A pedigree-based prediction model identifies carriers of deleterious de novo mutations in families with Li-Fraumeni syndrome.

De novo mutations (DNMs) are increasingly recognized as rare disease causal factors. Identifying DNM carriers will allow researchers to study the likely distinct molecular mechanisms of DNMs. We developed Famdenovo to predict DNM status (DNM or familial mutation [FM]) of deleterious autosomal dominant germline mutations for any syndrome. We introduce Famdenovo.TP53 for Li-Fraumeni syndrome (LFS) and analyze 324 LFS family pedigrees from four US cohorts: a validation set of 186 pedigrees and a discovery set of 138 pedigrees. The concordance index for Famdenovo.TP53 prediction was 0.95 (95% CI: [0.92, 0.98]). Forty individuals (95% CI: [30, 50]) were predicted as DNM carriers, increasing the total number from 42 to 82. We compared clinical and biological features of FM versus DNM carriers: (1) cancer and mutation spectra along with parental ages were similarly distributed; (2) ascertainment criteria like early-onset breast cancer (age 20-35 yr) provides a condition for an unbiased estimate of the DNM rate: 48% (23 DNMs vs. 25 FMs); and (3) hotspot mutation R248W was not observed in DNMs, although it was as prevalent as hotspot mutation R248Q in FMs. Furthermore, we introduce Famdenovo.BRCA for hereditary breast and ovarian cancer syndrome and apply it to a small set of family data from the Cancer Genetics Network. In summary, we introduce a novel statistical approach to systematically evaluate deleterious DNMs in inherited cancer syndromes. Our approach may serve as a foundation for future studies evaluating how new deleterious mutations can be established in the germline, such as those in TP53.

Programmed cell death-ligand 1 expression in hepatocellular carcinoma and its correlation with clinicopathological characteristics.

BACKGROUND AND AIM: Programmed cell death-ligand 1 (PD-L1) immunohistochemistry score has been approved as the predictive biomarker for anti-PD1/PD-L1 therapy in several advanced malignancies. Although its predictive role remained inconclusive in hepatocellular carcinoma, ongoing study of anti-PD1/PD-L1 therapy showed promising results. However, less is known about the PD-L1 immunohistochemistry score and factors correlated with it in hepatocellular carcinoma. We investigated PD-L1 immunohistochemistry scores in a large cohort of hepatocellular carcinoma, as well as its correlation with various clinical and genomic factors. METHODS: Immunohistochemistry was performed to detect the expression of PD-L1 protein in 315 hepatocellular carcinoma tissues. All slides were independently reviewed by three senior pathologists. Next-generation YS panel (450 genes) sequencing was performed on 309 patients. RESULTS: Higher PD-L1 expression as measured by combined positive score (CPS) was associated with increased Edmondson-Steiner grade (grade III vs II, P = 0.041) and TP53 mutations (P = 0.021). PD-L1 CPS had no correlation with tumor mutational burden (Spearman's correlation coefficient 0.067). PD-L1 CPS was not significantly associated with hepatitis B virus infection. CONCLUSIONS: Our data indicated that patients with higher Edmondson-Steiner grade (grade III) had significantly higher PD-L1 CPS than patients with lower Edmondson-Steiner grade (grade II). Patients with TP53 mutations had significantly higher PD-L1 expression.

Trans-acting non-synonymous variant of FOXA1 predisposes to hepatocellular carcinoma through modulating FOXA1-ERα transcriptional program and may have undergone natural selection.

Interplay of pioneer transcription factor forkhead box A1 (FOXA1) and estrogen receptor has been implicated in sexual dimorphism in hepatocellular carcinoma (HCC), but etiological relevance of its polymorphism was unknown. In the case control study (1152 patients versus1242 controls), we observed significant increase in HCC susceptibility in hepatitis B virus carriers associated with a non-synonymous Thr83Ala variant of FOXA1 (odds ratio [OR], 1.28; 95% confidence interval [CI], 1.11-1.48, for Ala83-containing genotype, after validation in an independent population with 933 patients versus 1030 controls), a tightly linked (CGC)5/6or7 repeat polymorphism at its promoter (OR 1.32; 95% CI 1.10-1.60, for (CGC)6or7-repeat-containing genotype), and their combined haplotype (OR 1.50; 95% CI 1.24-1.81, for (CGC)6or7-Ala83 haplotype). The susceptible FOXA1-Ala83 impairs its interaction with ERα, attenuates transactivation toward some of their dual target genes, such as type 1 iodothyronine deiodinase, UDP glucuronosyltransferase 2 family, polypeptide B17 and sodium/taurocholate cotransporting polypeptide, but correlates with strengthened cellular expression of α-fetoprotein (AFP) and elevated AFP serum concentration in HCC patients (n = 1096). The susceptible FOXA1 cis-variant with (CGC)6or7 repeat strengthens the binding to transcription factor early growth response 1 and enhances promoter activity and gene expression. Evolutionary population genetics analyses with public datasets reveal significant population differentiation and unique haplotype structure of the derived protective FOXA1-Thr83 and suggest that it may have undergone positive natural selection in Chinese population. These findings epidemiologically highlight the functional significance of FOXA1-ERα transcriptional program and regulatory network in liver cancer development.

