Nanomedicine and Personalized Treatments
Nanomedicine is simply the medical application of nanotechnologies. The idea is the involvement the use of nanoparticles to improve the behaviour of drug substances. The goal is to achieve improvement over conventional chemotherapies. Customized treatments will be required to overcome the issues raised by clinical patient and disease heterogeneity. As one might expect, the same drug will accumulate in tumors at varying concentrations in patients with different cancers. But this also happens in patients with the same kind of cancer. It has to be ensured that drug nanocarriers are really accumulating in the specific tissues to better treat patients. This brings in the necessity of a treatment prediction tool to select the patients most likely to accumulate high amounts of the nanomedicine of interest and hence benefit from nanomedicinal treatment.
Positron Emission Tomography (PET) is such a noninvasive quantitative imaging tool with excellent sensitivity and spatial/temporal resolution required at the whole-body level. Radiolabeling of liposomal nanomedicines with single-photon emission computed tomography (SPECT) radionuclides has been successfully used to study their biodistribution in preclinical and clinical studies, but SPECT imaging suffers from lower sensitivity and temporal/spatial resolution than PET. However, an ideal PET radiolabeling method viable for both preclinical and clinical imaging wasn’t explored before. Rafael T. M. de Rosales, Alberto Gabizon and colleagues at King’s College London and the Shaare Zedek Medical Center sought to address this challenge.
Edmonds, S. et al. Exploiting the Metal-Chelating Properties of the Drug Cargo for In Vivo Positron Emission Tomography Imaging of Liposomal Nanomedicines. ACS Nano (2016). doi:10.1021/acsnano.6b05935
The following Mediso systems were used to conduct the animal imaging studies: nanoScan PET/CT and NanoSPECT/CT Silver upgrade. Both systems are equipped with the MultiCell animal handling and monitoring system , thus enabling a combined PET-CT/SPECT-CT imaging strategy. Interestingly both PET and SPECT were performed in the same animals (by moving the same bed from scanner from scanner, while the animals were anesthetized in fixed position) that allowed to image the tumour cells with SPECT and the nanomedicine with PET.
Liposomal Drug PET Radiolabeling Method Development
The researchers introduced a simple and efficient PET radiolabeling method exploiting the metal-chelating properties of certain drugs (e.g., bisphosphonates such as alendronate and anthracyclines such as doxorubicin) and widely used ionophores radiolabeled with long half-life metallic PET isotopes, such as 89Zr, 52Mn and 64Cu. The labels — and thus the liposomal drugs — could then be tracked using positron emission tomography (PET) to see where they go within the body. The article discusses in details the feasibility and effectiveness of their method, as well as its advantages and limitations, and show its utility for detecting and quantifying the biodistribution of a liposomal nanomedicine containing an aminobisphosphonate in vivo.
In a model of metastatic breast cancer, the researchers demonstrated that their technique allows quantification of the biodistribution of a radiolabeled stealth liposomal nanomedicine. Alendronate (ALD), an aminobisphosphonate, was selected as the radionuclide-binding drug of choice to develop this method for two reasons: (i) known ability to act as metal chelator to form inert coordination complexes with zirconium, copper, and manganese; and (ii) demonstrated anticancer activity and γ−δ T-cell immunotherapy sensitizing properties. The used liposomal formulation is referred to as PLA in the article.
