Maternal pre-pregnancy BMI, offspring epigenome-wide DNA methylation, and childhood obesity: findings from the Boston Birth Cohort.

BACKGROUND: Maternal pre-pregnancy obesity is an established risk factor for childhood obesity. Investigating epigenetic alterations induced by maternal obesity during fetal development could gain mechanistic insight into the developmental origins of childhood obesity. While obesity disproportionately affects underrepresented racial and ethnic mothers and children in the USA, few studies investigated the role of prenatal epigenetic programming in intergenerational obesity of these high-risk populations. METHODS: This study included 903 mother-child pairs from the Boston Birth Cohort, a predominantly urban, low-income minority birth cohort. Mother-infant dyads were enrolled at birth and the children were followed prospectively to age 18 years. Infinium Methylation EPIC BeadChip was used to measure epigenome-wide methylation level of cord blood. We performed an epigenome-wide association study of maternal pre-pregnancy body mass index (BMI) and cord blood DNA methylation (DNAm). To quantify the degree to which cord blood DNAm mediates the maternal BMI-childhood obesity, we further investigated whether maternal BMI-associated DNAm sites impact birthweight or childhood overweight or obesity (OWO) from age 1 to age 18 and performed corresponding mediation analyses. RESULTS: The study sample contained 52.8% maternal pre-pregnancy OWO and 63.2% offspring OWO at age 1-18 years. Maternal BMI was associated with cord blood DNAm at 8 CpG sites (genome-wide false discovery rate [FDR] < 0.05). After accounting for the possible interplay of maternal BMI and smoking, 481 CpG sites were discovered for association with maternal BMI. Among them 123 CpGs were associated with childhood OWO, ranging from 42% decrease to 87% increase in OWO risk for each SD increase in DNAm. A total of 14 identified CpG sites showed a significant mediation effect on the maternal BMI-child OWO association (FDR < 0.05), with mediating proportion ranging from 3.99% to 25.21%. Several of these 14 CpGs were mapped to genes in association with energy balance and metabolism (AKAP7) and adulthood metabolic syndrome (CAMK2B). CONCLUSIONS: This prospective birth cohort study in a high-risk yet understudied US population found that maternal pre-pregnancy OWO significantly altered DNAm in newborn cord blood and provided suggestive evidence of epigenetic involvement in the intergenerational risk of obesity.

Healthy lifestyle, DNA methylation age acceleration, and incident risk of coronary heart disease.

BACKGROUND: DNA methylation clocks emerged as a tool to determine biological aging and have been related to mortality and age-related diseases. Little is known about the association of DNA methylation age (DNAm age) with coronary heart disease (CHD), especially in the Asian population. RESULTS: Methylation level of baseline blood leukocyte DNA was measured by Infinium Methylation EPIC BeadChip for 491 incident CHD cases and 489 controls in the prospective China Kadoorie Biobank. We calculated the methylation age using a prediction model developed among Chinese. The correlation between chronological age and DNAm age was 0.90. DNA methylation age acceleration (Δage) was defined as the residual of regressing DNA methylation age on the chronological age. After adjustment for multiple risk factors of CHD and cell type proportion, compared with participants in the bottom quartile of Δage, the OR (95% CI) for CHD was 1.84 (1.17, 2.89) for participants in the top quartile. One SD increment in Δage was associated with 30% increased risk of CHD (OR = 1.30; 95% CI 1.09, 1.56; Ptrend = 0.003). The average number of cigarette equivalents consumed per day and waist-to-hip ratio were positively associated with Δage; red meat consumption was negatively associated with Δage, characterized by accelerated aging in those who never or rarely consumed red meat (all P < 0.05). Further mediation analysis revealed that 10%, 5% and 18% of the CHD risk related to smoking, waist-to-hip ratio and never or rarely red meat consumption was mediated through methylation aging, respectively (all P for mediation effect < 0.05). CONCLUSIONS: We first identified the association between DNAm age acceleration and incident CHD in the Asian population, and provided evidence that unfavorable lifestyle-induced epigenetic aging may play an important part in the underlying pathway to CHD.

Hsa_circ_0007312 Promotes Third-Generation Epidermal Growth Factor Receptor-Tyrosine Kinase Inhibitor Resistance through Pyroptosis and Apoptosis via the MiR-764/MAPK1 Axis in Lung Adenocarcinoma Cells.

