Jianbo Tian

A functional polymorphism in lnc-LAMC2-1:1 confers risk of colorectal cancer by affecting miRNA binding

Genome-wide association studies (GWASs) have identified multiple susceptibility loci of colorectal cancer (CRC), however, causative polymorphisms have not been fully elucidated. Long non-coding RNAs (lncRNAs) are a recently discovered class of non-protein coding RNAs that involved in a wide variety of biological processes. We hypothesized that single nucleotide polymorphisms (SNPs) in lncRNA may associate with the CRC risk by influencing lncRNA functions. To evaluate the effects of SNPs on CRC susceptibility in Chinese populations, we first screened out all potentially functional SNPs in exons of lncRNAs located in CRC susceptibility loci identified by GWAS. Eight SNPs were selected and genotyped in 875 CRC cases and 855 controls and replicated in an independent case-control study consisting of 768 CRC cases and 768 controls. Analyses showed that CG and GG genotypes of the rs2147578 were significantly associated with increased risk for CRC occurrence in both case-control studies [combined analysis OR = 1.29; 95% confidence interval (CI) = 1.11-1.51, P = 0.001] compared to the rs2147578 CC genotype. Bioinformatics analyses showed that rs2147578 is located in the transcript of lnc-LAMC2-1:1 and could influence the binding of lnc-LAMC2-1:1/miR-128-3p. Further luciferase reporter assays demonstrated that the construct with the risk rs2147578G allele had relatively high expression activity compared with that of the rs2147578C allele. Expression quantitative trait loci analyses also showed that rs2147578 is correlated with the expression of a well established oncogene LAMC2 (laminin subunit gamma 2). These findings indicated that rs2147578 in lnc-LAMC2-1:1 might be a genetic modifier for the development of CRC.

A low-frequency variant in SMAD7 modulates TGF-β signaling and confers risk for colorectal cancer in Chinese population

The TGF‐β pathway plays an essential role in regulating cell proliferation and differentiation. GWASs and candidate approaches have identified a battery of genetic variants in the TGF‐β pathway contributing to colorectal cancer (CRC). However, most of the significant variants are common variants and their functions remain ambiguous. To identify causal variants with low‐frequency in the TGF‐β pathway contributing to CRC susceptibility in Chinese population, we performed targeted sequencing of 12 key genes in TGF‐β signaling in CRC patients followed by a two‐stage case‐control study with a total of 5109 cases and 5169 controls. Bioinformatic annotations and biochemical experiments were applied to reveal the potential functions of significant variants. Seven low‐frequency genetic variants were captured through targeted sequencing. The two stage association studies showed that missense variant rs3764482 (c. 83C>T; p. S28F) in the gene SMAD7 was consistently and significantly associated with CRC risk. Compared with the wild type, the ORs for variant allele were 1.37 (95%CI: 1.10‐1.70, P = 0.005), 1.55 (95%CI: 1.30‐1.86, P = 1.15 × 106), and 1.48 (1.29‐1.70, P = 2.44 × 10;8) in stage 1, stage 2, and the combined analyses, respectively. Functional annotations revealed that the minor allele T of rs3764482 was more effective than the major allele C in blocking the TGF‐β signaling and inhibiting the phosphorylation of receptor‐regulated SMADs (R‐SMADs). In conclusion, low‐frequency coding variant rs3764482 in SMAD7 is associated with CRC risk in Chinese population. The rs3764482 variant may block the TGF‐β signaling via impeding the activation of downstream genes, leading to cancer cell proliferation, thus contributing to CRC pathogenesis.

A functional polymorphism located at transcription factor binding sites, rs6695837 near LAMC1 gene, confers risk of colorectal cancer in Chinese populations.

