Dimensional Advantages And Structure-Activity Relationship For Anti-Tumor Nano-Medicines With Sophisticated Configuration

Background:With the recent development of nanoscience,the application of such discipline in biomedical fields,including antitumor therapy,has been deeply explored.However,compared with the few nano-medicines that can really be used in the clinical settings,most nano-medicines are only in the research stage.One of the reasons for that is because many nano-medicines do not have sophisticated configuration for the study of structure-activity relationship,although some of them showed excellent anti-tumor properties.The relationship between the structure and the relevant efficacy has not been found,which leads to the failure of linking the nano-medicines to some therapeutic targets.Therefore,people are not able to acquire enough nano-medicines for anti-tumor applications.If researchers want to design and synthesize more effective anti-tumor nano-medicines,the urgent problem is to explore the structure-activity relationship between nano-medicine for anti-tumor therapy and anti-tumor efficacy.Thus,in order to better understand the structure-activity relationship of nanomaterials and design better anti-tumor nanomedicines,researchers must have a clearer understanding of nanomaterials regarding the effects and advantages at the nanometer level.The dimensional advantages at nanoscale is a kind of size-related effect or structure-activity relationship that is rarely mentioned in the field of nanotechnology but is of great significance for anti-tumor therapy.Nanomaterials in different dimensions have their own characteristics and dimensional advantages regarding anti-tumor applications.Purpose:1.In this study,representative types of zero-dimensional,two-dimensional and three-dimensional nanomaterials were selected respectively,and the advantages of each dimension were fully utilized to design and select suitable nanomaterials with sophisticated configuration according to their characteristics.As one-dimensional nanomaterials do not show good dimensional advantages,they are not discussed in this study.2.Zero-dimensional nanomaterials refer to nanomaterials with relatively simple structural components in all three dimensions at the nanometer size.Due to their simple structure,they often become components of higher-dimensional nanomaterials.The individual application of zero-dimensional nanomaterials has the characteristics of predictable function and controllable degradation.In this study,a kind of nanocluster Cu6NPs was selected as the representative zero-dimensional nanomaterial for chemodynamic anti-tumor therapy.3.Two-dimensional nanomaterials refer to the nanoscale materials with two dimensions far exceeding another dimension.This kind of materials has a large surface and thus has a stronger availability regarding the active area,and it has a unique interaction mode with the cell membranes.Such interaction also makes it have unique function regarding cell membranes.In this study,a two-dimensional conjugated organic framework material CPF-Cu was selected as a representative two-dimensional material to synthesize for photothermal therapy and fast cell staining performance.4.Three-dimensional nanomaterials refer to the nanomaterials composed of zero-dimensional,one-dimensional or two-dimensional materials in nanometer size.Therefore,three-dimensional nanomaterials often emphasize the application of composite functional materials.The complex system in the biological environment often makes composite three-dimensional materials show unparalleled advantages.In this study,the porous organic cage Pt-in-(HR)CC3 was synthesized as the representative of three-dimensional nanomaterials for tumor radiosensitization.Part 1:Zero-dimension nanocluster Cu6NPs towards anti-tumor chemodynamic therapyMethods:1.Cu6NPs was synthesized via a one-pot route by mixing 2-mercaptopyrimidine and Cu(NO3)2·3H2O in dimethylformamide.2.The obtained Cu6NPs was characterized by means including SCXRD,scanning electron microscopy,and transmission electron microscopy.3.The cytotoxicity of Cu6NPs was evaluated against different tumor cell lines and normal cell lines by CCK8 kit.4.In order to further investigate the anti-tumor mechanism of Cu6NPs,related functional experiments were completed such as apoptosis detection,dead and alive staining,and cell cycle detection in vitro.5.The animal experiments were also completed in vivo to verify the anti-tumor effect of Cu6NPs.Different concentrations were used to observe the tumor size by intraperitoneal injection in nude mice with established xenograft model.We have also detected related biological toxicity Cu6NPs in vivo.6.We tracked the UV-Vis absorption spectra and PXRD of Cu6NPs in weak acid solution,and observed the changes of Cu6NPs under tumor microenvironment by scanning electron microscopy and energy dispersive spectroscopy for morphological analysis.7.The chemodynamic anti-tumor mechanism of Cu6NPs was verified by detecting the Fenton-like reaction ability with Cu6NPs and the intracellular ROS by ROS probes.Results:1.Cu6NPs was synthesized and measured their atomically precise structures through the obtained single crystals.2.Cu6NPs was measured to exhibit good stability and good luminescence properties in a non-acidic environment,which indicates that they have potential applications in medical imaging and biological targeting.3.The average particle size of Cu6NPs components measured in aqueous solution is within the range of effective tumor absorption,indicating that it has the potential for anti-tumor applications in terms of size.Whereas,the morphology of Cu6NPs showed a uniform distribution,and its size was smaller than the filtration threshold of the kidney,indicating its minimal potential side effects.4.Our results demonstrate that Cu6NPs exhibits high cytotoxicity to tumor cells,while their ligand components have little cytotoxicity.Meanwhile,Cu6NPs showed lower cytotoxicity to normal cardiomyocytes.5.The tumor cells were treated with different concentrations of Cu6NPs,and it was proved that Cu6NPs