Conversion to complete resection with mFOLFOX6 with bevacizumab or cetuximab based on K-RAS status for unresectable colorectal liver metastasis (BECK study): Long-term results of survival.

BACKGROUND/PURPOSE: To investigate the long-term outcome and entire treatment course of patients with technically unresectable CRLM who underwent conversion hepatectomy and to examine factors associated with conversion to hepatectomy. METHODS: Recurrence and survival data with long-term follow-up were analyzed in the cohort of a multi-institutional phase II trial for technically unresectable colorectal liver metastases (the BECK study). RESULTS: A total of 22/12 patients with K-RAS wild-type/mutant tumors were treated with mFOLFOX6 + cetuximab/bevacizumab. The conversion R0/1 hepatectomy rate was significantly higher in left-sided primary tumors than in right-sided tumors (75.0% vs 30.0%, P = .022). The median follow-up was 72.6 months. The 5-year overall survival (OS) rate in the entire cohort was 48.1%. In patients who underwent R0/1 hepatectomy (n = 21), the 5-year RFS rate and OS rate were 19.1% and 66.3%, respectively. At the final follow-up, seven patients had no evidence of disease, five were alive with disease, and 20 had died from their original cancer. All 16 patients who achieved 5-year survival underwent conversion hepatectomy, and 11 of them underwent further resection for other recurrences (median: 2, range: 1-4). CONCLUSIONS: Conversion hepatectomy achieved a similar long-term survival to the results of previous studies in initially resectable patients, although many of them experienced several post-hepatectomy recurrences. Left-sided primary was found to be the predictor for conversion hepatectomy.

Polymorphic edge detection (PED): two efficient methods of polymorphism detection from next-generation sequencing data.

BACKGROUND: Accurate detection of polymorphisms with a next generation sequencer data is an important element of current genetic analysis. However, there is still no detection pipeline that is completely reliable. RESULT: We demonstrate two new detection methods of polymorphisms focusing on the Polymorphic Edge (PED). In the matching between two homologous sequences, the first mismatched base to appear is the SNP, or the edge of the structural variation. The first method is based on k-mers from short reads and detects polymorphic edges with k-mers for which there is no match between target and control, making it possible to detect SNPs by direct comparison of short-reads in two datasets (target and control) without a reference genome sequence. The second method is based on bidirectional alignment to detect polymorphic edges, not only SNPs but also insertions, deletions, inversions and translocations. Using these two methods, we succeed in making a high-quality comparison map between rice cultivars showing good match to the theoretical value of introgression, and in detecting specific large deletions across cultivars. CONCLUSIONS: Using Polymorphic Edge Detection (PED), the k-mer method is able to detect SNPs by direct comparison of short-reads in two datasets without genomic alignment step, and the bidirectional alignment method is able to detect SNPs and structural variations from even single-end short-reads. The PED is an efficient tool to obtain accurate data for both SNPs and structural variations. AVAILABILITY: The PED software is available at: https://github.com/akiomiyao/ped .

Global transcriptome analysis of pig induced pluripotent stem cells derived from six and four reprogramming factors.

Pigs are important, both for agriculture and as animal models for human diseases. However, due to the lack of embryonic stem cells, the possibility of genetic modification is quite limited. To overcome this limitation, induced pluripotent stem (iPS) cells have been derived from pigs. Despite the public availability of a large number of expression datasets from mice, rats, and primates-derived iPS cells, the expression profile of pig-derived iPS cells is quite limited. Furthermore, there is no dataset focused on the profiling of pig-derived iPS cell with six reprogramming factors (Oct3/4, Sox2, Klf4, c-Myc, Lin28, and Nanog). Here, we used Illumina RNA sequencing platform to characterize the mRNA expression of four-factor derived and six-factor derived pig iPS cells. We observed that the expression levels of whole genes in our established six factors derived iPS cells and parent fibroblast, and compared with that of iPS cells with four factors in public database. These data are valuable in understanding species difference in the reprogramming process of stem cells, and could help identify the key regulating genes involved in the process.

Multi-institutional phase II study on the feasibility of liver resection following preoperative mFOLFOX6 therapy for resectable liver metastases from colorectal cancers.

