Seung-Jun Seo

Enhanced proton treatment in mouse tumors through proton irradiated nanoradiator effects on metallic nanoparticles

The impact of protons on metallic nanoparticles (MNPs) produces the potent release of MNP-induced secondary electrons and characteristic x-rays. To determine the ability of secondary radiations to enhance proton treatment, the therapeutic irradiation of tumors was investigated in mice receiving 100-300 mg MNPs/kg intravenously prior to single dose, 10-41 Gy, proton irradiation. A proton beam was utilized to irradiate nanoparticles with a single Bragg peak set to occur inside a tumor volume (fully absorbed) or to occur after the beam had traversed the entire body. The dose-dependent increase in complete tumor regression (CTR) was 37-62% in the fully-absorbed irradiation group or 50-100% in the traversing irradiation group, respectively, compared with the proton-alone control mice (p < 0.01). One year survival was 58-100% versus 11-13% proton alone. The dose-dependent increase of intracellular reactive oxygen species level was 12-36% at 10 Gy compared with the proton-alone control cell. Therapeutic effective drug concentration that led to 100% CTR with a proton dose of 31 Gy was measured either 41 µg Au/g tissue or 59 µg Fe/g tissue. MNP-based proton treatment increased not only percent CTR and survival in vivo but also ROS generation in vitro, suggesting tumor dose enhancement from secondary radiation as one potent pathway of therapeutic enhancement.

Comment on ‘Therapeutic application of metallic nanoparticles combined with particle-induced x-ray emission effect’

Metallic nanoparticles (MNP) are able to release localized x-rays when activated with a high energy proton beam by the particle-induced x-ray emission (PIXE) effect. The exploitation of this phenomenon in the therapeutic irradiation of tumors has been investigated. PIXE-based x-ray emission directed at CT26 tumor cells in vitro, when administered with either gold (average diameter 2 and 13 nm) or iron (average diameter 14 nm) nanoparticles (GNP or SNP), increased with MNP solution concentration over the range of 0.1-2 mg ml(-1). With irradiation by a 45 MeV proton therapy (PT) beam, higher concentrations had a decreased cell survival fraction. An in vivo study in CT26 mouse tumor models with tumor regression assay demonstrated significant tumor dose enhancement, thought to be a result of the PIXE effect when compared to conventional PT without MNP (radiation-only group) using a 45 MeV proton beam (p < 0.02). Those receiving GNP or SNP injection doses of 300 mg kg(-1) body weight before proton beam therapy demonstrated 90% or 75% tumor volume reduction (TVR) in 20 days post-PT while the radiation-only group showed only 18% TVR and re-growth of tumor volume after 20 days. Higher complete tumor regression (CTR) was observed in 14-24 days after a single treatment of PT with an average rate of 33-65% for those receiving MNP compared with 25% for the radiation-only group. A lower bound of therapeutic effective MNP concentration range, in vivo, was estimated as 30-79 µg g(-1) tissue for both gold and iron nanoparticles. The tumor dose enhancement may compensate for an increase in entrance dose associated with conventional PT when treating large, solid tumors with a spread-out Bragg peak (SOBP) technique. The use of a combined high energy Bragg peak PT with PIXE generated by MNP, or PIXE alone, may result in new treatment options for infiltrative metastatic tumors and other diffuse inflammatory diseases.

Sonodynamically Induced Antitumor Effects of 5-Aminolevulinic Acid and Fractionated Ultrasound Irradiation in an Orthotopic Rat Glioma Model

