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Study question Is it possible to determine the difference between 2.1 pronuclear (2.1PN) zygotes and tripronuclear (3PN) zygotes from time-lapse images? Summary answer A pronucleus of less than 15 μm in diameter can be considered the micro pronucleus (micro PN), and it is possible to classify 2.1PN zygotes. What is known already 2.1PN zygotes are defined as zygotes with two pronuclei and one smaller pronucleus. Capalbo et al. (2017) reported that most of the 2.1PN-derived blastocysts were diploid by preimplantation genetic testing for aneuploidy (PGT-A), including single-nucleotide polymorphisms (SNPs) analysis. Thus, the treatment with 2.1 PN zygotes should be performed with chromosome testing. In Japan, where PGT-A is not available in principle, 2.1PN zygotes are rarely used in the embryo transfer. On the other hand, the size of the micro pronucleus in 2.1PN zygotes has not been clearly defined, and it is difficult to determine differences between 2.1PN and 3PN zygotes. Study design, size, duration The study was performed retrospectively on 2463 cycles of in vitro fertilization (IVF) conducted at our clinic between August 2020 and December 2021. A total of 3073 embryos underwent conventional-IVF (c-IVF) or intracytoplasmic sperm injection (ICSI) and were cultured in the time-lapse incubator, of which 221 zygotes with three pronuclei were used in the study. Participants/materials, setting, methods The diameter of the three PNs at one hour before syngamy from time-lapse images; 2.1 PN and 3PN zygotes were classified in the report by Capalbo et al. (2017). The age of the patients and the method of insemination between the groups were compared, and the diameter of the micro PN was analysed. Moreover, logistic regression analysis was performed to investigate the predictor of 2.1PN zygotes from the morphological characteristics of oocytes at ICSI. Main results and the role of chance The mean age of each patient was 42.9 years for 2.1PN zygotes and 39.8 years for 3PN zygotes, significantly higher for 2.1PN zygotes (P =0.003). On the other hand, when comparing the stage of oocyte maturation at the time of oocyte retrieval, there was no significant difference (P =0.749). According to the insemination method, the incidence of 2.1PN zygotes was significantly higher in ICSI (including rescue-ICSI) compared to c-IVF: 32.9% [95%CI: 22.5-44.6%] vs 2.4% [95%CI: 0.1-12.9%] (P Limitations, reasons for caution In this study, we have not investigated whether 2.1 PN zygotes become blastocysts. It will be necessary to further examine the criteria for 2.1PN along with chromosome testing to investigate the use of 2.1PN-derived blastocysts. Wider implications of the findings A pronucleus of less than 15 μm in diameter can be considered a micro PN. Compared to 3PN zygotes, 2.1PN zygotes were more frequently observed in older patients and in ICSI-derived zygotes. However, it is difficult to predict the incidence of 2.1PN zygotes from the oocytes’ morphological characteristics at ICSI. Trial registration number not applicable

Study question The aim of this study is to analyse the association between blastocyst diameter and TVs development, and to examine the efficacy of AH. Summary answer Blastocysts with a diameter of more than 170 μm leads to high incidence of TVs and AH applied from the incidence should be effective. What is known already TVs are protrusion of trophectoderm cells often observed in expanding blastocyst stages. TVs can be observed in expanding blastocysts regardless of Intracytoplasmic sperm injection (ICSI) and Conventional-IVF (C-IVF), when the internal pressure of blastocysts increase. The rate of TVs incidence in blastocysts inseminated by ICSI is higher than that by C-IVF, due to penetration of the needle into the zona pellucida. Moreover, it has been reported that TVs may inhibit blastocyst hatching. However, the developmental timing of TVs is still unclear, and there is no study that has analysed the association between blastocyst diameter and the incidence of TVs. Study design, size, duration 1) Diameters and TVs incidence of blastocysts by ICSI and C-IVF were measured, and the cut-off value and the area under the curve (AUC) of the receiver operating characteristic (ROC) curve were calculated to estimate the timing of TV incidence. 2) We analysed the clinical pregnancy rates of blastocysts with TVs treated by AH compared to those of blastocysts by C-IVF not subjected to AH. Participants/materials, setting, methods This study included 821 transferred frozen blastocysts ranging from March 2018 to November 2019. The embryos were cultured in a dry incubator after insemination by ICSI or C-IVF. Blastocyst freezing conditions were set at day5 to day7 with a diameter of more than 150 μm in inner diameter of zona pellucida, and this was measured before freezing. The ROC curve was performed using EZR statistical analysis software. Main results and the role of chance 1) The incidence of TVs in blastocysts by ICSI and C-IVF was 27.5% (117/424) and 14.6% (58/397) respectively. The rate of the incidence of TVs in blastocysts inseminated by ICSI and C-IVF; 8.6% (12/140) and 0.95% (1/105) in 150–159 μm, 12.7% (14/110) and 8.2% (6/73) in 160–169 μm, 40.6% (28/69) and 10.5% (6/57) in 170–179 μm, 55.6% (30/54) and 25.5% (13/51) in 180–189 μm, 66.7% (20/30) and 35.7% (10/28) in 190–199 μm, and 68.4% (13/19) and 26.8% (22/82) in the diameter of more than 200 μm. The cut-off value of the ROC curve was respectively 170 μm (sensitivity 78.6% and specificity 73.0%) and 176 μm (sensitivity 84.5% and specificity 59.6%) in the diameter; the AUC was 0.8 [95%CI:0.752–0.848] and 0.74 [95%CI:0.687–0.793] respectively. 2) The clinical pregnancy rate of TVs blastocyst vs C-IVF blastocyst was 52.7% (88/167) vs 57.8% (37/64) respectively. There is no significant difference between the two clinical pregnancy rates (P = 0.556). Limitations, reasons for caution The findings of this study have to be seen in light of some limitations. Since this study aimed to analyse the incidence of TVs based on blastocyst size, we did not take into account the grade according to the Gardner classification and the number of trophectoderm cells. Wider implications of the findings: Blastocysts inseminated by ICSI and C-IVF were highly likely to have TVs above 170 μm and 176 μm respectively. The clinical pregnancy rates of the blastocyst with TV treated by AH was similar to those of the C-IVF blastocyst. Trial registration number Not applicable

