Supplementary Figures 1-9 from Novel Chemical Enhancers of Heat Shock Increase Thermal Radiosensitization through a Mitotic Catastrophe Pathway

Gold nanomaterials have been shown to augment radiation therapy both in vitro and in vivo. However, studies on these materials are mostly phenomenological due to nanoparticle heterogeneity and the complexity of biological systems. Even accurate quantification of the particle dose still results in bulk average biases; the effect on individual cells is not measured but rather the effect on the overall population. To perform quantitative nanobiology, we coated glass coverslips uniformly at varying densities with Au nanoparticle preparations with different morphologies (45 nm cages, 25 nm spheres, and 30 nm rods). Consequently, the effect of a specific number of particles per unit area in contact with breast cancer cells growing on the coated surfaces was ascertained. Gold nanocages showed the highest degree of radiosensitization on a per particle basis, followed by gold nanospheres and gold nanorods, respectively. All three materials showed little cytotoxic effect at 0 Gy, but clonogenic survival decreased proportionally with the radiation dose and particle coverage density. A similar trend was seen in vivo in the combined treatment antitumor response in 4T1 tumor-bearing animals. The presence of gold affected the type and quantity of reactive oxygen species generated, specifically superoxide and hydroxyl radicals, and the concentration of nanocages correlated with the development of more numerous double-stranded DNA breaks and increased protein oxidation as measured by carbonylation. This work demonstrates the dependence on morphology and concentration of radiation enhancement by gold nanomaterials and may lead to a novel method to differentiate intra- and extracellular functionalities of gold nanomedicine treatment strategies. It further provides insights that can guide the rational development of gold nanomaterial-based radiosensitizers for clinical use.

Targeted delivery of drugs or other therapeutic agents through internal or external triggers has been used to control and accelerate the release from liposomal carriers in a number of studies, but relatively few utilize energy of therapeutic X-rays as a trigger. We have synthesized liposomes that are triggered by ionizing radiation (RTLs) to release their therapeutic payload. These liposomes are composed of natural egg phosphatidylethanolamine (PE), 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), cholesterol, and 1,2-disteroyl-sn-glycero-3-phosphoethanolamine-N-[methoxy (polyethylene glycol)-2000] (DSPE-PEG-2000), and the mean size of the RTL was in the range of 114 to 133 nm, as measured by nanoparticle tracking analysis (NTA). The trigger mechanism is the organic halogen, chloral hydrate, which is known to generate free protons upon exposure to ionizing radiation. Once protons are liberated, a drop in internal pH of the liposome promotes destabilization of the lipid bilayer and escape of the liposomal contents. In proof of principle studies, we assessed RTL radiation-release of fluorescent tracers upon exposure to a low pH extracellular environment or exposure to X-ray irradiation. Biodistribution imaging before and after irradiation demonstrated a preferential uptake and release of the liposomes and their cargo at the site of local tumor irradiation. Finally, a potent metabolite of the commonly used chemotherapy irinotecan, SN-38, was loaded into RTL along with near infrared (NIR) fluorescent dyes for imaging studies and measuring tumor cell cytotoxicity alone or combined with radiation exposure, in vitro and in vivo. Fully loaded RTLs were found to increase tumor cell killing with radiation in vitro and enhance tumor growth delay in vivo after three IV injections combined with three, 5 Gy local tumor radiation exposures compared to either treatment modality alone.

