Genome-Wide Association Analyses Identify CATSPERE as a Mediator of Colorectal Cancer Susceptibility and Progression.

Genome-wide association studies (GWAS) have revealed numerous genetic loci associated with colorectal cancer risk, but the mechanisms underlying these loci have not been comprehensively elucidated. In this study, we performed a GWAS meta-analysis with a two-stage replication strategy by combining eight colorectal cancer cohorts encompassing 7,186 cases and 8,512 controls in Chinese populations, accompanied by an evaluation encompassing 29,832 cases and 406,694 controls in European populations. The genetic variant rs505706 A>G, located at chr1q44 in the upstream region of catsper channel auxiliary subunit epsilon (CATSPERE), was associated with colorectal cancer risk and exhibited genome-wide significance (OR, 0.73; 95% confidence interval, 0.67-0.80; P = 9.75 × 10-12). Cell line and animal models were applied to assess the biological function of the genetic risk variant and the corresponding susceptibility gene. Genetically, the G allele of rs505706 resulted in long-range regulatory effects, reducing the binding affinity of POU2F1 for the CATSPERE promoter and thus abolishing the inhibitory effect of POU2F1 on CATSPERE transcription. Phenotypically, CATSPERE upregulation attenuated tumor growth in both colorectal cancer cells and xenograft models. Mechanistically, CATSPERE promoted calcium ion influx and apoptotic pathway activity. In zebrafish models, CATSPERE exerted pleiotropic effects, enhancing the progression of colorectal cancer. Collectively, these findings highlight a colorectal cancer susceptibility locus that acts to remotely modulate the activity of CATSPERE, a gene that mediates multiple functions involved in colorectal tumorigenesis and progression. SIGNIFICANCE: A GWAS meta-analysis identifies a novel susceptibility locus harboring a genetic risk variant that mediates pleiotropic biological effects in colorectal tumorigenesis and progression.

Association of genetic variants in autophagy-lysosome pathway genes with susceptibility and survival to prostate cancer.

BACKGROUND: Previous studies have indicated the connections between autophagy-lysosome pathway genes dysfunction and prostate cancer, but few studies have investigated whether single nucleotide polymorphisms (SNPs) in autophagy-lysosome pathway genes are implicated in prostate cancer risk and survival. MATERIALS AND METHODS: Logistic regression analysis and stepwise Cox regression analysis were conducted in 4,662 cases and 3,114 controls from the Prostate, Lung, Colorectal and Ovarian (PLCO) Cancer Screening Trial. The false positive rate probability (FPRP) method was applied to correct for multiple comparisons. Gene-based analysis was calculated by versatile gene-based association study approach. RESULTS: We found that SLC11A1 rs7573065 significantly increased the risk of prostate cancer [adjusted odds ratio (OR) = 1.24, 95% confidence interval (CI) = 1.06-1.46, P = 7.02 × 10-3, FPRP = 0.082]. Furthermore, rs7573065 was confirmed as the independent predicator of overall survival (OS) for prostate cancer patients [Hazard ratio (HR) = 1.30, 95% CI = 1.01-1.66, P = 0.041]. The significant association between SLC11A1 and prostate cancer risk was calculated by gene-based analysis (P = 0.030). We also observed that the mRNA of SLC11A1 in prostate tumor tissues was significantly over-expressed than that in normal tissues. CONCLUSION: This study suggested that rs7573065 in SLC11A1 was associated with an increased risk and poor OS of prostate cancer. Our findings may provide evidence for genetic variants in autophagy-lysosome pathway as the risk and prognostic biomarkers for prostate cancer.

Genetic variations in the CTLA-4 immune checkpoint pathway are associated with colon cancer risk, prognosis, and immune infiltration via regulation of IQCB1 expression.

The programmed cell death-1 (PD-1)/cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) immune checkpoint pathways serve as targets of immunotherapy for colorectal cancer. However, the associations between genetic variations in these pathways and colorectal cancer risk, prognosis, and immune status remain unclear. The associations between single-nucleotide polymorphisms (SNPs) and colorectal cancer risk and survival were evaluated in a case-control study comprising 1150 cases and 1342 controls along with 287 cases with overall survival information. We found that individuals with the A allele of B7-2 rs2681416 in CTLA-4 immune checkpoint pathway had a significantly increased risk of colorectal cancer [odds ratio (OR) = 1.37, P = 3.17 × 10-4] than those with G allele under the dominant model, which had a predominant site-specific effect in colon cancer (OR = 1.55, P = 3.11 × 10-5). In addition, rs2681416 significantly decreased the overall survival time of patients with colon cancer [hazard ratio (HR) = 1.96, P = 1.10 × 10-2], but not of patients with rectal cancer (P = 0.271). Moreover, rs2681416 had an expression quantitative trait locus effect on the B7-2 flanking gene IQCB1 in colon tissues, which contributed to colon cancer risk by regulating genome organization and influenced the expression of IQCB1 in an allele-specific manner. IQCB1 expression was upregulated in colorectal cancer tissues compared with normal tissues, accounting for various critical carcinogenic states in colon cancer and promoting immune infiltration of Th17 cells in the tumor microenvironment. Our study highlights the important roles of genetic variations in immune checkpoint pathways and provides new insight into potential site-specific independent biomarkers for colorectal cancer susceptibility, prognosis, and tumor immune status.

