Design Of Modular Peptide Probes Based On Aggregation-induced Emission And Their Application In Cancer Therapy

Due to the complexity,diversity and heterogeneity of cancer,many types of cancer have not been completely eradicated.To this end,researchers have developed many multimodule probes to treat cancer.In order to further improve the bioavailability of multimodule probes and reduce their side effects,it is urgent to develop safe and efficient multi-module probes.Aggregation-induced emission(AIE)molecules are playing an increasingly important role in the biomedical field because of their excellent resistance to photobleaching,high quantum yield,and low background fluorescence.At the same time,the excellent biodegradability,inherent good biological activity,excellent biocompatibility and unique diversity of peptides have laid a solid foundation for their biological applications.Therefore,combining the advantages of AIE molecules and peptides,aiming at improving the utilization rate of probes,starting from improving internalization efficiency,delivery efficiency and time efficiency,and studying the differences between cis/trans isomers of probes,this thesis fully constructs a series of AIE-based modular peptide probes for efficient and safe cancer treatment including the following four aspects:(1)To improve the internalization efficiency of each module of the probes,the modular peptide probes based on AIE can be cleaved into two by matrix metalloproteinase 2(MMP-2)before entering the cell,which reduces the hydrophobicity and particle size of probes.These two parts are efficiently internalized by cells with optimal macropinocytosis and caveolae-mediated endocytosis.The improvement of the cellular internalization efficiency of each module of the probes significantly enhances the probes utilization.Finally,the triple combination therapy of nanofiber therapy,gene interference and photodynamic therapy successfully down-regulates the expression of anti-apoptotic proteins and effectively inhibits the growth of cancer.(2)After being internalized into cells,probes still face the problem of lysosomal degradation that affects delivery efficiency.In order to improve the delivery efficiency of each module of the probes,AIE-based modular peptide probes can bypass lysosomes and directly reach mitochondria through self-regulating endocytic pathways.When the probes reach cancer tissue,MMP-2 can cleave them into two parts.The internalization kinetics of two parts are significantly different by fluorescence monitoring of AIE.The fastentering part can form nanofibers in the lysosome,reduce the expression of protein phosphatase 2A,and change the endocytic pathway of the slow-entering part,so that it can directly reach the mitochondria without passing through the lysosome,realizing the mitochondria-targeted efficient treatment.We also extend the application of this system to the delivery of si RNA and doxorubicin.(3)After reaching the intracellular target site,the probes may still be cleared by the cell due to its prolonged action.Therefore,it is crucial to improve the time efficiency of the probes.To this end,AIE-based modular peptide probes FSP can significantly improve probe utilization by causing rapid cell death through multiple phase separation.After the probes reach the vicinity of cancer cells,they are divided into two parts F and SP by MMP-2.SP quickly binds to the cell membrane to form pores in the cell membrane,destroying the structure of the cell membrane,allowing F to directly enter the cell and combine with a large amount of adenosine triphosphate(ATP)in the cell,causing the phase separation of intracellular proteins and triggering cell function disorder and rapid death.The probes act quickly,significantly improving the utilization of the probes and efficiently killing cancer cells.(4)The utilization of AIE-based modular peptide probes is greatly reformed by increasing internalization efficiency,delivery efficiency,and time efficiency.At the same time,the research on the cis/trans isomer structure of AIE molecules can further enhance the utilization of probes.In order to study the difference between cis/trans structures in biomedical applications,taking tetraphenylethylene(TPE)as an example,a probe RTP is synthesized by introducing the targeting peptide RGD and the structure Pal-RRRR that can be embedded in the cell membrane at both ends of the TPE.Subsequently,high performance liquid chromatography,mass spectrometry,nuclear magnetic resonance,ultraviolet spectroscopy and theoretical calculations confirm the successful synthesis and separation of cis/trans isomer probes(cis-RTP and trans-RTP).Isothermal titration calorimetry experiments show that compared with trans-RTP,cis-RTP can interact more stably with the cell membrane.In-depth study found that there are significant differences between cis-RTP and trans-RTP in terms of cell adhesion,migration,and intracellular pathway experiments.The elucidation of the structural differences between cis-trans isomers of AIE molecules can further improve the utilization of probes,which is conducive to the development of more efficient and better probes.