Activating CGAS-STING Pathway With Nanoparticles Delivering A Hybrid Prodrug For Enhanced Antitumor Immunotherapy

Background:Chemotherapy is the main measure of cancer treatment,and chemotherapeutic drugs have efficacy on both primary tumor and metastasis,but many chemotherapeutic drugs enter the body after being nonspecifically distributed resulting in low bioavailability,and the dose and efficacy of the final drugs are limited by systemic toxic side effects.In recent years,nanotechnology has brought a new dawn to cancer therapy.Drug delivery systems constructed based on nanotechnology have unique properties such as high solubility,high stability,tumor high targeting and low toxic side effects.Recent studies have shown that the chemotherapeutic drugs such as cisplatin and camptothecin（CPT）can activate the cGAS-STING pathway through DNA damage and induce immune responses.As an important intracellular immune response pathway,cGAS-STING pathway can effectively enhance antitumor immunity in the regulation of innate immunity and has attracted much attention in tumor immunotherapy.Some chemotherapeutic agents can activate cGAS-STING pathway through DNA damage to induce immune responses.In recent years,chemotherapy combined with immunotherapy,as an emerging treatment,highlights superior synergistic effects and currently plays an important role in promoting the transformation of"immune cold tumors"into"immune hot tumors".Objective:Construction of a platinum prodrug CPT-Pt（Ⅳ）nanodrug delivery system integrating camptothecin and cisplatin;To explore whether it can enhance the chemotherapeutic efficacy of platinum drugs;To analyze its correlation with cancer immunotherapy and the cGAS-STING pathway,in an attempt to elaborate the molecular mechanism that CPT-Pt（Ⅳ）exerts its effect.Methods:In this study,a prodrug CPT-Pt（Ⅳ）nanosystem integrating camptothecin and cisplatin was constructed by the method of oxidative valence;Prodrugs were promoted to self-assemble with ROS sensitive polymers（P1）and m PEG_2k-DSPE by sonication method to form nanoparticles（NPs）with ROS responsiveness;The antitumor activity of NPs was analyzed by MTT assay,Annexin V-FITC/PI double staining,and clonogenic assay in vitro;CT26 colon cancer mice subcutaneous tumor model was constructed and IVIS was applied to monitor the in vivo distribution and tumor suppressive effect of NPs;The contents of related cytokines in mouse peripheral serum were detected by ELISA;The numbers of relevant immune cells in mouse lymph,spleen,and tumor tissues were determined by flow cytometry;A variety of methods including Western blot technique,laser scanning confocal microscope combined with metabolomics were applied to analyze the antitumor mechanism of NPs.Results:This study obtained the following results:（1）A platinum prodrug CPT-Pt（Ⅳ）nanodrug delivery system integrating camptothecin and cisplatin,a kind of ROS responsive nanoparticles（NPs）,was successfully constructed;（2）The results demonstrated that the NPs entered tumor cells via endocytosis and could release cisplatin and CPT in response to ROS,resulting in dual damage to DNA of tumor cells;（3）NPs could effectively inhibit the growth of many kinds of tumor cells such as cisplatin resistant tumor cells;（4）The experimental results showed that the NPs aggregated at the tumor site in a mouse model of colorectal cancer and released cisplatin and CPT in response to ROS;（5）NPs increased tumor infiltration by CD8⁺T cells,induced DC cell maturation and promoted the conversion of"immune cold tumor"to"immune hot tumor",and in vivo experiments confirmed the antitumor effect of immune action;（6）NPs activated the cGAS-STING pathway to exert antitumor effects through a combination of chemotherapy and immunotherapy;（7）The NPs targeted accumulation and trapped in the tumor body of CT26 tumor bearing mice,exhibiting the characteristics of high efficacy and low toxicity.Conclusion:In this study,nanoparticles（NPs）with ROS responsiveness were successfully designed:in vitro,NPs could effectively inhibit the growth of a variety of tumor cells and activate the cGAS-STING pathway via DNA double damage;In vivo,NPs significantly inhibited tumor growth and induced DC cell maturation and tumor infiltration of CD8⁺T cells in a mouse model of colorectal cancer,exhibiting better antitumor efficacy.This study demonstrates that commonly used DNA targeting drugs delivered via nanodelivery system can activate the cGAS-STING pathway in situ and enhance chemo-immunotherapeutic efficacy,thus providing a novel strategy for clinical antitumor therapy.