Hepatocellular carcinoma (HCC) is both a devastating and common disease. Every year in the United States, about 24,500 men and 10,000 women are diagnosed with HCC, and more than half of those diagnosed patients die from this disease. Thus far, conventional therapeutics have not been successful for patients with HCC due to various underlying comorbidities. Poor survival rate and high incidence of recurrence after therapy indicate that the differences between the redox environments of normal surrounding liver and HCC are valuable targets to improve treatment efficacy. Parthenolide (PTL) is a naturally found therapeutic with anti-cancer and anti-inflammatory properties. PTL can alter HCC's antioxidant environment through thiol modifications leaving tumor cells sensitive to elevated reactive oxygen species (ROS). Investigating the link between altered thiol mechanism and increased sensitivity to iron-mediated lipid peroxidation will allow for improved treatment of HCC. HepG2 (human) and McARH7777 (rat) HCC cells treated with PTL with increasing concentrations decrease cell viability and clonogenic efficiency in vitro. PTL increases glutathione (GSH) oxidation rescued by the addition of a GSH precursor, N-acetylcysteine (NAC). In addition, this elevation in thiol oxidation results in an overall increase in mitochondrial dysfunction. To elucidate if cell death is through lipid peroxidation, using a lipid peroxidation sensor indicated PTL increases lipid oxidation levels after 6 h. Additionally, western blotting reveals glutathione peroxidase 4 (GPx4) protein levels decrease after treatment with PTL suggesting cells are incapable of preventing lipid peroxidation after exposure to PTL. An elevation in lipid peroxidation will lead to a form of cell death known as ferroptosis. To further establish ferroptosis as a critical mechanism of death for HCC in vitro, the addition of ferrostatin-1 combined with PTL demonstrates a partial recovery in a colony survival assay. This study reveals that PTL can induce tumor cell death through elevations in intracellular oxidation, leaving cells sensitive to ferroptosis.

The combination of biological variation and nanomaterial heterogeneity makes elucidating the mechanisms of interactions between cells and nanoparticles extremely complicated. Accurate nanoparticle quantification can be extremely challenging, and cellular response can change based on the location of the nanoparticle and the cell type under investigation. These complications are only amplified by the addition of external stimuli. These limitations have yielded a wide range of studies that show effects, but often provide little mechanistic insight. Gold (Au) nanomaterials were stably immobilized onto glass coverslips treated with mercaptosilane to control both the average number of nanoparticles that interact with cells and their spatial orientation relative to the cell membrane. Surfaces were characterized optically and by electron microscopy to confirm their surface density and uniformity. The thermal response of Au nanocage-coated surfaces to near infrared laser irradiation was measured in cell culture medium and modeled computationally. The modeling showed a vastly higher thermal dose than would be predicted by bulk temperature measurements. Adherent or non-adherent cell lines were cultured directly on the nanocage-coated surface or in the medium, respectively, in culture wells and laser irradiation was applied. Survival of cells growing in suspension correlated with the bulk temperature increase in the culture medium, as measured by viability assay. Conversely, adherent cells exhibited a much greater susceptibility than expected from the bulk temperature measurement, which is ostensibly related to the close interaction with the nanoparticles on their growth substrate and induction of substantially greater thermal dose upon laser exposure. This platform is designed to be a new tool to determine how many particles need to be in contact with a cell to induce desired physical or biological effects. Here we demonstrate the delivery of precise thermal doses following laser irradiation. The anticipated biological effects based on bulk measurements vastly underestimated the effects that were observed, which is ascribed to the proximity of the nanoparticle to the cell and the extraordinary high surface temperature of the particle. This platform could be expanded to a variety of nanoparticles, external stimuli, and cell types to enable more deliberate and optimized application of nanomedicine.

Glioblastoma (GBM) is highly resistant to treatment and invasion into the surrounding brain is a cancer hallmark that leads to recurrence despite surgical resection. With the emergence of precision medicine, patient-derived 3D systems are considered potentially robust GBM preclinical models. In this study, we screened a library of 22 anti-invasive compounds (i.e., NF-kB, GSK-3-B, COX-2, and tubulin inhibitors) using glioblastoma U-251 MG cell spheroids. We evaluated toxicity and invasion inhibition using a 3D Matrigel invasion assay. We next selected three compounds that inhibited invasion and screened them in patient-derived glioblastoma organoids (GBOs). We developed a platform using available macros for FIJI/ImageJ to quantify invasion from the outer margin of organoids. Our data demonstrated that a high-throughput invasion screening can be done using both an established cell line and patient-derived 3D model systems. Tubulin inhibitor compounds had the best efficacy with U-251 MG cells, however, in ex vivo patient organoids the results were highly variable. Our results indicate that the efficacy of compounds is highly related to patient intra and inter-tumor heterogeneity. These results indicate that such models can be used to evaluate personal oncology therapeutic strategies.

