Effects of WBM feeding on body weight, liver weight, ALT serum levels and uterus weight.

<p>Seven-week old female C57B1/6J mice were ovariectomized. These mice are a model of postmenopausal women (OVX, n = 16). Control mice (sham, n = 16) were subjected to sham operation survival surgery. Both sham and OVX mice were fed a HFD (n = 8/per group) or HFD with WBM (n = 8/per group) diet for 3 months. Body weight was measured once a week. (A) Body weight measurement of sham mice. (B) Body weight measurement of OVX mice. (C) Liver weight. (D) ALT serum levels. (E) Uterus weight. Values are expressed as mean and standard error for 8 mice. * Statistical significance was defined as <i>P</i><0.05.</p

Comparison of the effects of various species of mushroom on the fatty acid synthesis pathway.

<p>(A) <i>FAS</i> and (B) <i>ELOVL6</i> gene expressions in HepG2 cells treated with various species of mushroom extract (WBM, shiitake mushroom, enoki mushroom and oyster mushroom) for 24 hours. Real time PCR analysis was performed for the gene, <i>ELOVL6 and FAS</i>. Gene expression was normalized with the <i>β-actin</i> housekeeping gene. Values are expressed as mean and standard error. We analyzed the data for each dose separately by ANOVA, followed by Tukey's multiple comparison test. Statistical significance was defined as <i>P</i><0.05 compared to control.</p

LXR and ER activity in HepG2 cells treated with WBM extract.

<p>(A) HepG2 cells were transfected with the pCMX-VP16-hLXRa and LXR-responsive rCYP7A-DR-4x3-tk-LUC, and following were incubated with vehicle control (ethanol), T0901317 (10 µM) and/or WBM extract (1, 2 and 5 µl/ml) and assayed for luciferase activity. (B) For evaluation of WBM effects on ER activity, HepG2 cells were transiently transfected with the pSG5-ER and pGL₃(ERE)₃-Luc. 24 hours posttransfection, cells were treated with E2 (0.1 nM) or with WBM extract (5 and 10 µl/ml) for 24 hours. Data is expressed as relative luciferase unit/protein content. Values are expressed as mean and standard error. For LXR activity, we analyzed the data for each dose separately by ANOVA, followed by Tukey's multiple comparison test. * <i>P</i><0.05 compared indicated treatment, a; <i>P</i><0.05 compared to T0901314 treatment. For ER activity, we analyzed the data for each treatment by ANOVA, followed by comparison of all treatment groups with the control group (Dunnett's test). Statistical significance was defined as <i>P</i><0.05.</p

Composition of experimental diets.

<p>Powder was mixed to a modified AIN-93G diet enriched in fat (HF).</p>a<p>Carbohydrate: 43.30%, fiber: 13.20% protein: 28.80%, fat: 4.50%</p

WBM intake decreased fat accumulation in the liver and improved glucose clearance ability in OVX mice.

<p>(A) Macroscopic appearance of liver from mice fed the HFD and HFD+WBM for 3 months in each sham and OVX group. The tissues were fixed with 10% formalin overnight and then embedded in paraffin. Five-micrometer sections were cut and stained with hematoxylin and eosin, and examined by light microscope. Representative H&E staining of livers from each group are shown. (B) Serum glucose concentration after glucose injection in mice fed with HFD or HFD+WBM diet for 2 months. The mice were challenged with 1.5 mg glucose/g body weight glucose load. The glucose levels pre, 30, 60, 120 and 180 min post injection were measured using a glucometer. Values are expressed as mean and standard error for 8 mice. * Statistical significance was defined as <i>P</i><0.05.</p

Effects of methanol fractions from WBM extract on <i>FAS</i> and <i>ELOVL6</i> gene expressions and aromatase activity in HepG2 cells.

