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With the development of massive parallel sequencing technology, it has become easier to establish new model organisms that are ideally suited to the specific biological phenomena of interest. Considering the history of research using classical model organisms, we believe that the efficient construction and sharing of gene mutation libraries will facilitate the progress of studies using these new model organisms. Using C. elegans, we applied the TMP/UV mutagenesis method to animals lacking function in the DNA damage response genes atm-1 and xpc-1. This method produces genetic mutations three times more efficiently than mutagenesis of wild-type animals. Furthermore, we confirmed that the use of next-generation sequencing and the elimination of false positives through machine learning could automate the process of mutation identification with an accuracy of over 95%. Eventually, we sequenced the whole genomes of 488 strains and isolated 981 novel mutations generated by the present method; these strains have been made available to anyone who wants to use them. Since the targeted DNA damage response genes are well conserved and the mutagens used in this study are also effective in a variety of species, we believe that our method is generally applicable to a wide range of animal species.

Background The balance between ischemic and bleeding events and their association with platelet reactivity in patients receiving antiplatelet therapy after percutaneous coronary intervention ( PCI ), which differs among regions, is not fully evaluated for East Asians. We examined ischemic/bleeding events and platelet reactivity in Japanese patients undergoing PCI and determined associations between high/low platelet reactivity and clinical outcomes. Methods and Results PENDULUM (Platelet Reactivity in Patients with Drug Eluting Stent and Balancing Risk of Bleeding and Ischemic Event) is a prospective, multicenter registry of Japanese patients with PCI . Primary end points were incidence of first major adverse cardiac and cerebrovascular events ( MACCE ) and first major bleeding events at 12 months post‐ PCI . Platelet reactivity (P2Y 12 reaction unit [PRU ] value) was measured at 12 to 48 hours post‐ PCI ; patients were grouped as having high PRU (>208), optimal PRU (>85 to ≤208), and low PRU (≤85). MACCE and major bleeding occurred in 4.4% and 2.8% of 6267 patients, respectively. The mean±SD PRU value was 182.1±77.1. MACCE was significantly higher in the high PRU (5.7%; n=2227) versus the optimal PRU group (3.6%; n=3002). The hazard ratio (HR) for high PRU versus optimal PRU level was significantly higher for MACCE (adjusted HR, 1.53; 95% CI, 1.14–2.06 [ P =0.004]); stent thrombosis followed the same trend. Incidence of major bleeding did not differ significantly between groups. A high PRU level was significantly associated with MACCE in both patients with and patients without acute coronary syndrome. Conclusions These real‐world data suggest an association between high platelet reactivity and cardiovascular events in Japanese patients undergoing PCI . The trend was the same in both patients with and patients without acute coronary syndrome. REGISTRATION URL : https://www.umin.ac.jp/ctr . Unique identifier: UMIN 000020332.

// Naoki Ikari 1, 2, * , Shota Aoyama 1, * , Akiyoshi Seshimo 1 , Yuji Suehiro 3 , Tomoko Motohashi 3 , Shohei Mitani 3 , Sawako Yoshina 3 , Etsuko Tanji 2 , Akiko Serizawa 1 , Takuji Yamada 1 , Kiyoaki Taniguchi 1 , Masakazu Yamamoto 1 and Toru Furukawa 2, 4, 5 1 Department of Surgery, Institute of Gastroenterology, Tokyo Women’s Medical University, Tokyo, Japan 2 Institute for Integrated Medical Sciences, Tokyo Women’s Medical University, Tokyo, Japan 3 Department of Physiology, Tokyo Women’s Medical University School of Medicine, Tokyo, Japan 4 Department of Surgical Pathology, Tokyo Women’s Medical University Hospital, Tokyo, Japan 5 Department of Histopathology, Tohoku University Graduate School of Medicine, Sendai, Japan * These authors have contributed equally to this work Correspondence to: Toru Furukawa, email: toru.furukawa@med.tohoku.ac.jp Keywords: early gastric cancer; lymph node metastasis; lymphangiogenesis; NBN ; PAX8 Received: May 18, 2017 Accepted: January 13, 2018 Published: January 22, 2018 ABSTRACT Background and aim: Intramucosal gastric adenocarcinoma of the well-moderately differentiated type only exhibits lymph node metastasis in extremely rare cases. We encountered such case and investigated both the lymphangiogenic properties and somatic mutations in the cancer to understand the prometastatic features of early-stage gastric cancer. Methods: We quantitatively measured the density of lymphatic vessels and identified mutations in 412 cancer-associated genes through next-generation target resequencing of DNA extracted from tumor cells in a formalin-fixed and paraffin-embedded tissue. Functional consequence of the identified mutation was examined in vitro by means of gene transfection, immunoblot, and the quantitative real-time polymerase chain reaction assay. Results: The intramucosal carcinoma was accompanied by abundant lymphatic vessels. The metastatic tumor harbored somatic mutations in NBN , p.P6S, and PAX8 , p.R49H. The PAX8 R49H showed significantly higher transactivation activity toward E2F1 than the wild-type PAX8 (P< 0.001). Conclusions: Our data suggest that increased lymphangiogenesis and somatic mutations of NBN and/or PAX8 could facilitate lymph node metastasis from an intramucosal gastric carcinoma. These findings may potentially inform evaluations of the risk of developing lymph node metastasis in patients with intramucosal gastric cancer.

