Genetic Variation Of Gastrointestinal Neuroendocrine Neoplasms And The Associations Between CRKL,MCL1 Proteins Expression And Clinicopathologic Features

Objective: To investigate the genetic characteristics of Gastrointestinal-Neuroendocrine Neoplasms(GI-NENs)and to analyze the genomic differences between neuroendocrine tumors(NETs)and neuroendocrine carcinomas(NECs).The expressions of CRKL and MCL1 proteins in GI-NENs were observed to analyze the relationship between the expressions and clinicopathological features and prognosis.Methods: Next Generation Sequencing(NGS)was used to detect the changes of 425 genes in GI-NENs,including 4 NET G1,3 NET G2,3 NET G3 and 3 NEC.Immunohistochemistry(IHC)staining was performed to detected the expression of CRKL and MCL1 in 140 GI-NENs,including 93 G1,14 G2,7 G3 and 26 NEC.Results: Gene alterations were more common in NECs than NETs.NECs contained mutations involved TP53,MTOR,DDR2,ERBB4,BRCA1,BRCA2,ATM and SMAD4.Deep loss of SMAD4,SDHB,RB1,BRCA2 and high-level amplification of CRKL,CCNE1 and MCL1.The Tumor mutational burden(TMB)of NECs was significantly higher than that of NETs.3 pathways were found to be frequently altered in GI-NENs.The first pathway was involved in regulation of DNA repair and cell cycle.The most common genetic alteration was TP53 mutation.The second signaling pathway was Phosphatidylinositol3-kinase(PI3K)/protein kinase B(AKT)/mammalian target of rapamycin(m TOR).IGF1 R,CRKL,DDR2,ERBB4 and m TOR are the important factors of this pathway.The third pathway was the TGF-β/SMAD4 signaling pathway.Alterations of SMAD4 have been found in NECs.The expression of CRKL and MCL1 in GI-NENs tumor tissues was higher than that in paratumoral tissues.The high expression of CRKL was related to patient age,tumor diameter,primary site,histological classification/grading,depth of invasion,clinical stage,lymph node metastasis and vascular invasion.High expression of MCL1 protein was associated with tumor diameter,histological classification/grade,depth of invasion,clinical stage,and lymph node metastasis.Linear-regression analysis showed that MCL1 protein expression was negatively correlated with Overall survival(OS).Patients with high CRKL and MCL1 protein expression had a worse prognosis,and CRKL and MCL1 proteins were independent predictors of prognosis in patients with GI-NENs in this study.Conclusion:1.Genetic variation is more common than that of NETs,such as gene mutation of TP53,deletion of SMAD4 and amplification of CRKL and MCL1.The TMB value of NECs is significantly higher than that of NETs.2.Genetic alterations in GI-NECs may involve three signaling pathways,including DNA repair and cell cycle regulation,the PI3K/AKT/m TOR and the TGF-β/SMAD4 signaling pathway.3.The expression levels of CRKL and MCL1 proteins in tumor tissues of GI-NENs patients were significantly higher than those in paratumoral tissues,and their expression levels were correlated with tumor diameter,histological classification/grading,depth of invasion,clinical stage,lymph node metastasis and other pathological characteristics.4.Patients with high expression of CRKL and MCL1 proteins had worse prognosis and were independent predictors of prognosis in patients with GI-NENs.