Prognosis Of Surgically Resected Early-stage ALK-positive Lung Adenocarcinoma And Mechanism Of ALK Rearrangement Regulating PD-L1 Expression

Research background and purposes: Lung cancer has long been the most important cause of cancer-related deaths in China.Many key oncogene changes play an important role in the development and progression of lung adenocarcinoma(LAC).The lung cancer-associated tumor-driven genes include epidermal growth factor receptor(EGFR)gene mutation,echinoderms microtubule-associated protein-like 4 gene and anaplastic lymphoma kinase gene fusion(EML4-ALK),and murine sarcoma viral oncogene(KRAS).The EML4-ALK fusion gene exhibits potent carcinogenic activity both in vivo and in vitro.Although the small molecule tyrosine kinase inhibitor(TKI)Crizotinib for EML4-ALK is approved for the treatment of ALK-positive advanced non-small cell lung cancer(NSCLC)due to its good therapeutic effect and safety in clinical trials.However,the prognostic significance of ALK-positive LAC patients without Crizotinib after surgical resection is unclear.Previous studies have shown that the expression of programmed cell death protein ligand 1(PD-L1)in NSCLC patients is related to EGFR and is regulated by EGFR.The correlation and molecular mechanism of EML4-ALK interaction with PD-L1 is still unclear.Our previous study showed that the pathological tissues of ALK-positive patients and ALK-positive cell lines had high PD-L1 expression,and ALK had a positive regulation of PD-L1 expression.The purpose of the first part of the study was to detect the expression of ALK gene in LAC patients by Ventana immunohistochemistry(IHC)method,and to explore the clinicopathological features and prognostic significance of ALK rearrangement in patients with early LAC.The second part explores the specific mechanism of action of EML4-ALK fusion gene regulation of PD-L1 expression in LAC.Materials and Method: 1.Between 2011 and 2015,a total of 635 patients underwent radical resection and pathological stage I-IIIA LAC in Tianjin Medical University Cancer Hospital.The expression of ALK gene in tumor tissues was detected by D5F3 Ventana IHC.The clinical and pathological data and follow-up results of all enrolled patients were collected.The clinicopathological features of ALK fusion gene-positive LAC patients and their relationship with prognosis were analyzed by statistical methods.2.IHC was used to detect the expression of PD-L1 in cancer tissues.Western Blot was used to detect the expression of PD-L1 in each representative cell line.ALK positive cell lines were treated with four key signaling molecule inhibitors downstream of ALK,and then the expression of PD-L1 was detected by Western Blot.The molecular mechanism of EML4-ALK regulation of PD-L1 expression was investigated.Result: 1.The ALK positive rate in early LAC population was 11.1% detected by D5F3 Ventana immunohistochemistry.The three states of EGFR,KRAS mutation and ALK rearrangement are mutually exclusive.ALK positive was more common in younger female non-smoking patients with later pathological stages and higher lymph node metastasis rate,and pathological subtypes were more common in mucin production invasive adenocarcinoma solid type and invasive mucinous adenocarcinoma.It was more common in postoperative patients receiving adjuvant therapy,but no significant association was found between the ALK fusion gene and the size and family history of the tumor stage.ALK positivity was significantly associated with shorter disease-free survival.Multivariate analysis showed that ALK positive was a poor prognostic factor for DFS.2.There is a correlation between ALK positive and PD-L1 expression.After gradient treatment of H3122 cells with ERK inhibitor,the expression level of PD-L1 decrease with the descent of ERK protein expression,which indicates that the EML4-ALK gene regulates the expression of PD-L1 through the ERK signaling pathway.Conclusion: The clinicopathological features of ALK-positive lung adenocarcinoma patients are similar but different from those of EGFR-mutant patients.Positive ALK fusion gene is a factor in the poor prognosis of patients undergoing surgical resection of LAC.The EML4-ALK gene regulates the expression of PD-L1 via the ERK signaling pathway.