Purposes: Osimertinib is a third-generation epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI) used for patients with gefitinib (first-generation EGFR-TKI) resistance, but osimertinib resistance inevitably occurs. Therefore, it is necessary to explore the mechanisms of osimertinib resistance. Materials and Methods: We performed quantitative real-time polymerase chain reaction to detect hsa_circ_0007312 (circ7312), miR-764, and MAPK1 expressions in tissues and cells. Western blotting was used to detect protein levels in cells. Cell Counting Kit-8, apoptotic, and Transwell assays were used to explore biological functions. Luciferase assays were used to identify the interactions between circ7312 and miR-764, MAPK1 and miR-764. A xenograft experiment was performed to clarify the role of circ7312 in vivo. Public datasets were used to identify the relation between circ7312 expression and the cell half maximal inhibitory concentration value of osimertinib in 41 lung adenocarcinoma cell lines. The Student t-test, Kaplan-Meier analysis, and Pearson correlation analysis were used in data analysis. Results: We found that circ7312 knockdown increased miR-764 expression and decreased MAPK1 expression, and circ7312 regulated MAPK1 by sponging miR-764. In addition, high circ7312 expression has significant positive correlation with osimertinib IC50 values, circ7312 knockdown decreased the cell half maximal inhibitory concentration value of osimertinib and increased pyroptosis and apoptosis by sponging the miR-764/MAPK1 axis. We also found that circ7312 and MAPK1 were highly expressed in tumor tissues and related to poor prognosis. Xenograft experiments revealed that circ7312 knockdown decreased osimertinib resistance in vivo. Conclusion: We demonstrated that the inhibition of circ7312 decreased osimertinib resistance by promoting pyroptosis and apoptosis via the miR-764/MAPK1 axis, providing a novel target for osimertinib resistance therapy.

circ_0000567/miR-421/TMEM100 Axis Promotes the Migration and Invasion of Lung Adenocarcinoma and Is Associated with Prognosis.

Purpose: Due to the high metastatic ability and poor prognosis of lung adenocarcinoma (LUAD), we identified novel non-coding RNAs, which constitute approximately 60% of human transcripts, as prognostic biomarkers and potential therapeutic targets for LUAD. Methods: In this study, we downloaded and analyzed microRNA (miRNA) datasets from The Cancer Genome Atlas (TCGA) to identify dysregulated miRNAs correlating with the overall survival (OS) of LUAD patients. miR-421, circ_0000567, and TMEM100 expression levels were examined by quantitative real-time polymerase chain reaction (qRT-PCR) in NSCLC tissues from 73 patients and adjacent normal tissues. Cell migration and invasion were assayed using wound healing and transwell assays. miR-421 target predictions were conducted using starBase, CircInteractome, circBank, TargetScan, miRanda, MirDB, miRpath, and Gene Expression Omnibus (GEO) databases. The circular structure and stability of circ_0000567 were verified by RNase R digestion and qRT-PCR using oligo(dT) and random primers. A luciferase reporter assay was used to evaluate the relationship between miR-421, circ_0000567, and TMEM100. Results: The miRNA panel associated with OS in patients with LUAD was screened according to the hazard ratio (HR) of miRNAs from high to low. Based on the correlation between these miRNAs and OS, as well as miRNA expression levels, miR-421 was selected for further outcome analysis. High miR-421 expression was an independent risk factor for shorter OS in 73 patients collected from our department. Bioinformatic analyses, luciferase reporter assays, and functional assays showed that circ_0000567 could act as a sponge for miR-421 and prevent it from directly targeting the 3'-untranslated region of TMEM100 mRNA and further degrading it in LUAD. miR-421 expression was significantly upregulated, while circ_0000567 and TMEM100 were downregulated in tumor tissues of LUAD, compared to their counterparts in normal tissues. Gain- and loss-of-function assays showed that miR-421 promoted LUAD cell migration and invasion. Overexpression of circ_0000567 inhibited migration and invasion, whereas co-transfection of circ_0000567 and miR-421 mimics partly counteracted this effect. TMEM100 was upregulated by enhanced circ_0000567 in LUAD cells, and the expression of TMEM100 was inversely proportional to miR-421, whereas it was directly proportional to circ_0000567 in 73 LUAD specimens, which confirmed the competitive endogenous RNA (ceRNA) network. Conclusion: Our findings suggest that miR-421 promotes the migration and invasion of lung adenocarcinoma via circ_0000567/miR-421/TMEM100 signaling and could be a prognostic biomarker for LUAD.

Construction of a circRNA-miRNA-mRNA Regulated Pathway Involved in EGFR-TKI Lung Adenocarcinoma Resistance.

