Publications by authors named "Haruo Ikeda"

179 Publications

Unified magnifying endoscopic classification for esophageal, gastric and colonic lesions: a feasibility pilot study.

Endosc Int Open 2021 Sep 16;9(9):E1306-E1314. Epub 2021 Aug 16.

Digestive Diseases Center, Showa University Koto Toyosu Hospital, Tokyo, Japan.

Image-enhanced magnifying endoscopy allows optimization of the detection and diagnosis of lesions found in the gastrointestinal tract. Current organ-specific classifications are well-accepted by specialized endoscopists but may pose confusion for general gastroenterologists. To address this, our group proposed the Unified Magnifying Endoscopic Classification (UMEC) which can be applied either in esophagus, stomach, or colon. The aim of this study was to evaluate the diagnostic performance and clinical applicability of UMEC. A single-center, feasibility pilot study was conducted. Two endoscopists with experience in magnifying narrow band imaging (NBI), blinded to white-light and non-magnifying NBI findings as well as histopathological diagnosis, independently reviewed and diagnosed all images based on UMEC. In brief, UMEC is divided into three categories: non-neoplasia, intramucosal neoplasia, and deep submucosal invasive cancer. The diagnostic performance of UMEC was assessed while using the gold standard histopathology as a reference. A total of 303 gastrointestinal lesions (88 esophageal squamous lesions, 90 gastric lesions, 125 colonic lesions) were assessed. The overall accuracy for both endoscopists in the diagnosis of esophageal squamous cell cancer, gastric cancer, and colorectal cancer were 84.7 %, 89.5 %, and 83.2 %, respectively. The interobserver agreement for each organ, Kappa statistics of 0.51, 0.73, and 0.63, was good. UMEC appears to be a simple and practically acceptable classification, particularly to general gastroenterologists, due to its good diagnostic accuracy, and deserves further evaluation in future studies.
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http://dx.doi.org/10.1055/a-1499-6638DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8367430PMC
September 2021

Diagnosis of congenital esophageal stenosis in adults and treatment with peroral endoscopic myotomy.

Ann Gastroenterol 2021 Jul-Aug;34(4):493-500. Epub 2021 Mar 23.

Digestive Diseases Center, Showa University Koto Toyosu Hospital, Tokyo, Japan (Haruo Ikeda, Haruhiro Inoue, Mary Raina Angeli Abad, Yusuke Fujiyoshi, Yohei Nishikawa, Akiko Toshimori, Mayo Tanabe, Yuto Shimamura, Kazuya Sumi, Yugo Iwaya, Manabu Onimaru).

Background: Congenital esophageal stenosis (CES) in adults is a rare disorder that can present as achalasia, particularly in the distal esophagus. We describe the salient features of CES in adults and identify the feasibility and short-term outcomes of peroral endoscopic myotomy (POEM) for CES.

Methods: In this retrospective, single-center case series, we included 6 patients with a "misdiagnosis" of achalasia established elsewhere, ultimately diagnosed with CES and referred to our institution for POEM. Symptom improvement (clinical success rate), defined as an Eckardt Symptom Score (ESS) of <3 at 2-month follow up was assessed.

Results: Six patients (median age: 40 [range: 18-58] years; 4 males) were included. A long-standing history of dysphagia, ring-shaped stenosis on endoscopic examination, "lopsided hourglass" sign on barium esophagogram, and high-resolution manometry findings indicated by a compartmentalized intrabolus pressure pattern with distinction between the stenotic area and the lower esophageal sphincter were the salient features identified. POEM could not be completed in the first 2 cases due to technical challenges. All subsequent 4 patients who underwent successful POEM, exhibited improved ESS of ≤3 (clinical success rate 100%) at 2 months post-POEM.

Conclusions: Along with identification of salient features on several diagnostic modalities, a differential diagnosis of CES in adults must be considered in patients presenting with long-standing history of dysphagia arising from childhood and persisting into adulthood. Although favorable short-term outcomes of POEM were achieved, further evaluation is still warranted, and an inexperienced operator should not attempt POEM on CES patients due to its technical difficulties.
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http://dx.doi.org/10.20524/aog.2021.0618DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8276369PMC
March 2021

Utility of a new automated diagnostic program in high-resolution esophageal manometry.

J Gastroenterol 2021 Jul 13;56(7):633-639. Epub 2021 May 13.

Department of Gastroenterology and Hepatology, Gunma University Graduate School of Medicine, 3-39-15 Showa-machi, Maebashi, Gunma, 371-8511, Japan.

Background: A new automated diagnostic program for high-resolution esophageal manometry (HREM) has been developed. This diagnostic program could detect locations of landmarks and could make final diagnoses automatically. However, the accuracy of the program is not known. The aim of this study was to evaluate the accuracy of the automated diagnostic program for HREM.

Methods: A total of 445 studies were enrolled. An HREM system (Starlet®) was used, and esophageal motility was diagnosed using the Chicago classification v3.0. First, the locations of the upper esophageal sphincter, transition zone, lower esophageal sphincter, esophago-gastric junction, crural diaphragm and stomach were determined, and each swallow was checked manually. Then, the parameters of the Chicago classification were calculated using an analytic program of the Starlet, and diagnoses were made by three experts. Second, all study raw data were analyzed again by the automated diagnostic program. Diagnoses made by the program were compared to those made by experts to evaluate the accuracy of the diagnoses.

Results: The new diagnostic program could identify the landmarks of each swallow, calculate the parameters and make a final diagnosis within 10 s. The diagnoses made by the automated diagnostic program were not matched to those made by experts in only 10 studies, and the overall accuracy of the new automated diagnostic program thus reached 97.8% (435/445).

Conclusions: The new automated diagnostic program for HREM is clinically useful in terms of high diagnostic accuracy and time-saving.
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http://dx.doi.org/10.1007/s00535-021-01794-zDOI Listing
July 2021

Hemiacetal-less rapamycin derivatives designed and produced by genetic engineering of a type I polyketide synthase.

Sci Rep 2021 05 11;11(1):9944. Epub 2021 May 11.

National Institute of Advanced Industrial Science and Technology (AIST), 2-4-7 Aomi, Koto-ku, Tokyo, Japan.

