The transcriptional architecture of early human hematopoiesis identifies multilevel control of lymphoid commitment.

Understanding how differentiation programs originate from the gene-expression 'landscape' of hematopoietic stem cells (HSCs) is crucial for the development of new clinical therapies. We mapped the transcriptional dynamics underlying the first steps of commitment by tracking transcriptome changes in human HSCs and eight early progenitor populations. We found that transcriptional programs were extensively shared, extended across lineage-potential boundaries and were not strictly lineage affiliated. Elements of stem, lymphoid and myeloid programs were retained in multilymphoid progenitors (MLPs), which reflected a hybrid transcriptional state. By functional single cell analysis, we found that the transcription factors Bcl-11A, Sox4 and TEAD1 (TEF1) governed transcriptional networks in MLPs, which led to B cell specification. Overall, we found that integrated transcriptome approaches can be used to identify previously unknown regulators of multipotency and show additional complexity in lymphoid commitment.

Genome-wide methylation profiles reveal quantitative views of human aging rates.

The ability to measure human aging from molecular profiles has practical implications in many fields, including disease prevention and treatment, forensics, and extension of life. Although chronological age has been linked to changes in DNA methylation, the methylome has not yet been used to measure and compare human aging rates. Here, we build a quantitative model of aging using measurements at more than 450,000 CpG markers from the whole blood of 656 human individuals, aged 19 to 101. This model measures the rate at which an individual's methylome ages, which we show is impacted by gender and genetic variants. We also show that differences in aging rates help explain epigenetic drift and are reflected in the transcriptome. Moreover, we show how our aging model is upheld in other human tissues and reveals an advanced aging rate in tumor tissue. Our model highlights specific components of the aging process and provides a quantitative readout for studying the role of methylation in age-related disease.

Identifying potential DNA methylation markers in early-stage colorectal Cancer.

Colorectal cancer (CRC) is the second leading malignancy worldwide. Accurate screening is pivotal to early CRC detection, yet current screening modality involves invasive colonoscopy while non-invasive FIT tests have limited sensitivity. We applied a DNA methylation assay to identify biomarkers for early-stage CRC detection, risk stratification and precancerous lesion screening at tissue level. A model of biomarkers SFMBT2, ITGA4, THBD and ZNF304 showed 96.1% sensitivity and 87.0% specificity in CRC detection, with 100.0% sensitivity for advanced precancerous lesion and stage I CRC. Performances were further validated with TCGA data set, which showed a consistent AUC of 0.99 and exhibited specificity against other cancer types. KCNJ12, VAV3-AS1 and EVC were further identified for stage stratification (stage 0-I versus stage II-IV), with AUC of 0.87, 83.0% sensitivity and 71.2% specificity. Additionally, dual markers of NEUROD1 and FAM72C showed 83.2% sensitivity and 77.4% specificity in differing non-advanced precancerous lesions from inflammatory bowel diseases.

COL18A1 is a candidate eye iridocorneal angle-closure gene in humans.

Primary angle-closure glaucoma (PACG) is a common form of glaucoma in the Far East. Its defining feature is iridocorneal angle closure. In addition to PACG, indications of angle closure are included in the diagnostic criteria of related conditions primary angle-closure suspect (PACS) and primary angle closure (PAC). To the best of our knowledge, a causative gene for iridocorneal angle closure in humans has not been identified. This study aimed to identify the genetic cause of iridocorneal angle closure in a pedigree with at least 10 individuals diagnosed with PACS, PAC or PACG. Results of linkage analysis, segregation analysis of 44 novel variations, whole exome sequencing of 10 individuals, screenings of controls and bioinformatics predictions identified a mutation in COL18A1 that encodes collagen type XVIII as the most likely cause of angle closure in the pedigree. The role of COL18A1 in the etiology of Knobloch syndrome (KS) that is consistently accompanied by optic anomalies, available functional data on the encoded protein and the recognized role of collagens and the extracellular matrix in glaucoma pathogenesis supported the proposed role of the COL18A1 mutation in the pedigree. Subsequent identification of other COL18A1 mutations in PACS affected individuals of two unrelated families further supported that COL18A1 may affect angle closure. These PACS individuals were parents and grandparents of KS-affected children. In conclusion, a gene that affects angle closure in humans, a critical feature of PACG, has been identified. The findings also reinforce the importance of collagens in eye features and functions.