Monitoring Liposomal Nanomedicine Distribution
The biodistribution of the radiolabeled liposomes was monitored using PET imaging with 89Zr-PLA in a metastatic mammary carcinoma mouse model established in immunocompromised NSG mice. This cancer model is also traceable by SPECT imaging/fluorescence due to a dual-modality reporter gene, the human sodium iodide symporter (hNIS-tagRFP), that allows sensitive detection of viable cancer tissues (primary tumor and metastases) using SPECT imaging with 99mTc-pertechnetate and fluorescence during dissection and histological studies. The imaging protocol was as follows: first, mice were injected with 89Zr-PLA (4.6 ± 0.4 MBq) at t = 0 followed by nanoScan PET-CT imaging (liposome biodistribution). The same mice were then injected with 99mTc-pertechnetate (30 MBq) and imaged by SPECT-CT. The SPECT injection was repeated at t = 24 h, 72 h, and 168 h. It was confirmed by separate phantom studies that the presence of 99mTc was not affecting the quality/quantification of the PET study. CT images revealed a significant increase in tumor volume during the imaging study. Using the tumor volumes from SPECT and CT, the researchers calculated the percentage of necrotic tumor tissue over time, by subtracting the hNIS-positive volume (SPECT) to the total tumor volume (CT). A PET-CT study was also performed using 64Cu-PLA in an ovarian cancer model (SKOV-3/SCID-Beige) over 48 h to test the versatility and capability of the radiolabeling method.
The common MultiCell animal handling and monitoring system (developed by Mediso) on both imaging systems gave the possibility to easily co-register the PET-CT/SPECT-CT and PET/SPECT studies as the animals were moved in co-registered position between the systems.
MIP video (3D, rotating along z-axis) showing co-registration of PET (red signal, 89Zr-PLA) and SPECT (green signal, 99mTcO4-, hNIS positive viable tumour tissue) of representative tumor from the mutimodal PET/SPECT study in the 3E.Δ.NT/NSG model. Both signals/radiotracers accumulate predominantly at the rim of the tumour and areas of low colocalization as well as high co-localization (yellow) are evident.
Imaging with PET in mouse models of breast and ovarian cancer showed the drugs accumulated in tumors and metastatic tissues in varying concentrations and at levels well above those in normal tissues, the researchers report. In one mouse strain, the nanomedicines unexpectedly showed up in uteruses, a result that wouldn’t have been detected without conducting the imaging study, according to the researchers.
The results establish that preformed liposomal nanomedicines, including some currently in clinical use, can be efficiently labeled with PET radiometals and tracked in vivo by exploiting the metal affinity and high concentration of the encapsulated drugs. Importantly, the technique allows radiolabeling of preformed liposomal nanomedicines, without modification of their components and without affecting their physicochemical properties.
The versatility, efficiency, simplicity, and GMP compatibility of this method may enable submicrodosing imaging studies of liposomal nanomedicines containing chelating drugs in humans and may have clinical impact by facilitating the introduction of image-guided therapeutic strategies in current and future nanomedicine clinical studies. The ultimate goal is to use non-invasive imaging data to predict how much drug will be delivered to cancer tissues in specific patients, and whether the nanomedicine is reaching all the patient’s tumors in therapeutic concentrations.
Many thanks for Rafael T. M. de Rosales, the last author of the original article.
In the first published article from MSKCC (Carney, B. et al. Non-invasive PET Imaging of PARP1 Expression in Glioblastoma Models. Mol Imaging Biol 1–7 (2015)), using the nanoScan PET/MRI (1T) small animal imaging system, in vivo whole body PET/MRI imaging of [18F]PARPi in orthotopic brain tumor-bearing mice is referenced.
[18F]PARPi is a selective PARP1 imaging agent that can be used to visualize glioblastoma in xenograft and orthotopic mouse models with high precision and good signal/noise ratios offering new opportunities to non-invasively image tumor growth and monitor interventions.
Figure 6 in the article shows coronal views of contrast-enhanced MRI, [18F]PARPi PET images, and fused PET/MRI of orthotopic U251 MG tumor-bearing mice. In the top row the mouse receivied only [18F]PARPi, in the bottom row the mouse receivied [18F]PARPi after a 500-fold excess of olaparib.