Objectives: Epidermal growth factor receptor-tyrosine kinase inhibitors are widely used for lung epidermal growth factor receptor-positive lung adenocarcinomas, but acquired resistance is inevitable. Although non-coding RNAs, such as circular RNA and microRNA, are known to play vital roles in epidermal growth factor receptor-tyrosine kinase inhibitor resistance, comprehensive analysis is lacking. Thus, this study aimed to explore the circular RNA-microRNA-messenger RNA regulatory network involved in epidermal growth factor receptor-tyrosine kinase inhibitor resistance. Methods: To identify differentially expressed genes between the epidermal growth factor receptor-tyrosine kinase inhibitor sensitive cell line PC9 and resistant cell line PC9/ epidermal growth factor receptor-tyrosine kinase inhibitor resistance(PC9/ER), circular RNA, microRNA and messenger RNA microarrays were performed. Candidates were then identified to construct a circular RNA-microRNA-messenger RNA network using bioinformatics. Additionally, Gene Oncology and Kyoto Encyclopedia of Genes and Genomes pathway analyses were conducted to evaluate the network messenger RNA, setting up a protein-protein interaction network for hub-gene identification. Afterwards, RNA immunoprecipitation was performed to enrich microRNA, and quantitative real-time PCR was used to estimated gene expression levels. Results: In total, 603, 377, and 1863 differentially expressed circular RNA, microRNA, messenger RNAs, respectively, were identified using microarray analysis, constructing a circular RNA-microRNA-messenger RNA network containing 18 circular RNAs, 17 microRNAs and 175 messenger RNAs. Moreover, Gene Oncology and Kyoto Encyclopedia of Genes and Genomes pathway analyses showed that the most enriched biological process terms and pathways were related to epidermal growth factor receptor-tyrosine kinase inhibitor resistance, including Wnt and Hippo signaling pathways. Based on the competing endogenous RNA and protein-protein interaction network, circ-0007312 was showed to interact with miR-764 and both circ-0003748 and circ-0001398 were shown to interact with miR-628; both these microRNAs targeted MAPK1. Furthermore, circ-0007312, circ-0003748, circ-0001398, and MAPK1 were up-regulated, whereas miR-764 and miR-628 were downregulated in PC9/ER cells as compared to parental PC9 cells. We also found that circ-0007312 and miR-764 were positively expressed in plasma. Conclusions: Our original study associated with mechanism of target therapy in lung cancer provided a systematic and comprehensive regulation of circular RNA, microRNA and messenger RNA in epidermal growth factor receptor-tyrosine kinase inhibitor resistance. It was found that circ-0007312- miR-764-MAPK1, circ-0003748-miR-628-MAPK1, and circ-0001398-miR-628-MAPK1 axis may play key roles in epidermal growth factor receptor-tyrosine kinase inhibitor resistance.

HIF1A-AS2 induces osimertinib resistance in lung adenocarcinoma patients by regulating the miR-146b-5p/IL-6/STAT3 axis.

Although epidermal growth factor receptor tyrosine kinase inhibitors (TKIs) show efficacy in lung adenocarcinoma (LUAD) patients, TKI resistance inevitably develops, limiting long-term results. Thus, there is an urgent need to address drug resistance in LUAD. Long non-coding RNA (lncRNA) HIF1A-AS2 could be a critical mediator in the progression of various tumor types. We examined the function of HIF1A-AS2 in modifying tumor aggravation and osimertinib resistance in lung adenocarcinoma. Using clinical samples, we showed that HIF1A-AS2 was upregulated in LUAD specimens, predicting poorer overall survival and disease-free survival. HIF1A-AS2 silencing inhibited the proliferation, migration, and tumorigenesis of LUAD cells and therapeutic efficacy of osimertinib against tumor cells in vitro and in vivo. RNA precipitation assays, western blotting, luciferase assays, and rescue experiments demonstrated that HIF1A-AS2 sponged microRNA-146b-5p (miR-146b-5p), promoting interleukin-6 (IL-6) expression, activating the IL-6/STAT3 pathway, and leading to LUAD progression. miR-146b-5p and IL-6 levels were correlated with the prognosis of LUAD patients. Our results indicated that HIF1A-AS2 functions as an oncogenic factor in adenocarcinoma cells by targeting the miR-146b-5p/IL-6/STAT3 axis and may be a prognostic indicator of survival. Moreover, it can be a potential therapeutic target to enhance the efficacy of osimertinib in LUAD patients.

Epigenome-wide analysis of DNA methylation and coronary heart disease: a nested case-control study.

