Prediction of cell cycle distribution after drug exposure by high content imaging analysis using low-toxic DNA staining dye.

Interference in cell cycle progression has been noted as one of the important properties of anticancer drugs. In this study, we developed the cell cycle prediction model using high-content imaging data of recipient cells after drug exposure and DNA-staining with a low-toxic DNA dye, SiR-DNA. For this purpose, we exploited HeLa and MCF7 cells introduced with a fluorescent ubiquitination-based cell cycle indicator (Fucci). Fucci-expressing cancer cells were subjected to high-content imaging analysis using OperettaCLS after 36-h exposure to anticancer drugs; the nuclei were segmented, and the morphological and intensity properties of each nucleus characterized by SiR-DNA staining were calculated using imaging analysis software, Harmony. For the use of training, we classified cells into each phase of the cell cycle using the Fucci system. Training data (n = 7500) and validation data (n = 2500) were randomly sampled and the binary classification prediction models for G1, early S, and S/G2/M phases of the cell cycle were developed using four supervised machine learning algorithms. We selected random forest as the model with the best performance through 10-fold cross-validation; the accuracy rate was approximately 75%-87%. Regarding feature importance, variables expected to be biologically related to the cell cycle, for example, signal intensity and nuclear size, were highly ranked, suggesting the validity of the model. These results showed that the cell cycle can be predicted in cancer cells by simply exploiting the current prediction model using fluorescent images of DNA-staining dye, and the model could be applied for the use of future ex vivo drug sensitivity diagnosis.

Induction of stearoyl-CoA desaturase confers cell density-dependent ferroptosis resistance in melanoma.

Ferroptosis is a form of regulated cell death that is induced by inhibiting glutathione peroxidase 4 (GPX4), which eliminates lipid peroxidation. Ferroptosis induction is influenced by the cell environment. However, the cellular states altering ferroptosis susceptibility remain largely unknown. We found that melanoma cell lines became resistant to ferroptosis as cell density increased. Comparative transcriptome and metabolome analyses revealed that cell density-dependent ferroptosis resistance was coupled with a shift toward a lipogenic phenotype accompanied by strong induction of stearoyl-CoA desaturase (SCD). Database analysis of gene dependency across hundreds of cancer cell lines uncovered a negative correlation between GPX4 and SCD dependency. Importantly, SCD inhibition, either pharmacologically or through genetic knockout, sensitized melanoma cells to GPX4 inhibition, thereby attenuating ferroptosis resistance in cells at high density. Our findings indicate that transition to an SCD-inducing, lipogenic cell state produces density-dependent resistance to ferroptosis, which may provide a therapeutic strategy against melanoma.

Functional evaluation of BRCA1/2 variants of unknown significance with homologous recombination assay and integrative in silico prediction model.

Numerous variants of unknown significance (VUSs) exist in hereditary breast and ovarian cancers. Although multiple methods have been developed to assess the significance of BRCA1/2 variants, functional discrepancies among these approaches remain. Therefore, a comprehensive functional evaluation system for these variants should be established. We performed conventional homologous recombination (HR) assays for 50 BRCA1 and 108 BRCA2 VUSs and complementarily predicted VUSs using a statistical logistic regression prediction model that integrated six in silico functional prediction tools. BRCA1/2 VUSs were classified according to the results of the integrative in vitro and in silico analyses. Using HR assays, we identified 10 BRCA1 and 4 BRCA2 VUSs as low-functional pathogenic variants. For in silico prediction, the statistical prediction model showed high accuracy for both BRCA1 and BRCA2 compared with each in silico prediction tool individually and predicted nine BRCA1 and seven BRCA2 variants to be pathogenic. Integrative functional evaluation in this study and the American College of Medical Genetics and Genomics and the Association for Molecular Pathology (ACMG/AMP) guidelines strongly suggested that seven BRCA1 variants (p.Glu272Gly, p.Lys1095Glu, p.Val1653Leu, p.Thr1681Pro, p.Phe1761Val, p.Thr1773Ile, and p.Gly1803Ser) and four BRCA2 variants (p.Trp31Gly, p.Ser2616Phe, p.Tyr2660Cys, and p.Leu2792Arg) were pathogenic. This study demonstrates that integrative evaluation using conventional HR assays and optimized in silico prediction comprehensively classified the significance of BRCA VUSs for future clinical applications.