The animals were injected with 200 µCi of [18F]-PARPi and a 20 minutes static PET scan was acquired 2 hours post injection. 200 µL of diluted gadopentate dimegumine in saline solution was administered intravenously one minute prior to MRI acquisition. Tumor regions were identified on anatomic images acquired using a post-contrast T-weighted spin-echo (SE) acquisition. The co-localization of [18F]PARPi and tumor in PET/MRI studies was confirmed by ex vivo autoradiography. In PET/MRI fusion images, accumulation in the tumor was co-aligned with the orthotopic tumor on MRI. In mice receiving an injection of olaparib ahead of the radiotracer, the [18F]PARPi tumor uptake was negligible.
It's important to note that no further or manual co-registration was required at all as the PET/MRI studies performed on the nanoSCan PET/MRI are co-registered by nature due to the common gantry and automated acquisition system. The very same images are displayed in the viewer when the dual-modality study is loaded from the DICOM server after reconstruction. This gives scientists confidence when evaluating multi-modal data; changing animal physiology and data obtained at different times won't distort the findings.
This post summarizes the results on a research of a new Zr89 PET tracer for cell labeling. The open access article was published last month in the European Journal of Nuclear Medicine and Molecular Imaging journal:
Charoenphun, P. et al. [89Zr]Oxinate4 for long-term in vivo cell tracking by positron emission tomography. EJNMMI (2014)
The preclinical PET/CT images were acquired on a nanoScan PET/CT in vivo small animal imaging system at King’s College London.
Increasing sensitivity of cell tracking by changing labeling and detection from SPECT to PET
Cell tracking by gamma imaging with radionuclides has been performed clinically for over 30 years and is used for tracking autologous leukocytes to detect sites of infection/inflammation. The standard radiolabelling methodology has been non-specific assimilation of lipophilic, metastable complexes of indium-111 (with oxine) or technetium-99m (with HMPAO). Regenerative medicine and immune cell-based therapies are creating new roles for clinical tracking of these cells. Conventional cell radiolabelling methods have been applied for some of these cell types, but for clinical use new applications will require detection of small lesions and small numbers of cells beyond the sensitivity of traditional gamma camera imaging with In-111 or Tc-99m (e.g. coronary artery disease, diabetes, neurovascular inflammation and thrombus), creating a need for positron-emitting radiolabels to exploit the better sensitivity, quantification and resolution of clinical PET.
So far the search for positron emitting (PET) radiolabels for cells has met with limited success. The near-ubiquitous presence of glucose transporters allows labelling with [18F]-FDG but labelling efficiencies are highly variable, the radiolabel is prone to rapid efflux, and the short half-life (110 min) of F-18 allows only brief tracking. Copper-64 offers a longer (12 h) half-life and efficient cell labelling using lipophilic tracers but rapid efflux of label from cells is a persistent problem and a still longer half-life would be preferred. A “PET analogue” of In-111 oxine, capable of cell tracking over 7 days or more, would be highly desirable but is not yet available.
Zr-89 Oxine: a PET cell radiolabelling agent for long term in vivo cell tracking
This paper describes the first synthesis of Zr-89 oxine, and comparison with In-111 oxine for labelling several cell lines, human leukocytes and tracking of the cancer cell line GFP-5T33 cells in mice. The new lipophilic, metastable complex of Zr-89 can radiolabel a range of cells, independently of specific phenotypes, providing a long-sought solution to the unmet need for a long half-life positron-emitting radiolabel to replace In-111 for cell migration imaging. In addition to the expected advantages (enhanced sensitivity, resolution and quantification) of cell tracking with PET rather than scintigraphy or SPECT, Zr-89 shows less efflux from cells in vitro and in vivo than In-111. GFP-5T33 is a syngeneic murine multiple myeloma model originating from the C57Bl/KaLwRij strain, engineered to express green fluorescent protein (GFP). It was chosen for this work because the fate of the cells after i.v. inoculation is known from the literature. Intravenously injected cells migrate exclusively to the liver, spleen and bone marrow. Furthermore as the radiolabelled cells were GFP positive it was possible to validate the non-invasive images by using flow sorting of the GFP positive cells and negative cells. After flow sorting the authors were able to show that after 7 days in vivo the Zr-89 Oxine cells remained viable for the duration of the study, and that ~95% of radioactivity was present in viable GFP+ cells. The excellent in vivo survival and retention of radioactivity by the cells at 7 days, coupled with the demonstrated ability to acquire useful PET images up to 14 days, significantly extend the typical period over which cells can be tracked by radionuclide imaging with directly labelled cells.