Background: Identifying environmentally responsive genetic loci where DNA methylation is associated with coronary heart disease (CHD) may reveal novel pathways or therapeutic targets for CHD. We conducted the first prospective epigenome-wide analysis of DNA methylation in relation to incident CHD in the Asian population. Methods: We did a nested case-control study comprising incident CHD cases and 1:1 matched controls who were identified from the 10 year follow-up of the China Kadoorie Biobank. Methylation level of baseline blood leukocyte DNA was measured by Infinium Methylation EPIC BeadChip. We performed the single cytosine-phosphate-guanine (CpG) site association analysis and network approach to identify CHD-associated CpG sites and co-methylation gene module. Results: After quality control, 982 participants (mean age 50.1 years) were retained. Methylation level at 25 CpG sites across the genome was associated with incident CHD (genome-wide false discovery rate [FDR] < 0.05 or module-specific FDR < 0.01). One SD increase in methylation level of identified CpGs was associated with differences in CHD risk, ranging from a 47 % decrease to a 118 % increase. Mediation analyses revealed 28.5 % of the excessed CHD risk associated with smoking was mediated by methylation level at the promoter region of ANKS1A gene (P for mediation effect = 0.036). Methylation level at the promoter region of SNX30 was associated with blood pressure and subsequent risk of CHD, with the mediating proportion to be 7.7 % (P = 0.003) via systolic blood pressure and 6.4 % (P = 0.006) via diastolic blood pressure. Network analysis revealed a co-methylation module associated with CHD. Conclusions: We identified novel blood methylation alterations associated with incident CHD in the Asian population and provided evidence of the possible role of epigenetic regulations in the smoking- and blood pressure-related pathways to CHD risk. Funding: This work was supported by National Natural Science Foundation of China (81390544 and 91846303). The CKB baseline survey and the first re-survey were supported by a grant from the Kadoorie Charitable Foundation in Hong Kong. The long-term follow-up is supported by grants from the UK Wellcome Trust (202922/Z/16/Z, 088158/Z/09/Z, 104085/Z/14/Z), grant (2016YFC0900500, 2016YFC0900501, 2016YFC0900504, 2016YFC1303904) from the National Key R&D Program of China, and Chinese Ministry of Science and Technology (2011BAI09B01).

Small Cell Size Circulating Aneuploid Cells as a Biomarker of Prognosis in Resectable Non-Small Cell Lung Cancer.

OBJECTIVE: The size distribution of circulating aneuploid cells (CACs) and its clinical significance were investigated in resectable non-small cell lung cancer (NSCLC). PATIENTS AND METHODS: A total of 50 patients with resectable NSCLC were enrolled in this study. Blood samples (50 pre-surgery and 35 post-surgery) were collected and used for the detection of CAC chromosome 8 heteroploidy through the subtraction enrichment and immunostaining fluorescence in situ hybridization (SE-iFISH) method. RESULTS: Less than 20% small cell size and more than 80% large cell size CACs were detected. Karyotypes, including triploid, tetraploid, and multiploid, had varying distributions. The triploid subtype accounted for the majority of small cell size CACs, whereas the multiploid subtype accounted for the majority of large cell size CACs. We found that total small cell size and triploid small cell size CACs, but not large cell size CACs, derived from pre-surgery samples, were associated with shorter disease-free survival. Moreover, total small cell size and triploid small cell size CACs were associated with higher TNM stage and recurrence. Nevertheless, the variation between pre- and post-surgery CACs was not related to survival among patients with resectable NSCLC. CONCLUSIONS: Pre-surgery small cell size CACs, especially the triploid subtype, could be regarded as a potential prognostic biomarker for patients with resectable NSCLC.

A transdisciplinary approach to understand the epigenetic basis of race/ethnicity health disparities.

Health disparities correspond to differences in disease burden and mortality among socially defined population groups. Such disparities may emerge according to race/ethnicity, socioeconomic status and a variety of other social contexts, and are documented for a wide range of diseases. Here, we provide a transdisciplinary perspective on the contribution of epigenetics to the understanding of health disparities, with a special emphasis on disparities across socially defined racial/ethnic groups. Scientists in the fields of biological anthropology, bioinformatics and molecular epidemiology provide a summary of theoretical, statistical and practical considerations for conducting epigenetic health disparities research, and provide examples of successful applications from cancer research using this approach.

Improved lipidomic profile mediates the effects of adherence to healthy lifestyles on coronary heart disease.

Adherence to healthy lifestyles is associated with reduced risk of coronary heart disease (CHD), but uncertainty persists about the underlying lipid pathway. In a case-control study of 4681 participants nested in the prospective China Kadoorie Biobank, 61 lipidomic markers in baseline plasma were measured by targeted nuclear magnetic resonance spectroscopy. Baseline lifestyles included smoking, alcohol consumption, dietary habit, physical activity, and adiposity levels. Genetic instrument was used to mimic the lipid-lowering effect of statins. We found that 35 lipid metabolites showed statistically significant mediation effects in the pathway from healthy lifestyles to CHD reduction, including very low-density lipoprotein (VLDL) particles and their cholesterol, large-sized high-density lipoprotein (HDL) particle and its cholesterol, and triglyceride in almost all lipoprotein subfractions. The statins genetic score was associated with reduced intermediate- and low-density lipoprotein, but weak or no association with VLDL and HDL. Lifestyle interventions and statins may improve different components of the lipid profile.

Long non-coding RNA H19 confers resistance to gefitinib via miR-148b-3p/DDAH1 axis in lung adenocarcinoma.

