Circulating tumor cell enumeration for improved screening and disease detection of patients with colorectal cancer.

BACKGROUND: The immunochemical fecal occult blood test (iFOBT) for colorectal cancer (CRC) screening and the serum carcinoembryonic antigen (CEA) assay for disease detection of CRC is associated with a high false-positive rate and a low detection sensitivity, respectively. There is an unmet need to define additional modalities to complement these assays. Different subsets of circulating tumor cells (CTCs) are present in the peripheral blood of cancer patients. Whether or not CTCs testing supplements these clinical assays and is valuable for patients with CRC was investigated. METHODS: CTCs were enriched from pre-operative patients with CRC (n = 109) and the non-cancerous controls (n = 65). CTCs expressing either epithelial cell adhesion molecule (EpCAM) or podoplanin (PDPN, the marker associated with poor cancer prognosis) were defined by immunofluorescence staining and were analyzed alone or in combination with iFOBT or serum CEA. RESULTS: Patients with early or advanced stage of CRC can be clearly identified and differentiated from the non-cancerous controls (p < 0.001) by EpCAM+-CTC or PDPN+-CTC count. The sensitivity and specificity of EpCAM+-CTCs was 85.3% and 78.5%, respectively, when the cutoff value was 23 EpCAM+-CTCs/mL of blood; and the sensitivity and specificity of PDPN+-CTCs was 78.0% and 75.4%, respectively, when the cutoff value was 7 PDPN+-CTCs/mL of blood. Combined analysis of iFOBT with the EpCAM+-CTC and PDPN+-CTC count reduced the false-positive rate of iFOBT from 56.3% to 18.8% and 23.4%, respectively. Combined analysis of serum CEA with the EpCAM+-CTC and PDPN+-CTC count increased the disease detection rate from 30.3% to 89.9% and 86.2%, respectively. CONCLUSION: CTC testing could supplement iFOBT to improve CRC screening and supplement serum CEA assay for better disease detection of patients with CRC.

Mental Health Disorders and Associated Risk Factors in Quarantined Adults During the COVID-19 Outbreak in China: Cross-Sectional Study.

BACKGROUND: People undergoing mass home- and community-based quarantine are vulnerable to mental health disorders during outbreaks of coronavirus disease (COVID-19), but few studies have evaluated the associated psychosocial factors. OBJECTIVE: This study aimed to estimate the prevalence of anxiety and depressive symptoms and identify associated demographic and psychosocial factors in the general Chinese population during the COVID-19 pandemic quarantine period. METHODS: Participants aged 18 years or above were recruited in a cross-sectional online survey using snowball sampling from February 26-29, 2020. The survey included questions on demographics, family relationships, chronic diseases, quarantine conditions, lifestyle, COVID-19 infection, and anxiety and depressive symptoms. Logistic regression analyses were conducted to identify factors associated with elevated anxiety or depressive symptoms. RESULTS: Out of 2331 participants, 762 (32.7%) experienced elevated anxiety or depressive symptoms. Nine risk factors associated with anxiety or depressive symptoms included younger age, reduced income, having cancer or other chronic diseases, having family members living with cancer, concerns related to COVID-19 infection for themselves or family members, living alone, having family conflicts, having <3 or >8 hours of sedentary time per day, and worsened sleep quality. CONCLUSIONS: The findings highlight an urgent need for psychological support for populations at high risk for elevated anxiety or depressive symptoms during the COVID-19 pandemic.

Comprehensive transcriptome profiling of Taiwanese colorectal cancer implicates an ethnic basis for pathogenesis.