A genome-wide search for signals of high-altitude adaptation in Tibetans.

Genetic studies of Tibetans, an ethnic group with a long-lasting presence on the Tibetan Plateau which is known as the highest plateau in the world, may offer a unique opportunity to understand the biological adaptations of human beings to high-altitude environments. We conducted a genome-wide study of 1,000,000 genetic variants in 46 Tibetans (TBN) and 92 Han Chinese (HAN) for identifying the signals of high-altitude adaptations (HAAs) in Tibetan genomes. We discovered the most differentiated variants between TBN and HAN at chromosome 1q42.2 and 2p21. EGLN1 (or HIFPH2, MIM 606425) and EPAS1 (or HIF2A, MIM 603349), both related to hypoxia-inducible factor, were found most differentiated in the two regions, respectively. Strong positive correlations were also observed between the frequency of TBN-dominant haplotypes in the two gene regions and altitude in East Asian populations. Linkage disequilibrium and further haplotype network analyses of world-wide populations suggested the antiquity of the TBN-dominant haplotypes and long-term persistence of the natural selection. Finally, a "dominant haplotype carrier" hypothesis could describe the role of the two genes in HAA. All of our population genomic and statistical analyses indicate that EPAS1 and EGLN1 are most likely responsible for HAA of Tibetans. Interestingly, one each but not both of the two genes were also identified by three recent studies. We reanalyzed the available data and found the escaped top signal (EPAS1) could be recaptured with data quality control and our approaches. Based on this experience, we call for more attention to be paid to controlling data quality and batch effects introduced in public data integration. Our results also suggest limitations of extended haplotype homozygosity-based method due to its compromised power in case the natural selection initiated long time ago and particularly in genomic regions with recombination hotspots.

Genomic dissection of population substructure of Han Chinese and its implication in association studies.

To date, most genome-wide association studies (GWAS) and studies of fine-scale population structure have been conducted primarily on Europeans. Han Chinese, the largest ethnic group in the world, composing 20% of the entire global human population, is largely underrepresented in such studies. A well-recognized challenge is the fact that population structure can cause spurious associations in GWAS. In this study, we examined population substructures in a diverse set of over 1700 Han Chinese samples collected from 26 regions across China, each genotyped at approximately 160K single-nucleotide polymorphisms (SNPs). Our results showed that the Han Chinese population is intricately substructured, with the main observed clusters corresponding roughly to northern Han, central Han, and southern Han. However, simulated case-control studies showed that genetic differentiation among these clusters, although very small (F(ST) = 0.0002 approximately 0.0009), is sufficient to lead to an inflated rate of false-positive results even when the sample size is moderate. The top two SNPs with the greatest frequency differences between the northern Han and southern Han clusters (F(ST) > 0.06) were found in the FADS2 gene, which associates with the fatty acid composition in phospholipids, and in the HLA complex P5 gene (HCP5), which associates with HIV infection, psoriasis, and psoriatic arthritis. Ingenuity Pathway Analysis (IPA) showed that most differentiated genes among clusters are involved in cardiac arteriopathy (p < 10(-101)). These signals indicating significant differences among Han Chinese subpopulations should be carefully explained in case they are also detected in association studies, especially when sample sources are diverse.

NLRC5/CITA expression correlates with efficient response to checkpoint blockade immunotherapy.

Checkpoint blockade-mediated immunotherapy is emerging as an effective treatment modality for multiple cancer types. However, cancer cells frequently evade the immune system, compromising the effectiveness of immunotherapy. It is crucial to develop screening methods to identify the patients who would most benefit from these therapies because of the risk of the side effects and the high cost of treatment. Here we show that expression of the MHC class I transactivator (CITA), NLRC5, is important for efficient responses to anti-CTLA-4 and anti-PD1 checkpoint blockade therapies. Melanoma tumors derived from patients responding to immunotherapy exhibited significantly higher expression of NLRC5 and MHC class I-related genes compared to non-responding patients. In addition, multivariate analysis that included the number of tumor-associated non-synonymous mutations, predicted neo-antigen load and PD-L2 expression was capable of further stratifying responders and non-responders to anti-CTLA4 therapy. Moreover, expression or methylation of NLRC5 together with total somatic mutation number were significantly correlated with increased patient survival. These results suggest that NLRC5 tumor expression, alone or together with tumor mutation load constitutes a valuable predictive biomarker for both prognosis and response to anti-CTLA-4 and potentially anti-PD1 blockade immunotherapy in melanoma patients.

