Construction Of Supramolecular Self-assembled Nanoparticles And The Application In Lymphatic Imaging And Targeted Cancer Therapy

Cancer is the second reason of death after cardiovascular disease,accounting for about one-sixth deaths globally.The vast majority of cancer-related deaths（about 90%）are caused by cancer metastasis.At present,the traditional treatments for all kinds of cancer mainly include surgery,chemotherapy and radiation therapy.Cancer with different stages and grades needs patient-specific treatments.And surgery is the most common remedy of local treatments while chemotherapy is the most common therapy of systemic treatments.As the most widely used treatment for solid tumors,radical surgery not only needs to remove tumor tissue and some adjacent normal tissues,but has to remove the surrounding lymph nodes to prevent cancer recurrence caused by tumor metastasis.The destruction of the lymphatic system leads to troublesome complications such as lymphatic leakage and lymphedema,affecting quality of life of patients severely.When the distant organ metastases occur,chemotherapy has become the most common treatment.However,the non-specific cytotoxicity of chemotherapeutic drugs leads severe systemic side effects,which limits the wide application of chemotherapy and brings great suffering to patients.At present,with the concept of precision medicine（PM）becoming widespread,people have begun to focus on the minimal iatrogenic injury and the maximum therapeutic effects based on accurate diagnosis and targeted treatment.In order to achieve precision medicine,sentinel lymph node biopsy（SLNB）has been used to avoid excessive lymph node（LNs）dissection during surgical treatment and targeted drugs has been high-profile in chemotherapy.SLNB needs to use imaging agents to confirm the specific location of the sentinel lymph nodes（SLNs）.Among the traditional imaging methods,near-infrared（NIR）fluorescence imaging is easy to operate,and has high spatial resolution without ionizing radiation.Compared with NIR,near-infrared second region（NIR-II）has stronger tissue penetration,less background noise and scattering,leading to better imaging results.However,there is no FDA-approved NIR-II imaging materials.Currently,indocyanine green（ICG）was found to be able to fluoresce in the NIR-II,which is a FDA-approved NIR fluorescence imaging material with the potential to be used in NIR-II imaging.However,ICG has inherent defects such as easy quenching and low fluorescence quantum yield,which limit its application.At present,the most optimization schemes for ICG were preparing ICG as a nanoprobe,to made ICG more stable.However,there is no ideal method to improve the luminescence properties of ICG.Furthermore,we enhance the specificity of chemotherapy drugs,improve efficacy and reduce side effects by preparing targeted nanomedicines.However,the traditional preparation methods use covalent bonds to link targeted ligands with complicated process and potential cytotoxicity,which greatly limit the exploration of multifunctional nanomedicines.It is necessary to improve the preparation method of targeted nanomedicines.In order to improve the luminescence properties of ICG and adopt a simple preparation method to prepare targeted nanomedicines,we utilize supramolecular strategies to design and construct two kinds of self-assembled nanoparticles.The researches and results are as follows:1.The novel strategy of“isolated cage”assist NIR-II fluorescence lymphatic imaging and intraoperative navigationHerein,we developed two kinds of fluorescence nanoprobes based on“isolated cage”strategy,in which clickable organic fluorophores（ICG-DBCO）were separated by the linear fluoroalkyl chains and alkyl chains.ICG-DBCO bound to the PEG-LRA chains stably via copper-free click chemistry.This strategy could maximize inhibition ofπ-πstacking interactions and internal rotation of ICG in order to minimize aggregation-caused quenching emerged and improve the NIR-II fluorescence imaging performance and photostability.All materials used in these nanoprobes are FDA-approved,so as to facilitate future clinical translation.Then,we demonstrated that polymer side-chain engineering was an effective method to enhance fluorescence quantum yields and fluorescence lifetime by inhibiting theπ-πstaking interactions and self-aggregation of ICG via modulating the polymer–dye molecules interactions in vitro and in vivo.Furthermore,the high-intensity NIR-II fluorescence signal of PCMI NPs clearly visualized the popliteal LNs within 60 min of continuous laser irradiation with an excellent spatiotemporal resolution in vivo.And the signal intensity of PCMI NPs showed an upward trend for 2 h since the nanoparticles entered the lymph node,which indicated that nanoparticle encapsulation of ICG improved the metabolic stability in vivo.Benefiting from the higher spatiotemporal resolution at NIR-II window,the route of lymphatic drainage in hind legs of rabbits could be visualized clearly.Moreover,the SLNs of breast cancer and popliteal LNs could be excised precisely under the guidance of the fluorescence imaging.More importantly,the patency of the lymphovenous anastomosis（LVA）could be presented intuitively with qualitative and quantitative analysis of NIR-II fluorescence signals after intradermal injection of PCMI NPs in the distal of operation region in rabbit.In conclusion,optimizing the performance of ICG through the"isolation cage"strategy based on supramolecular chemistry is of great significance for the development and application of NIR-II fluorescent imaging probes in clinical.2.PSMA-targeted supramolecular nanoparticles prepared from cucurbit[8]uril-based ternary host-guest recognition for prostate cancer therapyWe successfully utilized a supramolecular strategy to design a nanomedicine for targeted drug delivery through cucurbit[8]uril-based host-guest ternary complexation and successfully prepared prostate-specific membrane antigen（PSMA）-targeted supramolecular nanoparticles encapsulating doxorubicin（DOX）.We designed and synthesized two polymers as methylviologen linked with poly（ε-caprolactone）（PCL-MV）and naphthalene linked with polyethylene glycol（Nap-PEG）.PCL-MV and Nap-PEG formed a supramolecular amphiphile through ternary host-guest complexation between MV,Nap and CB[8].Intriguingly,PSMA-617 as a prostate cancer targeting ligand contains a naphthalene group,which allows it to insert into the cavity of CB[8]together with MV,thus endowing the supramolecular nanoparticle with targeting ability.According to the molecular recognition mentioned above,we have successfully prepared DOX-loaded nanoparticles（SNPs@DOX）and PSMA-targeted DOX-loaded nanoparticles（P-SNPs@DOX）.Then we proved both P-SNPs@DOX and SNPs@DOX were internalized by 22RV1 and PC3 cells,while P-SNPs@DOX existed a higher cellular uptake by 22RV1 as PSMA-positive cells.Moreover,in vitro experiments exhibited that P-SNPs@DOX emerged higher cytotoxicity against 22RV1cells,with lower IC₅₀.These studies suggested that the introduction of PSMA enabled nanomedicines to be specifically internalized by cancer cells overexpressing PSMA,thereby increasing the anticancer efficacy.In addition,we also verified that the supramolecular nanoparticles we designed can encapsulate other hydrophobic drugs,such as PTX,proving the system was universally applicable.In conclusion,supramolecular self-assembly is an easy and feasible strategy for building targeted nanomedicines which improves the targeted anticancer efficacy while reduces side effects of drugs.This strategy is an inspiration for the preparation of targeted nanomedicine carriers.In summary,we have constructed two types of nanoparticles by applying supramolecular self-assembly strategy including NIR-II fluorescent probes and targeted nanomedicines,which effectively overcome the inherent defects of materials,and provided a new thought and method for the design and evaluation of targeted nanomedicines and small molecule probes.Meanwhile,our studies have proved that supramolecular nanomaterials have broad application prospects in the field of clinical.