Colorectal cancer (CRC) is one of the most commonly diagnosed cancers worldwide. While both genetic and environmental factors have been linked to the incidence and mortality associated with CRC, an ethnic aspect of its etiology has also emerged. Since previous large-scale cancer genomics studies are mostly based on samples of European ancestry, the patterns of clinical events and associated mechanisms in other minority ethnic patients suffering from CRC are largely unexplored. We collected 104 paired and adjacent normal tissue and CRC tumor samples from Taiwanese patients and employed an integrated approach - paired expression profiles of mRNAs and microRNAs (miRNAs) combined with transcriptome-wide network analyses - to catalog the molecular signatures of this regional cohort. On the basis of this dataset, which is the largest ever reported for this type of systems analysis, we made the following key discoveries: (1) In comparison to the The Cancer Genome Atlas (TCGA) data, the Taiwanese CRC tumors show similar perturbations in expressed genes but a distinct enrichment in metastasis-associated pathways. (2) Recurrent as well as novel CRC-associated gene fusions were identified based on the sequencing data. (3) Cancer subtype classification using existing tools reveals a comparable distribution of tumor subtypes between Taiwanese cohort and TCGA datasets; however, this similarity in molecular attributes did not translate into the predicted subtype-related clinical outcomes (i.e., death event). (4) To further elucidate the molecular basis of CRC prognosis, we developed a new stratification strategy based on miRNA-mRNA-associated subtyping (MMAS) and consequently showed that repressed WNT signaling activity is associated with poor prognosis in Taiwanese CRC. In summary, our findings of distinct, hitherto unreported biosignatures underscore the heterogeneity of CRC tumorigenesis, support our hypothesis of an ethnic basis of disease, and provide prospects for translational medicine.

Targeted sequencing of cancer-related genes in nasopharyngeal carcinoma identifies mutations in the TGF-ß pathway.

Approximately, 25% of nasopharyngeal carcinoma (NPC) patients develop recurrent disease. NPC may involve relatively few genomic alterations compared to other cancers due to its association with Epstein-Barr virus (EBV). We envisioned that in-depth sequencing of tumor tissues might provide new insights into the genetic alterations of this cancer. Thirty-three NPC paired tumor/adjacent normal or peripheral blood mononuclear cell samples were deep-sequenced (>1000×) with respect to a panel of 409 cancer-related genes. Newly identified mutations and its correlation with clinical outcomes were evaluated. Profiling of somatic mutations and copy number variations (CNV) in NPC tumors identified alterations in RTK/RAS/PI3K, NOTCH, DNA repair, chromatin remodeling, cell cycle, NF-κB, and TGF-ß pathways. In addition, patients harbored CNV among 409 cancer-related genes and missense mutations in TGF-ß/SMAD signaling were associated with poor overall survival and poor recurrence-free survival, respectively. The CNV events were correlated with plasma EBV copies, while mutations in TGFBR2 and SMAD4 abrogate SMAD-dependent TGF-ß signaling. Functional analysis revealed that the new TGFBR2 kinase domain mutants were incapable of transducing the signal, leading to failure of phosphorylation of SMAD2/3 and activation of downstream TGF-ß-mediated cell growth arrest. This study provides evidence supporting CNV and dysregulated TGF-ß signaling contributes to exacerbating the NPC pathogenesis.

CoMutPlotter: a web tool for visual summary of mutations in cancer cohorts.

BACKGROUND: CoMut plot is widely used in cancer research publications as a visual summary of mutational landscapes in cancer cohorts. This summary plot can inspect gene mutation rate and sample mutation burden with their relevant clinical details, which is a common first step for analyzing the recurrence and co-occurrence of gene mutations across samples. The cBioPortal and iCoMut are two web-based tools that allow users to create intricate visualizations from pre-loaded TCGA and ICGC data. For custom data analysis, only limited command-line packages are available now, making the production of CoMut plots difficult to achieve, especially for researchers without advanced bioinformatics skills. To address the needs for custom data and TCGA/ICGC data comparison, we have created CoMutPlotter, a web-based tool for the production of publication-quality graphs in an easy-of-use and automatic manner. RESULTS: We introduce a web-based tool named CoMutPlotter to lower the barriers between complex cancer genomic data and researchers, providing intuitive access to mutational profiles from TCGA/ICGC projects as well as custom cohort studies. A wide variety of file formats are supported by CoMutPlotter to translate cancer mutation profiles into biological insights and clinical applications, which include Mutation Annotation Format (MAF), Tab-separated values (TSV) and Variant Call Format (VCF) files. CONCLUSIONS: In summary, CoMutPlotter is the first tool of its kind that supports VCF file, the most widely used file format, as its input material. CoMutPlotter also provides the most-wanted function for comparing mutation patterns between custom cohort and TCGA/ICGC project. Contributions of COSMIC mutational signatures in individual samples are also included in the summary plot, which is a unique feature of our tool. CoMutPlotter is freely available at http://tardis.cgu.edu.tw/comutplotter .