Polymorphisms of Placental Iodothyronine Deiodinase Genes in a Rural Area of Northern China with High Prevalence of Neural Tube Defects.

INTRODUCTION: We have reported that high total homocysteine and the coexistence of inadequate thyroid hormones in maternal serum increase the risk of fetal neural tube defects (NTDs). Placental iodothyronine deiodinases (DIOs: DIO1, DIO2, and DIO3) play a role in regulating the conversions between different forms of maternal thyroid hormones. This study hypothesized that single nucleotide polymorphisms (SNPs) in placental DIOs genes could be related to NTDs. METHODS: We performed a case-control study from 2007 to 2009 that included pregnant women from Lüliang, Shanxi Province, China. Nine distinct SNPs in DIOs genes were analyzed, and placental samples were obtained from 83 pregnant women with NTD fetuses and 90 pregnant women with normal fetuses. The nine SNPs were analyzed using the Cochran-Armitage test and the Fisher's exact test. RESULTS: There were no statistically significant differences between case and control in the nine SNPs of DIOs (p > 0.05). CONCLUSIONS: The results of this study suggested that SNPs of DIO genes in the placenta among pregnant women have no statistically significant difference between the two groups, suggesting that other factors might be involved in metabolism of maternal thyroid hormone provided to fetuses, such as epigenetic modification of methylation and homocysteinylation and genomic imprinting in the placenta. Further functional studies on placenta samples are necessary.

Prognostic implications of prostaglandin E-major urinary metabolite in resected non-small-cell lung cancer.

OBJECTIVES: Cyclooxygenase-2-derived prostaglandin E2 (PGE2) is highly involved in the promotion of cancer progression. The end product of this pathway, PGE-major urinary metabolite (PGE-MUM), is a stable metabolite of PGE2 that can be assessed non-invasively and repeatedly in urine samples. The aim of this study was to assess the dynamic changes in perioperative PGE-MUM levels and their prognostic significance in non-small-cell lung cancer (NSCLC). METHODS: Between December 2012 and March 2017, 211 patients who underwent complete resection for NSCLC were analysed prospectively. PGE-MUM levels in 2 spot urine samples taken 1 or 2 days preoperatively and 3-6 weeks postoperatively were measured using a radioimmunoassay kit. RESULTS: Elevated preoperative PGE-MUM levels were associated with tumour size, pleural invasion and advanced stage. Multivariable analysis revealed that age, pleural invasion, lymph node metastasis and postoperative PGE-MUM levels were independent prognostic factors. In matched pre- and postoperative urine samples obtained from patients who are eligible for adjuvant chemotherapy, an increase in PGE-MUM levels following resection was an independent prognostic factor (hazard ratio 3.017, P = 0.005). Adjuvant chemotherapy improved survival in patients with increased PGE-MUM levels after resection (5-year overall survival, 79.0 vs 50.4%, P = 0.027), whereas survival benefit was not observed in those with decreased PGE-MUM levels (5-year overall survival, 82.1 vs 82.3%, P = 0.442). CONCLUSIONS: Increased preoperative PGE-MUM levels can reflect tumour progression and postoperative PGE-MUM levels are a promising biomarker for survival after complete resection in patients with NSCLC. Perioperative changes in PGE-MUM levels may aid in determining the optimal eligibility for adjuvant chemotherapy.

Risk of low birthweight associated with the timing and frequency of antenatal care visits in Lao PDR: a retrospective cohort study.

BACKGROUND: Antenatal care (ANC) plays an important role in preventing low birthweight (LBW). Whereas the government of Lao People's Democratic Republic (Lao PDR) has committed to increasing the usage of ANC, little attention has been given to the early initiation of ANC. The present study assessed the influence of delayed and fewer ANC visits on LBW in the country. METHODS: This is a retrospective cohort study conducted at Salavan Provincial Hospital. Study participants were all pregnant women who gave birth at the hospital between 1 August 2016 and 31 July 31 2017. Data were collected from medical records. Logistic regression analyses were performed to quantify the relationship between ANC visits and LBW. We also investigated factors associated with inadequate ANC visits: first ANC visit after the first trimester or < 4 ANC visits. RESULTS: The mean birth weight was 2808.7 g [standard deviation: SD 455.6]. Among 1804 participants, 350 (19.4%) had babies with LBW, and 147 (8.2%) had inadequate ANC visits. In multivariate analyses, compared to participants with adequate ANC visits, those with ≥ 4 ANC visits and the first ANC visit after the second trimester, those with < 4 ANC visits, and those with no ANC visits had higher odds ratios (ORs) of LBW: 3.77 (95% confidence interval: CI = 1.66-8.57), 2.39 (95% CI = 1.18-4.83) and 2.22 (95% CI = 1.08-4.56), respectively. Younger maternal age (OR 1.42; 95% CI = 1.07-1.89), government subsidisation (OR 2.69; 95% CI = 1.97-3.68) and ethnic minority (OR 1.88; 95% CI = 1.50-2.34) were associated with increased risk of insufficient number of ANC visits after adjusting for covariates. CONCLUSIONS: Frequent and early initiation of ANC was associated with a reduction in LBW in Lao PDR. Encouraging childbearing-aged women to receive sufficient ANC at proper timing may lead to a reduction in LBW and improvement in short- and long-term health outcomes of neonates. Special attention will be needed for ethnic minorities and women in lower socioeconomic classes.