BACKGROUND: Although liver resection combined with preoperative chemotherapy is expected to improve outcomes of patients with resectable colorectal liver metastasis (CRLM), there is as yet insufficient clinical evidence supporting the efficacy of preoperative systemic chemotherapy. The aim of this phase II study was to assess the feasibility and efficacy of preoperative FOLFOX systemic chemotherapy for patients with initially resectable CRLM. METHODS: A prospective multi-institutional phase II study was conducted to evaluate the feasibility and efficacy of preoperative chemotherapy for resectable CRLM (ClinicalTrials.gov identifier number NCT00594529). Patients were scheduled to receive 6 cycles of mFOLFOX6 therapy before liver surgery. The primary endpoint was the macroscopic curative resection rate. RESULTS: A total of 30 patients were included in this study. Two patients who were diagnosed with hepatocellular and intrahepatic cholangiocellular carcinoma based on pathology were excluded from the analysis. More than half of the patients (57 %) had solitary liver metastasis. The completion rate of preoperative chemotherapy was 64.3 % and the response rate was 53.6 %. Two patients were unable to proceed to liver resections due to disease progression and severe postoperative complications following primary tumor resection. Macroscopic curative resection was obtained in 89.3 % of eligible patients. Postoperative mortality and severe complication (≥Gr. 3) rates were 0 and 11 %, respectively. The 3-year overall and progression-free survival rates were 81.9 and 47.4 %, respectively. CONCLUSION: Our phase II study demonstrated the feasibility of liver resection combined with preoperative mFOLFOX6 therapy in patients with initially resectable CRLM. Further study is warranted to address the oncological effects of preoperative chemotherapy.

Conversion to complete resection with mFOLFOX6 with bevacizumab or cetuximab based on K-ras status for unresectable colorectal liver metastasis (BECK study).

BACKGROUND: Patients with colorectal liver metastasis (CRLM) might be down-staged by chemotherapy from an initially unresectable stage to a resectable stage. Because the tumor response to preoperative chemotherapy has been correlated with resection rate, the improved efficacy from the concept that only the patients without K-ras mutations receive an anti-EGFR antibody might be expected to increase the conversion rate. The purpose of this study is to evaluate the conversion rate from unresectable CRLM to complete resection. METHODS: We conducted a multi-institutional phase II trial for unresectable CRLM. Patients received mFOLFOX6 with either bevacizumab (bev) or cetuximab (cet) based on K-ras status (UMIN000004310). Planned treatment was for six cycles during which tumors were assessed for resectability every three cycles. Patients whose disease was unresectable after six cycles switched their chemotherapy regimen from mFOLFOX6 to FOLFIRI. The primary endpoint was R0 resection rate. RESULTS: Thirty-five patients with unresectable CRLM were enrolled. A total of 22/12 patients with K-ras wild-type/mutant (wt/mt) were treated with mFOLFOX6 plus cet/bev, respectively. The overall response rate was 64.7% (wt/mt; 77.3%/41.7%, P = 0.04). In 20 patients (58.8%), hepatectomy was performed according to protocol treatment, and the conversion rate was 72.7%/33.3% in wt/mt patients, respectively (P = 0.03). Finally, 23 patients (67.6%) underwent hepatectomy, and the conversion rate was 77.2%/50.0% in wt/mt patients (P = 0.09). The overall R0 resection rate was 47.1% (wt/mt; 50.0%/41.7%, P = 0.36). CONCLUSIONS: This prospective study showed that combined chemotherapy based on K-ras status can facilitate conversion to resection in patients with unresectable CRLM.

SV40 VP1 major capsid protein in its self-assembled form allows VP1 pentamers to coat various types of artificial beads in vitro regardless of their sizes and shapes.

The icosahedral capsid structure of simian virus 40 (diameter, 45 nm) consists of 72 pentameric subunits, with each subunit formed by five VP1 molecules. Electron microscopy, immuno-gold labeling, and ζ-potential analysis showed that purified recombinant VP1 pentamers covered polystyrene beads measuring 100, 200, and 500 nm in diameter, as well as silica beads. In addition to covering spherical beads, VP1 pentamers covered cubic magnetite beads, as well as the distorted surface structures of liposomes. These findings indicate that VP1 pentamers could coat artificial beads of various shapes and sizes larger than the natural capsid. Technology based on VP1 pentamers may be useful in providing a capsid-like surface for enclosed materials, enhancing their stability and cellular uptake for drug delivery systems.