ADAR1 promotes robust hypoxia signaling via distinct regulation of multiple HIF-1α-inhibiting factors.

Adenosine deaminase acting on RNA (ADAR)-catalyzed adenosine-to-inosine RNA editing is potentially dysregulated in neoplastic progression. However, how this transcriptome recoding process is functionally correlated with tumorigenesis remains largely elusive. Our analyses of RNA editome datasets identify hypoxia-related genes as A-to-I editing targets. In particular, two negative regulators of HIF-1A-the natural antisense transcript HIF1A-AS2 and the ubiquitin ligase scaffold LIMD1-are directly but differentially modulated by ADAR1. We show that HIF1A-AS2 antagonizes the expression of HIF-1A in the immediate-early phase of hypoxic challenge, likely through a convergent transcription competition in cis ADAR1 in turn suppresses transcriptional progression of the antisense gene. In contrast, ADAR1 affects LIMD1 expression post-transcriptionally, by interfering with the cytoplasmic translocation of LIMD1 mRNA and thus protein translation. This multi-tier regulation coordinated by ADAR1 promotes robust and timely accumulation of HIF-1α upon oxygen depletion and reinforces target gene induction and downstream angiogenesis. Our results pinpoint ADAR1-HIF-1α axis as a hitherto unrecognized key regulator in hypoxia.

High Efficient Expression, Purification, and Functional Characterization of Native Human Epidermal Growth Factor in Escherichia coli.

Human epidermal growth factor (hEGF) is a small, mitotic growth polypeptide that promotes the proliferation of various cells and is widely applied in clinical practices. However, high efficient expression of native hEGF in Escherichia coli has not been successful, since three disulfide bonds in monomer hEGF made it unable to fold into correct 3D structure using in vivo system. To tackle this problem, we fused Mxe GyrA intein (Mxe) at the C-terminal of hEGF followed by small ubiquitin-related modifier (SUMO) and 10x His-tag to construct a chimeric protein hEGF-Mxe-SUMO-H10. The fusion protein was highly expressed at the concentration of 281 mg/L and up to 59.5% of the total cellular soluble proteins. The fusion protein was purified by affinity chromatography and 29.4 mg/L of native hEGF can be released by thiol induced N-terminal cleavage without any proteases. The mitotic activity in Balb/c 3T3 cells is proliferated by commercial and recombinant hEGF measured with methylthiazolyldiphenyl-tetrazolium bromide (MTT) assay which indicated that recombinant hEGF protein stimulates the cell proliferation similar to commercial protein. This study significantly improved the yield and reduced the cost of hEGF in the recombinant E. coli system and could be a better strategy to produce native hEGF for pharmaceutical development.

Estrogen enhances activity of Wnt signaling during osteogenesis by inducing Fhl1 expression.

Estrogen is a crucial hormone for osteoclast inhibition and for preventing osteoporosis. However, the hormone's role in osteoblast growth and differentiation remains unclear. The complexity of estrogen's role in guiding osteoblast behavior arises partly from crosstalk with other signaling pathways, including Wnt signaling. In this study, we show that the Wnt agonist, LiCl, induced Fhl1 gene expression, which substantially enhanced osteoblast differentiation. Staining with alizarin red revealed that MC3T3-E1 mineralization was enhanced by overexpression of Fhl1. In addition, Fhl1 promoted the expression of the osteogenic markers, Runt-related transcription factor 2 (Runx2), osteocalcin (OCN), and osteopontin (OPN), whereas MC3T3-E1 cells with gene knockdown of Fhl1 exhibited limited mineralization and expression of Runx2, OCN, and OPN. We further demonstrate evidences from quantitative reverse transcription real-time polymerase chain reaction and reporter assay that Fhl1 expression was synergistically stimulated by estrogen (E2) and LiCl, but reduced by the estrogen-receptor inhibitor fulvestrant (ICI 182,780). However, estrogen could not enhance osteogenesis while Fhl1 expression was knocked down. Because estrogen and Wnt signaling frequently interact in developmental processes, we propose that Fhl1 can be an acting molecule mediating both signaling pathways during osteogenesis.