Preliminary Screening Of Immunoediting Related Genes And Immuno-related Abnormal Chromosome Regions In Nasopharyngeal Carcinoma

Nasopharyngeal carcinoma (nasopharyngeal carcinoma, NPC) is a malignancy derived from epithelium with high incidence in Southeast Asia and Southern China. The main treatment modality of NPC patients is radiotherapy. The epidemiological and etiological studies indicate that Epstein-Barr virus (EBV), certain chemical carcinogens and genetic susceptibility are closely associated with its occurrence and development.Nasopharyngeal carcinogenesis is a multistage, multifactor and multipathway process involving multiple gene alterations. But research on immunological mechanism of NPC is scarce. Cancer immunoediting was put forward by Schreiber and Dunn in 2002, which based on the development of modern immunology and experimental evidence. It is also called "three E's hypothesis", which explains three continuous immunological kinetic phases (elimination, equilibrium and escape) presented during the process of tumor development. This hypothesis has profound influence on the strategy to elucidate the mechanism of tumor occurrence and development as well as to explore cancer immunodiagno-sis and immunotherapy.Through gene chips study on NPC tissues and NPC cell lines, we have revealed the roles of many genes in the development of NPC. Based on comparative genomic hybridization (CGH) results from several laboratories, we have set up a tree model for explaining the pathogenesis of NPC. Furthermore, we have also explored the immunoediting events between CNE2 cells and NK cells recently.In this study, potential immunoediting related genes of NPC were screened by GenClip software, and the results were validated by SYBR real-time q-PCR. Moreover, we analyzed the positional feature of immuno-related genes in chromosomal regions of NPC, the relation among these chromosomal regions and fragile sites and microRNAs. This study provides the preliminary experimental evidence for immunological mechanism in the development of NPC. The experimental methods and results are as follows: I . Preliminary screening for immunoediting related genes in NPCDifferential expression criteria were set up for screening NPC gene chip data of our laboratory. 7 genes were found that they were up-regulated in NPC cell lines but down-regulated in NPC tissues, and 183 genes were found to be down-regulated in NPC cell lines but up-regulated in NPC tissues. According to gene ontology classification, 24 immuno-related genes of NPC were screened out from above-mentioned genes.A relational network of gene co-occurrences was constructed by using 11 genes that participating in the process of cancer immunoediting and 24 above-mentioned genes with the literature-mining software GenClip 3.2. There are 20 genes (IFNG, IFNA1, PRF1, TRAIL, STAT1, NKG2D, RAG2, RAG1, IL23A, IRF1, CXCL10, IFITM3, INDO, IL1B, CXCL9, CCL2, CDM, CCL18, IL2RG and FAS) in this relational network. Based on detailed analysis of literature, 6 genes were found to probably participated in the process of NPC immunoediting, which included IFNG, STAT1, INDO, CXCL10, CXCL9 and IRF1.Real-time PCR was performed to analyze these 6 genes, the results further suggests that IFNG, STAT1, INDO, CXCL10 and IRF1 are potential immunoediting related genes of NPC.II. Preliminary screening of immuno-related abnormal chromosome regions in NPCAccording to gene ontology classification, abnormal expression of immuno-related genes were found from the NPC gene chip data of our laboratory. 14 up-regulated immuno-related genes were discovered in NPC chromosomal amplification regions, in which 12 genes locate at the lq22-32 region (FCGR3A, FCGR2A, FCER1G, CD1D, CD1E, IFI16, FCGR2C, TNFSF4, MR1, RGS1 and CD46) , and 2 others locate at the 12q13-15 region (STAT6 and IFNG) , meanwhile, 5 down-regulated immuno-related genes are discovered in NPC chromosomal deletion regions, among them 4 locate at the 14q24-32 region (IGHM, IGHD, IGHG1 and IGHA1) , they are closely adjacent to each other, CHST4 at the 16q12-24 region. The positional features of immuno-related genes in the NPC chromosomal regions were studied according to the three stages suggested by in the tree model for pathogenesis of NPC which was constructed by 170 comparative genomic hybridization (CGH) samples, and related or nearby fragile sites were also searched out. We found 3 fragile sites in the immuno-related gene enriched area (lq22-32 and 14q24-32 region), such as FRA1G (1q25.1), FRA1K (1q31) and FRA14C (14q24.1).Human microRNAs probably related with the 1q22-32 and 14q24-32 regions were screened. We found 14 human microRNAs at two above mentioned regions, among which 8 microRNAs locate in 1q22-32 region ( including miR-009-1, miR-29c/miR-102, miR-213S/miR-181b, miR-214/miR-199a-2 and miR-205) , and the other 6 microRNAs locate at 14q24-32 region (including miR-127 / miR-136, miR-134 / miR-154 / miR-299 and miR-203) , We infer that these microRNAs are closely related to NPC.A relational network of genes was constructed by NPC immuno-related genes screened from all stages with the literature-mining software GenClip 3.2, and we screened 13 potential genes that are related to NPC immunoediting genes (IFNG, CD46, FCGR3A, FASLG, CD1D, CD1E, FCGR2C, FCGR2A, STAT6, MR1, TNFSF4, IGHA1 and IFI16).In summary, based on previous work of our laboratory, potential immunoediting related genes and immuno-related potential abnormal regions of chromosomes in NPC were screened by using bioinformatics method, and these data were validated preliminarily. The results suggest that IFNG, STAT1, INDO, CXCL10 and IRF1 are potential immunoediting related genes of NPC, and lq22-32 and 14q24-32 regions are potential immuno-related abnormal regions of chromosomes in NPC. The role of these genes in the pathogenesis of NPC remains to be elucidated.