Investigations of TiO2 NP as radiation dose enhancement agent: in vitro and phantom based studies

Radiotherapy is one of the basic methods for cancer treatments. This extremely valuable and effective technique deliver high therapeutic doses of ionising radiations to the malignant cells to shrink tumours and kill cancer cells in a way that is safer and more reliable. The source of this ionising radiation is typically high energy photon or electron beams, which are potentially carcinogenic and/or deadly to all cells at high dosage. Therefore, the major obstacle of planning and delivery of radiotherapy is the preservation of healthy surrounding tissues by limiting the delivered radiation dose to the tolerance levels of normal tissues while still ensuring the effective targeting of tumour volumes to eradicate it. Developments in the field of nanotechnology have potentially provided effective radiotherapy techniques through the use of high and medium atomic number (Z) nanostructure materials as radiosensitisation agents. The high Z nanoparticles (NPs) such as gold, bismuth compound, iodine and gadolinium have already been successfully utilised as radiosensitiser agents when applied to tumours for in vitro, in vivo and even in some preclinical trials. However, the high and medium Z materials are more toxic than low Z elements, therefore investigation of radiosensitasation induced by low Z elements have become more attractive. Several studies have been conducted to test the low Z nanoparticles as dose enhancing agents. Most of these studies were in the field of UV and X-rays of kilovoltage energy ranges. This thesis’ research extends thier application to include the most common form of radiotherapy i.e. using megavoltage range of X-rays. The aims of this thesis are focused on investigations of employing low Z materials and particularly anatase titanium dioxide nanoparticles (TiO2 NPs) as potential radiosensitisation agent and as imaging agent too. This research was conducted by two main ways, one by using phantoms (PRESAGE® dosimeters) and the other by in vitro using two types of cell lines, cultured human keratinocyte (HaCaT) and prostate cancer (DU145) cells, and both methods were aimed at determining the effects on NPs on the radiation dose enhancement at both low (kilovoltage) and high (megavoltage) radiotherapy X-ray beams. Furthermore, TiO2 NPs were activated via proton beam to investigate for their suitability as diagnostic agent hence this nano-compound qualifies to be true theranostic agent. Several characteristics of TiO2 NPs, which make them ideally suited for application in radiotherapy, are investigated throughout this research. Anatase TiO2 NPs were synthesised, characterised and functionalised to allow dispersion in culture medium for in vitro studies and halocarbons (PRESAGE® chemical compositions) for phantom based studies. The fabricated PRESAGE® dosimeters/phantoms were scanned to obtain the physical and radiological properties and further to determine the radiation dose enhancement induced by TiO2 NPs. Clonogenic and cell viability assays were employed to determine cells survival curves from which the dose enhancement levels “radiosensitisation” are deduced. The dose enhancement produced experimentally by 0.5, 1 and 4 mM TiO2 NPs concentrations for phantom and in vitro studies irradiated with 1-8 Gy ranges of radiation doses was quantified for kilovoltage and megavoltage energies of external X-ray radiation sources. Furthermore, the TiO2 NPs were activated via proton beam and the energy spectrums were acquired using Germanium detectors. The radiolabeled anatase TiO2 NPs were imaged using positron emission tomography (PET) scanner. One aspect of this research was to demonstrate that the TiO2 NPs were typically synthesised to achieve highly pure and uniform anatase nanocrystalline structure. This form of NPs creates more free radicals and effectively generates more reactive oxygen species (ROS) since it has larger surface to volume ratio. These features combined have a high impact in damaging DNA molecule of biological system during irradiation. In phantom studies, the radiation modifying effects of incorporating of PEG functionalised anatase TiO2 NPs in the formulation of the water equivalent 3D PRESAGE® dosimeter were explored. The dose enhancement factors (DEFs) were quantified and then the results were validated with the biological studies. The results clearly demonstrates that the sensitivity of the dosimeter increases to the radiation doses with the concentration of the TiO2 NPs incorporated in the composition of the PRESAGE® dosimeter. Furthermore, the measured DEF was significant at 80 kV compared to the negligible dose enhancement detected at 6 MV X-ray energy beams. The In vitro studies, TiO2 NPs proved to be cytocompatible to cells, even at very high concentrations. The DEFs were deduced from the data analysed in the form of cell survival curves. The result indicates that radiosensitisation induced by TiO2 NPs was significant at kilovoltage range of energy which the maximal dose enhancement was observed at 80 kV. Furthermore, significant radiosensitisation was observed for in vitro study at megavoltage energy beam. Higher radiosensitisation were obtained for low energy x-rays compared to the high energy ones. Generally, with the inclusion of TiO2 NPs in the target, same fraction of cells were destroyed with lower radiation doses compared to the case of absence of TiO2 NPs. This means if the TiO2 NPs are added to the biological target, a reduction of external dose of an order of magnitude can be achieved to deliver the same local control as without the inclusion of TiO2 NPs for treatments with kilovoltage and megavoltage X-rays beams. This reduction of delivered radiation dose to the target results in reducing the dose to the surrounding normal tissues during treatment that is the primary concern in all radiotherapy treatment procedures. Hence, TiO2 NPs are considered to be an efficient dose enhancer agent and have a great potential value for future clinical radiotherapy applications. In addition, the radiobiological effect of amino functionalised anatase TiO2 NPs on HaCaT and DU145 cell lines were investigated. The linear (α) and the quadratic (β) radiobiological-parameters were extracted from the cell survival curves in order to describe the DNA damage by radiation. The results clearly demonstrate that α value significantly increases with the inclusion of TiO2 NPs while β values do not show any predictable trend. This increase in α value indicates that the probability of double strand DNA breakage increases with the presence of NPs in the target. Accordingly, the DEF results for in vitro and phantom based studies showed good agreement with the hypothesis of α value increases as a consequence of inclusion TiO2 NPs in the target. There are measurable differences in the level of produced DEF between biological and phantom based studies at MV energy. The PRESAGE® dosimeters showed lower enhancements in radiosensitivity than cell in culture studies. This is due to PRESAGE® dosimeters is only able to detect the free electron generated as a result of photoelectric effect, Compton scatter and/or Auger effect and not being suited to detect the generated ROS (Biochemical effects) due to a lack of free water molecules in its structure, whereas cells can be affected by many other biochemical factors, such as the generated ROS which is added to the stress caused by generated electron free radicals, and this result in higher radiosensitivity. Therefore, the ROS generated from amino functionalised anatase TiO2 NPs upon exposed to radiotherapy X-ray energy beam was investigated. Aqueous solutions without and with the presence of TiO2 NPs was exposed to 6 MV beam. The result clearly shows that the level of generated ROS was proportionally dependent on the TiO2 NPs concentration. This explains that biochemical effects need to be considered as a key factor for enhancing the cellular radiosensitivity with the presence of nanoparticles, which would be an important consideration for in vitro and in vivo radiosensitivity measurements. Finally, the TiO2 NPs are proposed as a reliable potential candidate for producing nuclear medical radioisotopes via proton activation. The results demonstrate that intense peak was observed at 511 keV which correspond to the γ-ray resulted from electron-positron annihilation. This γ-ray peak is the most important radioisotope for potential nuclear medicine imaging applications using PET. Recently, several innovative for new radiopharmaceutical evolution potentially suggest β- and α emitters. Therefore, the produced 47Sc radionuclide is a promising therapeutic agent for preparing radiolabeled antibodies due to its favorable β- emission energy (162 keV) which decays to stable 47Ti (100% β- emission ), and to its moderate half-life (T1/2 = 3.35 d). To conclude, this research shows that TiO2 NPs improve the efficiency of dose delivery, which has implications for future radiotherapy treatments. The TiO2 NPs can also be used as a potential imaging agents hence with these findings renders these NPs as theranostic agents with dual effects (i.e. imaging and dose enhancer agent) simultaneously if it is in the targets

