Targeted Delivery Of Doxorubicin By Acid-Sensitive Nanoparticles Reverses Resistance To PD-L1 Checkpoint Inhibitors In Lung Cancer

Background:Immune checkpoint blockade(ICB),especially anti-PD-L1treatment,promotes new hope for cancer treatment.However,the acidic tumor microenvironment(TME)may lead to its treatment resistance and immune escape.Therefore,we designed acid-sensitive nanoparticles to activate anti-tumor immunity and reverse PD-L1 resistance by altering the acidic microenvironment combined with targeted delivery of doxorubicin.Methods:Doxorubicin(DOX)-loaded calcium carbonate nanomaterials(Nano-DCa)were prepared by double emulsion method and characterized by transmission electron microscope(TEM)and particle tracking analyzer(NTA).Their acid sensitivities were evaluated at different pH values by the multi-mode microplate reader.DOX release rates and cytotoxicity by CCK-8,flow cytometry,and western blot(WB).The acid targeting effect were observed by fluorescence spectrometer in vivo and the level of lactic acid in tumor.The anti-tumor effects of Nano-DCa combined with anti-PD-L1 treatment were investigated in subcutaneous lung cancer model.The immune activation mechanism was examined by H&E,TUNEL,flow cytometry,q PCR,CBA and immunohistochemistry.Lentivirus knocking out of PCSK9 gene expression in tumor cells was used to study the tumor growth results of Nano-DCa combined with PCSK9 knockout cells.Results:Nano-DCa trend to release DOX and cause greater cytotoxicity under acidic conditions.After intravenous injection,Nano-DCa effectively accumulates in the tumor and neutralizes the acidic TME.This acid--sensitive Nano-DCa activates the effect of anti-PD-L1 to suppress tumor growth and prolong the survival of mice by recovering the infiltration and function of CD8~+T cells in tumor and increasing the expression of inflammatory factors,such as IFN-γ,IL-4 and TNF,meanwhile reducing the infiltration of Treg cells and reversing the polarization of macrophages.PCSK9 knockout(PCSK9 KO)combined with Nano-DCa also significantly prolonged the survival of mice and inhibited tumor growth.Conclusion:These results reveal that Nano-DCa not only targets the delivery of chemotherapy drugs to enhance the immune response,but also breaks the blockade of PD-L1 efficacy mediated by immunosuppression in acidic environments.This study provides a reference for further improvement of the ICB effect.