BackgroundCombinatorial　immunotherapy　strategies　for　enhancing　the　responsiveness　of　immune　system　have　shown　great　promise　for　cancer　therapy.　Engineered　nanoformulation　incorporated　toll-like　receptor　(TLR)　9　agonist　CpG　ODN　has　shown　more　positive　results　in　suppressing　tumor　growth　and　can　significantly　enhance　other　immunotherapy　activity　with　combinatorial　effects　due　to　the　innate　and　adaptive　immunostimulatory　effects　of　CpG.ResultsIn　the　present　work,　protamine　sulfate　(PS)　and　carboxymethyl　beta-glucan　(CMG)　were　used　as　nanomaterials　to　form　nanoparticles　through　a　self-assembly　approach　for　CpG　ODN　encapsulation　to　generate　CpG　ODN-loaded　nano-adjuvant　(CNPs),　which　was　subsequently　mixed　with　the　mixture　of　mouse　melanoma-derived　antigens　of　tumor　cell　lysates　(TCL)　and　neoantigens　to　develop　vaccine　for　anti-tumor　immunotherapy.　The　obtained　results　showed　that　CNPs　was　able　to　effectively　deliver　CpG　ODN　into　murine　bone　marrow-derived　dendritic　cells　(DC)　in　vitro,　and　remarkably　stimulate　the　maturation　of　DC　cells　with　proinflammatory　cytokine　secretion.　In　addition,　in　vivo　analysis　showed　that　CNPs　enhanced　anti-tumor　activity　of　PD1　antibody　and　CNPs-adjuvanted　vaccine　based　on　the　mixture　antigens　of　melanoma　TCL　and　melanoma-specific　neoantigen　could　not　only　induce　anti-melanoma　cellular　immune　responses,　but　also　elicit　melanoma　specific　humoral　immune　responses,　which　significantly　inhibited　xenograft　tumor　growth.　Furthermore,　CD16　CAR-T　cells　were　generated　by　expressing　CD16-CAR　in　CD3(+)CD8(+)　murine　T　cells.ConclusionOur　results　eventually　showed　that　anti-melanoma　antibodies　induced　by　CNPs-adjuvanted　TCL　vaccines　were　able　to　collaborate　with　CD16-CAR-T　cells　to　generate　an　enhanced　targeted　anti-tumor　effects　through　ADCC　(antibody　dependent　cell　cytotoxicity)　approach.　CD16　CAR-T　cells　has　thus　a　great　potential　to　be　an　universal　promising　strategy　targeting　on　solid　tumor　synergistic　immunotherapy　via　co-operation　with　TCL-based　vaccine.