A prediction model for early systemic recurrence in breast cancer using a molecular diagnostic analysis of sentinel lymph nodes: A large-scale, multicenter cohort study.

BACKGROUND: The one-step nucleic acid amplification (OSNA) assay can quantify the cytokeratin 19 messenger RNA copy number as a proxy for sentinel lymph node (SN) metastasis in breast cancer. A large-scale, multicenter cohort study was performed to determine the prognostic value of the SN tumor burden based on a molecular readout and to establish a model for the prediction of early systemic recurrence in patients using the OSNA assay. METHODS: SN biopsies from 4757 patients with breast cancer were analyzed with the OSNA assay. The patients were randomly assigned to the training or validation cohort at a ratio of 2:1. On the basis of the training cohort, the threshold SN tumor burden value for stratifying distant recurrence was determined with Youden's index; predictors of distant recurrence were investigated via multivariable analyses. Based on the selected predictors, a model for estimating 5-year distant recurrence-free survival was constructed, and predictive performance was measured with the validation cohort. RESULTS: The prognostic cutoff value for the SN tumor burden was 1100 copies/µL. The following variables were significantly associated with distant recurrence and were used to construct the prediction model: SN tumor burden, age, pT classification, grade, progesterone receptor, adjuvant cytotoxic chemotherapy, and adjuvant anti-human epidermal growth factor receptor 2 therapy. The values for the area under the curve, sensitivity, specificity, and accuracy of the prediction model were 0.83, 63.4%, 81.7%, and 81.1%, respectively. CONCLUSIONS: Using the OSNA assay, the molecular readout-based SN tumor burden is an independent prognostic factor for early breast cancer. This model accurately predicts early systemic recurrence and may facilitate decision-making related to treatment.

Reforms of regulatory pathways for approval of new antineoplastic drugs in Japan from 2004 to 2019 and accompanying changes in pivotal clinical trial designs.

Background The Japanese Pharmaceuticals and Medical Devices Agency (PMDA) was established in 2004. Since then, various pieces of legislation, notices, and guidelines have been issued, and the regulatory approval pathways for domestic drugs have been diversified. However, the effects of these measures have not been fully examined. We examined the impact of these measures on the approval of antineoplastic drugs and the design of pivotal clinical trials for efficacy assessment by the PMDA. Methods We collected data on the antineoplastic drugs approved by the PMDA in fiscal years 2004-2019. We extracted the approval review pathways and the pivotal clinical trial designs from the PMDA review reports, and analyzed them to identify patterns. Results In total, 387 indications in oncology were approved by the PMDA in fiscal years 2004-2019, or 365 indications excluding multiple regulatory pathways. The number of approved indications generally increased year on year (p < 0.001). The largest number of approved indications was under the Orphan Drug Designation (31%, 114/365) and this continues to increase (p < 0.001). In the 288 indications for which clinical trial data were submitted for review, the pivotal clinical trial designs changed significantly (p < 0.001) after the guideline on clinical evaluation for antineoplastic drugs was revised in 2006. Conclusion The number of indications in oncology approved by the PMDA has been increasing over the past 16 years, alongside changes in regulatory pathways. The 2006 guideline on clinical evaluation had a particular impact on pivotal clinical trial designs.

Situation analyses of caregivers' knowledge on infant liquid formula in Japan: a cross-sectional questionnaire-based study.

