Yoshiki Kudo

Gene expression profile for predicting survival in advanced-stage serous ovarian cancer across two independent datasets.

Background Advanced-stage ovarian cancer patients are generally treated with platinum/taxane-based chemotherapy after primary debulking surgery. However, there is a wide range of outcomes for individual patients. Therefore, the clinicopathological factors alone are insufficient for predicting prognosis. Our aim is to identify a progression-free survival (PFS)-related molecular profile for predicting survival of patients with advanced-stage serous ovarian cancer. Methodology/Principal Findings Advanced-stage serous ovarian cancer tissues from 110 Japanese patients who underwent primary surgery and platinum/taxane-based chemotherapy were profiled using oligonucleotide microarrays. We selected 88 PFS-related genes by a univariate Cox model (p<0.01) and generated the prognostic index based on 88 PFS-related genes after adjustment of regression coefficients of the respective genes by ridge regression Cox model using 10-fold cross-validation. The prognostic index was independently associated with PFS time compared to other clinical factors in multivariate analysis [hazard ratio (HR), 3.72; 95% confidence interval (CI), 2.66–5.43; p<0.0001]. In an external dataset, multivariate analysis revealed that this prognostic index was significantly correlated with PFS time (HR, 1.54; 95% CI, 1.20–1.98; p = 0.0008). Furthermore, the correlation between the prognostic index and overall survival time was confirmed in the two independent external datasets (log rank test, p = 0.0010 and 0.0008). Conclusions/Significance The prognostic ability of our index based on the 88-gene expression profile in ridge regression Cox hazard model was shown to be independent of other clinical factors in predicting cancer prognosis across two distinct datasets. Further study will be necessary to improve predictive accuracy of the prognostic index toward clinical application for evaluation of the risk of recurrence in patients with advanced-stage serous ovarian cancer.

Computer-aided system for measuring the mandibular cortical width on dental panoramic radiographs in identifying postmenopausal women with low bone mineral density

IntroductionMandibular inferior cortical width manually measured on dental panoramic radiographs may be useful for identifying postmenopausal women with low skeletal bone mineral density (BMD). Automatic measurement of cortical width may enable us to identify a large number of postmenopausal women with suspected low skeletal BMD. The purposes of this study were to develop a computer-aided system for measuring mandibular cortical width on dental panoramic radiographs and clarify the diagnostic efficacy of this system.MethodsPanoramic radiographs of 100 postmenopausal women who had had BMD assessments of the lumbar spine and the femoral neck were used in this study. Experienced oral radiologist determined the position of the mental foramen on 100 digitized dental panoramic radiographs. After determination of the mental foramen, mandibular cortical width below the mental foramen was measured automatically with a computer-aided system by identifying the area of interest, enhancing the original image, determining inner and outer margins of the cortex, and selecting an appropriate point. Cortical width measured by this system was compared with BMD of the lumbar spine and the femoral neck.ResultsThere were statistically significant correlation between cortical width measured by the computer-aided system and spinal BMD (r=0.50) and femoral neck BMD (r=0.54). These correlations were similar with those between cortical width by manual measurement and skeletal BMD. Sensitivity and specificity for identifying postmenopausal women with low spinal BMD by the computer-aided system were about 88.0% and about 58.7%, respectively. Those for identifying postmenopausal women with low femoral neck BMD by this system were about 87.5% and about 56.3%, respectively.ConclusionOur results suggest that our computer-aided system may be useful for identifying postmenopausal women with low skeletal BMD.

Gene expression profiling of advanced-stage serous ovarian cancers distinguishes novel subclasses and implicates ZEB2 in tumor progression and prognosis

To elucidate the mechanisms of rapid progression of serous ovarian cancer, gene expression profiles from 43 ovarian cancer tissues comprising eight early stage and 35 advanced stage tissues were carried out using oligonucleotide microarrays of 18 716 genes. By non‐negative matrix factorization analysis using 178 genes, which were extracted as stage‐specific genes, 35 advanced stage cases were classified into two subclasses with superior (n = 17) and poor (n = 18) outcome evaluated by progression‐free survival (log rank test, P = 0.03). Of the 178 stage‐specific genes, 112 genes were identified as showing different expression between the two subclasses. Of the 48 genes selected for biological function by gene ontology analysis or Ingenuity Pathway Analysis, five genes (ZEB2, CDH1, LTBP2, COL16A1, and ACTA2) were extracted as candidates for prognostic factors associated with progression‐free survival. The relationship between high ZEB2 or low CDH1 expression and shorter progression‐free survival was validated by real‐time RT‐PCR experiments of 37 independent advanced stage cancer samples. ZEB2 expression was negatively correlated with CDH1 expression in advanced stage samples, whereas ZEB2 knockdown in ovarian adenocarcinoma SKOV3 cells resulted in an increase in CDH1 expression. Multivariate analysis showed that high ZEB2 expression was independently associated with poor prognosis. Furthermore, the prognostic effect of E‐cadherin encoded by CDH1 was verified using immunohistochemical analysis of an independent advanced stage cancer samples set (n = 74). These findings suggest that the expression of epithelial–mesenchymal transition‐related genes such as ZEB2 and CDH1 may play important roles in the invasion process of advanced stage serous ovarian cancer. (Cancer Sci 2009)

Characterization of amino acid transport systems in human placental brush-border membrane vesicles.