The sonodynamically induced selective antitumor effects of 5-aminolevulinic acid (5-ALA) on a C6 glioma that was implanted in a rat brain were evaluated. One week after the inoculation of the brains with the C6 rat glioma cells, glioma development was monitored using a 1.5 T MRI. Brains both with and without intravenous administration of 5-ALA (60 mg/kg body weight) or Radachlorin (40 mg/kg body weight) were insonated by a 1 MHz ultrasound at a dose of 2.65 W/cm(2). Irradiation was performed in a fractionated manner to avoid any thermal effects in the tissue due to the focused ultrasound; 16 min of irradiation were followed by a 3 min recess, then 4 min of resumed irradiation. Mean tumor sizes, measured after the rats were sacrificed 2 weeks post treatment, were 122.48 ± 39.64 mm(3) in sham-operated rats, 87.42 ± 21.40 mm(3) in rats receiving ultrasound without 5-ALA, 10.50 ± 8.20 mm(3) in rats receiving ultrasound with 5-ALA, and 56.42 ± 12.48 mm(3) in rats receiving ultrasound with Radachlorin. The tumor size was significantly smaller in the therapy group receiving sonodynamic 5-ALA than in any of the other groups (p < 0.05). This experimental rat model showed that sonodynamic therapy can be useful in the treatment of deep-seated malignant gliomas.

Photon activated therapy (PAT) using monochromatic Synchrotron x-rays and iron oxide nanoparticles in a mouse tumor model: feasibility study of PAT for the treatment of superficial malignancy

BackgroundX-rays are known to interact with metallic nanoparticles, producing photoelectric species as radiosensitizing effects, and have been exploited in vivo mainly with gold nanoparticles. The purpose of this study was to investigate the potential of sensitizing effect of iron oxide nanoparticles for photon activated therapy.MethodsX-rays photon activated therapy (PAT) was studied by treating CT26 tumor cells and CT26 tumor-bearing mice loaded with 13-nm diameter FeO NP, and irradiating them at 7.1 keV near the Fe K-edge using synchrotron x-rays radiation. Survival of cells was determined by MTT assay, and tumor regression assay was performed for in vivo model experiment. The results of PAT treated groups were compared with x-rays alone control groups.ResultsA more significant reduction in viability and damage was observed in the FeO NP-treated irradiated cells, compared to the radiation alone group (p < 0.04). Injection of FeO NP (100 mg/kg) 30 min prior to irradiation elevated the tumor concentration of magnetite to 40 μg of Fe/g tissue, with a tumor-to-muscle ratio of 17.4. The group receiving FeO NP and radiation of 10 Gy showed 80% complete tumor regression (CTR) after 15–35 days and relapse-free survival for up to 6 months, compared to the control group, which showed growth retardation, resulting in 80% fatality. The group receiving radiation of 40 Gy showed 100% CTR in all cases irrespective of the presence of FeO NP, but CTR was achieved earlier in the PAT-treated group compared with the radiation alone group.ConclusionsAn iron oxide nanoparticle enhanced therapeutic effect with relatively low tissue concentration of iron and 10 Gy of monochromatic X-rays. Since 7.1 keV X-rays is attenuated very sharply in the tissue, FeO NP-PAT may have promise as a potent treatment option for superficial malignancies in the skin, like chest wall recurrence of breast cancer.

Coulomb nanoradiator-mediated, site-specific thrombolytic proton treatment with a traversing pristine Bragg peak.

Traversing proton beam-irradiated, mid/high-Z nanoparticles produce site-specific enhancement of X-ray photon-electron emission via the Coulomb nanoradiator (CNR) effect, resulting in a nano- to micro-scale therapeutic effect at the nanoparticle-uptake target site. Here, we demonstrate the uptake of iron oxide nanoparticles (IONs) and nanoradiator-mediated, site-specific thrombolysis without damaging the vascular endothelium in an arterial thrombosis mouse model. The enhancement of low-energy electron (LEE) emission and reactive oxygen species (ROS) production from traversing proton beam-irradiated IONs was examined. Flow recovery was only observed in CNR-treated mice, and greater than 50% removal of the thrombus was achieved. A 2.5-fold greater reduction in the thrombus-enabled flow recovery was observed in the CNR group compared with that observed in the untreated ION-only and proton-only control groups (p < 0.01). Enhancement of the X-ray photon-electron emission was evident from both the pronounced Shirley background in the electron yield and the 1.2- to 2.5-fold enhanced production of ROS by the proton-irradiated IONs, which suggests chemical degradation of the thrombus without potent emboli.