Identification of low-frequency variants of UGT1A3 associated with bladder cancer risk by next-generation sequencing.

Although genome-wide association studies (GWASs) have successfully revealed many common risk variants for bladder cancer, the heritability is still largely unexplained. We hypothesized that low-frequency variants involved in bladder cancer risk could reveal the unexplained heritability. Next-generation sequencing of 113 patients and 118 controls was conducted on 81 genes/regions of known bladder cancer GWAS loci. A two-stage validation comprising 3,350 cases and 4,005 controls was performed to evaluate the effects of low-frequency variants on bladder cancer risk. Biological experiments and techniques, including electrophoretic mobility shift assays, CRISPR/Cas9, RNA-Seq, and bioinformatics approaches, were performed to assess the potential functions of low-frequency variants. The low-frequency variant rs28898617 was located in the first exon of UGT1A3 and was significantly associated with increased bladder cancer risk (odds ratio = 1.50, P = 3.10 × 10-6). Intriguingly, rs28898617 was only observed in the Asian population, but monomorphism was observed in the European population. The risk-associated G allele of rs28898617 increased UGT1A3 expression, facilitated UGT1A3 transcriptional activity, and enhanced the binding activity. In addition, UGT1A3 deletion significantly inhibited the proliferation, invasion, and migration of bladder cancer cells and xenograft tumor growth. Mechanistically, UGT1A3 induced LAMC2 expression by binding CBP and promoting histone acetylation, which remarkably promoted the progression of bladder cancer. This is the first targeted sequencing study to reveal that the novel low-frequency variant rs28898617 and its associated gene UGT1A3 are involved in bladder cancer development, providing new insights into the genetic architecture of bladder cancer.

UBE2N Regulates Paclitaxel Sensitivity of Ovarian Cancer via Fos/P53 Axis.

BACKGROUND: Chemo-resistance is still considered one of the key factors in the mortality of ovarian cancer. In this work, we found that ubiquitin-conjugating enzyme E2 N (UBE2N) is downregulated in paclitaxel-resistant ovarian cancer cells. It suggests UBE2N to be critical in the regulation of paclitaxel sensitivity in ovarian cancer. MATERIALS AND METHODS: Ovarian cancer cells with stably overexpressed UBE2N were injected into nude mice to assess tumor growth and paclitaxel sensitivity in vivo. The MTT assay was applied to observe the effect of UBE2N expression on paclitaxel sensitivity. A real-time PCR array, specific for human cancer drug resistance, was used to examine the potential downstream target genes of UBE2N. The expression of UBE2N and potential downstream target genes was determined by Western blotting. The analysis of Gene Ontology and protein-protein interactions of these differentially expressed genes (DEGs) was performed using online tools. To evaluate the prognostic value of hub genes expression for ovarian cancer patients treated with paclitaxel, we applied the online survival analysis tool. RESULTS: Overexpressed UBE2N enhanced the paclitaxel sensitivity of ovarian cancer cells in vitro and in vivo. Thirteen upregulated DEGs and 11 downregulated DEGs were identified when we knockdown UBE2N. Meanwhile, 9 hub genes with a high degree of connectivity were selected. Only Fos proto-oncogene, AP-1 transcription factor subunit (Fos), was overexpressed upon decreasing UBE2N levels, indicating a poor outcome for patients treated with paclitaxel. Moreover, reduced UBE2N could increase Fos expression and reduce P53. Furthermore, reversed regulation of Fos and P53 based on UBE2N reduction could reverse paclitaxel sensitivity, respectively. CONCLUSION: Our study suggests that UBE2N could be used as a therapeutic agent for paclitaxel-resistant ovarian cancer through Fos/P53 pathway. Further studies are needed to elucidate the specific mechanism.

Oncogenic role of ABHD5 in endometrial cancer.

