Design And Synthesis Of Intelligent Nanogels For Tumor Theranostics Applications

Cancer has become a global medical problem due to its high incidence and mortality.The “early detection and early treatment” has become an effective strategy to reduce its morbidity and mortality.In recent years,nanomedicine,which is a product of the combination of nanotechnology and pharmaceutical science,has brought new opportunities for accurate diagnosis and efficient treatment of cancer.For instance,two separate processes/functions of clinical diagnosis and treatment can be integrated into one nanocarrier to form a theranostic nanoplatform for real-time,accurate diagnosis and simultaneous treatment of various disease.The key technology is to build a suitable nanocarrier platform to integrate therapeutic agents and/or contrast agents and deliver them to the lesions efficiently.Among many various nanocarriers,nanogels（NGs）,which possess many advantages,such as high loading capacity,good stability,ease of functionalization,multiple responsiveness,and ease to be taken up by cells,have been widely applied in drug delivery and tissue engineering fields.As two typical representatives,polyethyleneimine（PEI）NGs and poly N-vinylcaprolactam（PVCL）NGs,possessing unique features like ease of synthesis,controllable size and easy to dope with inorganic nano-components to form a multifunctional hybrid system,have shown good application prospects in nano-biomedicine,especially in the field of tumor diagnosis and treatment.In this thesis,we used the polymeric NGs as nanocarriers to form a series of intelligent hybrid NGs platforms to explore strategies for accurate tumor diagnosis and efficient treatment.We first constructed PEI-based intelligent hybrid NG platform for magnetic resonance（MR）/photoacoustic（PA）imaging-guided tumor photothermal therapy（PTT）.Then,considering that the inverse microemulsion method lacks the flexibility in controlling the physical/chemical properties of PEI NGs,we used the precipitation polymerization method,which has better flexibility in tunning the physicochemical properties of NGs,to developed a series of PVCL NGs with different physicochemical properties and constructed an in vitro 3D tumor spheroid model to investigate the influence of physical/chemical properties of PVCL-based NGs on their penetration into 3D tumor spheroid.Finally,considering the radioresistance of mostsolid tumors,we further functionalize the PVCL NGs with good tumor penetration properties constructed in the previous section to develop an intelligent PVCL NGs platform for computed tomography（CT）/MR imaging and full-process sensitized radiotherapy（RT）of tumors.The main contents were detailed as follows:（1）The functionalization of PEI NGs and its application in MR/PA imaging-guided tumor-targeted PTTThe biological barriers often lead to the premature metabolic clearance of nanodrugs in vivo and low delivery efficiency at tumor sites.Zwitterion modification is beneficial to make the nanoplatform avoid the specific adsorption of various biomolecules during the blood circulation process,thereby prolonging their blood circulation time.In addition,targeted modification will help nanoplatforms efficiently identify tumor cells and accumulate in tumor sites.Basing on these,our study presents a Gd-and Cu S-integrated PEI NGs platform with folic acid（FA）and 1,3-propanesultone（1,3-PS）modification for tumor theranostic applications.This intelligent hybrid nanoplatform is expected to efficiently accumulate in the tumor site to achieve MR/PA dual-modal imaging and highly efficient PTT under the NIR-II laser irradiation.Firstly,we prepared PEI NGs via an inverse emulsion method,sequentially modified it with Gd chelates,targeting ligand FA and 1,3-PS,and finally used it as nanoreactor to synthesize in situ Cu S nanoparticles（NPs）inside to form hybrid NGs（Gd/Cu S@PEI-FA-PS NGs）for MR/PA imaging-guided tageted PTT of KB tumor.The as-synthesized Gd/Cu S@PEI-FA-PS NGs with a mean size of 85 nm exhibit good water dispersibility and protein resistance property,admirable r1 relaxivity（11.66 m M-1s-1）,excellent NIR-II absorption feature,high photothermal conversion efficiency（26.7%）,and FA-mediated targeting specificity to cancer cells overexpressing FA receptor（FAR）.In vivo experimental results show that the developed Gd/Cu S@PEI-FA-PS NGs can efficiently enrich in the tumor site and achieve enhanced MR/PA imaging effect.Meanwhile,combined with NIR-II（1064 nm）laser irradiation,the Gd/Cu S@PEI-FA-PS NGs can effectively ablate tumor and inhibit its growth.Therefore,by expanding the types of targeting ligands,the hybrid PEI NGs platform may be used for MR/PA imaging-guided efficient PTT of various tumors.