Strategies to increase the proportion of neural stem cells that differentiate into neurons are vital for therapy of neurodegenerative disorders. In vitro, the extracellular matrix composition and topography have been found to be important factors in stem cell differentiation. We have developed a novel artificial extracellular matrix (aECM) formed by attaching gold nanocages (AuNCs) to glass coverslips. After culturing rat neural stem cells (rNSCs) on these gold nanocage-coated surfaces (AuNC-aECMs), we observed that 44.6% of rNSCs differentiated into neurons compared to only 27.9% for cells grown on laminin-coated glass coverslips. We applied laser irradiation to the AuNC-aECMs to introduce precise amounts of photothermally induced heat shock in cells. Our results showed that laser-induced thermal stimulation of AuNC-aECMs further enhanced neuronal differentiation (56%) depending on the laser intensity used. Response to these photothermal effects increased the expression of heat shock protein 27, 70, and 90α in rNSCs. Analysis of dendritic complexity showed that this thermal stimulation promoted neuronal maturation by increasing dendrite length as thermal dose was increased. In addition, we found that cells growing on AuNC-aECMs post laser irradiation exhibited action potentials and increased the expression of voltage-gated Na+ channels compared to laminin-coated glass coverslips. These results indicate that the photothermal response induced in cells growing on AuNC-aECMs can be used to produce large quantities of functional neurons, with improved electrochemical properties, that can potentially be transplanted into a damaged central nervous system to provide replacement neurons and restore lost function.

Parthenolide (PTL) is a naturally occurring sesquiterpene lactone that has anti-tumor activity. The redox environment of hepatocellular carcinoma (HCC) becomes an important target due to the differences between cancer and normal tissue metabolism. We hypothesize that in vitro PTL will induce HCC cell death through increased thiol oxidation. Half maximal inhibitory concentrations (IC50) response to PTL was determined in human (HepG2, SNU-423) and rat (McARH7777) hepatoma cell lines using MTT assays for 24 h. Cellular respiration response to PTL was measured over 24 h using Seahorse Flux Analyzer. Intracellular glutathione (GSH) and oxidized glutathione (GSSG) levels were measured using spectrophotometry at 2 h, 4 h and 24 h. The overall oxidation status of cell lines was also determined using thiol redox-sensitive ratiometric sensor roGFP. PTL had over 90% cytotoxicity at high concentrations 20 μM for SNU-423, 54 μM for HepG2, and 40 μM for rat HCC after 24 h. IC50 concentrations of 6.6 μM for SNU-423, 18 μM for HepG2 and 13 μM for rat HCC were determined after 24 h. Cytotoxic doses of PTL induced a decline in cellular respiration of the HCC cell lines after 2 h. Furthermore, 2 h PTL treatment at IC50 or high concentration caused cyto-roGFP sensor to be oxidized 2.5- to 4-fold in human and rat HCC. PTL at IC50 doses resulted in increased percent GSSG as early as 2 h in human and rat cells. This increase was compensated by upregulation of total GSH levels to minimize cell toxicity in HepG2 cells, which appeared to be offset at high doses of PTL. In HepG2 cells increased percent GSSG levels also demonstrated a similar trend in both IC50 and high PTL doses. In vitro PTL decreased cellular respiration, increased overall oxidation status and increased glutathione oxidation in HCC as early as 2 h after treatment. In HepG2 cells, IC50 dose caused a compensatory upregulation of GSH levels to ostensibly overcome thiol oxidation induced cytotoxicity. However, higher concentrations of PTL overwhelmed this mechanism and caused significant cytotoxicity in all 3 cells lines suggesting PTL can potentially be delivered at high concentrations as locoregional therapy for HCC in vivo.