BACKGROUND: Abhydrolase domain containing 5 (ABHD5) functions as a tumor suppressor in colorectal and prostate cancers. The aim of this study was to investigate the roles of ABHD5 in endometrial cancer. MATERIALS AND METHODS: ABHD5 expression was detected in clinical samples by immunohistochemical staining. Cell proliferation and invasion were evaluated with the Cell Counting Kit-8 and Transwell assay, respectively. Western blotting was performed to analyze protein expression. Glucose uptake was assessed by 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]-2-deoxyglucose. Lactate production was detected by a lactate assay kit. RESULTS: In the present study, ABHD5 was overexpressed in endometrial cancer tissues, and its expression was closely correlated with the International Federation of Gynecology and Obstetrics (FIGO) stage and lymph node metastasis. In addition, we observed that the knockdown of ABHD5 inhibited cell proliferation, invasion, glucose uptake and lactate production in HEC-1A cells, which expressed high levels of ABHD5. Conversely, the opposite effects were observed when ABHD5 was ectopically expressed in Ishikawa cells, which had low levels of ABHD5. Furthermore, the changes in glycolysis regulators (enolase 1 [ENO1], glucose transporter 1 [GLUT1] and lactate dehydrogenase A [LDHA]) and epithelial-to-mesenchymal transition-related proteins (E-cadherin and Snail) in HEC-1A cells with ABHD5 knockdown were consistent with the effects of ABHD5 on glycolysis and cell invasion. Phosphatase and tensin homolog deleted on chromosome 10 (PTEN) was increased, while the phosphorylated AKT (p-AKT) was decreased when ABHD5 was downregulated. Notably, treatment with the allosteric AKT inhibitor MK-2206 completely abolished the effects caused by ABHD5 overexpression in Ishikawa cells. Finally, ABHD5 knockdown potently suppressed tumor growth in vivo. CONCLUSION: Overall, these results suggest that ABHD5 may play an oncogenic role in endometrial cancer via the AKT pathway.

Fatty Acid-Binding Protein 4 in Endometrial Epithelium Is Involved in Embryonic Implantation.

AIMS: To clarify the role of fatty acid-binding protein 4 (FABP4) of endometrial epithelial cell in the establishment and maintenance of pregnancy and the involvement in the pathogenesis of pregnancy loss. METHODS: The expression of FABP4 and uterine receptive factor (LIF, Integrin-ß3 and Claudin 4) was determined by Western blotting or quantitative PCR. FABP4 siRNA was used to silence FABP4 while FABP4 inhibitor was used to inhibit the function of FABP4 in endometrial epithelial cell. ICR mice were raised to evaluate the effect of FABP4 silence or inhibition on embryo implantation in vivo after FABP4 siRNA mixture or inhibitor was injected into uterus, and an embryonic adhesion system using trophoblast spheroids mimicking embryos was set up to assess the effect of FABP4 silence or inhibition on embryonic adhesion onto endometrial cell in vitro. RESULTS: The expression of FABP4 mRNA was significantly decreased in the deciduas of women with pregnancy loss compared with that of women with normal pregnancy. FABP4 siRNA significantly reduced the number of embryos implanted and FABP4 expression in ICR mice. FABP4 inhibition also significantly decreased the number of embryos implanted. Either silence or inhibition of FABP4 in endometrial epithelial cell abolished the expression of uterine receptive factors induced by the combination of estrogen and progesterone-induced, and reduced the number of trophoblast spheroids adhered onto endometrial cell. CONCLUSIONS: FABP4 regulates embryo implantation via altering uterine receptivity and decreased expression of FABP4 in endometrium may be linked with pregnancy loss, indicating FABP4 has biological role in the establishment and maintenance of pregnancy and subsequently is involved in pathogenesis of pregnancy loss.

Expression and function of fatty acid-binding protein 4 in epithelial cell of uterine endometrium.

The aims of this study were to delineate the expression of fatty-acid binding protein (FABP) 4 in human uterine endometrium and its function in the regulation of proliferation, migration and invasion of epithelial cells. Immunohistochenistry, immunofluorence and Western blotting were used to determine the expression and cellular localization of FABP4 in endometrium and endometrial epithelial cell lines. Interference of small ribonuclear acid (siRNA) and specific FABP4 inhibitor were used to inhibit FABP4. The proliferation, migration and invasion of epithelial cells were evaluated with CCK-8 assay, wound-healing test and transwell analysis respectively. We found that FABP4 was expressed by epithelial cells of proliferative endometrium and epithelial and stromal cells of secrectory endometrium. Epithelial cell lines Ishikawa and RL-952 expressed FABP4 and this expression was decreased by FABP4 siRNA. FABP4 siRNA and specific FABP4 inhibition significantly decreased the proliferation, migration and invasion of epithelial cell lines. We concluded that FABP4 is functionally expressed in endometrial epithelium and is necessary for maintaining the cell function of epithelial cells of endometrium.

