Salomeh Jelveh

Gold Nanoparticles as Radiation Sensitizers in Cancer Therapy

Abstract Among other nanoparticle systems, gold nanoparticles have been explored as radiosensitizers. While most of the research in this area has focused on either gold nanoparticles with diameters of less than 2 nm or particles with micrometer dimensions, it has been shown that nanoparticles 50 nm in diameter have the highest cellular uptake. We present the results of in vitro studies that focus on the radiosensitization properties of nanoparticles in the size range from 14–74 nm. Radiosensitization was dependent on the number of gold nanoparticles internalized within the cells. Gold nanoparticles 50-nm in diameter showed the highest radiosensitization enhancement factor (REF) (1.43 at 220 kVp) compared to gold nanoparticles of 14 and 74 nm (1.20 and 1.26, respectively). Using 50-nm gold nanoparticles, the REF for lower- (105 kVp) and higher- (6 MVp) energy photons was 1.66 and 1.17, respectively. DNA double-strand breaks were quantified using radiation-induced foci of γ-H2AX and 53BP1, and a modest increase in the number of foci per nucleus was observed in irradiated cell populations with internalized gold nanoparticles. The outcome of this research will enable the optimization of gold nanoparticle-based sensitizers for use in therapy.

Gold Nanostructures as a Platform for Combinational Therapy in Future Cancer Therapeutics

The field of nanotechnology is currently undergoing explosive development on many fronts. The technology is expected to generate innovations and play a critical role in cancer therapeutics. Among other nanoparticle (NP) systems, there has been tremendous progress made in the use of spherical gold NPs (GNPs), gold nanorods (GNRs), gold nanoshells (GNSs) and gold nanocages (GNCs) in cancer therapeutics. In treating cancer, radiation therapy and chemotherapy remain the most widely used treatment options and recent developments in cancer research show that the incorporation of gold nanostructures into these protocols has enhanced tumor cell killing. These nanostructures further provide strategies for better loading, targeting, and controlling the release of drugs to minimize the side effects of highly toxic anticancer drugs used in chemotherapy and photodynamic therapy. In addition, the heat generation capability of gold nanostructures upon exposure to UV or near infrared light is being used to damage tumor cells locally in photothermal therapy. Hence, gold nanostructures provide a versatile platform to integrate many therapeutic options leading to effective combinational therapy in the fight against cancer. In this review article, the recent progress in the development of gold-based NPs towards improved therapeutics will be discussed. A multifunctional platform based on gold nanostructures with targeting ligands, therapeutic molecules, and imaging contrast agents, holds an array of promising directions for cancer research.

Mitigation of radiation-induced lung injury by genistein and EUK-207.

Purpose: We examined the effects of genistein and/or Eukarion (EUK)-207 on radiation-induced lung damage and investigated whether treatment for 0–14 weeks (wks) post-irradiation (PI) would mitigate late lung injury. Materials and methods: The lungs of female Sprague-Dawley (SD) rats were irradiated with 10 Gy. EUK-207 was delivered by infusion and genistein was delivered as a dietary supplement starting immediately after irradiation (post irradiation [PI]) and continuing until 14 wks PI. Rats were sacrificed at 0, 4, 8, 14 and 28 wks PI. Breathing rate was monitored and lung fibrosis assessed by lung hydroxyproline content at 28 wks. DNA damage was assessed by micronucleus (MN) assay and 8-hydroxy-2-deoxyguanosine (8-OHdG) levels. The expression of the cytokines Interleukin (IL)-1α, IL-1β, IL-6, Tumor necrosis factor (TNF)-α and Transforming growth factor (TGF)-β1, and macrophage activation were analyzed by immunohistochemistry. Results: Increases in breathing rate observed in the irradiated rats were significantly reduced by both drug treatments during the pneumonitis phase and the later fibrosis phase. The drug treatments decreased micronuclei (MN) formation from 4–14 wks but by 28 wks the MN levels had increased again. The 8-OHdG levels were lower in the drug treated animals at all time points. Hydroxyproline content and levels of activated macrophages were decreased at 28 wks in all drug treated rats. The treatments had limited effects on the expression of the cytokines. Conclusion: Genistein and EUK-207 can provide partial mitigation of radiation-induced lung damage out to at least 28 wks PI even after cessation of treatment at 14 wks PI.