Brush-border microvillous plasma membrane vesicles were prepared from human full-term placental syncytiotrophoblasts and purified 33-fold from the homogenate with reference to a membrane marker enzyme, alkaline phosphatase (EC 3.1.3.1). Transport of alpha-(methylamino)isobutyrate by the membrane vesicles was stimulated in the presence of an Na+ gradient from the outside to the inside of the vesicles. The initial rate of uptake in a 10-s period was enhanced with increasing concentration of Na+ in the external medium. The level of alpha-(methylamino)isobutyrate transported into the vesicles reached a maximum 1 min after the start of incubation at 37 degrees C, and then decreased with time due to efflux. Extrapolation to infinite medium osmolarity showed no uptake, indicating transport of alpha-(methylamino)isobutyrate into membrane vesicles. The initial rate of uptake was dependent on temperature and pH: the highest rate occurred at 37 degrees C and the optimal pH was 8.0. When the alpha-(methylamino)isobutyrate concentration was varied, the initial rate of uptake dependent on an Na+ gradient (out greater than in) obeyed Michaelis-Menten kinetics with Km and Vmax values of 1.07 mM and 3.23 nmol/10 s per mg of protein, respectively. Cross-inhibition patterns indicated that at least three Na+-dependent and two Na+-independent carrier-mediated pathways existed in the human placental brush border. One Na+-dependent pathway interacted with all substrates tested. Another Na+-dependent route interacted with L-proline, alpha-(methylamino)isobutyrate, and L-methionine, while a third pathway was selective for L-methionine. One Na+-independent pathway was selective for L-cysteine, while the other pathway interacted with all substrates tested.

TALEN-mediated single-base-pair editing identification of an intergenic mutation upstream of BUB1B as causative of PCS (MVA) syndrome

Significance A single nucleotide substitution in an intergenic region upstream of BUB1B (encoding BUBR1) was identified as a candidate mutation for premature chromatid separation with mosaic variegated aneuploidy [PCS (MVA) syndrome], a cancer-prone genetic disorder. To prove that this is the causal mutation, we designed a unique genome editing strategy, transcription activator-like effector nuclease–mediated two-step single-base-pair editing, to biallelically introduce this substitution into cultured human cells. The cell clones showed chromosomal instability in the form of PCS and MVA, which are cellular hallmarks of the syndrome, suggesting that this is indeed the underlying mutation. This single-base-pair editing technique will be useful for investigations of noncoding variants of unknown functional relevance. Cancer-prone syndrome of premature chromatid separation with mosaic variegated aneuploidy [PCS (MVA) syndrome] is a rare autosomal recessive disorder characterized by constitutional aneuploidy and a high risk of childhood cancer. We previously reported monoallelic mutations in the BUB1B gene (encoding BUBR1) in seven Japanese families with the syndrome. No second mutation was found in the opposite allele of any of the families studied, although a conserved BUB1B haplotype and a decreased transcript were identified. To clarify the molecular pathology of the second allele, we extended our mutational search to a candidate region surrounding BUB1B. A unique single nucleotide substitution, G > A at ss802470619, was identified in an intergenic region 44 kb upstream of a BUB1B transcription start site, which cosegregated with the disorder. To examine whether this is the causal mutation, we designed a transcription activator-like effector nuclease–mediated two-step single-base pair editing strategy and biallelically introduced this substitution into cultured human cells. The cell clones showed reduced BUB1B transcripts, increased PCS frequency, and MVA, which are the hallmarks of the syndrome. We also encountered a case of a Japanese infant with PCS (MVA) syndrome carrying a homozygous single nucleotide substitution at ss802470619. These results suggested that the nucleotide substitution identified was the causal mutation of PCS (MVA) syndrome.

Intergender differences in histological architecture of the fascia pelvis parietalis: a cadaveric study.

The fascia pelvis parietalis (FPP) or endopelvic fascia is a well‐known structure, but few studies described the detailed histological architecture, including the composite fiber directions. We hypothesized a gender‐specific fiber architecture corresponding to the functional demand. For the first step to examine this hypothesis, we investigated specimens from 27 adult cadavers (10 males and 17 females) and 11 midterm fetuses (five males and six females) using immunohistochemistry and aldehyde‐fuchsin staining. The adult female FPP was a solid, thick monolayered structure that was reinforced by abundant elastic fibers running across the striated muscle fibers, but it contained little or no smooth muscles (SM). In contrast, the male FPP was multilayered with abundant SM. In midterm fetuses, SM originated from the inferior part of the bladder and extended inferiorly along the gender‐specific courses. Thus, we found a clear intergender difference in FPP architecture. However, the functional significance remained unknown because the basic architecture was common between nulliparous and multiparous women. Rather than for meeting the likely mechanical demands of pregnancy and vaginal delivery, the intergender difference of the FPP seemed to result from differences in the amount and migration course of bladder‐derived SM as well as in hormonal background. Clin. Anat. 24:469–477, 2011.