Engineering polyketide synthases is one of the most promising ways of producing a variety of polyketide derivatives. Exploring the undiscovered chemical space of this medicinally important class of middle molecular weight natural products will aid in the development of improved drugs in the future. In previous work, we established methodology designated 'module editing' to precisely manipulate polyketide synthase genes cloned in a bacterial artificial chromosome. Here, in the course of investigating the engineering capacity of the rapamycin PKS, novel rapamycin derivatives 1-4, which lack the hemiacetal moiety, were produced through the heterologous expression of engineered variants of the rapamycin PKS. Three kinds of module deletions in the polyketide synthase RapC were designed, and the genetically engineered vectors were prepared by the in vitro module editing technique. Streptomyces avermitilis SUKA34 transformed with these edited PKSs produced new rapamycin derivatives. The planar structures of 1-4 established based on 1D and 2D NMR, ESI-TOF-MS and UV spectra revealed that 2 and 3 had skeletons well-matched to the designs, but 1 and 4 did not. The observations provide important insights into the mechanisms of the later steps of rapamycin skeletal formation as well as the ketone-forming oxygenase RapJ.
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http://dx.doi.org/10.1038/s41598-021-88583-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8113240PMC
May 2021

Identification of functional cytochrome P450 and ferredoxin from Streptomyces sp. EAS-AB2608 by transcriptional analysis and their heterologous expression.

Appl Microbiol Biotechnol 2021 May 4;105(10):4177-4187. Epub 2021 May 4.

Technology Research Association for Next-Generation Natural Products Chemistry (N2PC), Aomi, Tokyo, Japan.

Bioconversion using microorganisms and their enzymes is an important tool in many industrial fields. The discovery of useful new microbial enzymes contributes to the development of industries utilizing bioprocesses. Streptomyces sp. EAS-AB2608, isolated from a soil sample collected in Japan, can convert the tetrahydrobenzotriazole CPD-1 (a selective positive allosteric modulator of metabotropic glutamate receptor 5) to its hydroxylated form at the C4-(R) position. The current study was performed to identify the genes encoding the enzymes involved in CPD-1 bioconversion and to verify their function. To identify gene products responsible for the conversion of CPD-1, we used RNA sequencing to analyze EAS-AB2608; from its 8333 coding sequences, we selected two genes, one encoding cytochrome P450 (easab2608_00800) and the other encoding ferredoxin (easab2608_00799), as encoding desirable gene products involved in the bioconversion of CPD-1. The validity of this selection was tested by using a heterologous expression approach. A bioconversion assay using genetically engineered Streptomyces avermitilis SUKA24 ∆saverm3882 ∆saverm7246 co-expressing the two selected genes (strain ES_SUKA_63) confirmed that these gene products had hydroxylation activity with respect to CPD-1, indicating that they are responsible for the conversion of CPD-1. Strain ES_SUKA_63 also showed oxidative activity toward other compounds and therefore might be useful not only for bioconversion of CPD-1 but also as a tool for synthesis of drug metabolites and in optimization studies of various pharmaceutical lead compounds. We expect that this approach will be useful for bridging the gap between the latest enzyme optimization technologies and conventional enzyme screening using microorganisms. KEY POINTS: • Genes easab2608_00800 (cyp) and easab2608_00799 (fdx) were selected by RNA-Seq. • Selection validity was evaluated by an engineered S. avermitilis expression system. • Strain ES_SUKA_63 showed oxidative activity toward CPD-1 and other compounds.
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http://dx.doi.org/10.1007/s00253-021-11304-zDOI Listing
May 2021

Hydroxylation, Epoxidation, and Dehydrogenation of Capsaicin by a Microbial Promiscuous Cytochrome P450 105D7.

Chem Biodivers 2021 Apr 3;18(4):e2000910. Epub 2021 Mar 3.

College of Life Sciences and Medicine, Zhejiang Sci-Tech University, Hangzhou, 310018, P. R. China.

Cytochrome P450 enzymes (P450s) are versatile biocatalysts, which insert a molecular oxygen into inactivated C-H bonds under mild conditions. CYP105D7 from Streptomyces avermitilis has been reported as a bacterial substrate-promiscuous P450 which catalyzes the hydroxylation of 1-deoxypentalenic acid, diclofenac, naringenin, compactin and steroids. In this study, CYP105D7 catalyzes hydroxylation, epoxidation and dehydrogenation of capsaicin, a pharmaceutical agent, revealing its functional diversity. The kinetic parameters of the CYP105D7 oxidation of capsaicin were determined as K =311.60±87.30 μM and k =2.01±0.33 min . In addition, we conducted molecular docking, mutagenesis and substrate binding analysis, indicating that Arg81 plays crucial role in the capsaicin binding and catalysis. To our best knowledge, this study presents the first report to illustrate that capsaicin can be catalyzed by prokaryotic P450s.
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http://dx.doi.org/10.1002/cbdv.202000910DOI Listing
April 2021

Long-term clinical results of per-oral endoscopic myotomy (POEM) for achalasia: First report of more than 10-year patient experience as assessed with a questionnaire-based survey.

Endosc Int Open 2021 Mar 19;9(3):E409-E416. Epub 2021 Feb 19.

Digestive Diseases Center, Showa University Koto Toyosu Hospital, Tokyo, Japan.

Since per-oral endoscopic myotomy (POEM) was introduced in 2010, it has become accepted as one of the standard treatments for esophageal achalasia worldwide. This study aimed to present long-term clinical results of POEM over 10 years and evaluate the technique and outcomes at the institution where it was first used in clinical settings. Questionnaire-based surveys were sent to patients who received POEM in our institution from September 2008 to May 2010. Patient demographics and procedural outcomes and open-ended questions were posed about the postoperative courses, including symptom improvement and recurrence, additional treatments, and post-POEM gastroesophageal reflux disease (GERD) symptoms. Achalasia symptoms and post-POEM GERD symptoms were evaluated with Eckhardt scores and GerdQ systems, respectively.  Thirty-six consecutive POEMs were performed in that period and 10-year follow-up data were obtained from 15 patients (41.7 %). Although four cases (26.7 %) required additional pneumatic balloon dilatation (PBD), reduction in post-Eckardt scores were observed in 14 cases (93.3 %). GerdQ score was positive in one patient (6.7 %). Proton pump inhibitors (PPI) were taken by four patients (26.7 %) and their symptoms were well-controlled.  Clinical results of POEM over 10 years were favorable regardless of various factors. Symptoms improved even in patients who required additional treatments, suggesting that POEM plays a significant role in treatment of achalasia.
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http://dx.doi.org/10.1055/a-1333-1883DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7895648PMC
March 2021

A novel methymycin analog, 12-ketomethymycin N-oxide, produced by the heterologous expression of the large pikromycin/methymycin biosynthetic gene cluster of Streptomyces sp. AM4900.

Biosci Biotechnol Biochem 2021 Mar;85(4):890-894

National Institute of Advanced Industrial Science and Technology, Tokyo, Japan.