BACKGROUND: Sales of the first domestic liquid infant milk formula in Japan started in March 2019, and some local governments have started to stockpile the formula in case of disasters. This study aimed to assess caregivers' knowledge of liquid formula to promote better utilization during disaster situations. METHODS: A cross-sectional, questionnaire-based study was conducted between August 26 and October 4, 2019, in Aiiku Hospital and Aiiku Clinic (Minato Ward, Tokyo, Japan). Caregivers of infants at their 1-, 3-4-, and 6-7-month medical check-ups participated. Caregivers' knowledge about the cup feeding method and handling of residual (left over) formula was evaluated. Caregivers were divided into high or low knowledge groups. Logistic regression analysis was performed to analyze the relation between caregivers' information sources on liquid formula and each caregiver's level of knowledge (high or low). RESULTS: Only 10.5% of caregivers were included in the high knowledge group regarding the cup feeding method, whereas 83.1% of caregivers were included in the high knowledge group regarding handling of residual liquid formula. Only 8.1% of caregivers were included in the high knowledge group regarding both cup feeding and handling of residual liquid formula. Logistic regression analysis showed that none of the information sources (hospital/clinic, TV/newspaper/magazine/website, milk company's website/advertisements, retail store, study meeting/event/social networking service, friend/infant's grandmother, workplace) affected the caregivers' knowledge. CONCLUSIONS: Caregivers' knowledge on the cup feeding method and handling of left over liquid formula was insufficient. Proactive and correct provision of information is required to improve caregivers' knowledge as this is an urgent infant safety issue in disaster situations.

Higher serum homocysteine and lower thyroid hormone levels in pregnant women are associated with neural tube defects.

BACKGROUND: This study tested the hypothesis that abnormal maternal metabolism of both homocysteine and thyroid hormone network in pregnant women is associated with neural tube defects (NTDs) in a part of China with high NTD prevalence. METHODS: A case-control study was performed between 2007 and 2009 in Lüliang Mountains, Shanxi Province. This study included 83 pregnant women who had fetuses with NTDs (cases) and 90 pregnant women with normal fetuses (controls). In addition, a cell model to illustrate the epidemiological findings was established. RESULTS: Fetuses of mother who had both high total homocysteine (tHcy) and inadequate free thyroxine were 3 times more at risk of developing NTDs (adjusted odds ratio = 3.5; 95 % confidence interval = 1.2-10.4; cases vs. controls) using multivariate logistic regression models. Furthermore, biological interaction between metabolisms of Hcy and thyroid hormones was demonstrated in vitro. In homocysteine thiolactone of a metabolite of Hcy-treated mouse embryonic neural stem NE4C cells, genes (Bmp7, Ctnnb1, Notch 1, Gli2, and Rxra) related to both neural tube closure and thyroid hormone network were shown to be regulated by H3K79 homocysteinylation, which increased their expression levels. CONCLUSIONS: The effect of maternal serum high tHcy on risk of developing NTDs is depended on maternal serum level of thyroxine. Meanwhile, a higher level of tHcy might also affect both maternal metabolism of thyroid hormone and neural tube closure in embryogenesis through homocysteinylation of histones.

Lamellarin 14, a derivative of marine alkaloids, inhibits the T790M/C797S mutant epidermal growth factor receptor.

The emergence of acquired resistance is a major concern associated with molecularly targeted kinase inhibitors. The C797S mutation in the epidermal growth factor receptor (EGFR) confers resistance to osimertinib, a third-generation EGFR-tyrosine kinase inhibitor (EGFR-TKI). We report that the derivatization of the marine alkaloid topoisomerase inhibitor lamellarin N provides a structurally new class of EGFR-TKIs. One of these, lamellarin 14, is effective against the C797S mutant EGFR. Bioinformatic analyses revealed that the derivatization transformed the topoisomerase inhibitor-like biological activity of lamellarin N into kinase inhibitor-like activity. Ba/F3 and PC-9 cells expressing the EGFR in-frame deletion within exon 19 (del ex19)/T790M/C797S triple-mutant were sensitive to lamellarin 14 in a dose range similar to the effective dose for cells expressing EGFR del ex19 or del ex19/T790M. Lamellarin 14 decreased the autophosphorylation of EGFR and the downstream signaling in the triple-mutant EGFR PC-9 cells. Furthermore, intraperitoneal administration of 10 mg/kg lamellarin 14 for 17 days suppressed tumor growth of the triple-mutant EGFR PC-9 cells in a mouse xenograft model using BALB/c nu/nu mice. Thus, lamellarin 14 serves as a novel structural backbone for an EGFR-TKI that prevents the development of cross-resistance against known drugs in this class.