Pancreatic cancer is the third leading cause of death in the US with a poor 5-year survival rate of 8.5%. A novel anti-cancer drug, dimethylamino parthenolide (DMAPT), is the water-soluble analog of the natural sesquiterpene lactone, parthenolide. The putative modes of action of DMAPT are inhibition of the Nuclear chain factor kappa-light-chain enhancer of activated B cells (NFB) pathway and depletion of glutathione levels; the latter causing cancer cells to be more susceptible to oxidative stress-induced cell death. Actinomycin-D (ActD) is a polypeptide antibiotic that binds to DNA, and inhibits RNA and protein synthesis by inhibiting RNA polymerase II. A phase 2 clinical trial indicated that ActD could be a potent drug against pancreatic cancer; however, it was not a favored drug due to toxicity issues. New drug entities and methods of drug delivery, used alone or in combination, are needed to treat pancreatic cancer more effectively. Thus, it was postulated that combining DMAPT and ActD would result in synergistic inhibition of Panc-1 pancreatic cancer cell growth because DMAPT's inhibition of NFB would enhance induction of apoptosis by ActD, via phosphorylation of c-Jun, by minimizing NFB inhibition of c-Jun phosphorylation. Combining these two drugs induced a higher level of cell death than each drug alone. A fixed drug ratio of DMAPT: ActD (1,200:1) was used. Data from metabolic (MTT) and colony formation assays were analyzed for synergism with CompuSyn software, which utilizes the Chou-Talalay equation. The analyses indicated synergism and moderate synergism at combination concentrations of DMAPT/ActD of 12/0.01 and 18/0.015M, respectively.

The core tenet of the IDTF is that the field of IR must shift the emphasis from supporting individual projects to the nurturing of investigators who demonstrate promise and commitment to IR research. This must be accomplished within a SIRF-supported longitudinal framework of support and feedback. We must cultivate a culture whereby developing the research skill set is valued. By ensuring prospective investment in a research culture, starting at the beginning of IR training, to support future success, IR will do more than survive: it will thrive and branch out into new areas of patient care. The end result will be increased awareness of our value among other specialties and hospital administrators, lower costs, expanded roles of IR in medicine, and, ultimately, improved outcomes for patients. Although many details need refinement, change is coming. The IDTF invites and encourages input from the membership of SIR on the issues and recommended adaptations articulated herein.

A simple and reproducible procedure was developed to measure the volume of liquid microinjected into cells. A calibration curve of droplet fluorescence intensity versus volume was constructed by injecting a fluorescent dextran solution through a 125-150 pm diameter micropipette into an oil-filled culture dish to create a spray of varied-sized droplets. The droplets retained a spherical shape because they were in an oil medium and they settled onto a glass surface coated with a superhydrophobic surface. Fluorescent micrographs of the droplets were obtained and analyzed with Image-J software to quantify the fluorescence intensity and radius of each spherical droplet to produce the calibration curve. Subsequently, Dut-145 human prostate carcinoma cells were microinjected with the same fluorescent dextran solution and fluorescent micrographs of the cells were obtained using the identical exposure conditions used to photograph the droplets. The measured fluorescence intensity-of the microinjected cells was entered into the formula for the regression line that was fit to the calibration curve allowing determination of the volume of solution injected into each cell. Thus, a mixture consisting of known concentrations of a test material of test material (macromolecules, drugs, etc.) and a fluorescent dextran, volumetric, tracer can be used to quantify the relationship between the amount of a microinjected material and subsequent effects on cells.

