Investigation On The Irradiation Activation Mechanism Of Rare Earth Nanomaterials For Tumor Therapeutic Efficacy

Cancer become the one of world’s leading cause of death,researchers have been exploring new methods and technologies for cancer diagnosis and therapy.Exogenous electromagnetic wave irradiation therapy（including radiotherapy,photodynamic therapy,etc.）,which use the characteristic of some micro/nanomaterials could be excited under specific electromagnetic wave irradiation,with the resultant emitted energy being able to cause spontaneous apoptosis of cells,and improve the efficiency and accuracy of cancer treatment,reduce side effects,and has become one of the hot spots in the frontier research field of tumor diagnosis and treatment.The clinical efficacy of this technology mainly depends on the mutual coupling between electromagnetic wave irradiation and nanofunctional materials,and also depends on the biological penetration of the irradiated electromagnetic wave that matched to micro/nanomaterials.Currently,the interconnection mechanism between some electromagnetic wave irradiation and nanofunctional materials is still unknown.Firstly,the identification of nanomaterials with desirable properties and optimal propensity to produce reactive oxygen species（ROS）under high penetration and low energy electromagnetic wave irradiation is still a stumbling block.For instance,photosensitizer materials utilized in photodynamic therapy with an ultraviolet-visible light excitation window,have a meager1 mm penetration depth for the human body.Near infrared light（NIR）has a larger penetration depth（about cm level）to organisms,in order to expand the clinical application of these materials for deeper underlying tissue therapy,upconversion nanoparticles were applied by loaded with traditional organic photosensitizers to produce ROS,however,this way is very inefficient and requires a high powered NIR laser for irradiation.Secondly,the radiosensitization mechanism of clinical tumor therapy needs to be clarified.Radiosensitization method utilizes the photoelectric effect and Compton effect generated by the interaction between the radiosensitized nanomaterials targeted at the tumor site and X-rays,increases the radiation energy absorption at the tumor site and elevates the efficiency of radiotherapy with reducing side effects of normal tissues.Since X-ray diagnosis and treatment equipment have been widely used in clinical departments,courtesy of its deep tissue penetration attributes,in that regard radiotherapy sensitization has a great clinical application value.From existing studies,there is consensus that when radiosensitizing materials interact with X-rays,those nanosensitizers liberate secondary electrons such as photoelectrons,Auger electrons and Compton electrons,etc.,which are the only factor can cause apoptosis of surrounding tumor cells or tissues.However,there are great discrepancies between theoretical calculation data and in vitro and in vivo experimental results,which reaffirms that the mechanism behind radiotherapy sensitization is not clear,and requires urgent and thorough probing in order to understand underlying mechanisms.Rare earth nanomaterials possess characteristics such as:high atomic number,rich energy levels and atomic orbital binding energy in the energy range of clinical radiotherapy.These properties make micro/nanomaterials favorable for the molecular design and preparation of electromagnetic wave energy conversion materials in cancer therapy;additionally,the good biocompatibility,stable optical properties,and easy to modify on the surfaces of rare earth nanomaterials to meet different targeting and therapeutic design requirements.Therefore,in this thesis,the two research hotspots of the mechanism of ROS production directly excited by near-infrared light and the sensitization mechanism of X-ray radiotherapy of rare earth nanomaterials are carried out as follows:（1）For seeking alternative ROS-generating micro/nanomaterials that can be stimulated by NIR light for photodynamic therapy applications,we reasoned that achieving a long half-life of electrons in the excited state of about 10^-3seconds may be the key factor for the production of reactive oxygen species by photosensitizer molecules,and considering that the excited triplet states of thulium ions in NIR absorption region have similar lifetime electrons and low fluorescence emission.In this study,the reactive oxygen species produced by Tm₂O₃nanoparticles under the irradiation of 808 nm low-power non-laser near-infrared light source were confirmed for the first time by using a variety of characterization experimental methods.The effect of surface modification of Tm₂O₃nanoparticles on the production of ROS under the excitation of laser or non-laser light sources with different wavelengths such as visible and near-infrared are studied,and find out the singlet oxygen quantum yield of Tm₂O₃nanoparticles is up to 37%.The near-infrared irradiated ROS generation mechanism of Tm₂O₃nanomaterials is expounded,and the key role of the ³H₄excited state with millisecond lifetime and low radiation transition characteristics of Tm ions would generate ROS via electron transfer or energy transfer is verified.Animal experiments have not only confirmed that the near-infrared photodynamic of Tm₂O₃nanoparticles has a significant tumor inhibitory effect,but also verified the great potential of low output power wide band halogen lamp in near-infrared photodynamic clinical treatment in the future.