（2）Study on the property modulation and tumor penetration capacity of PVCL-based NGsCurrent nanomedicine suffers from a big challenge since most of the nanocarrier systems lack the desired tumor penetration depth,thereby limiting their clinicaltranslation.The physical/chemical parameters of nanomaterials（e.g.,size,flexibility,and surface modification）are important ones affecting their tumor penetration properties.Basing on these,we report the synthesis of PVCL NGs with different physical/chemical properties by taking advantages of PVCL NGs such as ease of synthesis,convenient adjustment of surface physical/chemical properties and good biocompatibility.Additionally,3D tumor spheroid model was established to simulate the tumor environment in vivo to evaluate the tumor penetration capability of PVCL NGs.Using VCL as the main monomer,oligo（ethylene glycol）acrylate（OEGA）and glycidyl methacrylate（GMA）as comonomers,the PVCL/OEGA/GMA NGs with different hydrodynamic radii（Rh）,crosslinking densities,and chemical compositions were synthesized by adjusting the surfactant content,reaction temperature,cross linker content and comonomer types.Subsequently,the influence of different physical/chemical parameters of NGs on its penetration in tumor sites was investigated by constructing an in vitro 3D tumor spheroid model.The in vitro 2D cell experiment results indicate that the as-synthesized PVCL/OEGA/GMA NGs display the desired cytocompatibility,and the NGs with a smaller size display more enhanced cellular uptake than those having a larger size.Likewise,3D tumor spheroid penetration results indicate that the PVCL/OEGA/GMA NGs with Rh less than 200 nm can easily penetrate the 3D tumor spheroid,whereas the permeability of the NGs with Rh larger than 200 nm is greatly reduced.In addition,our data show that NGs with varying crosslinking densities,OEGA contents and OEGA chain lengths do not have any appreciable changes in terms of their cellular uptake and penetration in 3D tumor spheroid model.The findings are expected to provide theoretical guidance for the design of a novel NGs platform with excellent tumor penetration properties that can efficiently deliver contrast agents or therapeutic agents to the tumor.（3）The functionalization of PVCL NGs and its application in CT/MR imaging-guided full-process tumor radiosensitizationRegulating the tumor microenvironment and thus enhancing the sensitivity of tumors to radiation is an important method to improve the tumor RT effect.Herein,we report the construction of a multifunctional hybrid NG platform with Au and Mn O2 NPs co-loaded in the PVCL NGs,which utilize the unique features like good biocompatibility and easy to dope with inorganic nano-components of PVCL NGs,for tumor molecular imaging-guided “full-process” sensitized RT of tumors.Firstly,PVCL NGs were synthesized via precipitation polymerization asmentioned in Section 2 and in situ loaded with Au and Mn O2 NPs to form the intelligent hybrid NGs platform（PVCL-Au-Mn O2 NGs）.Finally,the PVCL-Au-Mn O2 NGs were used for CT/MR dual-modal imaging-guided “full-process” sensitized RT of tumors.The formed PVCL-Au-Mn O2 NGs are about 121.5 nm in size and exhibit good colloidal stability.In vitro and in vivo results indicate that the PVCL-Au-Mn O2 NGs can efficiently generate reactive oxygen species through a Fenton-like reaction to result in cell cycle distribution toward highly radiosensitive G2/M phase prior to X-ray irradiation,localize the radiation dose to sensitize the RT of cancer cells under X-ray irradiation,thus inducing significant DNA damage,and further prevent DNA-repairing process after RT through the continuous production of O2 to relieve tumor hypoxia.Likewise,PVCL-Au-Mn O2 NGs can perform CT/MR imaging of tumors,providing imaging guidance for precise radiotherapy.These findings provide a theoretical basis for the development of a new nanoplatform for efficient treatment of “full-process” radiosensitization.In summary,this thesis carries out different functionalizations of polymeric NGs from different perspectives,including theranostics（MR/PA imaging-guided PTT,CT/MR imaging-guided RT）,enhancement of the accumulation of nanomaterials in tumor sites（extension of the blood circulation time,receptor-mediated targeting capacity,and synthesis of nanomaterials with suitable sizes）,and multiple RT sensitization（regulation of tumor microenvironment,deposition of radiation energy in tumor cells,and prevention of DNA repair process）,to explore tumor precision diagnosis and efficient treatment strategies.These findings will strongly promote the application of polymeric NGs in biomedicine,especially the precision diagnosis and treatment of tumors,and provide new ideas for guiding clinical transformation.