Pathogenicity assessment of variants for breast cancer susceptibility genes based on BRCAness of tumor sample.

Genes involved in the homologous recombination repair pathway-as exemplified by BRCA1, BRCA2, PALB2, ATM, and CHEK2-are frequently associated with hereditary breast and ovarian cancer syndrome. Germline mutations in the loci of these genes with loss of heterozygosity or additional somatic truncation at the WT allele lead to the development of breast cancers with characteristic clinicopathological features and prominent genomic features of homologous recombination deficiency, otherwise referred to as "BRCAness." Although clinical genetic testing for these and other genes has increased the chances of identifying pathogenic variants, there has also been an increase in the prevalence of variants of uncertain significance, which poses a challenge to patient care because of the difficulties associated with making further clinical decisions. To overcome this challenge, we sought to develop a methodology to reclassify the pathogenicity of these unknown variants using statistical modeling of BRCAness. The model was developed with Lasso logistic regression by comparing 116 genomic attributes derived from 37 BRCA1/2 biallelic mutant and 32 homologous recombination-quiescent breast cancer exomes. The model showed 95.8% and 86.7% accuracies in the training cohort and The Cancer Genome Atlas validation cohort, respectively. Through application of the model for variant reclassification of homologous recombination-associated hereditary breast and ovarian cancer causal genes and further assessment with clinicopathological features, we finally identified one likely pathogenic and five likely benign variants. As such, the BRCAness model developed from the tumor exome was robust and provided a reasonable basis for variant reclassification.

Meta-analyses on progression-free survival as a surrogate endpoint for overall survival in triple-negative breast cancer.

PURPOSE: Earlier studies suggest progression-free survival (PFS) may be used as a surrogate endpoint for overall survival (OS) in metastatic breast cancer, which could shorten follow-up duration and speed up assessment of treatment effects. However, to our knowledge, the association between them is still unclear in advanced or metastatic triple-negative breast cancer (TNBC). METHODS: A literature-based meta-analysis followed by correlation analysis was conducted in advanced or metastatic TNBC. Weighted multiple regression analysis was then used to test the strength of the association between medians of PFS and OS, and the association between HRPFS and HROS. RESULTS: Fourteen randomized clinical trials published between January 2007 and August 2019, 31 median pairs for PFS and OS, and 17 pairs for HRPFS and HROS from 3,880 patients were selected. The Pearson correlation coefficient between medians of PFS and OS was 0.84 (95% confidence interval (CI) 0.68-0.92, p < 0.001), and the correlation coefficient between HRPFS and HROS was 0.86 (95% CI 0.63-0.95, p < 0.001). Weighted multiple regression analysis showed HRPFS was the most significant predictor of HROS among covariates analyzed (p < 0.001). Both the medians of PFS and OS correlation, and the HRPFS and HROS correlation were 0.79 (p < 0.001), 0.80 (p = 0.001), respectively, in the 11 trials excluding immunotherapy and bevacizumab-based therapy trials. CONCLUSIONS: Our analysis suggests PFS can be strongly correlated with OS and considered a valid surrogate endpoint for OS in advanced or metastatic TNBC.

Prostaglandin E-major urinary metabolite as a noninvasive surrogate marker for infantile necrotizing enterocolitis.

BACKGROUND: Early definitive diagnosis of necrotizing enterocolitis (NEC) based on Bell's staging criteria is difficult because there are few observable changes on abdominal imaging and blood chemistry tests at the onset of the disease. PURPOSE: To investigate whether prostaglandin E-2 major urinary metabolite (PGE-MUM) can be a useful surrogate marker reflecting the disease state and severity of NEC in infants. METHODS: Infants were enrolled in this study between January 2014 and December 2016. NEC diagnosis was based on Bell's staging criteria > Stage II or necrotic bowel observed at surgery. After diagnosis, PGE-MUM level was measured and compared with that of the other disease and healthy infant groups. RESULTS: Median PGE-MUM value was highest in the NEC group (576 [65-3672] µg/g•Cre/BSA × 1000), followed by the other disease group (94 [57-296] µg/g•Cre/BSA × 1000) and the healthy infant group (19 [10-44] µg/g•Cre/BSA × 1000) (sensitivity: 92.3%, specificity: 81.5%, accuracy: 85.0%; p < 0.01). PGE-MUM level correlated with improved status of NEC, length of necrotic intestine, and Bell's staging criteria. CONCLUSIONS: PGE-MUM level may be a useful surrogate biomarker reflecting the disease state of NEC. The method of urine sample collection is also advantageous, being noninvasive for infants. This is the first study reporting PGE-MUM level in NEC. TYPE OF STUDY: Study of diagnostic test. LEVEL OF EVIDENCE: LEVEL II.