（2）In order to explore the radiosensitization mechanism,according to the characteristic of K-edge effect of micro/nanomaterials in X-ray absorption,that is,when the photon energy of X-ray is close to or equal to the K-edge binding energy of the material,the probability of photoelectric effect will increase one order of magnitude.In this work,four rare earth nanoparticles with different K orbital electron binding energies are designed:Na Tm F₄,Na Tm_0.6Lu_0.4F₄,Na Tm_0.4Lu_0.6F₄and Na Lu F₄;After calculated by simulation,it is found that under the irradiation close to the 59.4 ke V energy,which is the atomic orbital binding energy of Tm element,the secondary electrons quantity were increased with percentage of Tm;however,when X-ray energy was closed to the atomic orbital binding energy of Lu element（63.3 ke V）,the secondary electron quantity of the four nanoparticles were in the opposite order.Due to the medical X-ray has intrinsic high intensity peaks at 59 ke V,under the ke V level X-ray irradiation of CT imaging equipment,the nanomaterials with higher proportion of Tm element present stronger CT image effect,indicating that the above four rare earth nanomaterials have different X-ray absorption capacity;The results of radiosensitization of cells showed that,among them,Na Tm F₄indicated a stronger DNA damage to U87 tumor cell,there were no differences in the results of U87 cells radiosensitization among the rest of three types of rare earth nanomaterials;Under the 4 Gy X-ray irradiation,there were also no differences in the amount of ROS produced by the above four rare earth nanomaterial,but the intracellular ROS generated by four nanosensitizers are different:Na Tm F₄rare earth nanomaterials are slightly high,and there is no significant difference between the other three rare earth nanomaterials.Based on the influence of four rare earth nanomaterials on the U87 cells’radiosensitization effect,this thesis infers that under ke V level X-ray irradiation,in addition to the radiosensitization effect of secondary electrons,reactive oxygen species are also the main factor in radiosensitization mechanism.（3）Due to the reactive oxygen species with limited effective distance and mainly attack mitochondria to cause apoptosis,for further verify the correlation between the rare earth nanomaterials’radiosensitization of the reactive oxygen species and secondary electrons,an animal experimental studies were settled to explore the relationship between radioenhancement effect and organelle targeting of rare earth nanoparticles.The results shown that the rare earth nanomaterials used in the experiment all have significant tumor growth inhibition effect.It is the first time to confirm that the rare earth nanomaterials（Na Gd F₄:Yb,Er）have radiosensitization effect;even the accumulation amount in tumor tissue of UCN-FA was higher than UCN-TPP and UCN-FA/TPP,but the number of nanomaterials（UCN-FA,UCN-TPP and UCN-FA/TPP）targeted in mitochondria were similar;Different surface modifications of rare earth nanomaterials Na Gd F₄:Yb,Er would lead to different results in phagocytosis and intracellular targeting of U87 cells,and then affect the differences in apoptosis pathways of U87 cells.Therefore,their radiobiological effects are shown as a comprehensive effect of secondary electrons and reactive oxygen species,that is,the rare earth nanomaterials in the cytoplasm mainly act on destroying DNA double bonds,however,rare earth nanomaterials targeting mitochondria mainly apoptotic U87 tumor cells through reactive oxygen species.It is further confirmed that the radiosensitizing effect of rare earth nanomaterials should be the result of the combination of their cellular uptake and intracellular targeting,and the radiosensitizing mechanism should include two key factors:secondary electrons and reactive oxygen species.In summary,the thulium oxide nanomaterials that could directly produce ROS under808 nm near-infrared light excitation were unearthed for the first time,followed by a high quantum efficiency.The mechanism of reactive oxygen production in the near-infrared band is related to the millisecond lifetime of the ³H₄excited state and low radiation transition of Tm ions.Moreover,it was found for the first time that the reactive oxygen species produced by rare earth nanomaterials also play an important role in the radiosensitization mechanism under ke V level X-ray irradiation.The above findings ushered in a new perspective to understand the enigma that is the mechanism for harnessing X-ray radiosensitization for efficient and accurate tumor therapy.