Study question Why do multinucleated blastomeres appear at high frequency in two-cell-stage embryos in humans? Summary answer Failure in microtubule assembly during the first mitotic spindle body formation by sperm centrosome-dependent microtubule organising centres (MTOCs) may lead to chromosomal instability. What is known already Unlike that in mice, multinucleated blastomeres appear at high frequency in two-cell-stage embryos in humans. However, the underlying mechanism remains elusive. In mice, multiple acentriolar MTOCs appear around the male and female pronuclei after pronuclear disappearance and contribute to dual-spindle formation, engulfing each parental chromosome. This spindle formation may ensure an error-free division, keeping the chromosomes stable during the first cleavage, as observed in mice, but it is unclear whether a similar mechanism exists in humans. Study design, size, duration To examine how sperm centrosomes contribute to MTOC formation in humans, two types of 3PN zygotes derived fromeither conventional in vitro fertilization (c-IVF, n = 30) or intracytoplasmic sperm injection (ICSI, n = 10) were used. The zygotes were collected from October 2018 to January 2020. MTOC and mitotic spindle formation at consecutive stages of development during the first cleavage were analysed under static and dynamic conditions using immunofluorescence assay and fluorescent live-cell imaging. Participants/materials, setting, methods Under ethics approval, 3PN zygotes were donated by infertile couples undergoing c-IVF or ICSI cycles at the Yamashita Shonan Yume Clinic in Japan. All participants provided informed consent. Immunofluorescence assay was performed using antibodies against α-tubulin, pericentrin, and H3K9me3 after fixation with MTSB-XF solution. Fluorescent live-cell imaging was performed using TagGFP2-H2B mRNA (chromosome marker) and FusionRed-MAP4 mRNA (microtubule marker). Main results and the role of chance Immunofluorescence revealed that while 3PN zygotes derived from c-IVF showed four pericentrin dots, those derived from ICSI exhibited two pericentrin dots. In pro-metaphase, an independent group of chromosomes derived from each pronucleus and MTOCs were formed by the sperm centrosome at the core. Microtubules from each MTOC extended toward the chromosomes in the early metaphase; a quadrupolar spindle was formed in the c-IVF-derived zygotes, and a bipolar spindle was formed in the ICSI-derived zygotes by the MTOCs at the zygote apex after chromosome alignment. In pro-metaphase, the microtubules extended from the MTOCs to the nearest chromosome. Since microtubule assembly was found on oocyte-derived chromosomes, we hypothesised that whether a chromosome is surrounded by microtubules depends on the location of the MTOCs, irrespective of its origin. Live-cell imaging of histone H2B and MAP4 revealed that four MTOCs appeared around the three pronuclei just before the disappearance of the pronuclear membrane; microtubules then extended from the MTOCs toward the chromosomes, beginning to form a mitotic spindle as the chromosomes moved to the centre of the oocyte. Interestingly, one of the three assembled chromosome groups showed no microtubule assembly in the pro-metaphase. Similar results were obtained in all six 3PN zygotes subjected. Limitations, reasons for caution We demonstrated the high risk of developing bare chromosomes not surrounded by microtubules during the formation of the first mitotic spindle, using human tripronuclear zygotes. However, owing to unavailability of normal fertilized oocytes for this study because of the clinical use, we were unable to confirm this in normal zygotes. Wider implications of the findings Although two sperm centrosome-dependent MTOCs are expected to be formed in normal fertilized oocytes, these MTOCs are not sufficient to completely enclose physically separated female and male chromosomes with the microtubules. This explains the high frequency of zygotic division errors that lead to unstable human chromosomes. Trial registration number not applicable

Systematic study of nano-fiber formation by He plasma irradiation on period 6 metals (Ta, W, Re, Ir, Pt, Au) was performed. Under the same He plasma irradiation conditions (flux, fluence, and ion energy), long nano-fibers were produced on W, Re, and Ir among which the nano-fibers on Re had the highest growth rate. On the other hand, Ta, Pt and Au did not show long nano-fibers (slight nano-fiber formation was seen on Pt). The transition metals in favor of the nano-fiber formation are stiff materials with high Young's and shear moduli. In addition, stiff materials tend to have high He binding energies with He clusters at vacancies. It is speculated that stiffness could contribute to generation of high stress field and high pressure He bubble formation, which are possibly important for nano-fiber formation.

Difference of helium (He) agglomeration energies between period 6 elements, tantalum (Ta), tungsten (W), iridium (Ir) and gold (Au), is illustrated by using first principles calculations based on density functional theory (DFT). It is found that He in W and Ir can agglomerate more easily than Ta and Au. We investigate a relationship between the He agglomeration tendency and the growth of nanostructure by He plasma irradiation. Thus, the four metals are exposed to He plasma irradiation. Each metal has different structures after the He plasma exposure. Surface nanostructures of W and Ir are fuzzy fiber-like while these structures are not observed in Ta and Au. In the meantime, W and Ir have a tendency to agglomerate He atoms at a vacancy or interstitial sites easily. This correlation suggests that the He agglomeration may play a role for understanding the fuzz formation mechanism. Keywords: Helium plasma induced structures, Fuzz, Nanostructures, Density functional theory