Genome-wide association studies (GWASs) have identified multiple single nucleotide polymorphisms (SNPs) associated with colorectal cancer (CRC) susceptibility. However, the elucidation of causal SNPs and the biological mechanisms behind are still limited. In this study, we initially performed systematic bioinformatics analyses on CRC GWAS-identified loci to seek for potential functional SNPs located at transcription factor binding sites (TFBSs), and then a two-stage case-control study comprised of 1353 cases and 1448 controls of Chinese populations and functional analyses were conducted. As a result, only one SNP rs6695837 out of the nine candidate SNPs survived after two-stage analyses by Bonferroni correction. In combined analyses, rs6695837 exhibited significant associations with CRC risk (TT: CC, odds ratio (OR) = 1.31, 95% confidence interval (CI) = 1.06-1.63; dominant model, OR = 1.21, 95% CI = 1.03-1.43; additive model, OR = 1.15, 95% CI = 1.03-1.28). Functional annotations by RegulomeDB and rSNPBase indicated its biological role and dual-luciferase reporter assays revealed a significant increase in luciferase expression for the reconstructed plasmid with rs6695837T allele, compared with the one with C allele (PSW480 = 0.0002, PLovo = 0.0003). Further gene expression analyses demonstrated significantly higher expression of LAMC1 gene in CRC tumor tissues than that in adjacent non-cancerous tissues (P = 0.0004). These findings strongly suggest that the functional SNP located at TFBSs, rs6695837 might contribute to CRC susceptibility, and the exact biological mechanism awaits further research.

A polymorphic MYC response element in KBTBD11 influences colorectal cancer risk, especially in interaction with an MYC-regulated SNP rs6983267

Background MYC is a well-established cancer driver gene regulating the expression of numerous genes, indicating that polymorphisms in MYC response elements could affect tumorigenesis through altering MYC regulation. We performed integrative multistage study to evaluate the effects of variants in MYC response elements and colorectal cancer (CRC) risk. Patients and methods We systematically integrated ChIP-Seq, DNase-Seq and transcription factor motif data to screen variants with potential ability to affect the MYC binding affinity. Then, we conducted a two-stage case-control study, totally consisting of 4830 CRC cases and 4759 controls in Chinese population to identify risk polymorphisms and interactions. The effects of risk variants were confirmed by functional assays in CRC LoVo, SW480 and HCT15 cells. Results We identified a novel polymorphism rs11777210 in KBTBD11 significantly associated with CRC susceptibility (P = 2.43 × 10-12). Notably, we observed a significant interaction between rs11777210 and MYC nearby rs6983267 (P-multi = 0.003, P-add = 0.005), subjects carrying rs6983267 GG and rs11777210 CC genotypes showing higher susceptibility to CRC (2.83-fold) than those carrying rs6983267 TT and rs11777210 TT genotypes. We further demonstrated that rs6983267 T > G increased MYC expression, and MYC bound to and negatively regulated KBTBD11 expression when the rs11777210 C risk allele was present. KBTBD11 was downregulated in tumor tissues, and KBTBD11 knockdown promoted cell proliferation and inhibited cell apoptosis. Conclusion The rs11777210 is a potential predictive biomarker of CRC susceptibility, and KBTBD11 functions as a putative tumor suppressor in tumorigenesis. Our study highlighted the high CRC risk of people carrying rs6983267 G and rs11777210 C alleles, and provided possible biological mechanism of the interaction.

A Rare Variant P507L in TPP1 Interrupts TPP1–TIN2 Interaction, Influences Telomere Length, and Confers Colorectal Cancer Risk in Chinese Population