Background Dual therapy with a direct oral anticoagulant (DOAC) plus a P2Y12 receptor inhibitor is recommended in patients with nonvalvular atrial fibrillation who undergo percutaneous coronary intervention. Thus, we evaluated the effects of DOAC edoxaban plus P2Y12 receptor inhibitor prasugrel on bleeding time (BT), and pharmacodynamics (PD) and pharmacokinetics (PK) of edoxaban in healthy elderly Japanese male subjects. Methods A single-center, clinical pharmacology study with randomized, open-label, repeated dosing enrolled 24 participants in two groups of 12 receiving 30 mg edoxaban once daily for 3 days; then 30 mg edoxaban plus 2.5 mg prasugrel (Group 1) or 30 mg edoxaban plus 3.75 mg prasugrel (Group 2) once daily for 5 days. Primary endpoint was BT by the Ivy method. Secondary endpoints were the PD parameters of prothrombin time (PT), activated partial thromboplastin time (aPTT), prothrombin fragment F1 + 2 (F1 + 2), and P2Y12 reaction units (PRU), and PK profiles of edoxaban alone and in combination with prasugrel. Results Geometric least squares mean of BT ratios (vs. baseline) for 3-day edoxaban treatment were 1.097 (90% confidence interval (CI) 0.911–1.321) in Group 1 and 1.327 (90% CI 1.035–1.703) in Group 2; for 5-day edoxaban plus 2.5 mg and 3.75 mg prasugrel, they were 1.581 (90% CI 1.197–2.087) and 1.996 (90% CI 1.482–2.690), respectively. Contributions of prasugrel to the effects (edoxaban + prasugrel/edoxaban) were 1.442 (90% CI 1.096–1.897) in Group 1 and 1.504 (90% CI 1.172–1.930) in Group 2. Edoxaban prolonged PT and aPTT and decreased F1 + 2. Adding on prasugrel did not appreciably change PT, aPTT, or F1 + 2. Prasugrel reduced PRU, whereas edoxaban had no effect on PRU. We recorded 26 adverse events; 23 were treatment-emergent (positive fecal occult blood test). All participants with adverse events recovered during follow-up. Conclusions Coadministration of 2.5 mg and 3.75 mg prasugrel with 30 mg edoxaban prolonged BT in healthy elderly Japanese male subjects. The effect was dependent on the dose of prasugrel. Prasugrel did not affect PD or PK profiles of edoxaban. Edoxaban had no effect on PD of prasugrel. Trial registration Japan Registry of Clinical Trials No. jRCTs071190006; registration date, 26-April-2019.

The association of an IgM-Fc receptor (FcμR) with chronic lymphocytic leukemia (CLL) was suggested more than 30 years ago, but its authenticity has never been formally addressed. We examined the expression of the recently identified FcμR by B and T cells in CLL patients using receptor-specific monoclonal antibodies. CLL B cells (CD5+/CD19+) expressed much higher levels of FcμR on their cell surface than B cells from healthy donors. Such enhanced expression was more evident in immunoglobulin heavy chain variable region (IGHV)–mutated, CD38− or early Rai-stage CLL than in IGHV-unmutated, CD38+, or advanced Rai-stage CLL. Intriguingly, surface FcμR levels also were significantly elevated in the non-CLL B cells (CD5−/CD19+) and T cells (CD5+/CD19−), especially in IGHV-mutated CLL. CLL patients also had high serum titers of FcμR compared with healthy donors, and serum FcμR levels correlated significantly with circulating lymphocyte numbers but not with the IGHV mutation status or Rai stage. The serum FcμR was resolved as an ∼ 40-kDa protein, distinct from the cell surface FcμR of ∼ 60 kDa, and it was produced by both CLL B and non-CLL B cells. Mass spectrometric analysis revealed that the serum FcμR is a soluble form of the receptor encoded by an alternatively spliced FcμR transcript. These findings indicate enhanced levels of both membrane-bound and soluble forms of FcμR in CLL patients.

3.5 Elevated Levels of Both Membrane-Bound and Soluble Forms of a Receptor Specific for the Fragment Crystallizable Region of Immunoglobulin M in Patients with CLL Fu Jun Li, Yoshiki Kubagawa, Matthew K. McCollum, Landon Wilson, Tomoko Motohashi, Luigi F. Bertoli, James C. Barton, Stephen Barnes, Randall S. Davis, Hiromi Kubagawa Departments of Medicine, Pathology, and Pharmacology, University of Alabama at Birmingham, AL; Brookwood Medical Center, Birmingham, AL