Background: The importance of tumor-infiltrating lymphocytes (TIL) in invasive breast carcinoma for tumor development and therapeutic response is widely accepted. However, the immune microenvironment of breast ductal carcinoma in situ (DCIS) has not been fully elucidated. Evasion of immune surveillance is a necessary step in tumor evolution. In DCIS, the tumor cells are relatively protected from the immune system due to an myoepithelial cell layer and basement membrane, and intraductal immune cells are rarely detected. In contrast, in invasive disease, cancer cells and immune cells are often intermingled. Thus, understanding the immune microenviroment of in situ to invasive carcinoma transition might be particularly important to identify novel targets for early stage of tumor invasion. Aims: The aim of this study is to evaluate the clinical importance of TILs in DCIS. Methods: TILs were assessed in 133 DCIS samples with or without microinvasive disease according to the proposed method from the International Immuno-Oncology Working Group on Breast Cancer. In addition, the relationship between TILs in DCIS and clinicopathological features was evaluated. Results: TILs are present in most DCIS in varying levels. The median proportion of TILs in DCIS was 14%. Only a minority of DCIS showed >50% TILs, which represented only 12.8% of all cases. High TILs in DCIS was significantly associated with comedo necrosis (p Conclusions: High TILs in DCIS was significantly associated with adverse histopathologic features. Further characterization of immune environment of DCIS may be essential for immunotherapy and breast cancer prevention. Citation Format: Yamashita N, Hisamatsu Y, Shigechi T, Tokunaga E, Saeki H, Oki E, Maehara Y. The immune microenvironment of ductal carcinoma in situ of the breast [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P5-07-07.

Objective: Ultrasonography has been prevalently used as a valid and objective modality for joint examination in patients with rheumatoid arthritis (RA). This study aimed to examine and compare the concordance between ultrasound, clinical, and patient-reported assessment of joint synovitis in RA. Methods: Fifty patients with RA (84% female; median age, 69 years; disease duration, 2.6 years; disease activity score of 28 joints, 3.8) completed the self-evaluation of joint symptoms including pain and considerable stiffness in the (proximal) interphalangeal, metacarpophalangeal, wrist, elbow, shoulder, knee, and ankle joints. These joints were also examined by a physician in order to assess the presence of tenderness or swelling; the presence of imaging synovitis was assesses by ultrasonography. Results: In a total of 1492 evaluated joints, symptoms (pain and stiffness), tenderness, and swelling were noted in 288 (19.3%), 182 (12.2%), and 220 (14.7%) joints, respectively, while ultrasound indicated synovitis in 317 (21.2%) joints. The overall concordance rate with ultrasound findings was lowest for joint tenderness (κ = 0.30), followed by concordance with symptoms (κ = 0.39) and swelling (κ = 0.43), irrespective of the evaluated joint, but excepting the elbow. Moreover, the percentages of inflamed joints detected only on the basis of symptoms, tenderness, or swelling were 18.6%, 2.2%, and 8.5%, respectively, of all joints with synovitis signs on ultrasonography. Conclusion: Joint swelling showed the best concordance with ultrasonography, followed by patient-reported joint symptoms, and joint tenderness. Joint symptoms, rather than tenderness evaluation, may provide a better clinical indicator of synovitis in RA patients. This article is protected by copyright. All rights reserved.

Backgrounds; The wild-type p53-induced phosphatase 1(Wip1) is a member of the serine/threonine protein phosphatases, and plays an important role in the nucleus as one of the key components in the DNA damage response network. Wip1 is encoded by the protein phosphatase magnesium dependent 1 delta (PPM1D), sited on locus 17q23. PPM1D gene amplification and/or Wip1 expression have been observed in numerous tumors, including breast cancer. PPM1D is referred to as oncogene, as Wip1 inhibits phosphorylation of p53 and work as a negative regulator in cell death. Inhibition of Wip1 may have an important therapeutic role in suppressing tumor growth and evolution. Aims; We evaluated the expression of Wip1 mRNA, Wip1 protein and PPM1D DNA copy number to clarify the relationship between Wip1 expression and the clinicopathological features and prognosis to determine the biological significance of Wip1. Materials and Methods; Breast cancer cell lines (MCF7, T47D, MDA-MB231, HCC1937, HS578T, BT20 and SKBr3) were used for Wip1 expression analysis and copy number analysis. The specimens were obtained from Japanese breast cancer patients who underwent surgery without neoadjuvant chemotherapy or endocrine therapy in our department. Wip1 mRNA expression was evaluated in 140 cases by quantitative RT-PCR and Wip1 protein expression was evaluated in 192 cases by immunohistochemistry (IHC). The PPM1D DNA copy number was analyzed by genomic PCR in 33 breast cancer cases and by SNP-CGH array (Illumina, Human Omni 2.5-8) in 12 cases. The effects on the cell growth of the Wip1 inhibitor (GSK2830371) were analyzed by the viability assay in MCF7. Results; Wip1 mRNA expression was significantly higher in MCF7, luminal type cell line. There was no significant correlation between Wip1 mRNA expression and prognosis. In IHC, positive nuclear Wip1 protein expression was detected in 21 cases (10.9%). There was no significant correlation between Wip1 mRNA expression and Wip1 protein expression. There was no significant association between the Wip1 protein expression levels and the clinicopathological factors and the prognosis. PPM1D DNA copy number significantly correlated with Wip1 protein expression (p=0.0035). Copy number gain at 17q23 was detected in 6 cases by SNP-CGH array, and all of these six cases showed positive nuclear Wip1 expression. PPM1D copy number gain was not observed in Wip1 negative cases. In the cell viability assay, the MCF7 cell growth was suppressed by Wip1 inhibitor administration. Conclusions;Wip1 protein expression in nucleus is important as oncogene, and its expression may be regulated by PPM1D copy number gain. Wip1 is considered to be the new therapeutic target for breast cancer patients. Citation Format: Inoue Y, Yamashita N, Tokunaga E, Kitao H, Tanaka K, Saeki H, Oki E, Maehara Y. The clinical importance of nuclear wild-type p53-induced phosphatase 1(Wip1) expression in breast cancer patients [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P1-08-13.