Genistein Can Mitigate the Effect of Radiation on Rat Lung Tissue

Abstract We investigated whether genistein could protect the lung from radiation-induced injury. We hypothesized that genistein would reduce the levels of inflammatory cytokines and ROS after irradiation and therefore lead to reduced DNA damage and functional deficits. Whole lungs of Sprague-Dawley rats were irradiated with 18 Gy at ∼0.5 Gy/min. At 28 weeks a micronucleus assay was used to examine DNA damage and, using immunohistochemical analysis, expression of IL-1α, IL-1β, IL-6, TNF-α and TGF-β, macrophage activation, oxidative stress (8-OHdG) and collagen levels were measured. A TBARS assay was used to measure the level of malondialdehyde. Functional damage was assessed by measuring the breathing rate of the rats over the course of the experiment. The increase in breathing rate after irradiation was damped in rats receiving genistein during the phase of pneumonitis (6–10 weeks), and there was a 50–80-day delay in lethality in this group. Genistein treatment also decreased the levels of the inflammatory cytokines TNF-α, IL-1β and TGF-β and led to a reduction in collagen content, a reduction in 8-OHdG levels, and complete protection against DNA damage measured in surviving rats at 28 weeks after irradiation. These results demonstrates that genistein treatment can provide partial protection against the early (pneumonitis) effects of lung irradiation and reduce the extent of fibrosis, although not sufficiently to prevent lethality at the radiation dose used in this study.

Investigations into the role of inflammation in normal tissue response to irradiation.

PURPOSE Radiation-induced inflammation and production of reactive oxygen species (ROS) play a critical role in normal tissue response. In this study we have examined some aspects of these effects in lung and skin. METHODS The superoxide dismutase (SOD) catalase mimetic, EUK-207, and genistein, an isoflavone with anti-inflammatory properties, were given post-irradiation and micronuclei (MN) formation was determined in cells derived from irradiated lung and skin. Changes in breathing rate were measured using a plethysmograph following irradiation of C57Bl6 mice knocked out for tumor necrosis factor (TNF)-alpha or its receptors, TNFR1/2, or treated with endotoxin (lipopolysaccharide - LPS). RESULTS Both EUK-207 and genistein given after irradiation caused a large reduction in MN levels observed in lung cells during 14 weeks post-irradiation but ceasing treatment resulted in a rebound in MN levels at 28 weeks post-irradiation. In contrast, treatment with EUK-207 was largely ineffective in reducing MN observed in skin cells post-irradiation. Knock-out of TNF-alpha resulted in a reduced increase in breathing rate (peak at 12 weeks post-irradiation) relative to wild-type and TNFR1/2 knock-out. Treatment with LPS 1 h post-irradiation also reduced the increase in breathing rate. CONCLUSIONS The increase in MN in lung cells after treatment with EUK-207 or genistein was stopped suggests that continuing ROS production contributes to DNA damage in lung cells over prolonged periods. That this effect was not seen in skin suggests this mechanism is less prominent in this tissue. The reduced level of radiation pneumonitis (as monitored by breathing rate changes) in animals knocked out for TNF-alpha suggests that this cytokine plays a significant role in inducing inflammation in lung following irradiation. The similar effect observed following LPS given post-irradiation suggests the possibility that such treatment modifies the long-term cyclic inflammatory response following irradiation in lungs.

