was observed in the lungs 1 h post injection of 99mTc-labeled DTPA and chitosan MBs, while a similar distribution to the lungs and the liver was seen after the administration of PMAA MBs. The highest counts of 99mTc-labeled NOTA-SPION and DTPA-SPION MBs were observed in the lungs, liver, and kidneys 1 h post injection. The highest counts were observed in the liver, spleen, and kidneys as confirmed by MRI 24 h post injection. Furthermore, the results obtained from organ measurements were in good agreement with those obtained from SPECT/CT. CONCLUSIONS: In conclusion, microbubbles functionalized by different ligands can be labeled with radiotracers and utilized for SPECT/CT imaging, while the incorporation of SPION in MB shells enables imaging using MR. Our investigation revealed that biodistribution may be modified using different ligands. Furthermore, using a single contrast agent with fusion SPECT/CT/MR multimodal imaging enables visualization of functional and anatomical information in one image, thus improving the diagnostic benefit for patients

Thermostable Luciferase from <i>Luciola cruciate</i> for Imaging of Carbon Nanotubes and Carbon Nanotubes Carrying Doxorubicin Using in Vivo Imaging System

In the present study, we introduce a novel method for in vivo imaging of the biodistribution of single wall carbon nanotubes (SWNTs) labeled with recombinant thermo-stable <i>Luciola cruciata</i> luciferase (LcL). In addition, we highlight a new application for green fluorescent proteins in which they are utilized as imaging moieties for SWNTs. Carbon nanotubes show great positive potential compared to other drug nanocarriers with respect to loading capacity, cell internalization, and biodegradability. We have also studied the effect of binding mode (chemical conjugation and physical adsorption) on the chemiluminescence activity, decay rate, and half-life. We have shown that through proper chemical conjugation of LcL to CNTs, LcL remained biologically active for the catalysis of d-luciferin in the presence of ATP to release detectable amounts of photons for in vivo imaging. Chemiluminescence of LcL allows imaging of CNTs and their cargo in nonsuperficial locations at an organ resolution with no need of an excitation source. Loading LcL-CNTs with the antitumor antibiotic doxorubicin did not alter their biological activity for imaging. In vivo imaging of LcL-CNTs has been carried out using “IVIS spectrum” showing the uptake of LcL-CNTs by different organs in mice. We believe that the LcL-CNT system is an advanced powerful tool for in vivo imaging and therefore a step toward the advancement of the nanomedicine field