Purpose: To investigate age-related differences in outcomes of critically ill patients with sepsis around the world. Methods: We performed a secondary analysis of data from the prospective ICON audit, in which all adult ( >16 years ) patients admitted to participating ICUs between May 8 and 18, 2012, were included, except admissions for routine postoperative observation. For this sub-analysis, the 10,012 patients with completed age data were included. They were divided into five age groups - 80 years. Sepsis was defined as infection plus at least one organ failure. Results: A total of 2963 patients had sepsis, with similar proportions across the age groups (80 = 30.9%). Hospital mortality increased with age and in patients >80 years was almost twice that of patients 70 years was independently associated with increased risk of dying. Conclusions: The odds for death in ICU patients with sepsis increased with age with the maximal rate of increase occurring between the ages of 71 and 77 years. (C) 2019 Elsevier Inc. All rights reserved.

Background The natural history of human T-cell leukemia virus type 1 (HTLV-1), which causes adult T-cell leukemia (ATL) or HTLV-1 associated myelopathy, after liver transplantation is unclear. Methods We conducted a nationwide survey to investigate the impact of HTLV-1 status on living donor liver transplantation (LDLT) in Japan. We analyzed the cases of 82 HTLV-1-positive recipients and six HTLV-1-negative-before-LDLT recipients who received a hepatic graft from HTLV-1-positive donors. Results Adult T-cell leukemia developed in five recipients who ultimately died. Of these five, two received grafts from HTLV-1-positive donors and three from HTLV-1-negative donors. The 1-, 3-, and 5-year ATL development rates were 4.5%, 6.5%, and 9.2%, respectively. Fulminant hepatic failure as a pre-transplant diagnosis was identified as an independent risk factor for ATL development (P = 0.001). The 1-, 3-, and 5-year survival rates for HTLV-1-positive recipients who received grafts from HTLV-1-negative donors were 79.9%, 66.1%, and 66.1%, and from HTLV-1-positive donors were 83.3%, 83.3%, and 60.8%, respectively. The 1-year survival rate for HTLV-1-negative recipients who received grafts from HTLV-1-positive donors was 33.3%. Conclusions Fulminant hepatic failure is an independent risk factor for ATL development in HTLV-1-positive recipients. Grafts from HTLV-1-positive living donors can be transplanted into selected patients.

Background: E-cadherin and vimentin are now regarded as major and conventional canonical markers of epithelial-mesenchymal transition (EMT). It is commonly assumed E-cadherin is uniformly lost during the process of EMT. We previously reported that the elevated expression of vimentin contributes to the aggressive phenotype in invasive breast cancer. On the other hand, the role of E-cadherin in breast cancer biology might be unclear and more complex. Although, cell cohesion during breast cancer invasion is often overlooked, accumulating evidences indicate breast tumor cells are typically cohesive and often display membrane-localized E-cadherin in both the primary tumor and distant metastases, termed collective invasion. Multiple mechanisms have emerged to address how epithelial breast tumors invade. Aims: The aim of this study is to reveal the clinical importance of the expression of E-cadherin and vimentin in breast cancer. Methods: The E-cadherin and vimentin protein expression were evaluated by immunohistochemistry (IHC) in 177 invasive breast cancer samples. Among these, E-cadherin and vimentin expression were evaluated in the set of primary breast cancer and metastatic lymph nodes in 65 cases. Results: The positive vimentin expression was highly correlated with poor disease-free survival (DFS) and overall survival (OS) (p=0.019 and p=0.0044), however, the E-cadherin expression alone did not correlate with prognosis. Interestingly, Both E-cadherin and vimentin positive tumor had the worst DFS and OS among all breast cancer (p=0.03 and p=0.0089). Vimentin expression was highly correlated between primary tumors and metastatic lymph nodes. However, E-cadherin expression levels were significantly elevated in metastatic lymph nodes (p=0.0017), Co-expression of E-cadherin and vimentin in the metastatic lymph nodes also showed worst DFS and OS (p=0.12 and p=0.027). Conclusions: Co-expression of E-cadherin and vimentin seems to be associated with the most aggressive phenotype and poorest prognosis in breast cancer, and positive E-cadherin expression may not always play roles for tumor suppression. Citation Format: Yamashita N, Tokunaga E, Inoue Y, Tanaka K, Ueo H, Saeki H, Oki E, Maehara Y. Epithelial paradox; clinical significance of co-expression of E-cadherin and vimentin in invasive breast cancer. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P2-05-12.

