Photothermal-immunotherapy Of Melanoma Based On Adenosine Blockade

Background: In recent years,tumor immunotherapy,including chimeric antigen receptor T cell therapy(CAR-T)and immune checkpoint blockade(ICB)therapy,has attracted worldwide attention by activating the immune system to kill tumors.Although clinical immunotherapy has achieved good results in some tumors,it still faces many challenges,such as low immune response and limited anti-tumor efficacy.The low response of the immune response is largely due to the immunosuppressive tumor microenvironment(TME).In the tumor microenvironment,the massive accumulation of cancer-associated fibroblasts(CAFs)and the densely cross-linked extracellular matrix(ECM,such as collagen)hinder the efficient penetration of cytotoxic T cells(CTLs).Immunosuppressive factors(such as adenosine,TGF-β,IL-10,etc.)inhibit the killing effect of T cells on tumor cells,resulting in a decrease in immune response.Adenosine,a key metabolite highly expressed in TME,inhibits tumor immunity and has become a potential metabolic immune checkpoint.Adenosine binds to the adenosine A2 A receptor(A2AR)on the surface of T cells,resulting in the creation of an immunosuppressive microenvironment.Blocking A2 AR can prevent T cell inactivation and promote anti-tumor immune responses.In preliminary clinical trials,A2 AR signaling has achieved good results as a potential immune metabolic regulatory target.Combining immunotherapy with other treatments(such as photothermal therapy)to improve the tumor immune microenvironment can greatly enhance the immune response.Objectives: This study aims to explore a new strategy for synergistic tumor therapy by constructing a new immunostimulatory nanomedicine that combines photothermal therapy with blocking the metabolic immune checkpoint adenosine.To investigate whether nanomedicine can weaken the tumor stromal barrier,enhance T cell infiltration and activity,and enhance anti-tumor immunity through the synergistic effect of photothermal and A2 AR blockade.We further explored whether photothermal immunotherapy based on A2 AR blockade could effectively inhibit tumor growth.Methods: The adenosine A2 A receptor inhibitor SCH442416(SCH)and the targeting aptamer AS1411(Apt)were rationally coupled to black phosphorus nanosheets(BPs)to construct a new immunostimulatory nanomedicine Apt@SCH@BPs.and targeted enrichment to the tumor region.Through the synergistic effect of photothermal effect and adenosine blockade generated by near-infrared laser excitation,it can ablate tumor cells,promote the maturation of dendritic cells,inhibit regulatory T cells,stimulate cytotoxic T cells,and weaken the tumor interstitial barrier(such as cancer related fibroblasts and collagen),simultaneously enhance the infiltration and activity of T cells at the tumor site,and improve the immune response.Results: A novel immunostimulatory nanomedicine,Apt@SCH@BPs,was successfully designed and synthesized,consisting of black phosphorus,adenosine inhibitor SCH442412 and nucleic acid aptamer AS1411.Black phosphorus can not only be used as a drug carrier,but also has good photothermal properties;the adenosine inhibitor SCH442412 can block the binding of adenosine to A2 A receptors,and the nucleic acid aptamer AS1411 induces targeted enrichment of nanodrugs in tumors.Through a series of characterization and cell experiments on Apt@SCH@BPs,it was found that it has high photothermal conversion efficiency and good PTT killing effect.Significant toxic side effects,and can induce immunogenic cell death.Through in vitro and in vivo targeting experiments,it was demonstrated that Apt@SCH@BPs can be efficiently enriched to melanoma tumor sites.By establishing a mouse xenograft melanoma model,we found that photothermal therapy and A2 AR blockade have synergistic effects,not only can effectively ablate tumor cells,but also greatly promote dendritic cell maturation,inhibit regulatory T cells,and reduce Cytotoxic T cells can be stimulated.In addition,Apt@SCH@BPs can effectively weaken the matrix barrier formed by cancer-related fibroblasts and collagen,and promote the infiltration and penetration of T cells.The synergistic therapy effectively inhibited the growth of melanoma.At the same time,Apt@SCH@BPs had no obvious liver and kidney toxicity and had good biocompatibility by H&E staining of mouse liver and kidney,as well as peripheral blood hematology and serum biochemical analysis.Conclusions: Through the synergistic effect of photothermal and A2 AR blockade,Apt@SCH@BPs can not only effectively ablate tumor cells,but also promote DCs maturation,Tregs inhibition,and enhance cytotoxic T cells;reduction of CAFs and collagen can attenuate the intertumoral stroma,leading to Increased infiltration of cytotoxic T cells.This synergistic treatment strategy produced strong immune stimulation and antitumor efficacy in melanoma,effectively inhibited the growth of melanoma,and had a favorable safety profile.As a novel nanomaterial,black phosphorus can be used for the development of immunomodulatory nanotherapeutics due to its high photothermal efficiency,biocompatibility,and degradability.