A novel methymycin analog, 12-ketomethymycin N-oxide, was produced by the heterologous expression of the pikromycin/methymycin biosynthetic gene cluster of Streptomyces sp. AM4900 together with 12-ketomethymycin, which was only isolated by the biotransformation of the synthetic intermediate before. Their structures were determined by the spectroscopic data and the chemical derivatization. 12-Ketomethymycin showed a weak cytotoxicity against SKOV-3 and Jurkat cells, although its N-oxide analog did not show any activity. Both showed no antibacterial activities against Escherichia coli and Micrococcus luteus.
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http://dx.doi.org/10.1093/bbb/zbaa111DOI Listing
March 2021

Simplified endoscopic pressure study integrated system for the diagnosis of gastroesophageal reflux disease.

Dig Endosc 2021 May 22;33(4):663-667. Epub 2021 Mar 22.

Digestive Diseases Center, Showa University Koto Toyosu Hospital, Tokyo, Japan.

Endoscopic pressure study integrated system (EPSIS) is a novel tool for the diagnosis of gastroesophageal reflux disease. It enables the evaluation of the function of the lower esophageal sphincter by monitoring intragastric pressure (IGP) while insufflating the stomach during esophagogastroduodenoscopy. EPSIS can predict abnormal acid reflux with high accuracy based on previous studies. IGP was measured by inserting through the working channel of the scope an intragastric catheter connected to a pressure measuring device. Herein, we assess the feasibility of an updated EPSIS system, which can be performed just by connecting a flush tube to the working channel. This method does not require inserting foreign objects in the stomach and spares catheter insertion in order to simplify the procedure and reduce costs. A single-center pilot study was conducted to evaluate the association between catheter-based EPSIS and the updated EPSIS. The results of EPSIS in 20 patients who underwent both methods were assessed. In all cases, the waveform pattern of IGP measured by catheter-based EPSIS and updated EPSIS was consistent with 15 uphill pattern and five flat pattern. Intraobserver agreement of waveform pattern was perfect between two examiners with kappa value = 1. Intraclass correlation coefficient (ICC) for intraobserver reliability for maximum IGP was excellent with 0.91 (95% confidence interval [CI] of 0.77 < ICC < 0.96) and for pressure gradient was also good with 0.89 (95% CI of 0.71 < ICC < 0.95). In conclusion, our study suggests that the updated EPSIS can be performed without the use of a catheter.
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http://dx.doi.org/10.1111/den.13947DOI Listing
May 2021

A novel endoscopic purse-string suture technique, "loop 9", for gastrointestinal defect closure: a pilot study.

Endoscopy 2021 Jan 20. Epub 2021 Jan 20.

Digestive Diseases Center, Showa University Koto Toyosu Hospital, Tokyo, Japan.

BACKGROUND : This study aimed to assess the feasibility and efficacy of the novel loop 9 method of gastrointestinal (GI) defect closure. METHODS : 20 patients underwent a GI procedure that required defect closure. Loop 9 can be delivered through a single instrument channel (3.2 mm) and released at the defect site. After it has been anchored by two clips positioned on opposite sides of the defect edge, the loop 9 is tightened by pulling the end of the suture intraluminally using biopsy forceps. Additional clips are placed to achieve complete closure. The primary outcome was complete closure rate. The secondary outcomes were closure time, sustained closure rate, and adverse events. RESULTS : Complete closure was achieved in 100 % of cases. The mean size of the mucosal defects was 17.5 mm (range 10-55 mm). The median closure time was 14 minutes. The sustained closure rate was 90 %. No adverse events were noted. CONCLUSIONS : The loop 9 technique is feasible and effective in achieving complete and sustained closure of therapeutic endoscopy-related GI defects.
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http://dx.doi.org/10.1055/a-1364-4160DOI Listing
January 2021

Diagnostic yield of fourth-generation endocytoscopy for esophageal squamous lesions using a modified endocytoscopic classification.

Dig Endosc 2020 Dec 15. Epub 2020 Dec 15.

Digestive Disease Center, Showa University Koto, Tokyo, Japan.

Objectives: Endocytoscopy (EC) is an ultra-high magnification endoscopy designed to provide in vivo histologic assessment. This study aimed to evaluate the diagnostic yield of the newly developed fourth-generation EC for esophageal squamous lesions by using a modified EC classification.

Methods: A total of 2548 EC images of 57 esophageal targeted areas between June 2015 and October 2017 were retrospectively collected. Two lesions with low-quality images were excluded. Only EC images were independently reviewed by two expert and two non-expert endoscopists. The lesions were classified according to a three-tier modified EC classification. We used a multilevel logistic regression to analyze the data.

Results: The sensitivity and specificity of diagnosing non-squamous cell cancer (SCC) vs SCC were 82.5% and 83.0% by the experts; 90.1% and 75.0% by non-experts. The interobserver agreement among the four raters was good (kappa statistic 0.59). The diagnostic accuracy of experts and non-experts was similar (P = 0.16 for specificity and P = 0.20 for sensitivity). The sensitivity and specificity of EC for non-neoplasia vs neoplasia were 88.7% and 74.6% by experts; 90.3 and 52.1% by non-experts. The interobserver agreement among the four raters was moderate (kappa statistic 0.44). The specificity of experts was higher compared to non-experts, although the difference did not reach statistical significance (P = 0.08 for specificity and P = 0.93 for sensitivity).

Conclusions: Fourth-generation EC offers acceptable diagnostic accuracy and reliability in both experts and non-experts, especially when diagnosing SCC lesions.
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http://dx.doi.org/10.1111/den.13914DOI Listing
December 2020

Autophagy-related 16-like 1 is influenced by human herpes virus 1-encoded microRNAs in biopsy samples from the lower esophageal sphincter muscle during per-oral endoscopic myotomy for esophageal achalasia.

Biomed Rep 2021 Jan 30;14(1). Epub 2020 Oct 30.

Division of Medicine and Clinical Science, Faculty of Medicine, Tottori University, Yonago, Tottori 683-8504, Japan.

Esophageal achalasia is characterized by abnormal peristalsis of the esophageal body and impaired relaxation of the lower esophageal sphincter (LES); however, its etiology remains unknown. One of the potential causes of esophageal achalasia is herpes simplex virus type 1 (HSV-1). Following infection with HSV-1, a complex interaction between the autoimmune and inflammatory responses is initiated. Viral microRNAs (miRNAs/miRs) serve a crucial role in this interaction. In the present study, the expression of E3 ubiquitin-protein ligase component n-recognition 1 () and autophagy-related 16-like 1 () was assessed in patients with sporadic and classic achalasia as potential targets of the viral miRNAs. We assessed the mRNA levels of target transcripts using reverse transcription-quantitative PCR. expression was slightly decreased, although the difference was not significant. However, expression was significantly decreased in the LES. In conclusion, expression was reduced in the LES of achalasia patients; therefore, might be a target of HSV1-miR-H1, and its reduction could be related to the disease mechanism.
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http://dx.doi.org/10.3892/br.2020.1383DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7678595PMC
January 2021

Heterologous Expression of the Biosynthetic Gene Cluster for Verticilactam and Identification of Analogues.