Background. The estrogen receptor (ER)/ GATA3/ Forkhead box A1 (FOXA1) network is necessary for the ERα functional signature specific to luminal type breast cancers. High expression of FOXA1 indicates a good prognosis in ER-positive breast cancer. However, little is known about the association between the expression of FOXA1 and GATA3, and the efficacy of neoadjuvant endocrine therapy (NAE). This study investigated their predictive potential for NAE and the changes of their expression after NAE. Methods. The expression of ER, progesterone receptor (PgR), Ki67, FOXA1, and GATA3 were analyzed by immunohistochemistry in 66 patients with hormone receptor-positive/ human epidermal growth factor receptor 2 (HER2)-negative breast cancer who had been treated with NAE between March 2003 and December 2012 at Kyushu University Hospital, National Kyushu Cancer Center, and Sagara Hospital. The association between the expression of biological marker and the efficacy of NAE, and their expression changes after NAE were evaluated. Results. The median age of the patients was 60 years (range, 30–84 years). Pre- and post-menopausal patients were 24 (36.4%) and 42 (63.6%). Endocrine agents that were administered are as follows: aromatase inhibitors (AIs) for 42 patients (63.6%), luteinizing hormone-releasing hormone (LHRH) agonist plus AI for 10 patients (15.2%), LHRH agonist plus tamoxifen for 13 patients (19.7%). NAE yielded a partial response (PR) in 21 patients (31.8%) and stable disease (SD) in 45 patients (68.2%). Breast conserving surgery was performed in 56 patients (84.8%) and mastectomy was performed in 10 patients (15.2%). Preoperative Endocrine Prognostic Index (PEPI) score was 0 in 10 patients (15.2%) and 1 or greater (score 1 ≤) in 56 patients (84.8%). Pre-treatment FOXA1 expression was positively correlated with GATA3 (P = 0.0003) and PgR (P = 0.0138). Post-treatment Ki67 expression was significantly lower in tumors, which achieved PR compared with those with SD (P = 0.0007). The expression of PgR, Ki67, and FOXA1 was significantly lower in post-treatment tumors compared with those in pre-treatment samples (p < 0.0001, p < 0.0001 and p < 0.0001, respectively). The expression of PgR, Ki67, and FOXA1 was significantly reduced in both tumors with PR and those with SD (PR: P = 0.0004, P < 0.0001, and P = 0.0417, respectively; SD: P < 0.0001, P = 0.0001, and P < 0.0001, respectively). The expression of PgR, Ki67, and FOXA1 was significantly decreased in post-treatment tumors in both patients with the PEPI score 0 and those with score 1 ≤ (score 0: P = 0.0078, P = 0.0059, and P = 0.0098, respectively; score 1 ≤: P < 0.0001, P < 0.0001, and P = 0.0002, respectively). In tumors with PgR > 20%, the expression of Ki67 and FOXA1 were significantly lower in post-treatment tumors (P < 0.0001 and P < 0.0001, respectively). Conclusions. FOXA1 expression correlated with PgR expression, and was reduced significantly after NAE. These results suggest that blocking the effect of estrogen might reduce FOXA1 expression. Citation Format: Tanaka K, Tokunaga E, Inoue Y, Ueo H, Yamashita N, Sagara Y, Ohi Y, Taguchi K, Ohno S, Okano S, Kitao H, Oki E, Oda Y, Maehara Y. The relationship between the expression of FOXA1 and GATA3 and the efficacy of neoadjuvant endocrine therapy. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P5-13-05.

Background Acneiform rash as an adverse event often affects the treatment by EGFR-TKIs. Since minocycline has been suggested to reduce the rash, we assessed the efficacy and safety of prophylactic administration of minocycline simultaneously during erlotinib treatment. Methods Patients of ECOG performance status 0-2 with advanced NSCLC, who had not been treated with EGFR-TKIs and would receive erlotinib treatment were randomized 1:1 into either group A, with minocycline or group B, without minocycline. The patients assigned to group A were started on minocycline 100mg/day orally for 8 weeks with erlotinib. Primary end point was the frequency of grade ≥2 rash acneiform by independent assessment in first 8 weeks. We expected the prophylactic minocycline decreased the incidence of grade ≥2 skin rash from 50% to 30%. The planned sample size was 280 patients with a = 0.025 (one-sided) and b = 0.10. Results Patients accrual was started in March 2015 and ended in June 2018 because of slow accrual. Ninety-four patients were finally enrolled and 93 were full-analysis set. The median age of the patients was 71 years old (range 45 to 89),58 patients were female. EGFR mutation status positive/negative/unknown=78/13/2 patients. The frequency of grade ≥2 rash acneiformby independent assessment was 33.3% [95%C.I. 20.0-49.0%] in group A vs. 44.2% [95%C.I. 29.1-60.1%] in group B (p = 0.296). The frequency by physicians’ evaluation was 31.3% [95%C.I. 18.7-48.8%] and 45.5% [95%C.I. 30.4-61.2%] (p = 0.161). However, the frequency of grade ≥2 rash acneiform on day 15 by physicians’ evaluation was significantly decreased (4.4% [95%C.I. 5.3-14.8%] in group A vs. 25.0% [95%C.I. 13.2-40.3%] in group B (p = 0.005)). As for toxicity, the incidence of any grade skin-related toxicity as pruritus (39.6% vs. 63.6%) and pain of skin (14.6% vs. 25.0%) was less common in group A. Hence, anorexia (41.7% vs 20.5%), nausea (25.0% vs 4.5%), and dysgeusia (22.9% vs 15.9%) occurred more frequently. Conclusions Although the frequency of acneiform rash tended to decrease, prophylactic administration of minocycline is not recommended because of increasing gastrointestinal toxicity. Legal entity responsible for the study The authors. Funding Has not received any funding. Disclosure T. Kozuki: Honoraria (self), Research grant / Funding (self): AstraZeneca; Honoraria (self), Research grant / Funding (self): Chugai Pharmaceutical Co.; Honoraria (self), Research grant / Funding (self): Eli-Lilly Japan; Honoraria (self): Taiho Pharmaceutical Co.; Honoraria (self): Ono Pharmaceutical Co.; Honoraria (self): Bristol-Myers Squibb; Honoraria (self), Research grant / Funding (self): MSD; Honoraria (self): Boehringer Ingelheim; Honoraria (self): Kyowa-Hakko Kirin; Honoraria (self): Pfizer; Honoraria (self): Nippon-kayaku; Research grant / Funding (self): Merck Biopharma. M. Takenoyama: Honoraria (self), Research grant / Funding (self): Chugai Pharmaceutical. All other authors have declared no conflicts of interest.