In search of the sequence of pathogenic events leading to glucocorticoid-induced osteonecrosis, we determined the molecular, biomechanical, cellular, and vascular changes in the femur of C57BL/6 mice receiving prednisolone for 14, 28, or 42 days. The femoral head, but not the distal femur, of mice treated for 14 days showed a decrease in the expression of the hypoxia-inducible factor (Hif)-1α and vascular endothelial growth factor (VEGF), the number of osteoblasts, and bone formation rate and strength and showed an increase in osteoclasts. These changes were accompanied by conversion of the normal dendritic vasculature to pools of edema as detected by magnetic resonance imaging, providing robust diagnostic evidence of early osteonecrosis. At that time point, there were no detectable changes in bone density, cortical or cancellous bone architecture, midshaft or distal cancellous bone, or osteocyte apoptosis. In mice treated for 28 days, femoral head cancellous density, cortical width, and trabecular thickness decreased, and by 42 days the femoral heads had full-depth cortical penetrations and cancellous tissue osteonecrosis. These results indicate that the femoral head is a particularly sensitive anatomical site to the adverse effects of glucocorticoid excess on bone and that decreases of Hif-1α and VEGF expression, bone vascularity, and strength precede the loss of bone mass and microarchitectural deterioration, thus rendering the femoral head vulnerable to collapse.

A series of novel, heteroaryl carboxylic acid conjugates of the sesquiterpene melampomagnolide-B (MMB, 3) has been evaluated as antitumor agents against an NCI panel of 64 human hematopoetic and solid tumor cell lines. The indole-3-acrylic acid conjugate 7j and the indole-3-carboxylic acid conjugate 7k were found to be the most potent analogs in the series. Compounds 7j and 7k exhibited remarkable growth inhibition, with GI(50) values in the range 0.03-030 mu M and 0.04-0.28 mu M, respectively, against the cell lines in the leukemia sub-panel, and GI(50) values of 0.05-0.40 mu M and 0.04 -0.61 mu M, respectively, against 90% of the solid tumor cell lines in the NCI panel. Compound 7a was particularly effective against the sub-panel of breast cancer cell lines with GI(50) values in the range <0.01 -0.30 mu M. Compounds 7j, 7a and its water soluble analog 7p also exhibited potent anticancer activity against rat 9L-SF gliosarcoma cells in culture. Compound 7j was the most potent compound in the series in the M9-ENL1 AML cell assay with a lethal dose concentration EC50 value of 720 nM, and exhibited the greatest cytotoxicity against a collection of primary AML stem cell specimens, which included a specimen that was unresponsive to PTL, affording EC50 values in the range 0.33-1.0 mu M in three out of four specimens. The results from this study provide further evidence that analogs of the sesquiterpene MMB can be designed to afford molecules with significantly improved anticancer activity. Thus, both 7j and 7k are considered potential lead molecules in the search for new anticancer agents that can be used as treatments for both hematopoetic and solid tumors. (C) 2017 Elsevier Masson SAS. All rights reserved.

IV injection of dodecafluoropentane emulsion (DDFPe) increases oxygen transportation and reduces brain infarct volume in a rabbit stroke model. Tissue distribution of the parent perfluorocarbon dodecafluoropentane (DDFP) is unknown but is critical to understanding the mechanism by which DDFPe is effective in treating ischemia and for determining safe dosing. Previous studies showed a DDFP blood half-life of <2 min yet therapeutic effects lasted >90 min after injection. We describe DDFP distribution in brain, kidney, liver, spleen, and lung following nine dosing regimens in New Zealand White (NZW) rabbits. Single and multi-dose schedules were administered to NZW rabbits (n = 27). A single DDFPe dose (0.6 ml/kg) group was sacrificed 2 min after dosing and eight multi-dose groups (4 doses of 0.3 or 0.6 ml/kg and 15 doses of 0.1, 0.3, or 0.6) were sacrificed 90 min after final injections. Tissues were flash frozen and analyzed with headspace sampling/GC-MS. DDFP brain concentration increased with increasing dose in the 15 dose groups (4.70, 8.34, and 14.3 μg/g) and indicative of linear pharmacokinetics within this dose range. The DDFP lung concentration was not reflective of increasing dose or dose frequency. The total clearance of DDFP was consistent with previous reports showing 98% of DDFP is cleared within 2 h of administration.