PosMed: Ranking genes and bioresources based on Semantic Web Association Study.

Positional MEDLINE (PosMed; http://biolod.org/PosMed) is a powerful Semantic Web Association Study engine that ranks biomedical resources such as genes, metabolites, diseases and drugs, based on the statistical significance of associations between user-specified phenotypic keywords and resources connected directly or inferentially through a Semantic Web of biological databases such as MEDLINE, OMIM, pathways, co-expressions, molecular interactions and ontology terms. Since 2005, PosMed has long been used for in silico positional cloning studies to infer candidate disease-responsible genes existing within chromosomal intervals. PosMed is redesigned as a workbench to discover possible functional interpretations for numerous genetic variants found from exome sequencing of human disease samples. We also show that the association search engine enhances the value of mouse bioresources because most knockout mouse resources have no phenotypic annotation, but can be associated inferentially to phenotypes via genes and biomedical documents. For this purpose, we established text-mining rules to the biomedical documents by careful human curation work, and created a huge amount of correct linking between genes and documents. PosMed associates any phenotypic keyword to mouse resources with 20 public databases and four original data sets as of May 2013.

Ulcerative colitis associated with isolated unilateral hypoglossal nerve palsy.

We herein describe a rare case of ulcerative colitis associated with unilateral hypoglossal nerve palsy. A 64-year-old woman developed severe active ulcerative colitis and was treated with prednisolone. The dose of oral prednisolone was reduced to 7.5 mg/day, following which the patient noticed slight dysphagia and a speech disturbance. She was diagnosed with unilateral hypoglossal nerve palsy, which was thought to be caused by mononeuritis. She was treated with intravenous methylprednisolone at a dose of 500 mg/day, which improved the neuropathy. When the neuropathy occurred, the patient was in a mildly active stage of ulcerative colitis. We concluded that the mononeuritis observed in the present case was likely an extraintestinal manifestation of ulcerative colitis.

Leber's Hereditary Optic Neuropathy with Olivocerebellar Degeneration due to G11778A and T3394C Mutations in the Mitochondrial DNA.

BACKGROUND: Leber's hereditary optic neuropathy (LHON) is a mitochondrial disorder with optic nerve atrophy. Although there are no other associated neurological abnormalities in most cases of LHON, cases of "LHON plus" have been reported. CASE REPORT: The proband was a 37-year-old man who had visual and gait disturbances that had first appeared at 10 years of age. He showed horizontal gaze palsy, gaze-evoked nystagmus, dysarthria, and cerebellar ataxia. Brain and orbit MRI disclosed atrophy of the optic nerve and cerebellum, and degenerative changes in the bilateral inferior olivary nucleus. Mutational analyses of mitochondrial DNA identified the coexistence of heteroplasmic G11778A and homoplasmic T3394C mutations. CONCLUSIONS: These results suggest that the combination of G11778A and T3394C mutations leads to an atypical LHON phenotype.

The RIKEN integrated database of mammals.

The RIKEN integrated database of mammals (http://scinets.org/db/mammal) is the official undertaking to integrate its mammalian databases produced from multiple large-scale programs that have been promoted by the institute. The database integrates not only RIKEN's original databases, such as FANTOM, the ENU mutagenesis program, the RIKEN Cerebellar Development Transcriptome Database and the Bioresource Database, but also imported data from public databases, such as Ensembl, MGI and biomedical ontologies. Our integrated database has been implemented on the infrastructure of publication medium for databases, termed SciNetS/SciNeS, or the Scientists' Networking System, where the data and metadata are structured as a semantic web and are downloadable in various standardized formats. The top-level ontology-based implementation of mammal-related data directly integrates the representative knowledge and individual data records in existing databases to ensure advanced cross-database searches and reduced unevenness of the data management operations. Through the development of this database, we propose a novel methodology for the development of standardized comprehensive management of heterogeneous data sets in multiple databases to improve the sustainability, accessibility, utility and publicity of the data of biomedical information.

[A case of acute cerebellar ataxia associated with serum anti-NH2 terminal of alpha-enolase (NAE) antibody].