The use of PET Zr-89 oxine for cell tracking could have a dramatic impact in the investigation of infection, inflammation and cell-based therapies in humans.
Gnad, T. et al. Adenosine activates brown adipose tissue and recruits beige adipocytes via A2A receptors. Nature advance online publication, (2014) Published online 15 October 2014
It’s rare when an Nature article is directly relied on in vivo imaging experiment. The ‘Adenosine activates brown adipose tissue and recruits beige adipocytes via A2A receptors’ article was published online in Nature on 15 October 2014. Dr. Peter Brust, Professor at Helmholtz-Zentrum Dresden - Rossendorf, Institute of Radiopharmaceutical Cancer Research, Research Site Leipzig participated in the design and data analysis of the PET/MRI studies published in article. In his very recent talk at the EANM 2014 Mediso Preclinical User Workshop he gave the insight for the audience that molecular biology and conventional laboratory test results were actually crowned by the results of the in vivo imaging experiments performed with our nanoScan PET/MRI.
Brown adipose tissue (BAT) is specialized in energy expenditure, making it a potential target for anti-obesity therapies. However current BAT therapies based on cold exposure or B-adrenergic agonists are clinically not feasible, therefore alternative strategies has to be explored for developing new therapy possibilities. The researchers showed that adenosine activates human and murine brown adipocytes at low nanomolar concentrations. and induces browning of WAT. In the light of the world-wide obesity pandemic, activators of BAT may be potential drug targets for anti-obesity therapies and as shown here, adenosine is a previously unappreciated activator of BAT.
Adenosine role in BAT activating
Adenosine is released in BAT during stimulation of sympathetic nerves as well as from brown adipocytes. Pharmacological blockade or genetic loss of A receptors in mice caused a decrease in BAT-dependent thermogenesis, whereas treatment with A2A agonists significantly increases energy expenditure. Moreover, pharmacological stimulation of A2A receptors or injection of lentiviral vectors expressing the A receptor into white fat induced brown-like cells—so-called beige adipocytes. Importantly, mice fed a high-fat diet and treated with an A agonist are leaner with improved glucose tolerance.
The detailed analysis required a suitable animal model that mimics the response of human BAT to adenosine. The in vivo imaging results validated the original hypothesis that adenosine receptors' agonist ligands really activate the activities of brown adipose tissue.
The PET/MRI studies of BAT activation were performed on nanoScan PET/MRI (Mediso Medical Imaging Systems, Hungary) using male anaesthetized C57BL/6 WTmice. Subcutaneous injection of vehicle, noradrenaline or PSB-0777 (the A2A agonist) (both 1 mg per kg body weight) was performed one minute before intraperitoneal injection of 14.7+/-0.4 MBq of [18F]FDG. The activity in the interscapular BAT region at 75 min post injection was expressed as mean standardized uptake value.
Stimulation with noradrenaline or AAA agonist caused a significantly higher uptake of [18F]FDG compared to vehicle treatment into murine BAT as measured with positron emission tomography/magnetic resonance imaging.
Taken together, the results demonstrated that adenosine–A2A signalling plays an unexpected physiological role in sympathetic BAT activation and protects mice from high-fat diet-induced obesity. Those findings reveal new possibilities for developing novel obesity therapies. The featured Mediso nanoScan fully integrated PET/MRI system is completely mature, reliable system and installed at fifteen sites currently, including Kayvan R. Keshari, PhD lab at Memorial Sloan Kettering Cancer Center in New York City, NY.