Nasopharyngeal Carcinoma Related Genes And Their Single Nucleotide Polymorphisms Screened Out From 6p21.3

Nasopharyngeal Carcinoma (NPC) is an endemic malignant tumor with an obvious ethnic aggregation and familial clustering. It has been speculated that some hereditary predispositions may be involved in NPC development. However, the effort to identify the specific NPC related genes have not been successful so far.There have been reports suggesting NPC susceptibility genes to be located in the region between class-I and III of MHC, nearing the D6S1624 microsatellite marker. Because neither large DNA fragment deletion nor amplification was detected, it was held that single nucleotide variation in that region, such as Single Nucleotide Polymorphisms (SNPs), could be associated with the susceptibility to NPC.In this study, the interesting genes being likely related to NPC and containing known cSNPs were picked out from those settling in the region between TNF a and D6S1624 by bio-informatics analysis. The DNA variations were detected through performance of DHPLC assay on PCR products of targeted fragments of interesting genes, and verified by sequencing. The statistical difference of the heterozygotic frequencies at a given locus between NPC matched and non-NPC peripheral blood samples was determined by association analysis.10 PCR primers were designed according to the sequences of 10 fragments of DNA within interesting 7 genes respectively. Total 2521 PCR products from the genomic DNA of NPC tissues, NPC and non-NPC control peripheral blood samples were subjected to DHPLC assay and 511 heterozygotes were detected. Among the heterozygotes, the heterozygoties of sequence No. 4 (belonged to gene PPP1R10) in NPC and non-NPC bloodsamples were 28.3% and 10.1% respectively with x2= 9.55,P=0.002, and OR = 3.5, P<0.01; 95%CI =1.53 ~ 7.99, meaning the risk of NPC in NPC patients is 3.5 times of that in control group.The sequencing of the heterozygotes discovered a new SNP locus, an A=>G transition, at 15110 of gene PPP1R10. With locating in the sequence of an intron, the SNP locus should have no effect on the protein sequence, hence on thereof structure. According to the principle of genetic linkage, it may be deduced that a NPC susceptibility gene should near this SNP locus. Taking the locus as a marker, perhaps, the effort to clone the hypothetical NPC susceptibility gene will be successful.Sequencing representatives of heterozygotes and homozygotes revealed 6 new SNP loci. Among them, 4 were located in intron region, while 1 in 5' untranslated region and 1 in an exon. However, the identification of their genotypes in the population studied needs more samples to be sequenced. The performance of sequencing also verified known SNP loci occurred in the analyzed fragments as reported by other studies.Through comparing DHPLC and sequencing results of NPC biopsy samples with matched blood ones, point mutations at SNP loci were found in nearly 48% NPC samples. It is suggested that genomic instability is occurred in NPC cells.In summary, the present work, as a part of the campaign to clone NPC related genes, has screened SNPs marker of the genes within 6p21.3 MHC region. In addition to identification of 6 new SNP loci with one of them associated with NPC, the result of this work may be used as a basis to clone NPC susceptibility gene in the future, and also provide some useful data to the SNP database of Chinese population.