<p>Cells were incubated with each methanol fraction from WBM extract for 24 hours. (A) <i>FAS</i> and <i>ELOVL6</i> gene expression was normalized with the β-actin housekeeping gene. (B) Microsome assays were performed to evaluate anti-aromatase effects of WBM using the substrate, [1-β-³H] androstenedione. Activity was calculated to measure supernatant containing [³H] H₂O as reaction product, and then was counted in a Scintillation Counter. Aromatase inhibition activity was calculated as the percentage of remaining activity from the reaction without mushroom fractions. Analyses were carried out in triplicate and data were expressed as the mean ± SE. We analyzed the data for each treatment by ANOVA, followed by comparison of all treatment groups with the control group (Dunnett's test). Statistical significance was defined as <i>P</i><0.05.</p

BRCAness as a Biomarker for Predicting Prognosis and Response to Anthracycline-Based Adjuvant Chemotherapy for Patients with Triple-Negative Breast Cancer

<div><p>Background</p><p>Triple-negative breast cancer (TNBC) is a heterogeneous tumor that encompasses many different subclasses of the disease. In this study, we assessed BRCAness, defined as the shared characteristics between sporadic and <i>BRCA1</i>-mutated tumors, in a large cohort of TNBC cases.</p><p>Methods</p><p>The BRCAness of 262 patients with primary TNBCs resected between January 2004 and December 2014 was determined through the isolation of DNA from tumor tissue. Classification of BRCAness was performed using multiple ligation-dependent probe amplification (MLPA). The tumor subtypes were determined immunohistochemically using resected specimens.</p><p>Results</p><p>Of the 262 TNBCs, the results of the MLPA assays showed that 174 (66.4%) tumors had BRCAness. Patients with BRCAness tumors were younger than patients with non-BRCAness tumors (<i>P</i> = 0.003). There was no significant difference between the two groups regarding their pathological stages. The BRCAness group had a significantly shorter recurrence-free survival (RFS) compared with the non-BRCAness group (<i>P</i> = 0.04) and had a shorter overall survival (OS) although this did not reach statistical significance. Adjuvant treatments with anthracycline-based regimens provided significantly greater benefits to the BRCAness group (<i>P</i> = 0.003 for RFS, and <i>P</i> = 0.03 for OS). Multivariate Cox proportional hazard model analysis showed that BRCAness was an independent negative prognostic factor, and the anthracycline-based adjuvant chemotherapy was an independent positive prognostic factor for both RFS and OS in TNBC.</p><p>Conclusions</p><p>The 66.4% patients of TNBCs showed BRCAness. BRCAness is essential as a biomarker in the subclassification of TNBCs and might be of use for predicting their prognosis. Furthermore, this biomarker might be a predictive factor for the effectiveness of anthracycline-based adjuvant chemotherapy for patients with TNBCs.</p></div

Patients and tumor characteristics (<i>n</i> = 262).

<p>Patients and tumor characteristics (<i>n</i> = 262).</p

Kaplan–Meier analysis of patients who received anthracycline-based adjuvant chemotherapy versus non-anthracycline-based adjuvant chemotherapy.

<p>(A) Recurrence-free survival (RFS) of BRCAness tumors (<i>n</i> = 126). (B) Overall survival (OS) of BRCAness tumors (<i>n</i> = 126). (C) RFS of non-BRCAness tumors (<i>n</i> = 53). (D) OS of non-BRCAness tumors (<i>n</i> = 53). Anthracycline or Anthra, anthracycline-based adjuvant chemotherapy; Non-Anthracycline or Non-Anthra, non-anthracycline-based adjuvant chemotherapy.</p

Image_1_Immunological analysis of hybrid neoantigen peptide encompassing class I/II neoepitope-pulsed dendritic cell vaccine.jpeg

Neoantigens/ are tumor-specific antigens that evade central immune tolerance mechanisms in the thymus. Long-term tumor-specific cytotoxic T lymphocyte activity maintenance requires class II antigen-reactive CD4+ T cells. We had previously shown that intranodal vaccination with class I neoantigen peptide-pulsed dendritic cells (DCs) induced a robust immune response in a subset of patients with metastatic cancer. The present study aimed to perform a detailed ex vivo analysis of immune responses in four patients receiving an intranodal hybrid human leukocyte antigen class II neoantigen peptide encompassing a class I neoantigen epitope (hybrid neoantigen)-pulsed DC vaccine. After vaccination, strong T-cell reactions against the hybrid class II peptide and the class I-binding neoantigen peptide were observed in all four patients. We found that hybrid class II neoantigen peptide-pulsed DCs stimulated CD4+ T cells via direct antigen presentation and CD8+ T cells via cross-presentation. Further, we demonstrated that hybrid class II peptides encompassing multiple class I neoantigen epitope-pulsed DCs could present multiple class I peptides to CD8+ T cells via cross-presentation. Our findings provide insight into the mechanisms underlying hybrid neoantigen-pulsed DC vaccine therapy and suggest future neoantigen vaccine design.</p