Decreased fetal pre-adipocyte factor-1 in pregnancies complicated by gestational diabetes mellitus.

BACKGROUND: Serum pre-adipocyte factor-1 (pref-1) is an inhibitor of adipocyte differentiation that increases in small for gestational age fetuses. It plays a role in adipose metabolism and is associated with an increased risk of metabolic diseases in adulthood. We hypothesized that preadipocyte factor-1 (pref-1) concentration is altered in fetuses born to women with gestational diabetes mellitus (GDM). METHODS: Umbilical cord blood pref-1 concentrations were determined by enzyme-linked immunosorbant assay in 37 fetuses from pregnancies complicated by GDM and 45 fetuses from normal pregnancies. RESULTS: Serum pref-1 concentrations were significantly lower in fetuses of women with GDM compared to normal pregnancies (16.12±6.48 vs. 22.09±7.22 µg/l, P=0.001). Birth weight was significantly higher in GDM fetuses compared to normal pregnancies (3567±544 vs. 3253±370 g, P=0.003). CONCLUSIONS: Pregnancies complicated by GDM have decreased fetal pref-1 concentrations compared to normal pregnancies. These differences may be significant in terms of their later development of metabolic conditions.

Potential role of mannose-binding lectin in intrauterine transmission of hepatitis B virus.

Intrauterine transmission of hepatitis B virus (HBV) is the main cause of the high prevalence of HBV in endemic areas; however, the mechanisms underlying intrauterine transmission of HBV remain unknown. To explore the role of mannose-binding lectin (MBL), a pattern recognition molecule of the innate immune system, in intrauterine transmission of HBV, we determined MBL levels using an enzyme-linked immunosorbent assay (ELISA) in cord serum of 7 intrauterine-infected neonates and 30 non-infected neonates born to HBV-positive mothers, and 30 control neonates born to HBV-negative mothers. We observed significant differences in cord serum MBL levels among the three groups (P < 0.001). Non-infected neonates had significantly higher MBL levels than controls (P < 0.001), and intrauterine-infected neonates had significantly lower serum MBL levels than non-infected neonates (P < 0.001). However, serum MBL levels were not significantly different between intrauterine-infected neonates and controls (P = 0.800). Our results indicate that maternal HBV infection induces an increase in fetal MBL levels and the absence of this increase is possibly associated with intrauterine transmission of HBV, suggesting that MBL plays a role in intrauterine transmission of HBV.

Serum preadipocyte factor-1 is increased in fetuses of pregnancy complicated with severe preeclampsia.

BACKGROUND: Preadipocyte factor-1 (Pref-1), an inhibitor of adipocyte differentiation, is increased in fetal blood of small for gestational age (SGA) and is considered a factor involved in determining adiposity and associated with high risk of metabolic diseases in adulthood. Preeclampsia is a condition closely associated with SGA, however, the alteration of Pref-1 of in fetuses of preeclampsia remains unknown. The aims of the current investigation were to clarify the alteration of serum Pref-1 in fetuses of preeclamptic pregnancy and to explore possible role of Pref-1 in metabolic diseases in late life. METHODS: Cord blood samples were taken at birth from 45 fetuses of normal pregnancy, 16 of gestational hypertension, 29 of mild preeclampsia and 29 of severe preeclampsia. Serum Pref-1 concentrations were measured with ELISA. RESULTS: There were significant differences in cord blood Pref-1 and neonatal birth weight among normal pregnancy, gestational hypertension, mild and severe preeclampsia (F=8.557, P<0.001 for Pref-1; F=38.405, P<0.001 for birth weight). Serum Pref-1 was significantly higher while birth weight were lower in severe preeclampsia than normal pregnancy, gestational hypertension and mild preeclampsia respectively (P<=0.001 for all). However, either serum Pref-1 or birth weight did not significantly differ among normal pregnancy, gestational hypertension and mild preeclampsia (P>0.05 for all). Fetal Pref-1 concentration was significantly negatively correlated with birth weight (R(2)=0.175, P=0.027 for severe preeclampsia; R(2)=0.209, P<0.001 for preeclampsia; R(2)=0.25, P<0.001 for all subjects). CONCLUSIONS: Increased serum Pref-1 was demonstrated in fetuses of preeclampsia-complicated pregnancy, and it may be proposed that Pref-1 is among the possible mediators leading to high risk of metabolic diseases in adulthood.