ATM Inhibitor And TGF-β Antibody-Loaded Copper Sulfide Nanoparticles For Low-Temperature Photothermal Therapy Of Hepatocellular Carcinoma

Background:Hepatocellular carcinoma(HCC)is the most common primary liver cancer,which is listed as the sixth most common tumor and the third most common cause of cancer-related death.Although surgical resection,radiofrequency ablation,transarterial chemical embolization,and tyrosine kinase inhibitor drug therapy have been proven to have survival benefits for patients with HCC,the therapeutic effect is still not ideal.Photothermal therapy(PTT)is a non-chemotherapy intervention for various types of tumors,which uses photothermal conversion agents to convert light energy into heat to ablate local tumor tissues,and has controllable radiation,low toxicity and high treatment specificity.PTT can combine with chemotherapy,immunotherapy and other therapies.Generally speaking,a temperature of more than 50 ℃ is required to completely ablate the tumor.However,high-temperature photothermal treatment may cause collateral damage to nearby healthy cells and tissues.Therefore,effective tumor ablation at a relatively low temperature(42-45 ℃)is essential for the future clinical application of PTT.However,due to insufficient heating at a low temperature,the thermal damage of tumor cells can be repaired by heat shock protein(HSP),which weakens the anti-tumor efficacy of low-temperature PTT.Copper sulfide nanoparticles(CuS NPs)have become suitable photothermal conversion agents and nano-carriers for drug delivery due to their hollow shell-core structure,high photothermal conversion efficiency and surface modification ability.Ataxia telangiectasia mutated(ATM)is a kinase that is responsible for the overall coordination of cell responses to DNA damage and is a therapeutic target for cancer.KU-60019 is an ATM kinase inhibitor.Compared with other ATM inhibitors,it has higher inhibitory efficiency,pharmacokinetics and bioavailability.An ongoing clinical trial of KU-60019 and two other clinical trials of ATM inhibitors suggest that ATM inhibitors have potential anti-tumor activity.More importantly,some studies have reported the regulatory relationship between ATM and HSP.Purpose:We designed and synthesized hollow copper sulfide nanoparticles(CuS-ATMi@TGF-βNPs)loaded with ATM inhibitor KU-60019 and surface-modified with tumor growth factorβ(TGF-β)antibody.Nanoparticles were studied in three aspects: chemical properties,in vitro function and in vivo therapeutic effect.The conceptual model of ATM inhibitor combined with TGF-β antibody-mediated low-temperature photothermal treatment of HCC proposed in this experiment provides a theoretical and experimental basis for clinical liver cancer treatment.Method:(1)CuS-ATMi@TGF-β NPs were prepared,and their morphology,particle size,potential,stability,dispersibility,photothermal conversion ability,photothermal stability,drug loading and release were studied.(2)Through cell experiments(cellular uptake,LIVE/DEAD cell staining,colony formation assay,cell migration assay,Western blot),we studied the targeting ability,cytotoxicity,cell killing ability,photothermal treatment effect,synergistic treatment effect and underlying molecular mechanism of CuS-ATMi@TGF-β NPs.(3)Through the establishment of H22 tumor-bearing mouse liver cancer model,we studied the biosafety,biodistribution,tumor growth inhibition,photothermal treatment effect,immune activation effect,heat shock protein expression of CuS-ATMi@TGF-β NPs.Result:(1)In this study,the CuS-ATMi@TGF-β NPs nano-photothermal therapeutic agent was successfully synthesized and prepared,which had a unique core/shell structure,and achieved drug loading and surface antibody modification.The large cavity and mesoporous surface structure of CuS-ATMi@TGF-β NPs increased the drug loading capacity.The strong near-infrared absorption capacity,high photothermal conversion efficiency and good photothermal stability made it a suitable carrier for PTT.(2)CuS-ATMi@TGF-β NPs had targeting abilities against tumor cells and were less toxic to normal cells.The ATM inhibitor and TGF-β antibody loaded CuS NPs had a synergistic effect.CuS-ATMi@TGF-β NPs treated with NIR showed strong cell killing ability on tumor cells and significantly inhibited the proliferation and migration of tumor cells.Loaded ATM inhibitors can improve the effect of low-temperature PTT by down-regulating the expression of P-AKT,HSP70 and HSP90.(3)In the H22 tumor-bearing mouse liver cancer model,the treatment of CuS-ATMi@TGF-β NPs + NIR showed good biosafety and the highest tumor growth inhibition rate.NIR treatment promoted the release of ATM inhibitors of nanoparticles and inhibited HSP,thereby improving the photothermal treatment effect at a low temperature.The modification of TGF-β antibody improved tumor targeting and activated the immune system to a certain extent.The combination of ATM inhibitor loading and TGF-β antibody modification synergistically enhanced the anti-liver cancer curative effect of CuS NPs.Conclusion:In summary,ATM inhibitor-mediated chemotherapy,CuS NPs-mediated PTT,and TGF-β antibody-mediated immunotherapy have a synergistic effect,significantly reducing the proliferation of HCC.The conceptual model of ATM inhibitor combined with TGF-βantibody-mediated low-temperature PTT of HCC proposed in this experiment has the potential for future clinical translational applications.