Prediction of individual response to platinum/paclitaxel combination using novel marker genes in ovarian cancers

We attempted to identify potent marker genes using a new statistical analysis and developed a prediction system for individual response to platinum/paclitaxel combination chemotherapy in ovarian cancer patients based on the hypothesis that expression analysis of a set of the key drug sensitivity genes for platinum and paclitaxel could allow us to predict therapeutic response to the combination. From 10 human ovarian cancer cell lines, genes correlative in the expression levels with cytotoxicities of cisplatin (CDDP) and paclitaxel were chosen. We first selected five reliable prediction markers for the two drugs from 22 genes already known as sensitivity determinants and then identified another 8 novel genes through a two-dimensional mixed normal model using oligomicroarray expression data. Using expression data of genes quantified by real-time reverse transcription-PCR, we fixed the best linear model, which converted the quantified expression data into an IC50 of each drug. Multiple regression analysis of the selected genes yielded three prediction formulae for in vitro activity of CDDP and paclitaxel. In the same way, using the same genes selected in vitro, we then attempted to develop prediction formulae for progression-free survival to the platinum/paclitaxel combination. We therefore constructed possible formulae using different sets of 13 selected marker genes (5 known and 8 novel genes): Utility confirmation analyses using another nine test samples seemed to show that the formulae using a set of 8 novel marker genes alone could accurately predict progression-free survival (r = 0.683; P = 0.042). [Mol Cancer Ther 2006;5(3):767–75]

IFPA Meeting 2010 Workshops Report II: Placental pathology; trophoblast invasion; fetal sex; parasites and the placenta; decidua and embryonic or fetal loss; trophoblast differentiation and syncytialisation

Workshops are an important part of the IFPA annual meeting. At IFPA Meeting 2010 diverse topics were discussed in twelve themed workshops, six of which are summarized in this report. 1. The placental pathology workshop focused on clinical correlates of placenta accreta/percreta. 2. Mechanisms of regulation of trophoblast invasion and spiral artery remodeling were discussed in the trophoblast invasion workshop. 3. The fetal sex and intrauterine stress workshop explored recent work on placental sex differences and discussed them in the context of whether boys live dangerously in the womb.4. The workshop on parasites addressed inflammatory responses as a sign of interaction between placental tissue and parasites. 5. The decidua and embryonic/fetal loss workshop focused on key regulatory mediators in the decidua, embryo and fetus and how alterations in expression may contribute to different diseases and adverse conditions of pregnancy. 6. The trophoblast differentiation and syncytialisation workshop addressed the regulation of villous cytotrophoblast differentiation and how variations may lead to placental dysfunction and pregnancy complications.

Characterization of amino-acid transport systems in guinea-pig intestinal brush-border membrane

The amino-acid transport systems have been characterized in brush-border membrane vesicles prepared from guinea-pig small intestine. Uptake of all amino acids tested was measured at the initial velocity for 5 s. L-Proline, alpha-(methylamino)isobutyrate, glycine, L-alanine and L-methionine were transported dependent solely on an Na+ gradient from the outside to the inside of the vesicles, and L-cysteine, L-phenylalanine and L-leucine were transported dependent largely on the Na+ gradient with a small fraction of Na+-independent transport. The transport of L-aspartic acid and L-lysine was independent of the Na+ gradient and L-lysine transport was somewhat inhibited by the presence of cations, including Na+, K+ and Li+. A cross-inhibition study of the uptake of these amino acids in the brush border of guinea-pig intestine revealed the presence of at least three Na+-dependent and three Na+-independent carrier-mediated systems. One Na+-dependent system interacted mainly with imino acid. Another Na+-dependent system interacted with neutral amino acids, while a third system was selective for glycine. One Na+-independent system is for acidic amino acids, another is responsible for neutral amino acids and a third for cationic amino acids. These transport systems of amino acids in guinea-pig small intestine are compared with those in rabbit and mouse intestine.

IFPA Meeting 2008 Workshops Report

Workshops are an important part of the IFPA annual meeting. At the IFPA meeting 2008 diverse topics were discussed in 12 themed workshops. Topics covered included: immunology of placentation; galectins and trophoblast invasion; signaling in implantation and invasion; markers to identify trophoblast subpopulations; placental pathology; placental toxicology; stereology; placental transport of fatty acids; placental mesenchymal stem cells; comparative placentation; trophoblast and neoplasia; trophoblast differentiation. This report is a summary of the various topics covered.