Feasibility of SPECT-CT Imaging to Study the Pharmacokinetics of Antisense Oligonucleotides in a Mouse Model of Duchenne Muscular Dystrophy.

Antisense oligonucleotides (AONs) are promising candidates for treatment of Duchenne muscular dystrophy (DMD), a severe and progressive disease resulting in premature death. However, more knowledge on the pharmacokinetics of new AON drug candidates is desired for effective application in the clinic. We assessed the feasibility of using noninvasive single-photon emission computed tomography-computed tomography (SPECT-CT) imaging to determine AON pharmacokinetics in vivo. To this end, a 2'-O-methyl phosphorothioate AON was radiolabeled with 123I or 111In, and administered to mdx mice, a rodent model of DMD. SPECT-CT imaging was performed to determine AON tissue levels, and the results were compared to data obtained with invasive analysis methods (scintillation counting and a ligation-hybridization assay). We found that SPECT-CT data obtained with 123I-AON and 111In-AON were qualitatively comparable to data derived from invasive analytical methods, confirming the feasibility of using SPECT-CT analysis to study AON pharmacokinetics. Notably, also AON uptake in skeletal muscle, the target tissue in DMD, could be readily quantified using SPECT-CT imaging, which was considered a particular challenge in mice, due to their small size. In conclusion, our results demonstrate that SPECT-CT imaging allows for noninvasive characterization of biodistribution and pharmacokinetics of AONs, thereby enabling quantitative comparisons between different radiolabeled AON drug candidates and qualitative conclusions about the corresponding unmodified parent AONs. This technology may contribute to improved (pre)clinical drug development, leading to drug candidates with optimized characteristics in vivo.

89 Zr- and Fe-Labeled Polymeric Micelles for Dual Modality PET and T1 -Weighted MR Imaging.

In this study, a new 89 Zr- and Fe3+ -labeled micelle nanoplatform (89 Zr/Fe-DFO-micelles) for dual modality position emission tomography/magnetic resonance (PET/MR) imaging is investigated. The nanoplatform consists of self-assembling amphiphilic diblock copolymers that are functionalized with 89 Zr-deferoxamine (89 Zr-DFO) and Fe3+ -deferoxamine (Fe-DFO) for PET and MR purposes, respectively. 89 Zr displays favorable PET imaging characteristics with a 3.3 d half-life suitable for imaging long circulating nanoparticles. The nanoparticles are modified with Fe-DFO as MR T1 -contrast label instead of commonly used Gd3+ -based chelates. As these micelles are cleared by liver and spleen, any long term Gd- related toxicity such as nephrogenic systemic fibrosis is avoided. As a proof of concept, an in vivo PET/MR study in mice is presented showing tumor targeting of 89 Zr/Fe-DFO-micelles through the enhanced permeability and retention (EPR) effect of tumors, yielding high tumor-to-blood (10.3 ± 3.6) and tumor-to-muscle (15.3 ± 8.1) ratios at 48 h post injection. In vivo PET images clearly delineate the tumor tissue and show good correspondence with ex vivo biodistribution results. In vivo magnetic resonance imaging (MRI) allows visualization of the intratumoral distribution of the 89 Zr/Fe-DFO-micelles at high resolution. In summary, the 89 Zr/Fe-DFO-micelle nanoparticulate platform allows EPR-based tumor PET/MRI, and, furthermore, holds great potential for PET/MR image guided drug delivery.

Bone metastasis treatment using magnetic resonance-guided high intensity focused ultrasound.

OBJECTIVES: Bone pain resulting from cancer metastases reduces a patient's quality of life. Magnetic Resonance-guided High Intensity Focused Ultrasound (MR-HIFU) is a promising alternative palliative thermal treatment technique for bone metastases that has been tested in a few clinical studies. Here, we describe a comprehensive pre-clinical study to investigate the effects, and efficacy of MR-HIFU ablation for the palliative treatment of osteoblastic bone metastases in rats. MATERIALS AND METHODS: Prostate cancer cells (MATLyLu) were injected intra-osseously in Copenhagen rats. Upon detection of pain, as determined with a dynamic weight bearing (DWB) system, a MR-HIFU system was used to thermally ablate the bone region with tumor. Treatment effect and efficacy were assessed using magnetic resonance imaging (MRI), single-photon emission computed tomography (SPECT) with technetium-99m medronate ((99m)Tc-MDP), micro-computed tomography (µCT) and histology. RESULTS: DWB analysis demonstrated that MR-HIFU-treated animals retained 58.6 ± 20.4% of limb usage as compared to 2.6 ± 6.3% in untreated animals (P=0.003). MR-HIFU delayed tumor specific growth rates (SGR) from 29 ± 6 to 13 ± 5%/day (P<0.001). Untreated animals (316.5 ± 78.9 mm(3)) had a greater accumulation of (99m)Tc-MDP than HIFU-treated animals (127.0 ± 42.7 mm(3), P=0.004). The total bone volume increase for untreated and HIFU-treated animals was 15.6 ± 9.6% and 3.0 ± 4.1% (P=0.004), respectively. Histological analysis showed ablation of nerve fibers, tumor, inflammatory and bone cells. CONCLUSIONS: Our study provides a detailed characterization of the effects of MR-HIFU treatment on bone metastases, and provides fundamental data, which may motivate and advance its use in the clinical treatment of painful bone metastases with MR-HIFU.

Novel analogs of antitumor agent calixarene 0118: Synthesis, cytotoxicity, click labeling with 2-[(18)F]fluoroethylazide, and in vivo evaluation.

Calixarene 0118 is a potent anti-angiogenic agent that effectively inhibited tumor growth in preclinical studies, and is currently being evaluated in a phase I clinical trial. We have designed two close mimetics of calixarene 0118 containing a terminal alkynyl-functional group, and developed an optimized semi-automated procedure for radiolabeling with 2-[(18)F]fluoroethylazide using click chemistry. Following semi-preparative HPLC purification and formulation, the lower-rim modified analog [(18)F]6 and the equatorially labeled [(18)F]13 were obtained in >97% radiochemical purity and overall decay-corrected isolated radiochemical yields of 18.7 ± 2.7% (n = 4) and 10.2 ± 5.0% (n = 4), respectively, in a total synthesis time of about 2 h. Preliminary in vivo studies in nude mice bearing human tumor xenografts revealed highest accumulation of both tracers in the liver, followed by spleen, kidney, lung and bone, with no substantial uptake in the tumor. Still, these first-in-class radiotracers are a valuable tool for pharmacokinetic profiling and improvement of calixarene-based anti-angiogenic therapeutics in the future, as similar radiolabeling strategies may be applied to other compounds in the calixarene series. The cold reference compounds of the radiotracers were characterized in terms of cytotoxicity and anti-proliferative effects on HUVEC cells and on MA148 human ovarian carcinoma cells, along with the respective precursors, a small series of 0118 analogs modified with short-chain linear alkyl substituents, and a PEG3-spaced calixarene dimer. While all of the new analogs proved at least equipotent to parent 0118, some of them inhibited HUVEC and MA148 cell growth almost 4- and 10-fold more effectively, rendering these analogs promising candidates for further evaluation in anti-angiogenic cancer therapy.