Intervention Of T Cell Function By Nanomedicine For Enhancing Cancer Immunotherapy

Compared with chemotherapies and radiotherapies,tumor immunotherapies have attracted much attention in recent years due to their high specificity and good therapeutic effect.Immune checkpoint antibody blockade therapies(such as CTLA-4 and PD-1/PD-L1 antibody therapies)and adoptive T cell transfer therapies(such as chimeric antigen receptor T cell therapies)have saved the lives of many patients with lethal cancers.In the past decade,a variety of tumor immunotherapies have been approved for the treatment of cancer patients.And more patients will benefit from tumor immunotherapies in the future.However,the response to immunotherapies requires the infiltration of the original or infused T cells to tumor sites.This thesis focuses on how to improve T cell infiltration and T cell function in the tumor microenviroment.The research contents mainly include the following two parts:1.The insufficient infiltration of T cells in solid tumors hinders the therapeutic efficacy of immune checkpoint blockade antibodies and chimeric antigen receptor(CAR)-T cells.Promoting T or CAR-T cells to efficiently and specifically migrate into solid tumors is critical for improving current immunotherapies,which remains a challenge.Herein,we proposed a CRISPR-based transcriptional activation(CRISPRa)nanomedicine for mobilizing tumor cells to recruit T cells by activating the expression of CXC-chemokine ligand(CXCL)9,CXCL10 and CXCL11.We demonstrated that the CRISPRa nanomedicine could dramatically upregulate the expression of T cell chemokines in tumor cells,thereby facilitating T cell migration directionally into solid tumors.Intratumoral injection of the CRISPRa nanomedicine inhibited the growth of different solid tumors including melanoma,pancreatic,colorectal,and breast cancers.Moreover,tumor-specific versions of the CRISPRa nanomedicine possessing the ability to upregulate the T cell chemokine expression only in tumors rather than normal tissues or cells were constructed.Systemic injection of the tumorspecific CRISPRa nanomedicine specifically recruited T or CAR-T cells into solid tumors and improved the therapeutic effects of immune checkpoint blockade antibodies(anti-PD-L1 or anti-CTLA-4)and CAR-T cells.Thus,the gene-activating nanomedicine provides a promising strategy for enhancing T cell infiltration to improve immunotherapies against solid tumors.2.Protein drugs such as monoclonal antibodies,targeting ligands and receptors are one of the most potential therapeutic drugs in tumor immunotherapies.However,the clinical performance of some protein drugs did not satisfied the expectations.Further research found that immunomodulatory molecules,such as the tumor necrosis factor receptor superfamily(TNFRSF),need to be in a multimerized state to better activate the downstream signaling.In this project,in order to enhance the function of T cells to eliminate tumors or directly induce tumor apoptosis,we developed a series of multimeric protein drugs based on a multivalent protein scaffold(MV)for inducing targeted receptor clustering and enhancing therapeutic antibody efficacy.These polyprotein drugs include MV-OX40 L,which enhances T cell activation,MV-TRAIL,which induces tumor cell apoptosis,and MV-Fc R,which promotes multimerization of therapeutical monoclonal antibodies(such as OX40 antibody and PD-L1antibody)to improve T cell function and tumor therapeutic effects.In the study,we constructed multimeric protein plasmid expression systems.For preparation of multimeric protein drugs,lentivirus was used to construct CHO-K1 stable expression cell lines and affinity chromatography was used to purify the products.And the obtained multimeric protein drugs were further conducted characterization and verified function.In summary,we developed a series of multimeric protein drugs and realized the small batch expression and purification of these protein drugs.Meanwhile we detected the function of these protein drugs both in vitro and in vivo.This work provides a platform and strategy to induce receptor especially TNFRSF aggregation and cross-linking for enhancing downstream cell signaling to directly or indirectly improve the function of T cells to eliminate tumors.