Epidermal growth factor receptor tyrosine kinase inhibitors therapy, such as gefitinib, have proven to be effective for lung adenocarcinoma with epidermal growth factor receptor-sensitive mutations. However, drug resistance remains inevitable and the underlying mechanisms are still elusive and poorly understood. In order to explore the mechanisms underlying tyrosine kinase inhibitors resistance, we used long non-coding RNA microarray analysis and found that long non-coding RNA H19 was highly expressed in gefitinib-resistant cell lines. In addition, knockdown of long non-coding RNA H19 was found to be able to decrease cell proliferation, half maximal inhibitory concentration (IC50) of gefitinib, migration and invasion. Mechanistically, we demonstrated that long non-coding RNA H19 positively regulated dimethylarginine dimethylaminohydrolase-1 expression via sponging miR-148b-3p. Furthermore, overexpression or inactivation of miR-148b-3p could enhance or reverse the inhibitory effect of long non-coding RNA H19 inhibition in lung adenocarcinoma cells, respectively. High expression of either long non-coding RNA H19 or dimethylarginine dimethylaminohydrolase-1 was associated with poorer overall survival in patients with lung adenocarcinoma, while high expression of miR-148b was associated with better overall survival. Overall, our data revealed that long non-coding RNA H19 confers resistance to gefitinib via miR-148b/dimethylarginine dimethylaminohydrolase-1 axis in lung adenocarcinoma, which offers a new insight into the epidermal growth factor receptor tyrosine kinase inhibitors therapy resistance.

Early famine exposure and adult disease risk based on a 10-year prospective study of Chinese adults.

OBJECTIVE: To comprehensively examine the potential impacts of prenatal experience of the Chinese Great Famine on chronic disease risks in the middle age. METHODS: This study included 92 284 participants aged 39-51 years from China Kadoorie Biobank born around the famine period and without major chronic diseases at baseline. We categorised participants into non-famine births (born between 1 October 1956 and 30 September 1958, and 1 October 1962 and 30 September 1964) and famine births (born between 1 October 1959 and 30 September 1961). The outcomes were incident cardiovascular disease, cancer and respiratory system disease. Cox regression was used to estimate adjusted HR and 95% CI for famine exposure. Subgroup analyses were performed according to baseline characteristics. RESULTS: During a median 10.1 years of follow-up, we identified 4626 incident ischaemic heart disease (IHD) cases, 7332 cerebrovascular disease cases, 3111 cancer cases and 16 081 respiratory system disease cases. In the whole population, prenatal famine exposure was not statistically associated with the risks of developing any chronic diseases in adulthood. However, for urban participants, compared with non-famine births, famine births had a higher risk of cerebrovascular disease (HR 1.18; 95% CI 1.09 to 1.28); such association was not shown for rural participants (p for interaction <0.001). Also, we observed the associations of prenatal famine exposure with IHD (HR 1.15; 95% CI 1.05 to 1.26) and cerebrovascular disease (HR 1.13; 95% CI 1.05 to 1.21) in participants with lower physical activity level, but not in those with higher ones (all p for interaction=0.003). CONCLUSION: Our findings indicate that prenatal exposure to the Chinese famine might be associated with an increased cardiovascular risk and such risk may be modified by adult lifestyle.

Shisa3 brakes resistance to EGFR-TKIs in lung adenocarcinoma by suppressing cancer stem cell properties.

BACKGROUND: Although EGFR tyrosine kinase inhibitors (EGFR-TKIs) are beneficial to lung adenocarcinoma patients with sensitive EGFR mutations, resistance to these inhibitors induces a cancer stem cell (CSC) phenotype. Here, we clarify the function and molecular mechanism of shisa3 as a suppressor that can reverse EGFR-TKI resistance and inhibit CSC properties. METHODS: The suppresser genes involved in EGFR-TKI resistance were identified and validated by transcriptome sequencing, quantitative real-time PCR (qRT-PCR) and immunohistochemistry. Biological function analyses, cell half maximal inhibitory concentration (IC50), self-renewal, and migration and invasion capacities, were detected by CCK8, sphere formation and Transwell assays. Tumorigenesis and therapeutic effects were investigated in nonobese diabetic/severe combined immunodeficiency (nod-scid) mice. The underlying mechanisms were explored by Western blot and immunoprecipitation analyses. RESULTS: We found that low expression of shisa3 was related to EGFR-TKI resistance in lung adenocarcinoma patients. Ectopic overexpression of shisa3 inhibited CSC properties and the cell cycle in the lung adenocarcinoma cells resistant to gefitinib/osimertinib. In contrast, suppression of shisa3 promoted CSC phenotypes and the cell cycle in the cells sensitive to EGFR-TKIs. For TKI-resistant PC9/ER tumors in nod-scid mice, overexpressed shisa3 had a significant inhibitory effect. In addition, we verified that shisa3 inhibited EGFR-TKI resistance by interacting with FGFR1/3 to regulate AKT/mTOR signaling. Furthermore, combinational administration of inhibitors of FGFR/AKT/mTOR and cell cycle signaling could overcome EGFR-TKI resistance associated with shisa3-mediated CSC capacities in vivo. CONCLUSION: Taken together, shisa3 was identified as a brake to EGFR-TKI resistance and CSC characteristics, probably through the FGFR/AKT/mTOR and cell cycle pathways, indicating that shisa3 and concomitant inhibition of its regulated signaling may be a promising therapeutic strategy for reversing EGFR-TKI resistance.