The aim of this study was to determine the uptake of intravenously administered N-[11CH3]-dimethylaminoparthenolide (DMAPT) into orthotopic 9LSF glioblastoma brain tumors in Fisher 344 rats from positron emission tomography (PET) imaging studies. [11C]methyl iodide (11CH3I) was utilized as a [11C]-labeling reagent to label the precursor methylaminoparthenolide (MAPT) intermediate. From PET imaging studies it was found that brain uptake of N-[11CH3]DMAPT into brain tumor tissue was rapid (30min), and considerably higher than that in the normal brain tissue.

PurposeIn vitro studies and preliminary screening in a rat liver tumor model demonstrated that transarterial chemoembolization (TACE) with the natural drug parthenolide (PTL) was effective for inhibiting the growth of liver tumor cells without intolerable liver toxicity. Consequently, a 24 rat study was initiated to determine if PTL-Lipiodol TACE treatment of Walker 256 rat liver tumors could inhibit tumor growth or induce tumor regression. The efficacy of the PTL-Lipiodol was also compared to that achieved with transarterial embolization (TAE) with Lipiodol only.MaterialsLuciferase expressing, Walker 256- tumor cells were inoculated into the left lateral liver lobe of male Wistar rats. Transarterial delivery into the tumors was accomplished by catheterizing the gastroduodenal artery and hand guiding the catheter into the left hepatic artery. Tumor-bearing rats were partitioned into three treatment groups: saline, Lipiodol and PTL dissolved in Lipiodol (PTL-Lipiodol: 80 mM PTL). Magnetic resonance imaging (MRI) and bioluminescence imaging were used to monitor and measure tumor growth noninvasively. Animal body weight and liver function blood biochemicals were measured to assess general health and detect toxicity in all 3 groups. Nine days following treatment rat livers were harvested and examined histologically (hematoxylin and eosin (H&E) staining).ResultsThe PTL-Lipiodol, TACE treated tumors exhibited a slight (10%), average decrease in tumor volume during a 9 day post treatment period while those embolized with saline or Lipiodol only exhibited an average tumor volume increase of 4.5-fold. The MRI images and blood enzyme/biochemical analyses showed no evidence of liver toxicity or systemic toxicity associated with the PTL-Lipiodol TACE treatment, even with such a high, localized concentration of PTL. Histopathology examinations are still ongoing.ConclusionsThe inhibition of tumor growth by the PTL-Lipiodol treatment was marked and significant (p= 0.0286) and can be attributed to the presence of PTL because the Lipiodol only treatment produced no significant decrease in tumor growth compared to that of the control group (p= 0.037). PurposeIn vitro studies and preliminary screening in a rat liver tumor model demonstrated that transarterial chemoembolization (TACE) with the natural drug parthenolide (PTL) was effective for inhibiting the growth of liver tumor cells without intolerable liver toxicity. Consequently, a 24 rat study was initiated to determine if PTL-Lipiodol TACE treatment of Walker 256 rat liver tumors could inhibit tumor growth or induce tumor regression. The efficacy of the PTL-Lipiodol was also compared to that achieved with transarterial embolization (TAE) with Lipiodol only. In vitro studies and preliminary screening in a rat liver tumor model demonstrated that transarterial chemoembolization (TACE) with the natural drug parthenolide (PTL) was effective for inhibiting the growth of liver tumor cells without intolerable liver toxicity. Consequently, a 24 rat study was initiated to determine if PTL-Lipiodol TACE treatment of Walker 256 rat liver tumors could inhibit tumor growth or induce tumor regression. The efficacy of the PTL-Lipiodol was also compared to that achieved with transarterial embolization (TAE) with Lipiodol only. MaterialsLuciferase expressing, Walker 256- tumor cells were inoculated into the left lateral liver lobe of male Wistar rats. Transarterial delivery into the tumors was accomplished by catheterizing the gastroduodenal artery and hand guiding the catheter into the left hepatic artery. Tumor-bearing rats were partitioned into three treatment groups: saline, Lipiodol and PTL dissolved in Lipiodol (PTL-Lipiodol: 80 mM PTL). Magnetic resonance imaging (MRI) and bioluminescence imaging were used to monitor and measure tumor growth noninvasively. Animal body weight and liver function blood biochemicals were measured to assess general health and detect toxicity in all 3 groups. Nine days following treatment rat livers were harvested and examined histologically (hematoxylin and eosin (H&E) staining). Luciferase expressing, Walker 256- tumor cells were inoculated into the left lateral liver lobe of male Wistar rats. Transarterial delivery into the tumors was accomplished by catheterizing the gastroduodenal artery and hand guiding the catheter into the left hepatic artery. Tumor-bearing rats were partitioned into three treatment groups: saline, Lipiodol and PTL dissolved in Lipiodol (PTL-Lipiodol: 80 mM PTL). Magnetic resonance imaging (MRI) and bioluminescence imaging were used to monitor and measure tumor growth noninvasively. Animal body weight and liver function blood biochemicals were measured to assess general health and detect toxicity in all 3 groups. Nine days following treatment rat livers were harvested and examined histologically (hematoxylin and eosin (H&E) staining). ResultsThe PTL-Lipiodol, TACE treated tumors exhibited a slight (10%), average decrease in tumor volume during a 9 day post treatment period while those embolized with saline or Lipiodol only exhibited an average tumor volume increase of 4.5-fold. The MRI images and blood enzyme/biochemical analyses showed no evidence of liver toxicity or systemic toxicity associated with the PTL-Lipiodol TACE treatment, even with such a high, localized concentration of PTL. Histopathology examinations are still ongoing. The PTL-Lipiodol, TACE treated tumors exhibited a slight (10%), average decrease in tumor volume during a 9 day post treatment period while those embolized with saline or Lipiodol only exhibited an average tumor volume increase of 4.5-fold. The MRI images and blood enzyme/biochemical analyses showed no evidence of liver toxicity or systemic toxicity associated with the PTL-Lipiodol TACE treatment, even with such a high, localized concentration of PTL. Histopathology examinations are still ongoing. ConclusionsThe inhibition of tumor growth by the PTL-Lipiodol treatment was marked and significant (p= 0.0286) and can be attributed to the presence of PTL because the Lipiodol only treatment produced no significant decrease in tumor growth compared to that of the control group (p= 0.037). The inhibition of tumor growth by the PTL-Lipiodol treatment was marked and significant (p= 0.0286) and can be attributed to the presence of PTL because the Lipiodol only treatment produced no significant decrease in tumor growth compared to that of the control group (p= 0.037).