J Nat Prod 2020 12 20;83(12):3598-3605. Epub 2020 Nov 20.

RIKEN Center for Sustainable Resource Science, Natural Product Biosynthesis Research Unit, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan.

Verticilactam and the new geometric isomers, verticilactams B and C, were produced by heterologous expression of the biosynthetic gene cluster for verticilactam using the SUKA17 strain. Only verticilactam, a compound with a characteristic β-ketoamide unit within a 16-membered polyketide macrolactam conjugated with an octalin skeleton, had been previously reported having been isolated from JC-8444. In this report, minor verticilactam derivatives were isolated from the transformed strain, and their structures elucidated by spectral analysis. Verticilactam B was a geometric isomer at Δ and Δ, and verticilactam C was the Δ and Δ isomer. In addition, the absolute configuration of verticilactam was confirmed by ECD analysis and NMR chemical shifts. The stereochemistry assignments of the hydroxy groups at C-10 and C-12 were supported by the domain organization of the polyketide synthase identified in the verticilactam gene cluster. Verticilactam showed moderate activity against the malaria parasite 3D7 strain with no significant cytotoxicity or antimicrobial effects.
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http://dx.doi.org/10.1021/acs.jnatprod.0c00755DOI Listing
December 2020

Characterization of intragastric pressure waveform in endoscopic pressure study integrated system: Novel diagnostic device for gastroesophageal reflux disease.

Dig Endosc 2021 Jul 8;33(5):780-787. Epub 2020 Dec 8.

Digestive Diseases Center, Showa University Koto Toyosu Hospital, Tokyo, Japan.

Objectives: Endoscopic pressure study integrated system (EPSIS) is a novel diagnostic tool for gastroesophageal reflux disease (GERD). EPSIS has been developed to evaluate lower esophageal sphincter function by monitoring the intragastric pressure (IGP) while insufflating the stomach during gastroscopy. Based on previous data, EPSIS could diagnose GERD with good accuracy by assessing IGP waveform pattern. This study aimed to further characterize the waveform to improve the diagnostic yield of EPSIS.

Methods: We conducted a retrospective analysis of patients with typical GERD symptoms who underwent both EPSIS and 24-h impedance-pH monitoring (MII-pH) at a single tertiary referral center from October 2018 to May 2020. EPSIS was performed by using a through-the-scope catheter connected to the pressure measuring system (TR-W550, TR-TeH08, AP-C35; Keyence, Osaka, Japan) to monitor IGP. Abnormal acid reflux was defined as acid exposure time (AET) over 6.0%. Pressure waveform was characterized as follows: (i) Basal IGP, (ii) Maximum IGP, (iii) Pressure difference, (iv) Gradient of the waveform.

Results: A total of 57 patients with GERD symptoms were analyzed. Twenty-one patients presented abnormal AET on MII-pH. Among EPSIS parameters, pressure difference during insufflation correlated with AET (ρ = -0.66, P < 0.01) and showed the best diagnostic accuracy for AET with the cutoff value of 4.7 mmHg (area under the curve [AUC], 0.87). The gradient of EPSIS waveform also revealed good diagnostic accuracy for abnormal AET with the cutoff value of 0.07 mmHg/s (AUC, 0.81).

Conclusions: Endoscopic pressure study integrated system waveform parameters, especially pressure difference, presented high diagnostic accuracy for the presence of abnormal acid reflux.
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http://dx.doi.org/10.1111/den.13867DOI Listing
July 2021

Modifications to central carbon metabolism in an engineered Streptomyces host to enhance secondary metabolite production.

J Biosci Bioeng 2020 Dec 4;130(6):563-570. Epub 2020 Sep 4.

Laboratory of Microbial Engineering, Ōmura Satoshi Memorial Institute, Kitasato University, Sagamihara, Kanagawa 252-0373, Japan. Electronic address:

To improve the production of secondary metabolites by alternation of the carbon metabolic flux, two types of deletion mutants of the central metabolic pathway, the Embden-Meyerhof (EM) or pentose phosphate (PP) pathway, in the genetically engineered Streptomyces avermitilis were constructed. Double-deletion mutants of phosphofructokinase (ΔpfkA1ΔpfkA3) in the EM pathway carrying a gene cluster for chloramphenicol biosynthesis markedly increased chloramphenicol production synthesized through the shikimate pathway. Although the ΔpfkA1ΔpfkA3 double-deletion mutant grew more slowly, its specific productivity of chloramphenicol (per dry cell weight) was 2.0-fold higher than that of the engineered S. avermitilis strain. However, the productivity of chloramphenicol was lower by the double-deletion mutant of transaldolase in the PP pathway, which supplies the precursor of the shikimate pathway. A carbon-flux analysis of the EM and PP pathways using [1-C] glucose revealed that carbon flux in the ΔpfkA1ΔpfkA3 double-deletion mutant increased through the PP pathway, which enhanced the production of chloramphenicol. These results suggest that a metabolic modification approach has the potential to increase the titers and yields of valuable secondary metabolites.
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http://dx.doi.org/10.1016/j.jbiosc.2020.08.006DOI Listing
December 2020

In vitro Cas9-assisted editing of modular polyketide synthase genes to produce desired natural product derivatives.

Nat Commun 2020 08 11;11(1):4022. Epub 2020 Aug 11.

National Institute of Advanced Industrial Science and Technology (AIST), 2-4-7 Aomi, Koto-ku, Tokyo, Japan.

One major bottleneck in natural product drug development is derivatization, which is pivotal for fine tuning lead compounds. A promising solution is modifying the biosynthetic machineries of middle molecules such as macrolides. Although intense studies have established various methodologies for protein engineering of type I modular polyketide synthase(s) (PKSs), the accurate targeting of desired regions in the PKS gene is still challenging due to the high sequence similarity between its modules. Here, we report an innovative technique that adapts in vitro Cas9 reaction and Gibson assembly to edit a target region of the type I modular PKS gene. Proof-of-concept experiments using rapamycin PKS as a template show that heterologous expression of edited biosynthetic gene clusters produced almost all the desired derivatives. Our results are consistent with the promiscuity of modular PKS and thus, our technique will provide a platform to generate rationally designed natural product derivatives for future drug development.
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http://dx.doi.org/10.1038/s41467-020-17769-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7419507PMC
August 2020

Importance of second-look endoscopy after per-oral endoscopic myotomy for safe postoperative management.