Reactive oxygen species-based measurement of the dependence of the Coulomb nanoradiator effect on proton energy and atomic Z value

Abstract Purpose: The Coulomb nanoradiator (CNR) effect produces the dose enhancement effects from high-Z nanoparticles under irradiation with a high-energy ion beam. To gain insight into the radiation dose and biological significance of the CNR effect, the enhancement of reactive oxygen species (ROS) production from iron oxide or gold NPs (IONs or AuNPs, respectively) in water was investigated using traversing proton beams. Methods and materials: The dependence of nanoradiator-enhanced ROS production on the atomic Z value and proton energy was investigated. Two biologically important ROS species were measured using fluorescent probes specific to •OH or in a series of water phantoms containing either AuNPs or IONs under irradiation with a 45- or 100-MeV proton beam. Results: The enhanced generation of hydroxyl radicals (•OH) and superoxide anions () was determined to be caused by the dependence on the NP concentration and proton energy. The proton-induced Au or iron oxide nanoradiators exhibited different ROS enhancement rates depending on the proton energy, suggesting that the CNR radiation varied. The curve of the superoxide anion production from the Au-nanoradiator showed strong non-linearity, unlike the linear behavior observed for hydroxyl radical production and the X-ray photoelectric nanoradiator. In addition, the 45-MeV proton-induced Au nanoradiator exhibited an ROS enhancement ratio of 8.54/1.50 ( / •OH), similar to that of the 100-KeV X-ray photoelectric Au nanoradiator (7.68/1.46). Conclusions: The ROS-based detection of the CNR effect revealed its dependence on the proton beam energy, dose and atomic Z value and provided insight into the low-linear energy transfer (LET) CNR radiation, suggesting that these factors may influence the therapeutic efficacy via chemical reactivities, transport behaviors, and intracellular oxidative stress.

Dark-Field Imaging: Recent developments and potential clinical applications

This paper describes an X-ray phase contrast imaging technique using analyzer-based optics called X-ray Dark-Field Imaging that has been under development for the past 10years. We describe the theory behind XDFI, the X-ray optics required for implementing it in practice, and algorithms used for 2D, 2.5D, and 3D image reconstruction. The XDFI optical chain consists of an asymmetrically cut, Bragg-type monochromator-collimator that provides a planar monochromatic X-ray beam, a positioning stage for the specimens, a Laue-case angle analyzer, and one or two cameras to capture the dark and bright field images. We demonstrate the soft-tissue discrimination capabilities of XDFI by reconstructing images with absorption and phase contrast. By using a variety of specimens such as breast tissue with cancer, joints with articular cartilage, ex-vivo human eye specimen, and others, we show that refraction-based contrast derived from XDFI is more effective in characterizing anatomical features, articular pathology, and neoplastic disease than conventional absorption-based images. For example, XDFI of breast tissue can discriminate between the normal and diseased terminal duct lobular unit, and between invasive and in-situ cancer. The final section of this paper is devoted to potential future developments to enable clinical and histo-pathological applications of this technique.

Bone dynamics in the upward direction after a maxillary sinus floor elevation procedure: serial segmentation using synchrotron radiation micro-computed tomography

Objective Maxillary sinus floor augmentation has been shown to be the most predictable surgical technique for enhancing the bone volume in the posterior area of the maxilla. The purpose of this study was to analyze the serial slice image segmentation of newly formed bone and bone substitutes after sinus floor elevation using synchrotron radiation X-ray micro-computed tomography (SR-μCT). Materials and methods Bone biopsy specimens were collected after 6 months of sinus floor augmentation. From the six bone biopsy specimens, the cross-sectional images at every 8 μm along the apical direction from the inferior border using serial segmentation from three-dimensional reconstructed X-ray images were analyzed. The amount of new bone and bone substitutes were measured at each slicing image (300–430 images per specimen). Results The bone dynamics between the new bone and bone substitutes along the inferior–superior direction in humans after maxillary sinus floor elevation (MSFE) were analyzed using the whole sample region. Although these observations suggest that the specimens are structurally inhomogeneous, sinus floor elevation was confirmed to be a reliable surgical procedure for increasing the amount of bone. Conclusion SR-μCT is highly effective for obtaining high-resolution images. An analysis of biological specimens using SR-μCT is quite reliable and this technique will be an important tool in the wide field of tissue engineering.

