Induction　of　antigen-specific　immune　response　by　maturing　dendritic　cells　(DCs)　is　a　promising　strategy　for　anti-tumor　immunotherapy.　However,　the　immunosuppressive　tumor　microenvironment　(TME)　and　the　ineffective　tumor-associated　antigen　(TAAs)　presentation　of　DCs　have　hindered　the　efficacy　of　immunotherapy.　Herein,　we　developed　an　immunostimulatory　nanosphere　by　direct　self-assembly　of　cyanine5　labeled　CPG　oligodeoxynucleotide　and　doxorubicin　(Cy5-CPG/DOX).　The　introduction　of　Cy5　into　CPG　could　increase　the　interaction　of　Cy5-CPG　and　DOX,　accelerating　the　formation　of　Cy5-CPG/DOX　nanosphere.　In　vivo,　DOX　can　effectively　downregulate　the　level　of　regulatory　T　cells　to　reverse　and　abrogate　immunosuppressive.　Meanwhile,　DOX　induce　immunogenic　cell　death　to　produce　TAAs,　which　combined　with　Cy5-CPG　promote　the　maturation　of　DCs,　improving　the　TAAs　presentation　ability　of　DCs.　Thus,　the　Cy5-CPG/DOX　could　awaken　the　innate　immune　system,　sensitizing　tumors　to　immune　checkpoint　inhibitors　mediated　by　the　programmed　death-ligand　1　(PD-L1)　antibody.　In　this　way,　the　Cy5-CPG/DOX　combined　with　PD-L1　can　decrease　the　exhaustion　of　cytotoxic　T　lymphocytes,　eliciting　a　durable　systemic　anti-tumor　immunity　to　eradicate　cancer.　Our　work　establishes　a　stable　approach　for　constructing　immunostimulatory　nanodrug,　promoting　the　development　of　progressive　therapeutic　systems.　©　2022　The　Author(s)