We reported a 61-year-old man who had developed acute cerebellar ataxia in the trunk and the lower limbs. His chemical blood analysis showed very mild hypothyroidism and the presence of serum anti-thyroid peroxidase (TPO) antibody and anti-NH2 terminal of alpha-enolase (NAE) antibody. While cerebellar atrophy was not evident on magnetic resonance imaging (MRI) of the brain, 99mTc-ECD SPECT using the easy Z-score imaging system (eZIS) showed decreased regional cerebral blood flow (rCBF) in the vermis of cerebellum. His cerebellar ataxia improved spontaneously within three weeks. The present case is very rare and suggests that anti-NAE autoantibody may be associated with actue cerebellar ataxia.

Prevalence of dementia in the rural island town of Ama-cho, Japan.

BACKGROUND: With the striking increase in the number of elderly people in Japan, dementia has not only become a medical but also a social issue. METHODS: We studied the prevalence of dementing disorders in a rural island town of Japan (Ama-cho), using a door-to-door 2-phase design. RESULTS: Of the 120 persons screened as having cognitive impairment, 104 people were diagnosed as having dementia. The prevalence (cases/100 persons aged 65 years and older) was 11.0 for all types of dementia, 7.0 for Alzheimer's disease, 1.7 for vascular dementia, 0.53 for dementia with Lewy bodies, 0.74 for Parkinson's disease dementia, 0.21 for progressive supranuclear palsy, 0.11 for frontotemporal lobar degeneration and 0.74 for other dementia. The overall prevalence was higher in women for Alzheimer's disease and Parkinson's disease dementia, and in men, for vascular dementia and dementia with Lewy bodies. CONCLUSION: We confirmed the overall prevalence of dementia in the elderly population aged 65 years and older to be 11.0. This finding is higher compared with previous reports in Japan.

Adherent monomer-misfolded SOD1.

BACKGROUND: Multiple cellular functions are compromised in amyotrophic lateral sclerosis (ALS). In familial ALS (FALS) with Cu/Zn superoxide dismutase (SOD1) mutations, the mechanisms by which the mutation in SOD1 leads to such a wide range of abnormalities remains elusive. METHODOLOGY/PRINCIPAL FINDINGS: To investigate underlying cellular conditions caused by the SOD1 mutation, we explored mutant SOD1-interacting proteins in the spinal cord of symptomatic transgenic mice expressing a mutant SOD1, SOD1(Leu126delTT) with a FLAG sequence (DF mice). This gene product is structurally unable to form a functional homodimer. Tissues were obtained from both DF mice and disease-free mice expressing wild-type with FLAG SOD1 (WF mice). Both FLAG-tagged SOD1 and cross-linking proteins were enriched and subjected to a shotgun proteomic analysis. We identified 34 proteins (or protein subunits) in DF preparations, while in WF preparations, interactions were detected with only 4 proteins. CONCLUSIONS/SIGNIFICANCE: These results indicate that disease-causing mutant SOD1 likely leads to inadequate protein-protein interactions. This could be an early and crucial process in the pathogenesis of FALS.

A novel cell transplantation protocol and its application to an ALS mouse model.

Amyotrophic lateral sclerosis (ALS) is a lethal neurodegenerative disease, which selectively affects motor neurons throughout the central nervous system. The extensive distribution of motor neurons is an obstacle to applying cell transplantation therapy for the treatment of ALS. To overcome this problem, we developed a cell transplantation method via the fourth cerebral ventricle in mice. We used mouse olfactory ensheathing cells (OECs) and rat mesenchymal stem cells (MSCs) as donor cells. OECs are reported to promote regeneration and remyelination in the spinal cord, while MSCs have a capability to differentiate into several types of specific cells including neural cells. Furthermore both types of cells can be relatively easily obtained by biopsy in human. Initially, we confirmed the safety of the operative procedure and broad distribution of grafted cells in the spinal cord using wild-type mice. After transplantation, OECs distributed widely and survived as long as 100 days after transplantation, with a time-dependent depletion of cell number. In ALS model mice, OEC transplantation revealed no adverse effects but no significant differences in clinical evaluation were found between OEC-treated and non-transplanted animals. After MSC transplantation into the ALS model mice, females, but not males, showed a statistically longer disease duration than the non-transplanted controls. We conclude that intrathecal transplantation could be a promising way to deliver donor cells to the central nervous system. Further experiments to elucidate relevant conditions for optimal outcomes are required.