DNA methylation profiles in African American prostate cancer patients in relation to disease progression.

This study examined whether differential DNA methylation is associated with clinical features of more aggressive disease at diagnosis and prostate cancer recurrence in African American men, who are more likely to die from prostate cancer than other populations. Tumor tissues from 76 African Americans diagnosed with prostate cancer who had radical prostatectomy as their primary treatment were profiled for epigenome-wide DNA methylation levels. Long-term follow-up identified 19 patients with prostate cancer recurrence. Twenty-three CpGs were differentially methylated (FDR q≤0.25, mean methylation difference≥0.10) in patients with vs. without recurrence, including CpGs in GCK, CDKL2, PRDM13, and ZFR2. Methylation differences were also observed between men with metastatic-lethal prostate cancer vs. no recurrence (five CpGs), regional vs. local pathological stage (two CpGs), and higher vs. lower tumor aggressiveness (one CpG). These results indicate that differentially methylated CpG sites identified in tumor tissues of African American men may contribute to prostate cancer aggressiveness.

A four-gene transcript score to predict metastatic-lethal progression in men treated for localized prostate cancer: Development and validation studies.

BACKGROUND: Molecular studies have tried to address the unmet need for prognostic biomarkers in prostate cancer (PCa). Some gene expression tests improve upon clinical factors for prediction of outcomes, but additional tools for accurate prediction of tumor aggressiveness are needed. METHODS: Based on a previously published panel of 23 gene transcripts that distinguished patients with metastatic progression, we constructed a prediction model using independent training and testing datasets. Using the validated messenger RNAs and Gleason score (GS), we performed model selection in the training set to define a final locked model to classify patients who developed metastatic-lethal events from those who remained recurrence-free. In an independent testing dataset, we compared our locked model to established clinical prognostic factors and utilized Kaplan-Meier curves and receiver operating characteristic analyses to evaluate the model's performance. RESULTS: Thirteen of 23 previously identified gene transcripts that stratified patients with aggressive PCa were validated in the training dataset. These biomarkers plus GS were used to develop a four-gene (CST2, FBLN1, TNFRSF19, and ZNF704) transcript (4GT) score that was significantly higher in patients who progressed to metastatic-lethal events compared to those without recurrence in the testing dataset (P = 5.7 × 10-11 ). The 4GT score provided higher prediction accuracy (area under the ROC curve [AUC] = 0.76; 95% confidence interval [CI] = 0.69-0.83; partial area under the ROC curve [pAUC] = 0.008) than GS alone (AUC = 0.63; 95% CI = 0.56-0.70; pAUC = 0.002), and it improved risk stratification in subgroups defined by a combination of clinicopathological features (ie, Cancer of the Prostate Risk Assessment-Surgery). CONCLUSION: Our validated 4GT score has prognostic value for metastatic-lethal progression in men treated for localized PCa and warrants further evaluation for its clinical utility.

Characterizing transcriptional heterogeneity through pathway and gene set overdispersion analysis.

The transcriptional state of a cell reflects a variety of biological factors, from cell-type-specific features to transient processes such as the cell cycle, all of which may be of interest. However, identifying such aspects from noisy single-cell RNA-seq data remains challenging. We developed pathway and gene set overdispersion analysis (PAGODA) to resolve multiple, potentially overlapping aspects of transcriptional heterogeneity by testing gene sets for coordinated variability among measured cells.