Dig Endosc 2021 Mar 2;33(3):364-372. Epub 2020 Sep 2.

Digestive Diseases Center, Showa University Koto Toyosu Hospital, Tokyo, Japan.

Objectives: Per-oral endoscopic myotomy (POEM) is a safe and effective treatment for achalasia and esophageal motility disorders. The role of second-look endoscopy (SE) on postoperative day 1 has not been examined. This study aimed to evaluate the findings and need of SE after POEM.

Methods: This is a single-center, retrospective study. All consecutive patients who underwent POEM and SE on postoperative day 1 between December 2017 and September 2019 were included. The primary endpoint was the rate of newly-detected adverse events (nAE) during SE that required endoscopic intervention or deviation from the normal postoperative course. Multivariate logistic regression was used to identify predictors of nAE.

Results: Four-hundred-ninety-seven patients (mean age, 50.3 years; female, 49.9%) were included. SE identified abnormal findings in a total of 71 patients (14.3%). nAE which required endoscopic intervention or deviation from the normal postoperative course were identified in 12 patients (2.4%): eight (1.6%) entry site dehiscence; two (0.4%) submucosal hemorrhage or hematoma; and two (0.4%) dehiscence of an intraoperative perforation site after endoclip closure. Other findings such as mucosal thermal damage without perforation and small submucosal hematoma were found in 54 patients (10.9%) and five patients (1.0%), respectively. Multivariate analysis showed that longer operation time and intraoperative adverse events (AE) were associated with clinically significant nAE during SE.

Conclusions: Second-look endoscopy can detect and treat nAE that may lead to severe AE. Thus, SE should be highly considered before starting oral ingestion in all cases, and especially in those who present an intraoperative AE and longer operation time.
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http://dx.doi.org/10.1111/den.13770DOI Listing
March 2021

Identification, cloning and heterologous expression of biosynthetic gene cluster for desertomycin.

J Antibiot (Tokyo) 2020 09 26;73(9):650-654. Epub 2020 May 26.

National Institute of Advanced Industrial Science and Technology, 2-4-7 Aomi, Koto-ku, Tokyo, 135-0064, Japan.

From our in-house microbial genome database of secondary metabolite producers, we identified a candidate biosynthetic gene cluster for desertomycin from Streptomyces nobilis JCM4274. We report herein the cloning of the 127-kb entire gene cluster for desertomycin biosynthesis using bacterial artificial chromosome vector. The entire biosynthetic gene cluster for desertomycin was introduced in the heterologous host, Streptomyces lividans TK23, with an average yield of more than 130 mg l.
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http://dx.doi.org/10.1038/s41429-020-0319-0DOI Listing
September 2020

Endoscopic treatment of proton pump inhibitor-refractory gastroesophageal reflux disease with anti-reflux mucosectomy: Experience of 109 cases.

Dig Endosc 2021 Mar 11;33(3):347-354. Epub 2020 Aug 11.

Digestive Diseases Center, Showa University Koto-Toyosu Hospital, Tokyo, Japan.

Objectives: Some patients with gastroesophageal reflux disease (GERD) are refractory to proton pump inhibitor (PPI) therapy. Anti-reflux mucosectomy (ARMS) is a minimally invasive endoscopic procedure for treatment of GERD. In this study, we retrospectively evaluated the outcomes of ARMS performed in patients with PPI-refractory GERD at our institution.

Methods: A total of 109 patients with PPI-refractory GERD who underwent ARMS were retrospectively reviewed. Pre- and post-ARMS questionnaire scores, acid exposure time (AET), DeMeester score, proximal extent, and PPI discontinuation rate were compared.

Results: There was a significant improvement in the symptom score (P < 0.01) and 40-50% of patients were able to discontinue PPI after ARMS. In patients who were followed up for 3 years, sustained improvement in subjective symptoms was observed. AET and DeMeester score significantly improved after ARMS (P < 0.01); however, there was no significant improvement in proximal extent (P = 0.0846).

Conclusions: Anti-reflux mucosectomy is an effective minimally invasive therapy for patients with PPI-refractory GERD. The therapeutic efficacy is attributable to suppression of acid backflow due to contraction of the scar tissue in cardia.
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http://dx.doi.org/10.1111/den.13727DOI Listing
March 2021

Endoscopic pressure study integrated system reflects gastroesophageal junction competence in patients with erosive esophagitis and Barrett´s esophagus.

Dig Endosc 2020 Nov 29;32(7):1050-1056. Epub 2020 Mar 29.

Digestive Diseases Center, Showa University Koto Toyosu Hospital, Showa University, Tokyo, Japan.

Objectives: The endoscopic pressure study integrated system (EPSIS) is a novel diagnostic tool for gastroesophageal reflux disease (GERD) by monitoring intragastric pressure (IGP). Evaluation of the lower esophageal sphincter (LES) function may be achieved endoscopically by utilizing this newly developed diagnostic tool. This study aimed to evaluate the association between EPSIS results and gastroesophageal reflux-related diseases, e.g., erosive esophagitis (EE) and Barrett's esophagus (BE).

Methods: This was a retrospective, single-center study. All patients who underwent EPSIS between November 2016 and July 2018 were included. EPSIS was performed during esophagogastroduodenoscopy with a dedicated electronic device and a through-the-scope catheter. The maximum IGP (IGP-max) and IGP waveform pattern (flat or uphill) were recorded with this system. Evaluation of an EE and BE was based on the Los Angeles classification and Prague classification, respectively.

Results: A total of 104 patients were enrolled; 29 (28%) had EE and 42 (40%) had BE. Patients with EE had lower IGP-max values (16.0 vs 18.8 mmHg, P = 0.01) and an EPSIS flat pattern was seen more frequently (82.8% vs 37.3%, P < 0.001). Similarly, patients with BE displayed a lower IGP-max (15.7 vs 19.6 mmHg, P < 0.001) and presented with an EPSIS flat pattern in a higher proportion (69% vs 37.1%, P < 0.001). These differences remained significant on multivariate analysis.

Conclusions: The EPSIS, as a novel diagnostic tool, was shown to exhibit a relation with EE and BE, implying that EPSIS is a promising modality to evaluate gastroesophageal reflux-related diseases and LES function endoscopically.
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http://dx.doi.org/10.1111/den.13644DOI Listing
November 2020

Anti-reflux mucosal ablation (ARMA) as a new treatment for gastroesophageal reflux refractory to proton pump inhibitors: a pilot study.