Comprehensive genomic profiling of high-grade serous ovarian carcinoma from Chinese patients identifies co-occurring mutations in the Ras/Raf pathway with TP53.

High-grade serous ovarian carcinoma (HGSOC) is a major form of ovarian epithelial tumor that is often diagnosed only at an advanced stage when it is already highly aggressive. We performed comprehensive genomic profiling using an analytically validated clinical next-generation sequencing assay to identify genomic alterations in 450 cancer-related genes in a cohort of 88 Chinese HGSOC patients. Overall, we detected 547 genomic alterations with an average of 6.2 alterations per tumor. Most of these HGSOC tumors had low tumor mutation burden and were microsatellite stable. Consistent with earlier studies, TP53 mutations were present in the majority (96.6%) of the tumors studied, and mutations in BRCA1/2 that affect DNA repair were also detected frequently in 20.5% of the tumors. However, we observed a 10.2% of mutated genes in the Ras/Raf pathway, all co-occurring with TP53 mutations in the same tumor, which was unrecognized previously. Our results show that in HGSOC patients, there may be an unrecognized co-occurrence of TP53 mutations with mutations in Ras/Raf pathway.

Degradation of chlorpyrifos alone and in combination with chlorothalonil and their effects on soil microbial populations.

In practice, pesticides are usually applied simultaneously or one after another for crop protection, and this type of pesticide application often leads to a combined contamination of pesticide residues in the soil environment. A laboratory study was conducted to investigate the influence of chlorothalonil on chlorpyrifos degradation and its effects on soil bacterial, fungal, and actinomycete populations. Under the experimental conditions here, the half-lives of chlorpyrifos alone, and in combination with chlorothalonil, at the recommended and double dosages, were measured to be 3.24, 2.77, and 2.63 d, respectively. Chlorpyrifos degradation was not significantly altered by its combination with chlorothalonil. However, the inhibitory effect of chlorpyrifos on soil microorganisms was increased by its combination with chlorothalonil, and the increase was related to the levels of chlorothalonil added. Compared to those in the controls, the populations of bacteria, fungi, and actinomycetes were significantly reduced by 44.1%, 61.1%, and 72.8%, respectively, on the first day after treatment (DAT) by chlorpyrifos alone. With the addition of chlorothalonil, the inhibition was increased to 55.2%, 79.3%, and 85.8% at the recommended dosage, and 86.0%, 94.1%, and 90.8% at the double dosage, at one DAT, respectively. The results suggested that combined effects should be taken into account to assess the actual impacts of pesticide applications.

Fine population structure analysis method for genomes of many.

Fine population structure can be examined through the clustering of individuals into subpopulations. The clustering of individuals in large sequence datasets into subpopulations makes the calculation of subpopulation specific allele frequency possible, which may shed light on selection of candidate variants for rare diseases. However, as the magnitude of the data increases, computational burden becomes a challenge in fine population structure analysis. To address this issue, we propose fine population structure analysis (FIPSA), which is an individual-based non-parametric method for dissecting fine population structure. FIPSA maximizes the likelihood ratio of the contingency table of the allele counts multiplied by the group. We demonstrated that its speed and accuracy were superior to existing non-parametric methods when the simulated sample size was up to 5,000 individuals. When applied to real data, the method showed high resolution on the Human Genome Diversity Project (HGDP) East Asian dataset. FIPSA was independently validated on 11,257 human genomes. The group assignment given by FIPSA was 99.1% similar to those assigned based on supervised learning. Thus, FIPSA provides high resolution and is compatible with a real dataset of more than ten thousand individuals.

A missense mutation in TCN2 is associated with decreased risk for congenital heart defects and may increase cellular uptake of vitamin B12 via Megalin.

