Polymer/lipid Composite Nanocarrier Delivering SiRNA For Cancer Immunotherapy

Immunotherapy is the most promising cancer treatment strategy today and has been hailed as the third revolution in cancer therapy.Among them,immune checkpoint blockade is currently the main approach of the immunotherapy for solid tumors.Immune checkpoint antibodies work principally at the protein level by blocking immune checkpoint molecules such as PD-L1,CTLA-4,PD-1,and CD47 on the surface of tumors or immune cells.According to the central law,besides the protein level,people can also intervene in these targets at the genetic level.Likewise,it may also achieve the modulation of immune checkpoint signaling pathways and the activation of antitumor immune response.In this thesis,two nanocarriers were designed and constructed using clinically approved polymers such as polyethylene glycol-poly（lactic-co-glycolic acid）and lipids,which were able to efficiently load nucleic acid drugs and regulate immune-related genes in vivo,enabling new strategies for nucleic acid drug-mediated tumor immunotherapy.The research content of this thesis is mainly divided into two parts.1.We constructed a polymer/lipid composite vesicle-like nanoparticle VNP by the double emulsification methond using clinically approved amphiphilic block copolymer m PEG-b-PLGA and cationic lipid DOTAP to achieve efficient codelivery of si RNA/MTO complexes.si CD47 encapsulated in VNP_{si CD47/MTO}could effectively silence CD47 gene to inhibit the“don’t eat me”signal mediated by CD47-SIRPαaxis on the surface of tumor cells,while MTO inside the nanoparticle induced surface exposure of CRT to upregulate the“eat me”signal,which would synergistically enhance the phagocytosis of tumor cells by macrophages.In the melanoma and colon cancer mouse models,VNP_{si CD47/MTO} could increase macrophage infiltration in tumor tissues,promote DC maturation,increase the number of CD8⁺T cells in tumors,leading to a strong antitumor immune response,eventually achieving significant tumor growth inhibition.This VNP nanocarrier provides a new way to synchronize the delivery of nucleic acid/chemotherapy drugs and achieve tumor immunotherapy.2.We designed and developed a universal polymer/lipid composite nucleic acid drugs delivery system NPs,using three carrier materials including clinically approved amphiphilic block copolymer m PEG-b-PLGA,cationic lipid DOTAP and ionizable lipid DLin-MC3-DMA,through a rational and optimized combination strategy.The NPs system could efficiently bind different types of nucleic acid drugs,such as si RNA,m RNA and CRISPR-Cas9 plasmids.As the si RNA delivery system in vivo,NP_{si RNA} could effectively silence CD47 and PD-L1 genes in mouse melanoma tumor model and elicite a robust antitumor immune response,thus significantly suppressing tumor growth.Moreover,the delivery of m RNA encoding OVA using the NPs system could successfully achieve in vivo translation of OVA protein,which would be processed by DC cells to present SIINFEKL antigenic peptide to activate antigen-specific CD8⁺T cells,thereby producing an excellent immunopreventive effect on OVA-expressing tumor.In this study,the NPs delivery system was prepared from clinically approved polymer and lipid materials,which avoided complicated material synthesis and purification process,and had good versatility for investigators to use and further optimize.