Endosc Int Open 2020 Feb 22;8(2):E133-E138. Epub 2020 Jan 22.

Digestive Diseases Center, Showa University Koto Toyosu Hospital, Tokyo, Japan.

 The incidence of proton pump inhibitor (PPI)-refractory gastroesophageal reflux disease (GERD) has been increasing. While surgical intervention with Laparoscopic Nissen Fundoplication remains the gold standard, less invasive anti-reflux interventions are desired. We have developed a minimally invasive anti-reflux mucosal ablation (ARMA) treatment. Herein, we report its technical details and describe its feasibility, safety, and efficacy in PPI-refractory GERD.  We conducted a prospective single-center single-arm interventional trial evaluating the outcome of ARMA in 12 patients with PPI-refractory GERD. GERD-Health Related Quality of Life Questionnaire (GERD-HRQL) evaluation, Frequency Scale for the Symptoms of GERD (FSSG) assessment, and impedance-pH monitoring were performed at baseline and at 2 months post-ARMA.  A total of 12 patients underwent ARMA with a median follow-up duration of 9 months (range: 6 - 14 months). Median GERD-HRQL score significantly improved from 30.5 to 12 (  = 0.002); median FSSG score significantly improved from 25 to 10.5 (  = 0.002), and median DeMeester score decreased from 33.5 to 2.8 (  = 0.049) at 2 months follow-up. No immediate complications were observed.  Our pilot study has shown that ARMA, a new endoscopic treatment for PPI-refractory GERD, is simple, safe, and improves GERD-related symptoms and objective acid reflux parameters.
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http://dx.doi.org/10.1055/a-1031-9436DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6976329PMC
February 2020

Novel macrolactam compound produced by the heterologous expression of a large cryptic biosynthetic gene cluster of Streptomyces rochei IFO12908.

J Antibiot (Tokyo) 2020 03 18;73(3):171-174. Epub 2019 Dec 18.

National Institute of Advanced Industrial Science and Technology, 2-4-7 Aomi, Koto-ku, Tokyo, 135-0064, Japan.

In the course of our studies on the heterologous expression of giant biosynthetic genes, we discovered a novel cryptic biosynthetic gene cluster in Streptomyces rochei IFO12908. During our efforts to express biosynthetic genes using the host SUKA strain derived from Streptomyces avermitilis, a novel polyene macrolactam compound designated as JBIR-156 was produced. We report herein the cloning and heterologous expression of the JBIR-156 biosynthetic gene cluster, and the isolation, structure determination, and cytotoxic activity of this novel compound.
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http://dx.doi.org/10.1038/s41429-019-0265-xDOI Listing
March 2020

Production of distinct labdane-type diterpenoids using a novel cryptic labdane-like cluster from Streptomyces thermocarboxydus K155.

Appl Microbiol Biotechnol 2020 Jan 5;104(2):741-750. Epub 2019 Dec 5.

Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, 04510, Ciudad de México, México.

Bioinformatic mining of the Streptomyces thermocarboxydus K155 genome predicted the presence of four synthases for the production of geosmin, hopene, albaflavenone, and a type B-type A diterpenoid system like that described for labdane-related diterpenoids (LRD). The lrd cluster was comprised by an operon of four genes (lrdABDC). This cluster seemed to be silent in the wild-type strain, as neither labdane nor terpene-like compounds were detected by UPLC-TOF-MS and GC-MS analyses in both culture supernatants and mycelial extracts. Heterologous expression of the lrdABDC cluster in a defective cyslabdan producer (Streptomyces cyslabdanicus K04-0144Δcld) generated 8,17-epoxy-7-hydroxy labda-12,14-diene and cyslabdan. The same cluster expressed in the strains Streptomyces coelicolor M1152, Streptomyces peucetius var. caesius, and Streptomyces avermitilis SUKA22 produced the general intermediary labda-8(17), 12(E),14-triene [(E)-biformene]. Besides (E)-biformene, S. coelicolor M1152 and S. avermitilis SUKA22 produced two and three different labdane-type diterpenoids, underlying the relevance of the genetic background of the Streptomyces host in product formation.
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http://dx.doi.org/10.1007/s00253-019-10240-3DOI Listing
January 2020

Combination of laparoscopic and endoscopic approaches for neoplasia with non-exposure technique (CLEAN-NET) for gastric submucosal tumors: updated advantages and limitations.

Ann Transl Med 2019 Oct;7(20):582

Digestive Diseases Center, Showa University Koto Toyosu Hospital, Tokyo, Japan.

Laparoscopic and endoscopic cooperative surgery (LECS) for gastric submucosal tumors (SMTs) has been developed under the concept of resecting gastric tumors with both complete curability and preserving organ functions. Precise resection is obtained by classical LECS, however, concerns regarding intraoperative bacterial infection and dissemination of the tumor cells into the abdominal cavity by LECS with exposure technique still remain. To prevent these concerns, several LECS-related procedures with non-exposure techniques, such as combination of laparoscopic and endoscopic approaches for neoplasia with non-exposure technique (CLEAN-NET) and non-exposed endoscopic wall-inversion surgery (NEWS), have been reported to be safe and feasible. Classical LECS, CLEAN-NET, and NEWS have the same concept, however, each has its own different characteristic procedures; exposure or non-exposure technique, inversion of the tumor into or outer the lumen, retrieval of tumor per oral or through the abdominal cavity, and dominance in the role of the endoscopist or the laparoscopic surgeon. Familiarization with these procedure details is important to understand their indications, advantages and limitations, resulting in providing a tailored minimally invasive surgery for patients. The main scope of this review article is to introduce readers to the clinical application, procedure, and results of CLEAN-NET, both from previous literatures and from our experience, as well as to offer a closer look at its advantages and limitations while comparing with other LECS procedures from the viewpoint of introducing CLEAN-NET first.
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http://dx.doi.org/10.21037/atm.2019.09.19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6861760PMC
October 2019

An Unusual Extender Unit Is Incorporated into the Modular Polyketide Synthase of Scopranones Biosynthesis.

Biochemistry 2019 12 6;58(50):5066-5073. Epub 2019 Dec 6.

Kitasato Institute for Life Sciences , Kitasato University , 1-15-1 Kitasato, Minami-ku , Sagamihara , Kanagawa 252-0373 , Japan.