Background Nanoparticle albumin-bound paclitaxel (nab-PTX) + carboplatin (Cb) therapy is one of the standard platinum-containing chemotherapy regimens for patients with advanced non-small cell lung cancer (NSCLC). Adding the anti-vascular endothelial growth factor antibody bevacizumab (BEV) to chemotherapy is an effective treatment option for non-squamous NSCLC. Because the efficacy and safety of the Cb + nab-PTX + BEV triplet regimen has not yet been assessed, we conducted a multicenter, open-label, phase I/II trial of Cb + nab-PTX + BEV therapy for patients with NSCLC. The phase II trial was based on the drug dose and schedule determined in the phase I trial. Methods In this phase II trial, the required number of patients was calculated to be 49 cases with α = 0.05 (one-sided) and β = 0.1 assuming a threshold response rate of 30% and an expected response rate of 50%. The patients were to receive 4–6 cycles of Cb (area under the curve = 6) + nab-PTX (100 mg/m2 on days 1, 8 and 15) + BEV (15 mg/kg on day 1) followed by a maintenance dose of nab-PTX + BEV every 3 weeks until disease progression. The primary endpoint was the overall response rate (ORR), and the secondary endpoints included overall survival (OS), progression free survival (PFS) time and toxicity. Results The trial was terminated early because of slow patient accrual. Finally, 47 cases were registered, and the main analysis was performed in 46 cases, excluding one case who was unqualified. The median age of the patients was 66 years. The transition percentage to maintenance therapy was 58.7%. The ORR based on central judgment was 56.5% (26/46 cases) with a 95% confidence interval (CI) of 42.2–70.8%, and the primary endpoint was met. The median PFS and OS were 7.79 months and 18.9 months, respectively. The main toxicity was myelosuppression, with grade 3–4 neutropenia (72.0%), anemia (28.0%), thrombocytopenia (14.0%) and febrile neutropenia (2.0%). The grade 3–4 sensory and motor neuropathy commonly seen with paclitaxel was 0%. All adverse events were manageable, and there was no treatment-related death. Conclusions Cb + nab-PTX + BEV therapy is a favorable and well-tolerated treatment for patients with advanced non-squamous NSCLC. Clinical trial identification UMIN000014560. Legal entity responsible for the study TORG/OLCSG. Funding Taiho Pharmaceutical CO., LTD. Disclosure T. Kubo: Honoraria (self): Taiho pharmaceutical; Honoraria (self): Chugai pharmaceutical; Honoraria (self): BMS. A. Bessho: Honoraria (self): Taiho pharmaceutical; Honoraria (self): Chugai pharmaceutical. A. Morita: Honoraria (self): Chugai pharmaceutical. T. Yokoyama: Honoraria (self): Taiho pharmaceutical; Honoraria (self): Chugai pharmaceutical. K. Kaira: Honoraria (self): Taiho pharmaceutical; Honoraria (self): BMS. T. Harada: Honoraria (self): Taiho pharmaceutical. T. Shimokawa: Research grant / Funding (institution): Taiho pharmaceutical; Research grant / Funding (institution): Chugai pharmaceutical; Research grant / Funding (institution): BMS. K. Kiura: Research grant / Funding (institution): Chugai pharmaceutical; Research grant / Funding (institution): BMS; Research grant / Funding (institution): Taiho pharmaceutical. H. Okamoto: Research grant / Funding (institution): Taiho pharmaceutical; Research grant / Funding (institution): BMS; Research grant / Funding (institution): Chugai pharmaceutical. All other authors have declared no conflicts of interest.

We appreciate the letter from Dr. Carlos Antonio Moura and Dr. Carlos Geraldo Moura for their interest in our article [1]. As they noticed, that study was intended to evaluate the concordance of patient-reported joint symptoms, physician-examined arthritic signs, and ultrasound-detected synovitis in rheumatoid arthritis (RA), but not to evaluate the use of MTX. Although we described the median and range of RA duration (months) as 31 and 0-608, we should have clearly stated that 11 of 50 patients enrolled in that study were treatment-naive. This article is protected by copyright. All rights reserved.

Background Joint destruction in RA includes both bone and cartilage lesions. By X-ray examination, cartilage destruction is evaluated as a joint space narrowing (JSN). However, joint space narrowing is not a direct evaluation of cartilage. Objectives We aimed to examine the finger joint cartilage thickness (CT) by ultrasound (US) imaging and clarify its clinical significance in patients with RA. Methods We enrolled 121 RA patients in low disease activity or clinical remission (DAS28-CRP Results CT in MCP joints ranged from 0.0 to 1.1 mm (median 0.5 mm), and CT in PIP ranged from 0.0 to 0.6mm (median 0.3mm), respectively. The sum of total CT from 8 fingers ranged from 4.0 to 9.4 mm (median 6.9 mm), and there was a significant difference in total CT, but not in JSN score, between male and female patients (7.4 versus 6.7, p=0.006; and 11 versus 10, p=0.899, respectively). CT was well correlated with JSN (ρ=-0.589, p Conclusions A direct visualization and quantification of finger joint CT, especially MCP joints, by US is valid and useful in RA. Disclosure of Interest None declared