INVESTIGATION OF TUMOR CELL TOXICITY FROM PARTICLE INDUCED X-RAY EMISSION FROM A 45-MeV PROTON BEAM IRRADIATED FERRITE NANOPARTICLE

In order to investigate the potential cytotoxic effects of particle-induced x-ray emission (PIXE) on tumor cells, 45 MeV proton beam was irradiated on C6 glioma cell lines that had taken up alginate-coated ferrite nanoparticles (Alg-SNP). Cells were anchored in vertical 96-well dishes facing a horizontal beam where the Bragg peak was placed on the upper part of the 96-well dish. Experimental groups included cells without SNP as a control (No-SNP), and cells incubated with SNP for 6 hours (6hr-SNP) or overnight (ON-SNP). A 0 to 200 Gy proton beam from an MC50 cyclotron (Scanditronix, Sweden) at the Korea Cancer Center Hospital (Seoul, Korea) was used to irradiate each experimental group. Perinuclear Alg-SNP nanoparticle distribution was observed in glioma cells. The test groups (6hr-SNP or ON-SNP) showed an estimated 20-28% (ANOVA, P < 0.05) less cell survival compared to the control group based on MTT assay. Nuclear damage, indicating apoptosis, was present at a higher frequency in the 6hr-SNP and ON-SNP groups up to relatively low radiation dose of 100 Gy by fluorescence microscopy upon Hoechst 33342 and Acridine Orange staining. Ferrite nanoparticles alone were not cytotoxic at the experimental concentration of 0.15 mg/ml. Therefore ferrite nanoparticles may induce additional cytotoxicity from X-ray emission from potential PIXE effects. PIXE and metal nanoparticles may be developed as a therapeutic factor and prodrug for localized proton beam therapy without side effects of solid or disseminate tumors on the surrounding normal tissue.

Effects of Escherichia Coli-derived Recombinant Human Bone Morphogenetic Protein-2 Loaded Porous Hydroxyaptite-based Ceramics on Calvarial Defect in Rabbits

Background Recombinant human bone morphogenetic proteins (rhBMPs) have been widely used in regenerative therapies to promote bone formation. The production of rhBMPs using bacterial systems such as Escherichia coli (E. coli) is estimated to facilitate clinical applications by lowering the cost without compromising biological activity. In clinical practice, rhBMP-2 and osteoconductive carriers (e.g., hydroxyapatite [HA] and bovine bone xenograft) are used together. This study examined the effect of E. coli-derived rhBMP-2 combined with porous HA-based ceramics on calvarial defect in rabbits. Methods Six adult male New Zealand white rabbits were used in this study. The experimental groups were divided into the following 4 groups: untreated (NC), bovine bone graft (BO), porous HA (HA) and porous HA with rhBMP-2 (HA-BMP). Four transosseous defects of 8 mm in diameter were prepared using stainless steel trephine bur in the frontal and parietal bones. Histological and histomorphometric analyses at 4 weeks after surgery revealed significant new bone formation by porous HA alone. Results HA-BMP showed significantly higher degree of bone formation compared with BO and HA group (P<0.05). The average new bone formation % (new bone area per total defect area) of NC, BO, HA, and HA-BMP at 4-week after surgery were 12.65±5.89%, 29.63±6.99%, 28.86±6.17% and 49.56±8.23%, respectively. However, there was no statistical difference in the bone formation between HA and BO groups. Conclusions HA-BMP promoted more bone formation than NC, BO and HA alone. Thus, using E. coli-derived rhBMP-2 combined with porous HA-based ceramics can promote new bone formation.