Scopranones, produced by sp. BYK-11038, are the novel bone morphogenetic protein inhibitors characterized by atypical two scoop-like moieties and a 3-furanone moiety. Two scoop-like moieties connected to a 3-furanone have not previously been reported in natural products, and their biosynthesis must occur via a unique pathway. Feeding experiments using C-labeled precursors indicated that scopranones were synthesized from three acetates and three butyrates in polyketide-type biosynthesis. Genome mining of sp. BYK-11038 revealed that the candidate biosynthetic gene cluster contains 21 open reading frames (ORFs), including three modular polyketide synthases (PKSs; SprA, SprB, and SprC), which were composed of 4 modules with one loading module and 18 additional ORFs (SprD to SprU) spanning a distance of 55 kbp. The characterization of in-frame deletion mutants and feeding experiments with the predicted extender units indicated that two genes, and , encoding discrete 3-oxoacyl-ACP synthases, and a gene, , encoding crotonyl-CoA reductase, were involved in assembling an unusual C branched extender unit, 2-(2-ethylbutyl)malonyl-CoA. Additionally, three ORFs, , , and , encoding cytochrome P450s and a monooxygenase, are important tailoring enzymes in post-PKS modification. SprT is an essential enzyme for decarboxylative ring contraction via oxidation, which converts the 2-pyranone to a 3-furanone.
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http://dx.doi.org/10.1021/acs.biochem.9b00908DOI Listing
December 2019

A novel endoscopic assessment of the gastroesophageal junction for the prediction of gastroesophageal reflux disease: a pilot study.

Endosc Int Open 2019 Nov 23;7(11):E1468-E1473. Epub 2019 Oct 23.

Digestive Diseases Center, Showa University Koto Toyosu Hospital, Tokyo 135-8577, Japan.

 Hiatal hernia and lower esophageal sphincter (LES) dysfunction play major roles in gastroesophageal reflux disease (GERD) pathogenesis. We developed a novel endoscopic assessment to evaluate the gastroesophageal junction (GEJ). This study aims to evaluate the feasibility of this method for the diagnostic prediction of GERD.  A retrospective analysis of patients with GERD symptoms who underwent gastroscopy and esophageal pH-impedance monitoring was conducted. The novel assessment evaluated the following in retroflex view: 1) Cardiac Opening (CO): diameter of the opening of the cardia, 2) Sliding Hernia (SH): length from the diaphragmatic crus to the squamocolumnar junction, 3) Scope Holding Time% (SHT%): the percentage of time that the Scope Holding Sign (SHS) was observed out of 30 seconds. The SHS is defined as the lower esophagus holding the endoscope under excessive insufflation. The results of this assessment and that of pH-impedance monitoring were compared.  In total, 61 patients (mean age ± SD, 54.1 ± 16.4 years, 32 males) were enrolled. CO and SH were significantly correlated with acid exposure time (AET) (ρ = 0.36,  = 0.005, and ρ = 0.36,  = 0.004). The optimal cutoff of CO for AET > 6 % was 3 cm (Sensitivity = 72.4 %, Specificity = 46.9 %, AUC = 0.64) and that of SH was 2 cm (Sensitivity = 55.2 %, Specificity = 75.0 %, AUC = 0.70). When the population was stratified according to this cutoff, patients with CO > 3 cm and those with SH > 2 cm presented higher AET (15.1 vs 4.1 %,  = 0.037, and 23.0 vs 3.6 %,  = 0.026). Optimal cutoff of SHT% for the number of all reflux episodes > 80 was 75 % (Sensitivity = 81.8 %, Specificity = 54.6%, AUC = 0.67). Patients with SHT% < 75 % presented a higher number of all reflux episodes (88 vs 65,  = 0.014). Sensitivity, specificity, and accuracy of SHT% < 75 % for all reflux episodes > 80 were 81.8 % (95 %CI: 67.7 - 91.8), 54.5% (95 %CI: 40.4 - 64.5), and 68.2 % (95 %CI: 54.0 - 78.1).  This novel endoscopic assessment of GEJ significantly predicted the presence of GERD and merits further testing in future studies.
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http://dx.doi.org/10.1055/a-0990-9737DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6811351PMC
November 2019

Hydroxylation of Steroids by a Microbial Substrate-Promiscuous P450 Cytochrome (CYP105D7): Key Arginine Residues for Rational Design.

Appl Environ Microbiol 2019 12 14;85(23). Epub 2019 Nov 14.

College of Life Sciences and Medicine, Zhejiang Sci-Tech University, Hangzhou, Zhejiang, China

Our previous study showed that CYP105D7, a substrate-promiscuous P450, catalyzes the hydroxylation of 1-deoxypentalenic acid, diclofenac, naringenin, and compactin. In this study, 14 steroid compounds were screened using recombinant cells harboring genes encoding CYP105D7 and redox partners (Pdx/Pdr, RhFRED, and FdxH/FprD), and the screening identified steroid A-ring 2β- and D-ring 16β-hydroxylation activity. Wild-type CYP105D7 was able to catalyze the hydroxylation of five steroids (testosterone, progesterone, 4-androstene-3,17-dione, adrenosterone, and cortisone) with low (<10%) conversion rates. Structure-guided site-directed mutagenesis of arginine residues around the substrate entrance and active site showed that the R70A and R190A single mutants and an R70A/R190A double mutant exhibited greatly enhanced conversion rates for steroid hydroxylation. For the conversion of testosterone in particular, the R70A/R190A mutant's / values increased 1.35-fold and the conversion rates increased significantly by almost 9-fold with high regio- and stereoselectivity. Molecular docking analysis revealed that when Arg70 and Arg190 were replaced with alanine, the volume of the substrate access and binding pocket increased 1.08-fold, which might facilitate improvement of the hydroxylation efficiency of steroids. Cytochrome P450 monooxygenases (P450s) are able to introduce oxygen atoms into nonreactive hydrocarbon compounds under mild conditions, thereby offering significant advantages compared to chemical catalysts. Promiscuous P450s with broad substrate specificity and reaction diversity have significant potential for applications in various fields, including synthetic biology. The study of the function, molecular mechanisms, and rational engineering of substrate-promiscuous P450s from microbial sources is important to fulfill this potential. Here, we present a microbial substrate-promiscuous P450, CYP105D7, which can catalyze hydroxylation of steroids. The loss of the bulky side chains of Arg70 and Arg190 in the active site and substrate entrance resulted in an up to 9-fold increase in the substrate conversion rate. These findings will support future rational and semirational engineering of P450s for applications as biocatalysts.
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http://dx.doi.org/10.1128/AEM.01530-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6856325PMC
December 2019

Gastric myotomy length affects severity but not rate of post-procedure reflux: 3-year follow-up of a prospective randomized controlled trial of double-scope per-oral endoscopic myotomy (POEM) for esophageal achalasia.