Chronic hepatitis B virus infection and total and cause-specific mortality: a prospective cohort study of 0.5 million people.

OBJECTIVES: Chronic hepatitis B virus (HBV) infection is associated with a higher risk of liver diseases. Substantial uncertainty remains, however, about the associations of HBV infection with mortality from extrahepatic causes, especially from subtypes of cardiovascular diseases. We prospectively examined the association of chronic HBV infection with total and cause-specific mortality. DESIGN: Population-based prospective cohort study. SETTING: China Kadoorie Biobank in which participants from 10 geographically diverse areas across China were enrolled between 2004 and 2008. PARTICIPANTS: 475 801 participants 30-79 years of age without reporting major chronic diseases at baseline were enrolled. Hepatitis B surface antigen (HBsAg) was tested using an on-site rapid test strip at baseline. PRIMARY AND SECONDARY OUTCOME MEASURES: Total and cause-specific mortality. RESULTS: A total of 35 822 deaths were recorded during ~10 years of follow-up. In multivariable-adjusted analyses, compared with HBsAg-negative participants, HBsAg-positive participants had an increased risk of total mortality (HR=2.01, 95% CI: 1.91 to 2.12), which was higher in men (HR=2.16, 95% CI: 2.01 to 2.31) than in women (HR=1.74, 95% CI: 1.60 to 1.90). Presence of HBsAg was associated with increased mortality from liver cancer (1339 deaths, HR=13.95, 95% CI: 12.46 to 15.62), infections (410 deaths, HR=10.30, 95% CI: 8.21 to 12.94), digestive diseases (688 deaths, HR=6.83, 95% CI: 5.49 to 8.50), intracerebral haemorrhage (4077 deaths, HR=1.38, 95% CI: 1.14 to 1.68) and ischaemic heart diseases (4624 deaths, HR=1.31, 95% CI: 1.09 to 1.58). The positive association between HBsAg status and risk of death was stronger in participants younger than 50 years, smokers, physically active or non-hypertensive participants. CONCLUSIONS: Among Chinese adults, chronic HBV infection was associated with increased mortality from a range of hepatic and extrahepatic diseases.

Long-term survival of patients with locally advanced esophageal squamous cell carcinoma receiving esophagectomy following neoadjuvant chemotherapy: a cohort study.

PURPOSE: The role of neoadjuvant chemotherapy and subsequent adjuvant therapy in the treatment of patients with locally advanced esophageal squamous cell carcinomas (ESCC) is not well established. PATIENTS AND METHODS: We retrospectively reviewed 228 patients with locally advanced ESCC receiving esophagectomy following neoadjuvant chemotherapy from January 2007 through December 2016. The probabilities of disease-free survival (DFS) and overall survival (OS) were estimated by means of the Kaplan-Meier method and were compared with the use of the log-rank test. Univariate and multivariate analyses of predictors of DFS and OS were performed using a Cox proportional-hazards model. Propensity score matching analysis was performed for further analysis regarding the benefit of adjuvant therapy. RESULTS: The pathological complete response of neoadjuvant chemotherapy was achieved in 13 of 228 patients (5.7%). With a median follow-up of 59.6 months, the median DFS and OS were 35.4 and 45.4 months, respectively. The multivariate Cox model determined chemotherapy regimens (P=0.003) and ypT category (P=0.006) were significant independent predictors of DFS; and chemotherapy regimens (P=0.001), ypT category (P<0.001), and ypN category (P=0.013) were significant independent predictors of OS. Furthermore, patients who received adjuvant therapy seemed to be associated with poorer survival (both DFS and OS) compared with those who did not in full cohort (P=0.001 and P=0.184, respectively) and matched cohort (P=0.251 and P=0.374, respectively). CONCLUSION: Surgery following neoadjuvant chemotherapy was applicable. Chemotherapy regimens and ypT category were significant independent predictors of both DFS and OS and ypN category was also a significant independent predictor of OS. However, these patients did not seem to benefit from subsequent adjuvant therapy. The necessity of adjuvant therapy requires further investigation.

Genomic sequencing and editing revealed the GRM8 signaling pathway as potential therapeutic targets of squamous cell lung cancer.