The preoperative risk factors of postoperative self-extubation in elderly patients.

OBJECTIVE: Self-extubation of tubes and catheters causes various adverse events in postoperative patients. We investigated preoperative risk factors associated with self-extubation. DESIGN: A matched case-control study. SETTING: Teikyo University Hospital. PARTICIPANTS: Postoperative patients over 50 years old. METHODS: Sixty-five patients with a comment in the incident report about self-extubation within 7 postoperative days were recruited for the case group. One hundred ninety-five matched patients in the control group were randomly recruited from an electronic medical record. This group was three times larger than the case group. The matching factors were age, sex, type of tube, duration of tube insertion, and year of the incident. Conditional multiple logistic regression analysis was performed. RESULTS: Sixty-five self-extubation events occurred, and constituted 6.5% of 996 postoperative incident reports. Three significant preoperative risk factors were abdominal operation (odds ratio [OR], 3.21; 95% confidence interval [95% CI], 1.05-10.83), history of dementia (OR, 10.71; 95% CI, 1.45-132.55), and preoperative hemoglobin level (OR, 0.77 per 1.0 g/dL increase; 95% CI, 0.62-0.96). CONCLUSIONS: Elderly patients with a history of dementia and low preoperative hemoglobin are at a risk of postoperative self-extubation, especially after an abdominal operation. These predictors can contribute to the more effective prevention of perioperative self-extubation.

Aging-associated latent herpes viral infection in normal Japanese individuals and patients with Werner syndrome.

A series of our "inflammageing" study examining serum samples from a maximum of 217 healthy Japanese individuals aged between 1 and 100 years and mutation-proven 40 patients with Werner syndrome (WS) indicated normal aging-associated elevations of highly sensitive CRP (hsCRP) and matrix metalloproteinase-9 (MMP-9). To further study the contribution of environmental factors such as persistent herpes viral infection to inflammageing, IgG antibodies against varicella/zoster virus (VZV) and cytomegalovirus (CMV) were examined in the same serum samples as has been done for hsCRP and MMP-9 analyses. The mean levels of serum IgG viral antibodies were comparable between normal (mean ± SE: 31.0 ± 4.3 unit) and WS (38.6 ± 7.6) for CMV, and between normal (42.0 ± 12.2) and WS (29.8 ± 3.8) for VZV, respectively. Significant associations of aging with IgG anti-CMV antibody were in normal aging (p = 0.023) and WS (p = 0.037), but not with IgG VZV in both conditions. Aging-associated change of IgG anti-CMV antibody titer in WS increased significantly (1.32 times higher) compared with normal aging (p = 0.037). IgG anti-CMV level was significantly elevated in the male gender than female in both conditions (p = 0.006). Elevated hsCRP level was significantly associated with IgG anti-CMV (p = 0.016) and IgG anti-VZV (p = 0.008) antibodies in normal aging, but not in WS. Serum MMP-9 was significantly associated with IgG anti-CMV level (p = 0.0002) in normal aging, but not in WS. Persistent herpes viral infection may constitute a part of "inflammageing" in normal aging and WS.

Distribution of cardiac troponin I in the Japanese general population and factors influencing its concentrations.