Accessing Genetic Information with Liquid Biopsies.

Recent scientific advances in understanding circulating tumor cells, cell-free DNA/RNA, and exosomes in blood have laid a solid foundation for the development of routine molecular 'liquid biopsies'. This approach provides non-invasive access to genetic information--somatic mutations, epigenetic changes, and differential expression--about the physiological conditions of our body and diseases. It opens a valuable avenue for future genetic studies and human disease diagnosis, including prenatal and neurodegenerative disease diagnosis, as well as for cancer screening and monitoring. With the rapid development of highly sensitive and accurate technologies such as next-generation sequencing, molecular 'liquid biopsies' will quickly become a central piece in the future of precision medicine.

A five-CpG DNA methylation score to predict metastatic-lethal outcomes in men treated with radical prostatectomy for localized prostate cancer.

BACKGROUND: Prognostic biomarkers for localized prostate cancer (PCa) could improve personalized medicine. Our group previously identified a panel of differentially methylated CpGs in primary tumor tissue that predict disease aggressiveness, and here we further validate these biomarkers. METHODS: Pyrosequencing was used to assess CpG methylation of eight biomarkers previously identified using the HumanMethylation450 array; CpGs with strongly correlated (r >0.70) results were considered technically validated. Logistic regression incorporating the validated CpGs and Gleason sum was used to define and lock a final model to stratify men with metastatic-lethal versus non-recurrent PCa in a training dataset. Coefficients from the final model were then used to construct a DNA methylation score, which was evaluated by logistic regression and Receiver Operating Characteristic (ROC) curve analyses in an independent testing dataset. RESULTS: Five CpGs were technically validated and all were retained (P < 0.05) in the final model. The 5-CpG and Gleason sum coefficients were used to calculate a methylation score, which was higher in men with metastatic-lethal progression (P = 6.8 × 10-6 ) in the testing dataset. For each unit increase in the score there was a four-fold increase in risk of metastatic-lethal events (odds ratio, OR = 4.0, 95%CI = 1.8-14.3). At 95% specificity, sensitivity was 74% for the score compared to 53% for Gleason sum alone. The score demonstrated better prediction performance (AUC = 0.91; pAUC = 0.037) compared to Gleason sum alone (AUC = 0.87; pAUC = 0.025). CONCLUSIONS: The DNA methylation score improved upon Gleason sum for predicting metastatic-lethal progression and holds promise for risk stratification of men with aggressive tumors. This prognostic score warrants further evaluation as a tool for improving patient outcomes.

TP53 mutations, tetraploidy and homologous recombination repair defects in early stage high-grade serous ovarian cancer.

To determine early somatic changes in high-grade serous ovarian cancer (HGSOC), we performed whole genome sequencing on a rare collection of 16 low stage HGSOCs. The majority showed extensive structural alterations (one had an ultramutated profile), exhibited high levels of p53 immunoreactivity, and harboured a TP53 mutation, deletion or inactivation. BRCA1 and BRCA2 mutations were observed in two tumors, with nine showing evidence of a homologous recombination (HR) defect. Combined Analysis with The Cancer Genome Atlas (TCGA) indicated that low and late stage HGSOCs have similar mutation and copy number profiles. We also found evidence that deleterious TP53 mutations are the earliest events, followed by deletions or loss of heterozygosity (LOH) of chromosomes carrying TP53, BRCA1 or BRCA2. Inactivation of HR appears to be an early event, as 62.5% of tumours showed a LOH pattern suggestive of HR defects. Three tumours with the highest ploidy had little genome-wide LOH, yet one of these had a homozygous somatic frame-shift BRCA2 mutation, suggesting that some carcinomas begin as tetraploid then descend into diploidy accompanied by genome-wide LOH. Lastly, we found evidence that structural variants (SV) cluster in HGSOC, but are absent in one ultramutated tumor, providing insights into the pathogenesis of low stage HGSOC.