The study sought to explore novel genetic aberration driving squamous cell lung carcinoma (LUSC). The whole exome (WES), whole genome (WGS) and target region (TS) sequencings and CRISPR-Cas9 genome editing techniques were integrated to explore and validate novel targeting candidates from LUSC primary tumors and corresponding patient-derived xenografts (PDXs). Seven genes (FGFR2, GRM1,PIK3CG, PIK3CA,ZFHX4, CSMD3, GRM8) with high frequencies of both single nucleotide variants (SNVs) and copy number variants (CNVs), and two genes (CLDN1 and RIT1) only with CNVs were identified by bioinformatics analysis. The functions of these candidates were validated through CRISPR-Cas9 system in primary PDX cells. Furthermore, we focused on the genetic and functional analysis of Metabotropic glutamate receptor 8 (GRM8), whose transcriptional activation was elucidated to promote the survival of LUSC tumor cell through inhibiting cAMP pathway and activating MAPK pathway. The SNV identified in GRM8, A112G, activated downstream signaling pathway and induced cell proliferation, which could be reversed by cAMP stimulator and MEK inhibitor. In conclusion, the components of GRM8 signaling pathway could serve as potential targets of squamous cell lung cancer carrying GRM8 activating variants.

Chronic hepatitis B virus infection and risk of chronic kidney disease: a population-based prospective cohort study of 0.5 million Chinese adults.

BACKGROUND: Existing evidence remains inconclusive as to the association between chronic hepatitis B virus (HBV) infection and the risk of chronic kidney disease (CKD). We prospectively examined the association between chronic HBV infection and CKD risk, and the joint associations of HBV infection with established risk factors of several lifestyle factors and prevalent diseases on CKD risk. METHODS: Participants from the China Kadoorie Biobank were enrolled during 2004-2008 and followed up until 31 December 2015. After excluding participants with previously diagnosed CKD, cancer, heart disease, and stroke at baseline, the present study included 469,459 participants. Hepatitis B surface antigen (HBsAg) was qualitatively tested at baseline. Incident CKD cases were identified mainly through the health insurance system and disease and death registries. RESULTS: During a median follow-up of 9.1 years (4.2 million person-years), we documented 4555 incident cases of CKD. Cox regression yielded multivariable-adjusted hazard ratios (HRs) and 95% confidence intervals (CIs). Compared with HBsAg-negative participants, the multivariable-adjusted HR (95% CI) for CKD was 1.37 (1.18, 1.60) for HBsAg-positive participants. The association was stronger in men (HR = 1.77; 95% CI: 1.43, 2.20) than in women (HR = 1.10; 95% CI: 0.88, 1.36). HBsAg-positive participants, with or without hepatitis or cirrhosis, whether or not under treatment, all showed increased risk of developing CKD. We observed positive additive interactions of HBsAg positivity with smoking, physical inactivity, or diabetes on CKD risk. Compared with HBsAg-negative participants who were nonsmokers, more physically active, or did not have diabetes at baseline, the greatest CKD risk for HBsAg-positive participants was for those who were smokers (HR = 1.85; 95% CI: 1.44, 2.38), physically inactive (HR = 1.91; 95% CI: 1.52, 2.40), or diabetic (HR = 6.11; 95% CI: 4.47, 8.36). CONCLUSIONS: In countries with a high endemicity of HBV infection, kidney damage associated with chronic HBV infection should be a non-negligible concern. Our findings also highlight the importance of health advice on quitting smoking, increasing physical activity, improving glucose control, and early screening for CKD in people with chronic HBV infection.

Associations of egg consumption with cardiovascular disease in a cohort study of 0.5 million Chinese adults.

OBJECTIVE: To examine the associations between egg consumption and cardiovascular disease (CVD), ischaemic heart disease (IHD), major coronary events (MCE), haemorrhagic stroke as well as ischaemic stroke. METHODS: During 2004-2008, over 0.5 million adults aged 30-79 years were recruited from 10 diverse survey sites in China. Participants were asked about the frequency of egg consumption and were followed up via linkages to multiple registries and active investigation. Among 461 213 participants free of prior cancer, CVD and diabetes, a total of 83 977 CVD incident cases and 9985 CVD deaths were documented, as well as 5103 MCE. Stratified Cox regression was performed to yield adjusted hazard ratios for CVD endpoints associated with egg consumption. RESULTS: At baseline, 13.1% of participants reported daily consumption (usual amount 0.76 egg/day) and 9.1% reported never or very rare consumption (usual amount 0.29 egg/day). Compared with non-consumers, daily egg consumption was associated with lower risk of CVD (HR 0.89, 95% CI 0.87 to 0.92). Corresponding multivariate-adjusted HRs (95% CI) for IHD, MCE, haemorrhagic stroke and ischaemic stroke were 0.88 (0.84 to 0.93), 0.86 (0.76 to 0.97), 0.74 (0.67 to 0.82) and 0.90 (0.85 to 0.95), respectively. There were significant dose-response relationships of egg consumption with morbidity of all CVD endpoints (P for linear trend <0.05). Daily consumers also had an 18% lower risk of CVD death and a 28% lower risk of haemorrhagic stroke death compared to non-consumers. CONCLUSION: Among Chinese adults, a moderate level of egg consumption (up to <1 egg/day) was significantly associated with lower risk of CVD, largely independent of other risk factors.

