Pharmacodynamics And Mechanism Of BiRGD-siPD-L1 Conjugate Specific Targeted Therapy Against Melanoma In Vitro And In Vivo

[Background]Small cell lung cancer(SCLC)is a lung tumor with a very high degree of malignancy.Chemoresistance frequently occurs in SCLC,in which it leads to treatment failure.However,the mechanisms underlying this process remain largely unclear.We used a cDNA microarray to screen differentially expressed genes between drug-resistant and drug-sensitive cell lines,and the result showed that CDYL was expressed at high levels in chemoresistant SCLC cells.However,whether CDYL affects chemoresistance in SCLC remains unknown.[Methods and contents]1.The effects of chromodomain Y-like(CDYL)on chemoresistance in SCLC were determined by cDNA microarray,cell transfection,Western blot,immunohistochemistry,CCK8 analysis of the IC50 values for cytotoxic agents(cisplatin and etoposide),flow cytometric analysis of cell apoptosis and cell cycle,and tumorigenicity experiments.2.The underlying mechanisms of chemoresistance were investigated through mRNA sequencing,chromatin immunoprecipitation-qPCR,electrophoretic mobility shift assay,CO-immunoprecipitation,GST pull down,bisulfite sequencing PCR,enzyme linked immunosorbent assay,and bioinformatics analysis.[Results]1.A greater percentage of samples from chemoresistant patients showed high CDYL expression than samples from chemosensitive patients and that the CDYL expression is related to clinical stage and prognosis.2.The CDYL levels of chemoresistant SCLC cells are significantly higher than that of chemosensitive cells.We next established cell line models with up-regulation and down-regulation of CDYL.Up-regulation of CDYL in vitro increases IC50 to chemotherapy drugs(cisplatinum and etoposide)in chemosensitive SCLC cell lines,decreases cell apoptosis and G1 arrest,while down-regulation of CDYL in vitro decreases IC50 to chemotherapy drugs in chemoresistant SCLC cell lines,increases cell apoptosis and G1 arrest.3.Up-regulated CDYL reduced the growth inhibition of chemotherapy drugs on xenograft tumor in nude mice,while down-regulated CDYL increased the growth inhibition of chemotherapy drugs on xenograft tumor in nude mice.4.Mechanistically,CDYL targets CDKN1C directly in SCLC.CDKN1C was expressed at lower levels in chemoresistant SCLC cells than in chemosensitive SCLC cells,then we established cell line models with up-regulation and down-regulation of CDKN1C;Up-regulation of CDKN1C decreases IC50 to chemotherapy drugs in chemoresistant SCLC cell lines,increases cell apoptosis and G1 arrest,while down-regulation of CDKN1C increases IC50 to chemotherapy drugs in chemosensitive SCLC cell lines,decreases cell apoptosis and G1 arrest.Rescue experiments showed that CDYL regulated chemoresistance by directly repressing CDKN1C(cyclin-dependent kinase inhibitor 1C)transcription.5.CDYL recruits the enhancer of zeste homolog 2(EZH2)to regulate trimethylation of lysine 27 in histone 3(H3K27me3)at the CDKN1C promoter region and promotes transcriptional silencing.6.The EZH2 inhibitor GSK126 de-represses CDKN1C and diminishes CDYL-induced chemoresistance in SCLC in vivo and in vitro.[Conclusions and significance]1.High CDYL expression than samples from chemosensitive patients and that the CDYL expression is related to clinical stage and prognosis.CDYL promotes the chemoresistance of SCLC in vivo and in vitro,decreases cell apoptosis and G1 arrest.2.CDYL recruits the EZH2 to regulate H3K27me3 at the CDKN1C promoter region and promotes transcriptional silencing,resulting in the chemoresistance in SCLC.3.The EZH2 inhibitor diminishes CDYL-induced chemoresistance in SCLC.Overall,these results reveal that the CDYL/EZH2/H3K27me3/CDKN1C axis promotes SCLC chemoresistance.These markers represent promising therapeutic targets for overcoming SCLC chemoresistance.