RTEL1 tagging SNPs and haplotypes were associated with glioma development

The overall incidence of brain tumors for benign and malignant tumors combined is 18.71 per 100,000 person-years; 11.52 per 100,000 person-years for benign tumors and 7.19 per 100,000 person-years for malignant tumors [1]. Though the age- standardized incidence recently reported varied greatly than ever, non-malignant tumours still accounted for 66% of all newly diagnosed primary brain tumours with the age-standardized incidence rate of 3.57 per 100,000 person-years, while malignant tumours incidence rate was 1.82 per 100,000 person-years (crude incidence rates were 3.69 and 1.92 per 100,000 respectively) [2]. Gliomas, most aggressive malignant brain tumours (astrocytic, oligodendroglial, oligoastrocytic and ependymal origin), represent 20.8% of all brain tumours [2], and account for almost 80% of primary malignant brain tumors, usually resulting in poor survival compared to other types of brain tumors [3].

Current evidence suggests that inherited risks play a significant role in glioma susceptibility, as with other cancers, and a majority of the inherited risk is due to the co-inheritance of multiple low-risk variants. These variants are commonly seen gene variants and hence can be identified through association studies [4]. The epidemiology of glioma has focused on identifying factors that can be modified to prevent this disease [5–7]. Recently studies of genetic risk factors for brain tumors have expanded to genome-wide association studies, and have focused on identifying germline polymorphisms associated with the risk of glioma as well as using molecular markers to classify glial tumors in more homogenous groups [6, 7].

A research group from the University of Texas M.D. Anderson Cancer Center conducted a meta-analysis of two genome-wide association studies (GWAS) by genotyping 550 K tagging single nucleotide polymorphisms (tSNPs) in a total of 1,878 cases and 3,670 controls, with validation in three additional independent series totaling 2,545 cases and 2,953 controls. They identified five risk loci for glioma including rs6010620 intronic to RTEL1 gene (P = 2.52 × 10^-12) to be associated with glioma risk [6]. Another study in Chinese also identified rs60106203 for glioma risk (P = 2.793 × 10^-6), and the locus also associated with glioblastoma risk (P = 3.573 × 10^-7) [8]. The subsequent study found that rs6010620 was statistically significantly associated with glioma risk in US female population [9]. Recently, a new independent GWAS of glioma using 1,856 cases and 4, 955 controls has found evidence of strong replication for three of the seven previously reported associations at 20q13.33 (RTEL), 5p15.33 (TERT), and 9p21.3 (CDKN2BAS), and consistent association signals for the remaining four at 7p11.2 (EGFR both loci), 8q24.21 (CCDC26) and 11q23.3 (PHLDB1) [7]. These data tend to show that common susceptibility alleles contribute to the risk of developing glioma and provide insight into disease causation of this primary brain tumor.

As the Chinese Han population is by far the population with the largest number in the world, we comprehensively analysized in this study the associations between RTEL1 genotypes and haplotypes with glioma risk, to uncover how germ-line genetic variants of the RTEL1 gene play a complex role in the development of glioma, to offer important insights into the etiology of glioma in the certain Chinese Han population.

As known, biomarker detection and screening is an emerging field for oncology [18–23]. Especially for gliomas considerable progresses have been made in identifying, characterizing, and attempting to apply molecular markers, e.g. in the previous study, we initially found the increased expression of miR-372 in glioma tissues was significantly correlated with advanced tumor progression and aggressive clinicopathological features [24]. And subsequently a series of have determined the associations between lots of SNPs in ABCB 1, NR 1/2, VEGFR 3, etc. and therapy outcome [25–27].

In this case–control study in Han Chinese population, we found two susceptibility tSNPs in RTEL1 gene that were associated with increased risk of glioma (rs6010620 and rs2297440). The genotype “GG” of rs6010620 was the protective genotype for glioma, and the genotype “CC” of rs2297440 was the protective genotype in glioma. We also observed in the RTEL1 gene a haplotype “GCT” that was associated with a decreased the risk and a haplotype of “ATT” with an increased risk of developing glioma.

As described initially, however, we failed to determine the associations between genotype frequency of GG or CC, and other parameters including WHO grading, gender, age at diagnosis, or KPS status. Additionally, we also tried to elucidate the relationship of genotype frequency of GG or CC, with overall surviaval (OS) in the correlated patients. During the follow-up period, only 11 of the patients with genotype of GG or CC had died [6 (8%) from the 75 patients with genotype GG, and 5 (6.8%) from the 73 patients with genotype CC], and most of the patients are alive and are being traced continuously.

Albeit the correlated survival data in the present study are still accumulating, our findings in this study have provided new evidence for the association between common SNPs (or haplotypes) and the risk of glioma in the Chinese Han population, suggesting an important determinant of glioma development by RTEL1 gene. RTEL1 gene locates in 20q13.3 with the length of 40.889 kb, including 40 exons. Known functions of RTEL1 include nucleic acid binding, ATP-dependent DNA helicase activity, DNA repair, apoptosis and anti-apoptosis, and so on. Previous study proposed that RTEL1 maintains genomic stability by suppressing homologous recombination [28, 29], and implements the second level of meiotic crossover control by promoting non-crossovers [30, 31]. A recent review point out that RTEL1 was an essential helicase for telomere maintenance and the regulation of homologous recombination [32, 33]. RTEL1 didn’t involve any KEGG pathway (http://www. genome.jp/kegg/) so far. The frequencies of rs6010620 risk genotypes were highly correlated with high-grade disease (P < 0.001), indicating that genetic variations at the locus has subtype-specific effects on the risk of developing glioma [34]. RTEL1 gene was over expressed in human gastrointestinal tract tumors [35]. Polymorphism in the RTEL1 gene was associated with glioblastoma survival [36].

Some limitations are inherent in this case–control study and must be noted here. Glioma patients were not sub-grouped by age or gender, and gender-specific significant variants were not tested. We selected tSNPs with frequencies higher than 5% in HapMap Asian populations to affirm the statistical power was large enough for analyzing data. We also designed a haplotype-based study to ensure sufficiently high power to detect the risk of glioma associated with candidate tSNPs. Another potential concern was population admixture, which is a known confounding factor for association analysis and may also result in inflated type-I error (false positive). In this study, glioma patients and controls were used in the same hospital to avoid the possibility that one may have a more pronounced selection bias. However, this bias is unlikely to be of significance because they did not differ in the distributions of demographic variable and genotype frequencies. We limited all subjects’ ethnicity to Han Chinese, and a living area to Xi’an City and its surrounding area, thus there is no substantial population admixture in our study populations.

In the upcoming studies, our team will go on to follow up the subjects recruited into the present study, and carry out additional research with larger subject numbers and grade types to further characterize the relationship among grades within the individuals, clinical features and the mentioned RTEL1 tagging SNPs & haplotypes. Furthermore, to elucidate the role of the RTEL1 gene in gliomagenesis, serum RTEL1 expression levels between different mutations or haplotype groups will be compared. And, we will also investigate the association between germline RTEL1 variants and somatic RTEL1 mutations, and the relationship between serum RTEL1 expression and somatic RTEL1 expression in the same glioma subjects.

In conclusion, our comprehensive analysis of tSNPs suggests that the genotypes of “GG” of rs6010620 and “CC” of rs2297440 (rs6010620 and rs2297440) in the RTEL1 gene, together with two haplotypes of GCT and ATT, were identified to be associated with glioma development. And it might be used to evaluate the glioma development risks to screen the above RTEL1 tagging SNPs and haplotypes.