Background: Breast cancer is a heterogeneous disease and divided into five distinct groups (Luminal A, Luminal B, HER2-enriched, Basal-like, and Claudin-low subtypes). DNA copy number studies have suggested that chromosomal aberrations are different among subtypes. Basal-like, and Claudin-low subtypes are mainly ER-, PR- and HER2- (triple negative: TN). Recent study reported that the basal-like subtype was the most distinct with common losses of the regions containing tumor suppressor genes such as RB1, BRCA1, INPP4B, and the greatest overall genomic instability. On the other hand, Claudin-low tumors showed few copy number changes. Luminal A tumors show fewer chromosomal copy number changes than Luminal B and HER2-enriched subtypes. Aims: To investigate the extent of the chromosomal aberrations and to evaluate the relationships between chromosomal aberrations and breast cancer subtypes and other clinicopathological characteristics and prognosis. Methods: Specimens were obtained from 363 invasive breast cancer patients who underwent surgery in our department between 1994 and 2011. Four breast cancer subtypes were determined by the immunohistochemical analysis of ER, PR and HER2; hormone receptor (HR; ER and/or PR)+/HER2-, HR+/HER2+, HR-/HER2+(HER2) and TN. Loss of heterozygosity (LOH) at the tumor suppressor gene (TSGs), BRCA1, BRCA2, TP53, RB1, PTEN and INPP4B, was investigated with microsatellite markers using our microsatellite analysis system. Copy number aberrations (CNAs) were analyzed by SNP-CGH array (Illumina, HumanOmni2.5-8).% Defects, which represents the percent of genome region containing chromosomal aberration, was determined by Karyostudio Software v1.4 (Illumina). Results: The incidence of LOH at BRCA, BRCA2, TP53, RB1, PTEN and INPP4B locus was 37.4, 34.0, 57.1, 41.9, 25.8 and 18.0%, respectively. The incidence of LOH was lowest in hormone receptor (HR)+/HER2- subtype, and higher in HER2 and triple negative (TN) subtypes. LOH at INPP4B locus was significantly associated with TN subtype. The LOH at BRCA1 and TP53 loci was highest in HER2 subtype. The LOH at BRCA1, TP53 and INPP4B loci was significantly associated with poor prognosis in all cases; however, the impact of LOH at these TSGs on the prognosis was different among subtypes. Coexistence of LOH at both BRCA1 and TP53 loci was significantly associated with aggressive phenotype and chromosomal aberrations.%Defect was highest in the tumors with LOH at both BRCA1 and TP53 loci. Coexistence of LOH at both BRCA1 and TP53 loci was significantly associated with poor prognosis, especially in HR+/HER2- subtype. Many cases with TN subtype reveal gross chromosomal aberrations, and these tumors were considered to be basal-like. However, there are some cases with few chromosomal alterations in TN, whose prognosis was very poor. These tumors were considered to be Claudin-low subtype. Conclusions: The incidence and biological significance of LOH at TSGs are different among breast cancer subtypes. Coexistence of LOH at both BRCA1 and TP53 loci was associated with gross chromosomal aberrations and poor prognosis. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P2-08-04.

Background: Triple negative breast cancer (TNBC) is typically associated with aggressive tumor phenotype and poor prognosis. TNBC is also considered highly heterogeneous disease. A better understanding of molecular and histopathological features of TNBC is of great importance, in order to develop a new therapeutic strategy and to improve the prognosis of TNBC. TNBC has many similarities with basal-like breast cancer, and is also associated with BRCAness. The major role of BRCA1 is to respond to DNA damage by participating in cellular pathways for DNA repair, mRNA transcription, cell cycle regulation, and protein ubiquitination. BRCA1 function loss leads to impaired homologous recombination mediated DNA repair. A loss of BRCA1 may thus be a biomarker of responsiveness to DNA damaging agents such as PARP inhibitor and cis/carboplatin. Germline mutations in BRCA1, however, account for approximately 5% of breast cancer cases, somatic mutations in BRCA1 rarely occur. Instead, lower than normal expression of BRCA1 is reported to be an important contributing factor in sporadic tumors. The BRCA1 promoter methylation may thus play an important role in the BRCA1 function loss in sporadic breast cancer. Aim: To evaluate the clinical importance of BRCA1 promoter methylation in breast cancer. Materials and Methods: Specimens were obtained from 71 TNBC and 163 non-TNBC patients who underwent surgery without neoadjuvant therapy in our department between 1990 and 2011. BRCA1 promoter methylation was investigated by using combined bisulfate and restriction analysis (COBRA). The BRCA1 mRNA expression was evaluated by quantitive RT-PCR. The BRCA1 protein level was assessed by immnohistochemistry. Loss of heterozygosity(LOH) at the BRCA1 locus was analyzed with microsatellite markers (D17S855 and D17S579) using our microsatellite analysis system. Copy number variations(CNVs) were analysed by SNP-CGH array (Illumina, HumanOmni2.5-8). Results: We found 12 patients with BRCA1 promoter methylation and all of them were TNBC (p Conclusions: The BRCA1 promoter methylation is considered to be a specific feature of aggressive phenotype of TNBC and to be one of the mechanisms of BRCA1 dysfunction in breast cancer. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P2-06-03.

Living mulch is a type of sustainable farming system that consists of cover crops planted either before or with a main crop; a living mulch is maintained as a living ground cover throughout the growing season of the main crop. Microbial biomass and abundance of mesofauna (microarthropods and enchytraeids) are important soil biological parameters in relation to soil function, plant productivity, and nutrient cycling; however, the effects of living mulch on these parameters are not fully understood. In this study we examined the effects of living mulch treatment with nitrogen fertilizer (0, 40, 160, or 200 kg ha−1) on the abundance of soil microarthropods (Oribatida, Mesostigmata, Prostigmata, and Collembola) and the effects of living mulch treatment on the dynamics of the soil biota (mesofauna, microarthropods, enchytraeids, and microbial biomass nitrogen) from spring to autumn. Our results showed that living mulch treatment significantly (p

Chabazite-Ca deposited on dacite laccolith from Osodi Hill, Dunabogdany, Hungary, exhibited bluish-white luminescence under ultraviolet (UV) light. The photoluminescence (PL) and optical excitation spectra of chabazite-Ca were obtained at 300 K. The PL spectrum under 300-nm excitation consists of (1) a Ce3+ band with a peak at 340 nm, (2) a broad main band with a peak at 453 nm and (3) five narrow bands at 592, 616, 650, 700 and 734 nm due to Eu3+. The main band is spread over the entire visible-wavelength region. The excitation spectrum obtained by monitoring green luminescence at 520 nm consists of a band at wavelengths shorter than 200 nm and an extremely broad band with a peak at 385 nm. The extremely broad band is spread over not only the UV region but also the blue region. The features of PL and excitation spectra suggest that the origin of bluish-white luminescence is luminescent organic matter incorporated into chabazite-Ca crystals during growth.

The tritium release model to represent the release behavior of bred tritium from solid breeder materials has been developed by the blanket group of Kyushu University. It has been found that water is released to the purge gas from solid breeder materials and that this water affects the tritium release behavior. In this study, the amount of surface water released from Li4SiO4 is quantified by the experiment. In addition, the tritium release behavior from Li4SiO4 are estimated based on the tritium release model using parameters obtained in our studies under conditions of commercial reactor operation and ITER test blanket module operation. The effect of the surface water on tritium release behavior is discussed from the obtained results. Moreover, the tritium inventory of Li4SiO4 is discussed based on calculation under the unsteady state condition. Further, the effects of grain size and temperature on distribution of tritium inventory under the steady state condition are evaluated, and the optimal grain size is discussed from the view point of tritium release from Li4SiO4.