Surg Endosc 2020 07 28;34(7):2963-2968. Epub 2019 Aug 28.

Digestive Disease Center, Showa University Koto Toyosu Hospital, Tokyo, Japan.

Background: Since Inoue performed the first POEM in 2008, safety and efficacy have been well-established. Early studies focused on refining the technique and avoiding incomplete myotomy. Following the discovery that many patients with abnormal acid exposure are asymptomatic, the focus shifted to post-POEM reflux, but no studies have identified any associated procedural factors. In this study, we examined the intermediate-term results of our previous randomized controlled trial, with particular attention to post-POEM reflux.

Methods: Previously, 100 consecutive patients were randomized to either double- or single-scope POEM. Endoscopy was conducted 2 months post-POEM and annually thereafter. Patients were included in the present study if they completed endoscopy ≥ 6 months post-POEM, and the clinical results of both groups were analyzed with particular attention to clinical efficacy and post-POEM reflux.

Results: Median follow-up was 3 years, and most myotomies were performed in the posterior location. The final gastric myotomy length was longer in the double-scope group (3.3 vs. 2.6 cm). Clinical efficacy (≥ 80%) and rates of post-POEM reflux (~ 60%) were similar; however, there was a higher incidence of moderate esophagitis (Los Angeles Grade B) in the double-scope group (25% vs. 4%). There were no cases of severe esophagitis (Los Angeles Grade C/D). Among patients with normal endoscopy at 2 months, > 40% developed erosive esophagitis on intermediate-term follow-up.

Conclusions: This is the first study to demonstrate a procedural factor that increases post-POEM esophagitis. Gastric myotomy > 2.5 cm results in increased rates of moderate esophagitis without improving clinical efficacy. Some patients developed esophagitis in a delayed fashion, emphasizing the importance of ongoing surveillance. We also believe that preserving the gastric sling fibers may help to reduce reflux rates. The double-scope method may help to control myotomy length (2.0-2.5 cm) and direction (lesser curve to avoid the gastric sling) to help maximize clinical efficacy while minimizing post-POEM reflux.
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http://dx.doi.org/10.1007/s00464-019-07079-0DOI Listing
July 2020

Utilizing fourth-generation endocytoscopy and the 'enlarged nuclear sign' for in vivo diagnosis of early gastric cancer.

Endosc Int Open 2019 Aug 8;7(8):E1002-E1007. Epub 2019 Aug 8.

Digestive Diseases Center, Showa University Koto Toyosu Hospital, Tokyo, Japan.

 Fourth-generation endocytoscopy is an ultra-high magnification endoscopic technique designed to provide excellent quality in vivo histologic assessment of gastrointestinal lesions. This study aims to evaluate the diagnostic accuracy of endocytoscopy in early gastric cancer diagnosis.  A single-center, retrospective analysis of prospectively collected data from all gastric endocytoscopic examinations was conducted. Two expert endoscopists, blinded to white-light and narrow-band imaging findings as well as histopathologic diagnosis, independently reviewed and diagnosed all endocytoscopic images. A newly recognized "enlarged nuclear sign" was detected, and its implication in early gastric cancer diagnosis was evaluated. The diagnostic performance of fourth-generation endocytoscopy was assessed while using the gold standard histopathology as a reference.  Forty-three patients (mean age±SD, 72.6 ± 12.1 years; 31 males) were enrolled. Based on histopathology, 23 had well-differentiated adenocarcinomas, four adenomas, and 16 non-neoplastic lesions. The sensitivity, specificity, and accuracy of fourth-generation endocytoscopy for gastric cancer diagnosis were 87.0 % (95 % CI: 67.9 - 95.5), 80.0 % (95 % CI: 58.4 - 91.9), and 83.7 % (95 % CI: 70.0 - 91.9) by endoscopist A; and 91.3 % (95 % CI: 73.2 - 97.6), 75.0 % (95 % CI: 53.1 - 88.8), and 83.7 % (95 % CI: 70.0 - 91.9) by endoscopist B. The inter-observer agreement, statistic = 0.71 (95 % CI: 0.50 - 0.93), was good. The sensitivity, specificity, and accuracy of the enlarged nuclear sign for early gastric cancer diagnosis were 87.0 % (95 % CI: 67.9 - 95.5), 95.0 % (95 % CI: 76.4 - 99.1), and 90.7 % (95 % CI: 78.4 - 96.3) by endoscopist A; and 82.6 % (95 % CI: 62.9 - 93.0), 85.0 % (95 % CI: 64.0 - 94.8), and 83.7 % (95 % CI: 70.0 - 91.9) by endoscopist B. The inter-observer agreement, statistic = 0.68 (95 % CI: 0.51 - 0.89) was good. Fourth-generation endocytoscopy appears to aid in the diagnosis of early gastric cancer, particularly well-differentiated adenocarcinomas, due to its good diagnostic accuracy and identification of the "enlarged nuclear sign," and deserves further evaluation in future studies.
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http://dx.doi.org/10.1055/a-0957-2866DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6687508PMC
August 2019

Mechanism of Action of Prethioviridamide, an Anticancer Ribosomally Synthesized and Post-Translationally Modified Peptide with a Polythioamide Structure.

ACS Chem Biol 2019 08 31;14(8):1819-1828. Epub 2019 Jul 31.

Chemical Genomics Research Group , RIKEN Center for Sustainable Resource Science , Saitama 351-0198 , Japan.

Thioviridamide, prethioviridamide, and JBIR-140, which are ribosomally synthesized and post-translationally modified peptides (RiPPs) possessing five thioamide bonds, induce selective apoptosis in various cancer cells, especially those expressing the adenovirus oncogene E1A. However, the target protein of this unique family of bioactive compounds was previously unknown. To investigate the mechanism of action, we adopted a combined approach of genome-wide shRNA library screening, transcriptome profiling, and biochemical identification of prethioviridamide-binding proteins. An shRNA screen identified 63 genes involved in cell sensitivity to prethioviridamide, which included translation initiation factors, aminoacyl tRNA synthetases, and mitochondrial proteins. Transcriptome profiling and subsequent analysis revealed that prethioviridamide induces the integrated stress response (ISR) through the GCN2-ATF4 pathway, which is likely to cause cell death. Furthermore, we found that prethioviridamide binds and inhibits respiratory chain complex V (FFo-ATP synthase) in mitochondria, suggesting that inhibition of complex V leads to activation of the GCN2-ATF4 pathway. These results imply that the members of a unique family of RiPPs with polythioamide structure target mitochondria to induce the ISR.
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http://dx.doi.org/10.1021/acschembio.9b00410DOI Listing
August 2019
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