Background: Telomere dysfunction triggers cellular senescence and constitutes a driving force for cancer initiation. Genetic variants in genes involved in telomere maintenance may contribute to colorectal cancer susceptibility. Methods: In this study, we firstly captured germline mutations in 192 patients with colorectal cancer by sequencing the coding regions of 13 core components implicated in telomere biology. Five potential functional variants were then genotyped and assessed in a case–control set with 3,761 colorectal cancer cases and 3,839 healthy controls. The promising association was replicated in additional 6,765 cases and 6,906 controls. Functional experiments were used to further clarify the potential function of the significant variant and uncover the underlying mechanism in colorectal cancer development. Results: The two-stage association studies showed that a rare missense variant rs149418249 (c.C1520T and p.P507L) in the 11th exon of TPP1 (also known as ACD, gene ID 65057) was significantly associated with colorectal cancer risk with the ORs being 2.90 [95% confidence interval (CI), 1.04–8.07; P = 0.041], 2.50 (95% CI, 1.04–6.04; P = 0.042), and 2.66 (95% CI, 1.36–5.18; P = 0.004) in discovery, replication, and the combined samples, respectively. Further functional annotation indicated that the TPP1 P507L substitution interrupted TPP1–TIN2 interaction, impaired telomerase processivity, and shortened telomere length, which subsequently facilitated cell proliferation and promoted colorectal cancer development. Conclusions: A rare variant P507L in TPP1 confers increased risk of colorectal cancer through interrupting TPP1–TIN2 interaction, impairing telomerase processivity, and shrinking telomere length. Impact: These findings emphasize the important role of telomere dysfunction in colorectal cancer development, and provide new insights about the prevention of this type of cancer. Cancer Epidemiol Biomarkers Prev; 27(9); 1029–35. ©2018 AACR.

The Associations between RNA Splicing Complex Gene SF3A1 Polymorphisms and Colorectal Cancer Risk in a Chinese Population.

Background Aberrant alternative splicing included alterations in components of the mRNA splicing machinery often occurred in colon cancer. However, the role of SF3A1, one key component of the mRNA splicing machinery, on colorectal cancer (CRC) risk was still not elucidated. Method and Findings We performed a hospital-based case-control study containing 801 CRC patients and 817 cancer-free controls to examine the association between SF3A1 polymorphisms and CRC risk in a Chinese population. Four candidate SNPs (rs10376, rs5753073, rs2839998 and rs2074733) were selected based on bioinformatics analysis and previous findings. The results showed no significant associations between these SNPs and CRC risk (P > 0.05). Besides, the stratified analysis based on the smoking and alcohol use status obtained no statistically significant results. Conclusion Our study was the first one to investigate the association between SF3A1 polymorphisms and CRC risk. The results suggested these four SNPs in SF3A1 were not associated with CRC risk in a Chinese population, however, further more studies are needed to confirm our findings.

Identification of a functional polymorphism affecting microRNA binding in the susceptibility locus 1q25.3 for colorectal cancer

Genome‐wide association studies (GWASs) have identified dozens of susceptibility loci for colorectal cancer (CRC). However, most of them lack functional genetic variants and clear biological mechanisms. MicroRNAs (miRNAs) are small noncoding RNAs involved in a variety of physiological and tumorigenic processes. Here we hypothesized that single nucleotide polymorphisms (SNPs) that affect miRNAs biogenesis and binding, could contribute to CRC risk in the Chinese population. To locate miRNA‐related SNPs in established GWAS loci, we initially screened out five candidate SNPs using a systematic bioinformatics analysis. Then, we performed a two‐stage case‐control study consisting of 2347 cases and 3390 controls, and found a positive polymorphism rs1062044, which presented consistently significant associations with CRC in both stages, and with an odds ratio (OR) = 1.32 (95% confidence interval (95%CI) = 1.18‐1.49, P = 3.43E‐06) under the dominant model in the combined study. Further luciferase reporter gene assays indicated that the variant G allele obviously improved the specific binding between miR‐423‐5p and the gene LAMC1. These findings suggested that the functional SNP rs1062044 at 1q25.3 might be a genetic modifier for the occurrence and development of CRC.