BACKGROUND: The 99th percentile of cardiac troponin I level in the general population is accepted as the cut-off for the diagnosis of acute myocardial infarction (AMI). However, it is not clear whether the cut-offs derived in racially and geographically different populations are applicable in Japan. METHODS: Troponin I was determined using the Abbott ARCHITECT STAT high-sensitive troponin I immunoassay in 698 apparently healthy individuals who visited the Japanese Red Cross Medical Center for a health checkup. RESULTS: The 99th percentile of the hsTnI in the overall population was 22.5 (95% confidence interval (CI), 16.8-36.6) pg/mL, 17.7 (95% CI 12.0-22.8) pg/mL for females and 30.6 (95% CI 17.1-53.4) pg/mL for males. The median of the hsTnI in the overall population was 3.2 (95% CI, 3.0-3.3) pg/mL, 2.6 (95% CI 2.4-2.8) pg/mL for females and 4.0 (95% CI 3.8-4.3) pg/mL for males. The age and gender had a significant influence on these values. The troponin I level also showed significant associations with the body mass index (BMI), the gamma glutamyl transferase (GGT), lactate dehydrogenase (LDH), estimated glomerular filtration rate (eGFR), and cardiac abnormalities by electrocardiography (ECG) but not with the high-sensitive C-reactive protein (hsCRP) level. CONCLUSIONS: The 99th percentiles of the troponin I measured in the general population in Japan were comparable as the ones derived in the US, Germany, and Singapore. The troponin I level was dependent on the gender, age, BMI, and cardiac abnormalities found by ECG but not by the hsCRP level.

Pulmonary Carcinoids and Low-Grade Gastrointestinal Neuroendocrine Tumors Show Common MicroRNA Expression Profiles, Different from Adenocarcinomas and Small Cell Carcinomas.

BACKGROUND: It is still uncertain whether small cell lung carcinomas (SCLCs), pulmonary carcinoids, and the gastrointestinal neuroendocrine tumors (GI-NETs) have a common origin. MicroRNA (miRNA) expression may clarify their genetic relationships and origin. METHODS: First, we compared the miRNA expression signature of formalin-fixed paraffin-embedded (FFPE) samples with frozen samples to verify the applicability of microarray analysis. Second, we compared the comprehensive miRNA expression patterns of pulmonary carcinoids and GI-NETs as well as other types of tumors and normal tissues from each organ using FFPE samples. These data were analyzed by hierarchical clustering and consensus clustering with nonnegative matrix factorization. RESULTS: We confirmed that FFPE samples retained the miRNA signatures. In the first hierarchical clustering comparing carcinoids/NETs with adenocarcinomas and normal tissues, most of the carcinoids (48/50) formed 1 major cluster with loose subpartitioning into each organ type, while all the adenocarcinomas (9/9) and normal tissues (15/15) formed another major cluster. The nonnegative matrix factorization approach largely matched the classification of the hierarchical clustering. In the additional cluster analysis comparing carcinoids/NETs with SCLCs, most carcinoids/NETs (17/22) formed a major cluster, while SCLCs (9/9) grouped together with pulmonary adenocarcinomas (3/3) and normal tissues (6/6) in another major cluster. Furthermore, a subset of miRNAs was successfully identified that exhibited significant expression in carcinoids/NETs. CONCLUSION: Carcinoids/NETs had a characteristic pattern of miRNA expression, suggesting a common origin for pulmonary carcinoids and GI-NETs. The expression profiles of pulmonary carcinoids and SCLCs were quite different, indicating the distinct histogenesis of these neuroendocrine neoplasms.

Prognostic Prediction of Oral Squamous Cell Carcinoma by E-Cadherin and N-Cadherin Expression in Overall Cells in Tumor Nests or Tumor Cells at the Invasive Front.

Epithelial-mesenchymal transition (EMT) is a significant process in the invasion and metastasis of cancers including oral squamous cell carcinoma (OSCC), and the cadherin switch has been identified as one of the hallmarks of EMT. The aim of the present study was to evaluate the significance of the cadherin switch in the prognosis of OSCC and generate a model for prognostic predictions. Seventy-six biopsy and/or initial surgical specimens from OSCC patients were immunohistochemically analyzed for the expression of E-cadherin and N-cadherin in either overall OSCC cells in tumor nests or in OSCC cells at the invasive front. Among 76 OSCC cases, overall OSCC cells in tumor nests were negative for the expression of E-cadherin in 10 cases and positive for that of N-cadherin in 53 cases. Among 10 cases negative for the expression of E-cadherin, 4 cases were positive for that of N-cadherin. In OSCC cells at the invasive front, the expression of E-cadherin was negative in 62 cases, while that of N-cadherin was positive in 39 cases. Among 62 cases negative for the expression of E-cadherin, 33 cases were positive for that of N-cadherin. A logistic regression analysis showed that a model using the evaluation of N-cadherin expression in overall OSCC cells in tumor nests with a cut-off point of 70 years old was the best fit model. These results suggest that N-cadherin has significant value in prognostic predictions for OSCC patients.