Intervention Of Immune Checkpoint By Nucleic Acid Nanomedicine For Tumor Therapy

Immune checkpoints are a kind of molecules that regulate the function（activation or suppression）of immune cells（T cells,NK cells and myeloid cells）,such as PD-1,TIGIT,CD47and so on.These molecules inhibit the activity of immune cells by binding to their corresponding receptors（PD-L1,CD155,SIRPα,and so on）on tumor cells,macrophages and dendritic cells in tumor microenvironment,allowing tumors to escape immune system.Since Ipilimumab targeting CTLA-4 was approved by FDA in 2011,a number of antibodies targeting PD-1/PD-L1 or CTLA-4 have been got access to clinical usage.Although immune checkpoint antibodies had shown great clinical value,there were limited choices for clinical use except for PD-1,PD-L1 and CTLA-4.With the clinical application of immune checkpoint blockades,reaserchers have found that insufficient T cell infiltration on solid tumors was a key factor of low responsive rate.on the one hand,there is an urgent need to develop a new type of therapy based on immune checkpoint for expanding the clinical choice.On the other hand,promoting T cell infiltration could enhance the efficiency of immune checkpoint blockade.Nucleic acid drugs as a new class of drugs could be applied to intervene the function of target gene or express antibody blockinging immune checkpoint to increase the cytotoxity of immune cells after improving the inhibitory tumor microenvironment.However,delivery of nucleic acid drugs in vivo has great obstacles,such as nucleases in everywhere and cell membranes are electrically repellent to negatively charged nucleic acid.Recently,nanoparticles have been proven to protect nucleic acids from nuclease degradation by wrapping drugs into the core of particles.Meanwhile nanoparticles could be modified on the surface to efficiently and specifically deliver nucleic acid into target cells.With the development of nanotechnology,a variety of nucleic acid drugs have been approved for the treatment of different diseases,such as a si RNA targeting transthyretin carried by liposome（Patisiran）and a m RNA for COVID-19vaccine also carried by liposome（BNT162b2）.Based on the advantage and potential of nucleic acid drugs,this article designed two strategies to intervene the function of immune checkpoints for enhancing the efficiency of tumor immunotherapy.The topic is mainly divided into the following two parts:1)CD155,highly expressed on tumor cells and tumor-infiltrating macrophages,has a dual function that could promote the proliferation of tumor cells as an oncogene and inhibit the function of immune cells as an immune checkpoint,called“onco-immunologic molecule”.Blocking the function of CD155 could have a dual effect of inhibiting the proliferation of tumor cells and eliminating the inhibitory effect to immune cells.We used cationic lipid-assisted polymer nanoparticle（CLAN）to encapsulate si RNA targeting CD155（si CD155）,called“CLAN_{si CD155}”.CLAN_{si CD155} nanoparticles could efficiently deliver si CD155 into melanoma cells and melanoma-infiltrating macrophages,subsequently CD155 was silenced in the two cells.On one hand,the proliferation of tumor cells was suppressed when CD155 was knocked down.On the other hand,the cytotoxic function of immune cells increased because the inhibitory function of CD155 on T cells and NK cells was relieved.Owing to this dual function of CD155 was interrupted by CLAN_{si CD155},tumor microenvironment was improved and the growth of melanoma was significantly suppressed.This research provides a new intervention therapy of immune checkpoints.2)Low expression of chemokines is a key factor that affects the efficiency of immune checkpoint blockade by limiting the infiltration of T cells on solid tumors.Based on this,we constructed a plasmid,which could simultaneously express chemokine CXCL9 and anti-PD-1single-chain variable fragment（sc Fv）in tumor cells,called p PDCX.Our results showed that CLAN nanoparticle carrying p PDCX(CLAN_{p PDCX})could delivery plasmid into B16-F10 cells,and efficiently express CXCL9 andαPD-1 scfv.On one hand,B16-F10 cells transfected CLAN_{p PDCX} could secret CXCL9 to attract T cells to infiltrate into tumor.On the other hand,B16-F10 cells transfected CLAN_{p PDCX} could secretαPD-1 scfv to block the PD-1 to enhance cytotoxic function of T cells.Subsequently,the growth of melanoma was significantly suppressed by the two effects of CLAN_{p PDCX}.This research provides a new strategy for improving the efficiency of immune checkpoint blockade.