BPI-9016M, a c-Met inhibitor, suppresses tumor cell growth, migration and invasion of lung adenocarcinoma via miR203-DKK1.

Activation of c-Met plays a critical role in tumorigenesis, migration and invasion in lung cancer. Here, we explored the therapeutic efficacy of a novel small-molecule c-Met inhibitor (BPI-9016M) in lung adenocarcinoma and investigated the underlying molecular mechanisms. Method: BPI-9016M, a c-Met tyrosine kinase receptor inhibitor, was used to treat patient-derived xenografts (PDX) from lung adenocarcinoma in NOD/SCID mice. Immunohistochemistry and Western blot analysis were used to determine the expression of c-Met and its downstream signaling molecules. CCK8, wound healing, and trans-well assays were used to analyze cell proliferation, spreading, migration and invasion. RNA sequencing and quantitative real-time PCR (qPCR) was used to screen and validate the expression of downstream genes in lung adenocarcinoma cells treated with BPI-9016M. Luciferase reporter assay was used to detect the interaction between miRNA and the targeted gene. Results: BPI-9016M significantly suppressed growth in three out of four lung adenocarcinoma PDX models, particularly in the tumors with high expression of c-Met. In lung adenocarcinoma cell lines, BPI-9016M treatment resulted in increased miR203, which reduced migration and invasion and also repressed Dickkopf-related protein 1 (DKK1) expression. Forced overexpression of DKK1 or down-regulation of miR203 reversed the inhibitory effect of BPI-9016M on migration and invasion. C-Met was verified to positively and negatively associate with DKK1 and miR203, respectively. High expression of c-Met/DKK1 or low expression of miR203 related to poor outcome of lung adenocarcinoma patients. Furthermore, we observed significantly enhanced tumor cell growth inhibition upon combining BPI-9016M treatment with miR203 mimics or DKK1 siRNA. Conclusion: Our data indicated that BPI-9016M is an effective agent against lung adenocarcinoma, particularly in tumors with c-Met activation, and likely functions through upregulation of miR203 leading to reduced DKK1 expression.

CMTM1_v17 is associated with chemotherapy resistance and poor prognosis in non-small cell lung cancer.

BACKGROUND: Considering neoadjuvant chemotherapy (NAC) prior to surgery could shrink and reduce the primary tumor and distant micro-metastases to reduce the high relapses rates, NAC has been an accepted therapeutic management for patients with non-small cell lung cancer (NSCLC). CMTM1_v17 is highly expressed in human testis tissues and solid tumor tissues but relatively low expression was obtained in the corresponding normal tissues. This study aims to investigate the significance of CMTM1_v17 in NSCLC and its association with platinum-based NAC efficacy. METHODS: 31 pairs of tumor tissues before and after NAC and 78 resected tumor tissues after NAC were utilized for immunohistochemistry (IHC) staining of CMTM1_v17 protein. The correlation between CMTM1_v17 expression and chemotherapy efficacy was analyzed. The prognostic value of CMTM1_v17 index for disease-free survival (DFS) and overall survival (OS) was analyzed using Kaplan-Meier survival and multivariable Cox regression. RESULTS: CMTM1_v17 expression was related to treatment effect and outcome in tumor tissues after NAC not before NAC from 31 cases of NSCLC. We identified that high CMTM1_v17 expression was associated with low objective remission rate (ORR) (P = 0.008) and poor prognosis (the median OS: 35.1 months vs 65.6 months, P = 0.0045; the median DFS: 17.27 months vs 35.54 months, P = 0.0207) in the 31 patients. Next, we detected CMTM1_v17 expression to confirm correlation between this protein status and clinical characteristics in 78 NSCLC patients with NAC treatment. The upregulation of CMTM1_v17 had a higher SD rate (P = 0.007) and worse outcome (the median OS: 41.0 months vs 80.6 months, P = 0.0028; the median DFS: 33.4 vs 64.8 months, P = 0.0032). COX multivariate analysis indicated that CMTM1_v17 is an independent prognostic risk factor on patients who have received NAC (OS: HR = 3.642, P = 0.002; DFS:HR = 3.094, P = 0.002). CONCLUSIONS: CMTM1_v17 expression is significantly associated with chemoresistance and poor prognosis of the early stage NSCLC patients who have received NAC.