Liposomes are unilamellar spherical membranes made of lipids. Their ability to evade the immune system and to transport water soluble substances encased in their inner cavity make them excellent carriers for drug delivery, especially for cancer treatment. To increase their efficacy, liposomes must be endowed with mechanisms for fast and controlled release of incorporated material after reaching their target. Here we show that controlled drug release from liposomes may be achieved through the use of X-ray as a triggering stimulus. In a first experimental approach, the liposomes encased the drug, specially formulated nano-scintillators, a photo-active Ca2+ cage, and the phospholipase PLA2. Upon interaction with the therapeutic actinic beam, the nano-scintillators emitted UV photons that released Ca2+ ions from their photo-labile cage. The released ions activated the enzyme PLA2 which destabilized the lipid bilayer and allowed the incorporated drug to cross the liposome membrane in a matter of minutes. In a second experimental approach, the water-filled cavity of pH sensitive liposomes was loaded with a drug simulator and chloral hydrate, a substance known for inducing decrease of a water solution's pH upon exposure to ionizing radiation. The acidic pH inside the liposomes achieved after irradiation destabilized the permeability function of the membrane and allowed the incorporated drug simulator to leak out. This release mechanism was much slower than the one observed upon the action of activated PLA2 as it spanned several hours. However, it was faster than the current uncontrolled drug release mechanisms relaying on slow bio-degradation, which may take weeks. In conclusion, X-ray triggered delivery from liposomes may become a powerful clinical method by presenting the opportunity to simultaneously perform highly localized radio and chemotherapy. Given the supra-additive effects of the combined therapies, significant increase in clinical efficacy is expected for tumor treatment.