Mitigation of Radiation-Induced Lung Injury with EUK-207 and Genistein: Effects in Adolescent Rats

Exposure of civilian populations to radiation due to accident, war or terrorist act is an increasing concern. The lung is one of the more radiosensitive organs that may be affected in people receiving partial-body irradiation and radiation injury in lung is thought to be associated with the development of a prolonged inflammatory response. Here we examined how effectively damage to the lung can be mitigated by administration of drugs initiated at different times after radiation exposure and examined response in adolescent animals for comparison with the young adult animals that we had studied previously. We studied the mitigation efficacy of the isoflavone genistein (50 mg/kg) and the salen-Mn superoxide dismutase-catalase mimetic EUK-207 (8 mg/kg), both of which have been reported to scavenge reactive oxygen species and reduce activity of the NFkB pathway. The drugs were given by subcutaneous injection to 6- to 7-week-old Fisher rats daily starting either immediately or 2 weeks after irradiation with 12 Gy to the whole thorax. The treatment was stopped at 28 weeks post irradiation and the animals were assessed for levels of inflammatory cytokines, activated macrophages, oxidative damage and fibrosis at 48 weeks post irradiation. We demonstrated that both genistein and EUK-207 delayed and suppressed the increased breathing rate associated with pneumonitis. These agents also reduced levels of oxidative damage (50–100%), levels of TGF-β1 expression (75–100%), activated macrophages (20–60%) and fibrosis (60–80%). The adolescent rats developed pneumonitis earlier following irradiation of the lung than did the adult rats leading to greater severe morbidity requiring euthanasia (∼37% in adolescents vs. ∼10% in young adults) but the extent of the mitigation of the damage was similar or slightly greater.

Investigations of antioxidant-mediated protection and mitigation of radiation-induced DNA damage and lipid peroxidation in murine skin

Abstract Purpose: Radioprotection and mitigation effects of the antioxidants, Eukarion (EUK)-207, curcumin, and the curcumin analogs D12 and D68, on radiation-induced DNA damage or lipid peroxidation in murine skin were investigated. These antioxidants were studied because they have been previously reported to protect or mitigate against radiation-induced skin reactions. Methods: DNA damage was assessed using two different assays. A cytokinesis-blocked micronucleus (MN) assay was performed on primary skin fibroblasts harvested from the skin of C3H/HeJ male mice 1 day, 1 week and 4 weeks after 5 Gy or 10 Gy irradiation. Local skin or whole body irradiation (100 kVp X-rays or caesium (Cs)-137 γ-rays respectively) was performed. DNA damage was further quantified in keratinocytes by immunofluorescence staining of γ-histone 2AX (γ-H2AX) foci in formalin-fixed skin harvested 1 hour or 1 day post-whole body irradiation. Radiation-induced lipid peroxidation in the skin was investigated at the same time points as the MN assay by measuring malondialdehyde (MDA) with a Thiobarbituric acid reactive substances (TBARS) assay. Results: None of the studied antioxidants showed significant mitigation of skin DNA damage induced by local irradiation. However, when EUK-207 or curcumin were delivered before irradiation they provided some protection against DNA damage. In contrast, all the studied antioxidants demonstrated significant mitigating and protecting effects on radiation-induced lipid peroxidation at one or more of the three time points after local skin irradiation. Conclusion: Our results show no evidence for mitigation of DNA damage by the antioxidants studied in contrast to mitigation of lipid peroxidation. Since these agents have been reported to mitigate skin reactions following irradiation, the data suggest that changes in lipid peroxidation levels in skin may reflect developing skin reactions better than residual post-irradiation DNA damage in skin cells. Further direct comparison studies are required to confirm this inference from the data.

Targeting the Renin-angiotensin system combined with an antioxidant is highly effective in mitigating radiation-induced lung damage.

PURPOSE To investigate the outcome of suppression of the renin angiotensin system using captopril combined with an antioxidant (Eukarion [EUK]-207) for mitigation of radiation-induced lung damage in rats. METHODS AND MATERIALS The thoracic cavity of female Sprague-Dawley rats was irradiated with a single dose of 11 Gy. Treatment with captopril at a dose of 40 mg/kg/d in drinking water and EUK-207 given by subcutaneous injection (8 mg/kg daily) was started 1 week after irradiation (PI) and continuing until 14 weeks PI. Breathing rate was monitored until the rats were killed at 32 weeks PI, when lung fibrosis was assessed by lung hydroxyproline content. Lung levels of the cytokine transforming growth factor-β1 and macrophage activation were analyzed by immunohistochemistry. Oxidative DNA damage was assessed by 8-hydroxy-2-deoxyguanosine levels, and lipid peroxidation was measured by a T-BARS assay. RESULTS The increase in breathing rate in the irradiated rats was significantly reduced by the drug treatments. The drug treatment also significantly decreased the hydroxyproline content, 8-hydroxy-2-deoxyguanosine and malondialdehyde levels, and levels of activated macrophages and the cytokine transforming growth factor-β1 at 32 weeks. Almost complete mitigation of these radiation effects was observed by combining captopril and EUK-207. CONCLUSION Captopril and EUK-207 can provide mitigation of radiation-induced lung damage out to at least 32 weeks PI after treatment given 1-14 weeks PI. Overall the combination of captopril and EUK-207 was more effective than the individual drugs used alone.