promoted tumor cell apoptosis,thereby inhibiting cell proliferation.In particular,Cu6NPs can mediate cell arrest in the G2 phase.6.At the end of the animal experiment,it was found that the tumor volume of the control group increased rapidly;while in the drug-treated group using Cu6NPs,the tumor volume was significantly reduced compared with the control group.No major pathological abnormalities were detected in the major organs of Cu6NPs-treated mice,and no cardiotoxicity,renal toxicity,or hepatotoxicity of Cu6NPs was observed in blood samples.7.It was observed that the structure of Cu6NPs was slowly destroyed in acidity,and the results of EPR demonstrated that a large amount of ROS was generated during the destruction process.While the Fenton-like reactions at different pH values showed that the Fenton-like reactions were stronger at lower pH values.8.The increase of intracellular ROS was detected with the increase of Cu6NPs concentration.Con clusion:1.Based on the characteristics and dimensional advantages of zero-dimensional nanomaterials,we synthesized nanoclusters Cu6NPs for anti-tumor chemodynamic therapy.2.The nanomaterial has transient anti-tumor chemodynamic therapy effect generated by targeting the tumor microenvironment,and Cu6NPs immediately degrades into copper ions and ligands with minimal biotoxic effects.This zero-dimensional nanomaterial has the dimensional advantage of known functions and controllable degradation.Part 2:Two-dimension nano-sized covalent organic framework CPF-Cu towards ultrafast tumor cell staining and anti-tuor photothermic therapyMethods:1.CPF-Cu was synthesized using TCNB and DCH as monomers and CuCl2 as structure directing agent.2.Various material science characterization techniques were used including spherical aberration electron microscopy,PXRD and XPS to explore the structure and material characteristics of CPF-Cu.3.The performance of CPF-Cu was observed regarding ultra-fast tumor cell staining by confocal microscopy,and live cell imaging technology was used to dynamically observe the process of CPF-Cu entering tumor cells.4.A thermal imager was used to detect the heating effect of CPF-Cu under the illumination of light,and calculate the photothermal conversion efficiency.5.The anti-tumor photothermal therapy effect of CPF-Cu was further tested through cell experiments including live and dead staining and CCK8.Results:1.Different characterization techniques were used to demonstrate that CPF-Cu is a nanoscale crystalline framework composed of fused quasi-phthalocyanine centers with uniform elemental distribution and two-dimensional layered features,in which the copper element is single-atom arrangement structure.2.The fluorescence properties of CPF-Cu was observed and determined its fluorescence properties and strong fluorescence quantum yield in COFs.3.Through confocal microscopy,CPF-Cu can be stained within 1 minute,and we use live cell imaging technology to dynamically observe that CPF-Cu can quickly enter tumor cells and has strong stability.4.Through experiments,we measured that the photothermal conversion efficiency of CPF-Cu dissolved in DMEM at a concentration of 100μg mL-1 under the excitation of a light with a wavelength of 660 nm was about 39.3%.5.The anti-tumor photothermal therapy effect of CPF-Cu was further tested through cell experiments.The experimental results showed that after photothermal therapy,tumor cells died by inhibiting proliferation and mediating apoptosis.Conclusion:1.According to the characteristics and dimensional advantages of twodimensional nanomaterials,we synthesized conjugated organic framework CPF-Cu for tumor photothermal therapy.2.The nanomaterial exhibits a strong anti-tumor photothermal therapy effect,and can quickly penetrate the cell membrane of tumor cells for ultra-fast tumor cell staining.It has high material availability of two-dimensional nanomaterials and special interaction with cell membranes which is the dimensional advantage of two-dimensional nanomaterials.Part 3:Three-dimension nano-sized porous organic cage Pt-in-(HR)CC3 towards enhanced anti-tumor radiotherapyMethods:1.Pt-in-(HR)CC3 was synthesized with reference to the method in the previous literature.2.PXRD,SEM,TEM and other characterization methods were applied to characterize Pt-in-(HR)CC3.3.The water solubility of Pt-in-(HR)CC3 was characterized using zeta potential and other assays,and observed its dissolution state in different common biological solvents.4.The biotoxicity of Pt-in-(HR)CC3 was detected in different cell lines by CCK8.5.In vitro cell experiments were performed to verify the radiosensitization properties of Pt-in-(HR)CC3 by means of colony formation assay and apoptosis detection.6.In vitro animal experiments were further applied to detect the radiosensitization performance and biosafety of Pt-in-(HR)CC3.Results:1.The organic cage structure of the synthesized Pt-in-(HR)CC3 was well preserved which was proved by material characterization,and its particle size was in nanometer size.2.It was proved by different means that Pt-in-(HR)CC3 has good water solubility,which can be dissolved in many solvents and medium commonly used in biology.3.The low biotoxicity of Pt-in-(HR)CC3 was examined in different cell lines.4.The radiosensitization performance of Pt-in-(HR)CC3 was verified by in vitro cell experiments,and verified that its mechanisms include generating more reactive oxygen species,mediating tumor cell apoptosis,and inhibiting proliferation and migration.5.The radiosensitization performance was tested and better safety of Pt-in-(HR)CC3 by in vitro animal experiments.Conclusion:1.Based on the characteristics and dimensional advantages of three-dimensional nanomaterials,we synthesized a porous organic cage Pt-in-(HR)CC3 for tumor radiosensitization.2.The nanomaterial exhibits good water solubility and radiosensitization effect.The water solubility of Pt-in-(HR)CC3 comes from porous organic cages and the radiosensitization performance comes from platinum clusters,so Pt-in-(HR)CC3 has the dimensional advantage of the composite function regarding three-dimensional nanomaterials.