Loss of BLK expression as a potential predictor of poor prognosis and immune checkpoint blockade response in NSCLC and contribute to tumor progression

Abstract

BackgroundImmune checkpoint blockade (ICB) has been proved to have significant anti-tumor effect in the clinical treatment of non-small cell lung cancer (NSCLC). Therefore, biomarkers predicting ICB response can provide better treatment for patients with NSCLC. MethodsDifferential expression genes (DEGs) were identified by ImmuCellAI database. Copy number alteration (CNA) was analyzed by cBioPortal. The predicted efficiency of 4 genes on cancer immunotherapy was assessed by ROC analysis. The survival value of BLK was analyzed by Kaplan-Meier plotter and Prognoscan analysis. Clinical significance of BLK IHC-TMA score in NSCLC was also explored. The CCK-8 assay, wound healing assay, western blot assay in vitro and subcutaneous xenograft experiments in vivo were used for investigating the functions of BLK. The RNA-sequencing were performed to screen BLK regulated genes and conducted for GO/KEGG enrichment analysis. The transcriptional regulatory factor of BLK promoter region was predicted by ChIP-seq analysis. Results39 common DEGs between ICB Response (R) group and No Response (NR) group with NSCLC were identified, in which the CNA frequency of BLK deletion (> 6%) was found. The predicted efficiency of BLK on immunotherapy was performed best in NSCLC (AUC>0.7). Low expression of BLK was related to NSCLC with significantly poor prognosis. BLK overexpression can inhibit growth of NSCLC via activating apoptosis pathway, inhibiting the G2M checkpoint and Glycolysis pathway. The enrichment analysis indicated that BLK regulated genes related to oncogenic potential in NSCLC. Besides, BLK expression was inhibited via H3K27me3 modification in A549 and H1299 cells. BLK mRNA level was negatively correlated with methylation and positively correlated with the tumor purity in NSCLC. ConclusionOur study provides strong evidence that low expression of BLK may serve as a biomarker for poor prognosis in NSCLC, while response to ICB therapy and contributes to NSCLC tumor progression.