Breast cancer risk-associated variants at 6q25.1 influence risk of hepatocellular carcinoma in a Chinese population

The gender disparity observed in the incidence of hepatocellular carcinoma (HCC) suggests an important role of estrogens in HCC pathogenesis. In this study, we conducted a case-control study to investigate whether breast cancer risk-associated single nucleotide polymorphisms (SNPs) located at estrogens loci identified by genome-wide association studies (GWASs) also predispose to HCC in a Chinese population. Three candidate SNPs at 6q25.1 were genotyped in 2025 HCC cases and 2032 healthy controls. Differential expression analyses and expression quantitative trait loci (eQTL) analyses were conducted to further explore the potential function of significant SNPs and genes they reside in. Two of the three candidate SNPs (rs9383951 and rs9485372) were observed to be significantly associated with HCC risk. Under a dominant model, the odds ratios (OR) for rs9383951 and rs9485372 were 1.28 (95% CI: 1.10-1.49, P = 0.002) and 1.34 (95% CI: 1.17-1.53, P = 2.75 × 10-5), respectively. We also found a significant accumulative effect of these two SNPs and there was a gradual increase in OR with a greater number of hazard genotypes. Moreover, the association between rs9383951 and HCC risk was specific in males. Lower ESR1 and TAB2 expressions were investigated in hepatic tumor tissues than adjacent normal tissues. We found a significant association between rs9383951 and ESR1 expression (P = 0.047). Besides, ESR1 expression was significantly correlated with the expression of TAB2. Taken together, our study identified two genetic variants at 6q25.1 newly associated with HCC risk, suggesting ESR1 and estrogen signaling may play a role in mediating susceptibility to HCC in Chinese population.

CancerSplicingQTL: a database for genome-wide identification of splicing QTLs in human cancer

Abstract Alternative splicing (AS) is a widespread process that increases structural transcript variation and proteome diversity. Aberrant splicing patterns are frequently observed in cancer initiation, progress, prognosis and therapy. Increasing evidence has demonstrated that AS events could undergo modulation by genetic variants. The identification of splicing quantitative trait loci (sQTLs), genetic variants that affect AS events, might represent an important step toward fully understanding the contribution of genetic variants in disease development. However, no database has yet been developed to systematically analyze sQTLs across multiple cancer types. Using genotype data from The Cancer Genome Atlas and corresponding AS values calculated by TCGASpliceSeq, we developed a computational pipeline to identify sQTLs from 9 026 tumor samples in 33 cancer types. We totally identified 4 599 598 sQTLs across all cancer types. We further performed survival analyses and identified 17 072 sQTLs associated with patient overall survival times. Furthermore, using genome-wide association study (GWAS) catalog data, we identified 1 180 132 sQTLs overlapping with known GWAS linkage disequilibrium regions. Finally, we constructed a user-friendly database, CancerSplicingQTL (http://www.cancersplicingqtl-hust.com/) for users to conveniently browse, search and download data of interest. This database provides an informative sQTL resource for further characterizing the potential functional roles of SNPs that control transcript isoforms in human cancer.

Integrative functional genomics identifies regulatory genetic variant modulating RAB31 expression and altering susceptibility to breast cancer

Despite the successes of genome‐wide association study (GWAS) in identifying breast cancer (BC) risk‐associated variants, only a small fraction of the heritability can be explained. The greatest challenge in the post‐GWAS is to identify causal variants and underlying mechanisms responsible for BC susceptibility. In this study, we integrated functional genomic data from ENCODE ChIP‐seq, ANNOVAR, and the TRANSFAC matrix to identify potentially regulatory variants with modulating FOXA1‐binding affinity across the whole genome, and then conducted a two‐stage case‐control study including 2164 cases and 2382 controls to investigate the associations between candidate SNPs and BC susceptibility. We identified a BC susceptibility SNP, rs6506689 G>T, with an odds ratio (OR) of 1.23 (95% confidence interval = 1.07‐1.40, P = 0.003) under a dominant model in the combined study. Biological assays indicated that the germline G>T variation at rs6506689 creates a FOXA1‐binding site and up‐regulates the expression of RAB31, thus playing an important role in the development of BC. Our results highlight the importance of regulatory genetic variants in the development of BC by influencing TF‐DNA interaction and provide critical insights to pinpoint causal genetic variants.