Titanium dioxide nanoparticles as radiosensitisers: An in vitro and phantom-based study

Objective: Radiosensitisation caused by titanium dioxide nanoparticles (TiO2-NPs) is investigated using phantoms (PRESAGE® dosimeters) and in vitro using two types of cell lines, cultured human keratinocyte (HaCaT) and prostate cancer (DU145) cells. Methods: Anatase TiO2-NPs were synthesised, characterised and functionalised to allow dispersion in culture-medium for in vitro studies and halocarbons (PRESAGE® chemical compositions). PRESAGE® dosimeters were scanned with spectrophotometer to determine the radiation dose enhancement. Clonogenic and cell viability assays were employed to determine cells survival curves from which the dose enhancement levels "radiosensitisation" are deduced. Results: Comparable levels of radiosensitisation were observed in both phantoms and cells at kilovoltage ranges of x-ray energies (slightly higher in vitro). Significant radiosensitisation (~67 %) of control was also noted in cells at megavoltage energies (commonly used in radiotherapy), compared to negligible levels detected by phantoms. This difference is attributed to biochemical effects, specifically the generation of reactive oxygen species (ROS) such as hydroxyl radicals (•OH), which are only manifested in aqueous environments of cells and are non-existent in case of phantoms. Conclusions: This research shows that TiO2-NPs improve the efficiency of dose delivery, which has implications for future radiotherapy treatments. Literature shows that Ti2O3-NPs can be used as imaging agents hence with these findings renders these NPs as theranostic agents