It is observed that a fair amount of physical and chemical adsorbed water is released from solid breeder materials by introduction of dry N2 gas and that not a little amount of water is also continuously produced by the water formation reaction when the purge gas with hydrogen is applied. It is reported by present authors that the water released to the purge gas from solid breeder materials affects the tritium release behavior. The capacity and desorption rate of chemical adsorbed water, and the capacity and rate of water formation reaction for Li4SiO4, which has been supplied from FzK, are quantified in this study. It is found that the overall reaction rate of water formation on Li4SiO4 is larger than the rate observed for other solid breeder materials. Therefore, most hydrogen added to the blanket purge gas changes to water so far as the water formation capacity of Li4SiO4 remains. It is also found that water formation capacity of Li4SiO4 is almost the same as that of Li2TiO3. Tritium release be...

Five Na 2 SO 4 :RE 3+ phosphors activated with rare-earth (RE) ions (RE 3+ =Ce 3+ , Sm 3+ , Tb 3+ , Dy 3+ and Tm 3+ ) were synthesized by heating natural thenardite Na 2 SO 4 from Ai-Ding Salt Lake, Xinjiang, China with small amounts of rare-earth fluorides, CeF 3 , SmF 3 , TbF 3 , DyF 3 and TmF 3 , at 920 °C in air. The photoluminescence (PL) and optical excitation spectra of the obtained phosphors were measured at 300 and 10 K. In the PL spectrum of Na 2 SO 4 :Ce 3+ at 300 K, two overlapping bands with peaks at 335 and 356 nm due to Ce 3+ were first observed. Narrow bands observed in PL and excitation spectra of Na 2 SO 4 :RE 3+ (RE 3+ =Sm 3+ , Tb 3+ , Dy 3+ and Tm 3+ ) phosphors were well identified with the electronic transitions within the 4f n ( n =5, 8, 9 and 12) configurations of RE 3+ . The existence of excitation bands with high luminescence efficiency at wavelengths shorter than 230 nm is characteristic of Na 2 SO 4 :RE 3+ (RE 3+ =Sm 3+ , Tb 3+ , Dy 3+ and Tm 3+ ) phosphors. The obtained results suggest that these phosphors are unfavorable as the phosphor for usual fluorescence tubes, i.e., mercury discharge tubes, but may be favorable as the phosphor for UV-LED fluorescent tubes and as cathodoluminescence, X-ray luminescence and thermoluminescence phosphors.

Objectives The Mediterranean diet, with a high content of olive oil, is associated with a reduced risk of coronary artery disease. The aim of this study was to determine the effect of oleuropein, one of the polyphenols in olive oil, on vascular smooth muscle cell (SMC) proliferation in vitro . Design This was an experimental study. Materials and methods Bovine vascular SMCs were cultured in the presence of 100 μM of oleuropein. On days 1, 3 and 5, cell number was counted. Cell cycle analysis was performed by flow cytometry. Cell cycle regulators were assessed by immunoblotting. Results Cell proliferation in the presence of oleuropein was significantly inhibited by 92%. Cell cycle analysis revealed that oleuropein treatment blocked cells in the G1–S phase compared with the non-treated group. Among G1 phase regulators, retinoblastoma protein, cyclinD, p21 and p27 were not affected by oleuropein, but extracellular signal-regulated kinase 1/2 (ERK1/2) activation was inhibited. Growth of SMC treated with 100 μM of the mitogen-activated protein (MAP) kinase/ERK kinase 1 (MEK1) inhibitor PD98059 was also significantly inhibited by 70%. Conclusions Oleuropein inhibited SMC proliferation through a cell cycle block between the G1 and the S phases, which may be regulated by ERK1/2. These results suggest a mechanism by which olive oil consumption may have beneficial effects on cardiovascular mortality by inhibiting SMC proliferation.

The photoluminescence (PL) properties of four gypsums from three localities, namely, Inner Mongolia (China), Turkey, and Canada, are investigated at room temperature. Under 365-nm excitation, two types of gypsums from Inner Mongolia exhibit different luminescence colors, namely, yellow and cyan-white, and gypsums from Turkey and Canada exhibit yellow and bluish-white luminescence, respectively. The PL spectra of these gypsums consist of continuous sub-bands distributed in all visible wavelengths, and the excitation spectra of these gypsums consist of continuous sub-bands distributed in a wide range of wavelengths, i.e., 200-500 nm. The features of PL and excitation spectra suggest that the origin of luminescence from these gypsums may be luminescent organic substances included into crystals in the growth process rather than the impurity ions substituted for Ca2+.

Prediction of the postoperative course of esophagectomy is an important part of the strict perioperative management of patients undergoing surgery for esophageal cancer.To evaluate their clinical importance, peripheral blood values, including white blood cell count (WBC), lymphocyte count, and the levels of total protein, transferrin, factor XIII, D-dimer, fibrin, and fibrinogen degradation products (FDP) were measured before and after esophagectomy for esophageal cancer in 24 patients.The preoperative WBC and the pre- and postoperative lymphocyte count were decreased remarkably in patients who received preoperative chemoradiotherapy. The values of perioperative serum transferrin were significantly lower in patients with postoperative pneumonia than in those without. The activity of plasma factor XIII was suppressed on postoperative day (POD) 7 in patients with pneumonia and on POD 14 in patients with leakage.These results suggest that patients who receive preoperative chemoradiotherapy are potentially immunosuppressed, the preoperative serum transferrin level is a possible predictive marker of postoperative pneumonia, and suppression of factor XIII activity is related to anastomotic insufficiency.