The generation challenge programme platform: semantic standards and workbench for crop science.

The Generation Challenge programme (GCP) is a global crop research consortium directed toward crop improvement through the application of comparative biology and genetic resources characterization to plant breeding. A key consortium research activity is the development of a GCP crop bioinformatics platform to support GCP research. This platform includes the following: (i) shared, public platform-independent domain models, ontology, and data formats to enable interoperability of data and analysis flows within the platform; (ii) web service and registry technologies to identify, share, and integrate information across diverse, globally dispersed data sources, as well as to access high-performance computational (HPC) facilities for computationally intensive, high-throughput analyses of project data; (iii) platform-specific middleware reference implementations of the domain model integrating a suite of public (largely open-access/-source) databases and software tools into a workbench to facilitate biodiversity analysis, comparative analysis of crop genomic data, and plant breeding decision making.

Hepatic ischemia-reperfusion increases vascular endothelial growth factor and cancer growth in rats.

BACKGROUND: In liver surgery, the hepatic pedicle often is clamped to reduce blood loss, and later unclamped, representing hepatic ischemia and reperfusion (I/R) with induction of hypoxia. Vascular endothelial growth factor (VEGF) expression reportedly is induced by hypoxia; further, some cancer cells express the VEGF receptor (flt-1, flk-1/KDR). We hypothesized that I/R-induced VEGF expression could enhance growth of microscopic tumor via VEGF receptors on tumor cells, thus promoting liver metastasis in a rat model. MATERIALS AND METHODS: Time-dependent VEGF expression in liver and plasma was determined by enzyme-linked immunosorbent assay in rats subjected to 60 min of 70% hepatic I/R (I/R group). Other rats given an intrasplenic inoculation of a rat colon adenocarcinoma cell line (RCH-H4) were divided 3 days later into three groups: group A, untreated; group B, sham operation; group C, 70% I/R for 60 min. Liver metastasis was evaluated on day 14. Expression of flt-1 and flk-1/KDR was examined in RCN-H4 cells, and effects of exogenous VEGF on RCN-H4 cell proliferation were determined by MTT assays. RESULTS: Hepatic VEGF expression increased significantly in the I/R group compared to the control group. Liver metastasis was more extensive in group C than in groups A and B. RCN-H4 cells expressed flt-1 and flk-1/KDR, while exogenous VEGF increased RCN-H4 cell proliferation. CONCLUSION: Hepatic ischemia reperfusion leads to induction of VEGF and this is associated with increased tumor burden in an animal model of colon cancer metastasis.

Modified hemi-double-stapling technique combined with the temporal abdominal-wall-lift method for performing Billroth I anastomosis after laparoscopically assisted distal gastrectomy.

The authors have used a modified hemi-double-stapling (HDS) technique for reconstruction after laparoscopically assisted distal gastrectomy. The stomach is resected from the greater curvature side using a linear stapler inserted into the stomach from that side at a position vertical to the line of the greater curvature. Resection of the stomach is performed by extending the resection line to the lesser curvature using laparoscopic coagulating shears. The resected specimen is examined. After placement of a purse-string suture at the duodenal stump, an anvil is inserted into the stump, and an additional suture with 2-0 silk is placed over the purse-string suture. A curved intraluminal stapler (CDH25) is inserted into the stomach through the opening made on the lesser curvature side, and the center rod of the stapler is passed through the gastric wall on the corner of the resection line at the greater curvature. Ligation with 2-0 silk is added to the center rod by suturing the gastric tissue 5-8 mm from the center rod to encircle it. The authors call this the "one-knot setup HDS," and with this method, a large-caliber anastomosis is secured. In many cases, it is difficult to observe the anastomotic site through the small incisional opening. However, under laparoscopy with the temporal abdominal wall-lift method using the Multi Flap Gate, the anastomotic site can be easily and safely observed. One-knot setup HDS combined with the temporal abdominal wall-lift method is considered a safe and simple method for performing Billroth I anastomosis in laparoscopic distal gastrectomy.

Development of a novel data mining tool to find cis-elements in rice gene promoter regions.