Maximizing Soil Carbon Sequestration: Assessing Procedural Barriers to Carbon Management in Cultivated Tropical Perennial Grass Systems

The natural capacity of the terrestrial landscape to capture and store carbon from the atmosphere can be used in cultivated systems to maximize the climate change mitigation potential of agricultural regions. A combination of inherent soil carbon storage potential, conservation management, and rhizosphere inputs should be considered when making landscape‐level decisions about agriculture if climate change mitigation is an important goal. However, the ability to accurately predict soil organic carbon accumulation following management change in the tropics is currently limited by the commonly available tools developed in more temperate systems, a gap that must be addressed locally in order to facilitate these types of landscape‐level decisions. Here, we use a case study in Hawaii to demonstrate multiple approaches to measuring and simulating soil carbon changes after the implementation of zero‐tillage cultivation of perennial grasses following more than a century of intensive sugarcane cultivation. We identify advancements needed to overcome the barriers to potential monitoring and projection protocols for soil carbon storage at our site and other similar sites

Ex vivo culture of intact human patient derived pancreatic tumour tissue

The poor prognosis of pancreatic ductal adenocarcinoma (PDAC) is attributed to the highly fibrotic stroma and complex multi-cellular microenvironment that is difficult to fully recapitulate in pre-clinical models. To fast-track translation of therapies and to inform personalised medicine, we aimed to develop a whole-tissue ex vivo explant model that maintains viability, 3D multicellular architecture, and microenvironmental cues of human pancreatic tumours. Patient-derived surgically-resected PDAC tissue was cut into 1-2 mm explants and cultured on gelatin sponges for 12 days. Immunohistochemistry revealed that human PDAC explants were viable for 12 days and maintained their original tumour, stromal and extracellular matrix architecture. As proof-of-principle, human PDAC explants were treated with Abraxane and we observed different levels of response between patients. PDAC explants were also transfected with polymeric nanoparticles + Cy5-siRNA and we observed abundant cytoplasmic distribution of Cy5-siRNA throughout the PDAC explants. Overall, our novel model retains the 3D architecture of human PDAC and has advantages over standard organoids: presence of functional multi-cellular stroma and fibrosis, and no tissue manipulation, digestion, or artificial propagation of organoids. This provides unprecedented opportunity to study PDAC biology including tumour-stromal interactions and rapidly assess therapeutic response to drive personalised treatment.John Kokkinos, George Sharbeen, Koroush S. Haghighi, Rosa Mistica C. Ignacio, Chantal Kopecky, Estrella Gonzales-Aloy ... et al

A higher effort-based paradigm in physical activity and exercise for public health: making the case for a greater emphasis on resistance training

It is well known that physical activity and exercise is associated with a lower risk of a range of morbidities and all-cause mortality. Further, it appears that risk reductions are greater when physical activity and/or exercise is performed at a higher intensity of effort. Why this may be the case is perhaps explained by the accumulating evidence linking physical fitness and performance outcomes (e.g. cardiorespiratory fitness, strength, and muscle mass) also to morbidity and mortality risk. Current guidelines about the performance of moderate/vigorous physical activity using aerobic exercise modes focuses upon the accumulation of a minimum volume of physical activity and/or exercise, and have thus far produced disappointing outcomes. As such there has been increased interest in the use of higher effort physical activity and exercise as being potentially more efficacious. Though there is currently debate as to the effectiveness of public health prescription based around higher effort physical activity and exercise, most discussion around this has focused upon modes considered to be traditionally ‘aerobic’ (e.g. running, cycling, rowing, swimming etc.). A mode customarily performed to a relatively high intensity of effort that we believe has been overlooked is resistance training. Current guidelines do include recommendations to engage in ‘muscle strengthening activities’ though there has been very little emphasis upon these modes in either research or public health effort. As such the purpose of this debate article is to discuss the emerging higher effort paradigm in physical activity and exercise for public health and to make a case for why there should be a greater emphasis placed upon resistance training as a mode in this paradigm shift