Deregulation of folate and vitamin B12 (VB12) metabolism contributes to the risk of congenital heart defects (CHDs). Transcobalamin (TCN2) is essential for transporting VB12 from blood to cells as TCN2-bound VB12 (holo-TC) is the only form for somatic cellular uptake. In this study, we performed an association study between common polymorphisms in 46 one carbon metabolism genes and CHD in 412 CHDs and 213 controls. Only two significant association signals in coding regions were identified: FTCD c.1470C>T & TCN2 c.230A>T. The only missense mutation, TCN2 c.230A>T, was further validated in 412 CHDs and 1177 controls. TCN2 c.230T is significantly associated with reduced CHD risk in North Chinese (odds ratio = 0.67, P = 4.62e-05), compared with the 230A allele. Interestingly, the mean level of plasma holo-TC in women with the TA genotype was 1.77-fold higher than that in women with the AA genotype. Further analysis suggested that c.230A>T enhanced the cellular uptake of holo-TC via the LRP2 receptor. Our results determined that a functional polymorphism in TCN2 contributes to the prevalence of CHDs. TCN2 c.230A>T is significantly associated with a reduced CHD risk, likely due to TCN2 c.230T improving the interaction between holo-TC and its LRP2 receptor.

Association study of single nucleotide polymorphisms in mature microRNAs and the risk of thyroid tumor in a Chinese population.

MicroRNAs (miRNAs) are small non-coding RNA molecules that are involved in a variety of cellular functions. Single nucleotide polymorphisms (SNPs) have been identified in mature miRNAs (mmSNPs), some of which have been linked to cancer risk; however, it is unclear which mmSNPs contribute to the susceptibility to thyroid tumors. In the present study, we examined the influence of selected mmSNPs on the risk of thyroid tumor. After systematic in silico screening, seventeen mmSNPs were identified and genotyped in a Chinese population including 828 patients with papillary thyroid cancer (PTC), 488 patients with benign thyroid tumor (BN), and 1038 cancer-free controls. Multivariate logistic regression analyses were conducted to evaluate the association of SNP genotypes and alleles with the risk of developing PTC and BN. Three SNPs (rs67106263 in mir-3144, GA versus GG, OR = 1.35, 1.09-1.68; rs4919510 in mir-608, CC versus GG/GC, OR = 0.76, 0.60-0.97; and rs79402775 in mir-933, AA versus GG/GA, OR = 1.76, 1.00-3.12) were associated with PTC risk. A combined effect of unfavorable genotypes was observed to give increased PTC risk in a dose-dependent manner. In addition, three SNPs (rs10061133 in mir-449b, rs79402775 in mir-933 and rs4919510 in mir-608) showed at least borderline correlations with the risk of BN. False-positive report probability was assessed for significant findings. The rs67106263 SNP was associated with the expression level of mir-3144 in thyroid tissue. These results indicate that mmSNPs may contribute to genetic susceptibility to thyroid tumors. Large validation and functional studies are required to further explore the role of mmSNPs in carcinogenesis.

Y chromosomes of 40% Chinese descend from three Neolithic super-grandfathers.

Demographic change of human populations is one of the central questions for delving into the past of human beings. To identify major population expansions related to male lineages, we sequenced 78 East Asian Y chromosomes at 3.9 Mbp of the non-recombining region, discovered >4,000 new SNPs, and identified many new clades. The relative divergence dates can be estimated much more precisely using a molecular clock. We found that all the Paleolithic divergences were binary; however, three strong star-like Neolithic expansions at ∼6 kya (thousand years ago) (assuming a constant substitution rate of 1×10(-9)/bp/year) indicates that ∼40% of modern Chinese are patrilineal descendants of only three super-grandfathers at that time. This observation suggests that the main patrilineal expansion in China occurred in the Neolithic Era and might be related to the development of agriculture.

Changes in attitudes, knowledge and behavior associated with implementing a comprehensive school health program in a province of China.

After successful pilot projects, Zhejiang Province, China, decided to systematically scale-up health promoting schools (HPS) over the entire province of 47 million. This study describes the interventions and self-reported changes in attitudes, knowledge and behavior during the first phase of scaling-up. Group interviews were conducted with a sample of 191 participants (school administrators, teachers, students and parents) from nine schools with a total of approximately 15 200 students. Grounded theory guided data analysis. Schools implemented all HPS components (school health policy, physical school environment, psychosocial school environment, health education, health services, nutrition services, counseling/mental health, physical exercise, health promotion for staff and outreach to families and communities), adapted to local circumstances. Participants reported a range of changes in attitudes (paying more attention to health, attaining better 'psychological quality' and confidence, forming friendships between teachers and students and feeling more relaxed), knowledge and concepts (increasing knowledge about various health issues, developing a broader concept of health and gaining better understanding about the HPS concept) and behavior (actively participating, increasing physical activity, improving sanitary habits, reducing or quitting smoking, eating more nutritiously, increasing safety behavior, sustaining less injuries and improving parent-child communication). This qualitative study shows the feasibility and efficacy of implementing HPS in Zhejiang Province, China.