BACKGROUND: Information on more than 35 000 full-length Oryza sativa cDNAs, together with associated microarray gene expression data collected under various treatment conditions, has made it feasible to identify motifs that are conserved in gene promoters and may act as cis-regulatory elements with key roles under the various conditions. RESULTS: We have developed a novel tool that searches for cis-element candidates in the upstream, downstream, or coding regions of differentially regulated genes. The tool first lists cis-element candidates by motif searching based on the supposition that if there are cis-elements playing important roles in the regulation of a given set of genes, they will be statistically overrepresented and will be conserved. Then it evaluates the likelihood scores of the listed candidate motifs by association rule analysis. This strategy depends on the idea that motifs overrepresented in the promoter region could play specific roles in the regulation of expression of these genes. The tool is designed so that any biological researchers can use it easily at the publicly accessible Internet site http://hpc.irri.cgiar.org/tool/nias/ces. We evaluated the accuracy and utility of the tool by using a dataset of auxin-inducible genes that have well-studied cis-elements. The test showed the effectiveness of the tool in identifying significant relationships between cis-element candidates and related sets of genes. CONCLUSION: The tool lists possible cis-element motifs corresponding to genes of interest, and it will contribute to the deeper understanding of gene regulatory mechanisms in plants.

Mitochondrial changes in motor neurons of homozygotes of leucine 126 TT deletion SOD1 transgenic mice.

We investigated the time course of ultrastructural changes of mitochondria in the spinal cord of homozygotes of Leu126TTdel SOD1 (superoxide dismutase 1) with FLAG (signal sequence at the C-terminal protein) transgenic mice (DF-homo). Non-Tg mice and wild-type human SOD1 with FLAG epitope transgenic mice (WF) were investigated as controls for non-onset Tg mice. Expansion and vacuolation of the mitochondrial matrix was exhibited in motor neurons in the anterior horns of DF-homo Tg mice at the presymptomatic stage. Such mitochondrial degeneration became severe at the postsymptomatic stage. In contrast, expansion of the mitochondrial inner-membrane space was not evident even at the terminal stage. Microvacuoles of cytoplasm and fibrillar inclusions were rarely shown from the early symptomatic stage. WF mice showed expansion and vacuolation of the mitochondrial inner membrane space at old age. Non-Tgs showed no obvious change in mitochondria. Gold-labeled human SOD1 immunoreactivity showed small amount of gold deposits in the vacuolated mitochondria. These results suggest that the expansion and vacuolation of mitochondrial matrix in the spinal cord of DF-homo transgenic mice is the first pathological change, but that it is not directly caused by the aggregation of an abnormal human SOD1 protein in intermembrane space of mitochondria.

Gene organization in rice revealed by full-length cDNA mapping and gene expression analysis through microarray.

Rice (Oryza sativa L.) is a model organism for the functional genomics of monocotyledonous plants since the genome size is considerably smaller than those of other monocotyledonous plants. Although highly accurate genome sequences of indica and japonica rice are available, additional resources such as full-length complementary DNA (FL-cDNA) sequences are also indispensable for comprehensive analyses of gene structure and function. We cross-referenced 28.5K individual loci in the rice genome defined by mapping of 578K FL-cDNA clones with the 56K loci predicted in the TIGR genome assembly. Based on the annotation status and the presence of corresponding cDNA clones, genes were classified into 23K annotated expressed (AE) genes, 33K annotated non-expressed (ANE) genes, and 5.5K non-annotated expressed (NAE) genes. We developed a 60mer oligo-array for analysis of gene expression from each locus. Analysis of gene structures and expression levels revealed that the general features of gene structure and expression of NAE and ANE genes were considerably different from those of AE genes. The results also suggested that the cloning efficiency of rice FL-cDNA is associated with the transcription activity of the corresponding genetic locus, although other factors may also have an effect. Comparison of the coverage of FL-cDNA among gene families suggested that FL-cDNA from genes encoding rice- or eukaryote-specific domains, and those involved in regulatory functions were difficult to produce in bacterial cells. Collectively, these results indicate that rice genes can be divided into distinct groups based on transcription activity and gene structure, and that the coverage bias of FL-cDNA clones exists due to the incompatibility of certain eukaryotic genes in bacteria.