Effects of lipopolysaccharide on the response of C57BL/6J mice to whole thorax irradiation.

BACKGROUND AND PURPOSE Inflammatory and fibrogenic processes play a crucial role in the radiation-induced injury in the lung. The aim of the present study was to examine whether additive LPS exposure in the lung (to simulate respiratory infection) would affect pneumonitis or fibrosis associated with lung irradiation. MATERIAL AND METHODS Wildtype C57Bl/6J (WT-C57) and TNFα, TNFR1 and TNFR2 knockout ((-/-)) mice, in C57Bl/6J background, were given whole thorax irradiation (10 Gy) with or without post-irradiation intratracheal administration of LPS (50μg/mice). Functional deficit was examined by measuring breathing rate at various times after treatment. Real-time Reverse Transcription-Polymerase Chain Reaction (RT-PCR) and immunohistochemistry were used to analyze the protein expression and m-RNA of Interleukin-1 alpha (IL-1α), Interleukin-1 beta (IL-1β), Interleukin-6 (IL-6), Tumour Necrosis Factor alpha (TNFα) and Transforming Growth Factor beta (TGFβ) in the lung at various times after treatment. Inflammatory cells were detected by Mac-3 (macrophages) and Toluidine Blue (mast cells) staining. Collagen content was estimated by hydroxyproline (total collagen) and Sircol assay (soluble collagen). Levels of oxidative damage were assessed by 8-hydroxy-2-deoxyguanosine (8-OHdG) staining. RESULTS LPS exposure significantly attenuated the breathing rate increases following irradiation of WT-C57, TNFR1(-/-) and TNFR2(-/-)mice and to a lesser extent in TNFα(-/-) mice. Collagen content was significantly reduced after LPS treatment in WT-C57, TNFR1(-/-) and TNFα(-/-) mice and there was a trend in TNFR2(-/-) mice. Similarly there were lower levels of inflammatory cells and cytokines in the LPS treated mice. CONCLUSIONS This study reveals a mitigating effect of early exposure to LPS on injury caused by irradiation on lungs of C57Bl mice. The results suggest that immediate infection post irradiation may not impact lung response negatively in radiation-accident victims, however, further studies are required in different animal models, and with specific infectious agents, to confirm and extend our findings.

Mitigation of Lung Injury after Accidental Exposure to Radiation

There is a serious need to develop effective mitigators against accidental radiation exposures. In radiation accidents, many people may receive nonuniform whole-body or partial-body irradiation. The lung is one of the more radiosensitive organs, demonstrating pneumonitis and fibrosis that are believed to develop at least partially because of radiation-induced chronic inflammation. Here we addressed the crucial questions of how damage to the lung can be mitigated and whether the response is affected by irradiation to the rest of the body. We examined the widely used dietary supplement genistein given at two dietary levels (750 or 3750 mg/kg) to Fischer rats irradiated with 12 Gy to the lung or 8 Gy to the lung + 4 Gy to the whole body excluding the head and tail (whole torso). We found that genistein had promising mitigating effects on oxidative damage, pneumonitis and fibrosis even at late times (36 weeks) when drug treatment was initiated 1 week after irradiation and stopped at 28 weeks postirradiation. The higher dose of genistein showed no greater beneficial effect. Combined lung and whole-torso irradiation caused more lung-related severe morbidity resulting in euthanasia of the animals than lung irradiation alone.