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Recent research around the world has consistently reported that medical students experience a high rate of psychological morbidity, depersonalization, and low personal accomplishment. Resilience-enhancing programs have been proposed and implemented even in Japan. However, most of them remain extracurricular programs that are not specifically tailored to medical students. Additionally, they mostly mimic resiliency programs in North America, although studies have indicated that cultural perspective to the self, others, and context contribute to the capacity to respond to a stressful situation.In this context, the presenters investigated what factors might affect the similarities or differences in the perceptions of resilience among experienced palliative care physicians in Canada and Japan in 2017-2018 in order to propose a theory for a resiliency curriculum from a different cultural perspective. This study showed that Japanese physicians are more likely to rely on “Relationships” with other persons such as mentors, family, friends, or colleagues; in contrast, Canadian physicians tended to be more focused on individual factors such as “Autonomy” and “Confidence”.As a result, Showa University School of Medicine in Japan has developed a progressively advancing resiliency program for first through fourth year medical students as part of a new curriculum, implementation of which will begin in the spring of 2020. This represents one of the largest revisions in the school’s history. In this presentation, a blueprint for resiliency programs in a new curriculum will be presented, including course description, course content, educational objectives, learning resources, timetables, and instructional strategies.

Anti-HER2 antibody is molecular targeted antibody for cancer therapy. Approximately 20% of breast cancers are characterized by overexpression of HER2 protein. However, the recurrence rate was 30% and the metastasis rate was 18% one year after treatment of Anti-HER2 antibody for HER2 positive breast cancer. The resistance to antibody treatment is a major problem for patients. We previously reported that Anti-HER2 antibody and Gamma Interferon (IFN-γ) combined therapy showed higher anti-tumor effect than usual therapy in vitro and in vivo mouse experiments.In this study, we evaluated whether anti-HER2 antibody and IFN-γ combined therapy shows good synergistic effect against drug resistant HER2 positive breast cancer cells and higher antitumor effect than conventional clinical treatment. The resistant cell lines were made under the continuous presence of antibody until cell growth was not affected by the drug. We divided the resistant cells into the appropriate number of groups, which we and treated with anti-cancer therapy. We evaluated the antitumor effect for both in vitro study and in vivo mouse xenograft model prepared with the same immunogenicity. And we investigated the differences of immunofluorescence staining of CD8, Gr-1 and PDL-1 in tissues, especially related to immunity system.The combined therapy showed significantly higher anti-tumor effect than other groups in vitro and in vivo experiments. The combined therapy affects anti-tumor immunity in this immunofluorescence experiment. Taken together, we showed the possibility that combined therapy could be an effective treatment option for anti-HER2 antibody resistant breast cancer, helping patients suffering from cancer progression after developing treatment resistance.

Background/aim Benifuuki tea has recently been used as an alternative therapy for pollinosis, and it may be consumed with pharmaceutical drugs. This study aimed to examine cytochrome P450 (CYP)-mediated food-drug interactions with Benifuuki tea in rats. Materials and methods The inhibitory effects of Benifuuki tea and (-)-epigallocatechin-3-O-(3-O-methyl) gallate (EGCG3"Me) on CYP activities were evaluated in vitro. Midazolam pharmacokinetics was investigated after two treatments with Benifuuki tea. In an ex vivo study, CYP activities were determined after 1-week-treatment with the tea. Results Benifuuki tea and EGCG3"Me inhibited CYP2D and CYP3A activities in a concentration-dependent manner in vitro. However, MDZ metabolism did not change by Benifuuki treatment in vivo and ex vivo. In contrast, CYP2D activity was decreased ex vivo. Conclusion Normal intake of Benifuuki tea is not likely to cause food-drug interactions by CYP3A inhibition or induction. In contrast, Benifuuki tea consumption may lead to food-drug interactions through the inhibition of CYP2D.

Triple-negative breast cancer (TNBC), which does not show hormone sensitivity, is a poor prognosis disease without an established targeted treatment, so that establishing a therapeutic target for each subtype is desired. In addition, microRNA (miRNA), a non-cording RNA 19-25 nucleotide-longs in length, is known to be involved in regulating gene expression. We examined miRNA expression after exposure to eribulin, MDA-MB-231 cells, non-basal-like type of TNBC cell lines, and HCC1143 cells, basal-like type of TNBC cell lines. The activity of caspase-3 significantly increased compared to the control in MDA-MB-231, whereas no significant difference was observed in HCC1143. The expression level of 20-miRNAs significantly increased compared to the control in MDA-MB-231 after exposure to eribulin. The expression level of 6-miRNAs also significantly increased compared to the control in HCC1143. In these 2 cell types, miR-125b-1 and miR-195 were commonly expressed. While the expression level of miR-125b-1 decreased in both cells, the expression level of miR-195 increased in MDA-MB-231 and decreased in HCC1143. The expression level of miR-195 targeting Wnt3a significantly decreased compared to the control in MDA-MB-231, whereas it significantly increased in HCC1143. These results showed that exposure to eribulin highly increased the expression of miR-195 while it decreased the expression of Wnt3a in non-basal-like type of TNBC. Some miRNAs are known to regulate other signaling pathways involved in human pathogenesis by regulating the Wnt signaling pathway, and miRNA can act as a tumor-suppressing gene; therefore, miR-195 may serve as a therapeutic target in non-basal-like type of TNBC.

It is believed that hepatocellular damage in human chronic cholestatic liver diseases is caused by the accumulation of hydrophobic bile acids, such as glycochenodeoxycholic acid(GCDCA). Previous reports have shown that GCDCA-induced apoptosis is promoted by both mitochondria-mediated and endoplasmic reticulum(ER)stress-associated pathways in HepG2 cells and that these two pathways are linked by the action of caspase-8 on BAP31. However, the interdependence of multiple pathways remains poorly understood and the role of caspases is unclear. Thus, in the present study, we investigated the interactions among the executioner caspases in GCDCA-induced apoptotic HepG2 cells. Cells were treated for 1-24 h with GCDCA(300μM), in the presence or absence of inhibitors of caspase-9, -4 or -3(each at 30μM). Pretreatment of cells with the caspase-9 inhibitor significantly suppressed GCDCA-induced apoptosis ; however, pretreatment of cells with the caspase-4 inhibitor had no effect. Furthermore, pretreatment of cells with the caspase-9 inhibitor significantly reduced GCDCA-induced increases in caspase-3 and -4 activity, as well as the mRNA expression of BiP, a molecular chaperone located in the ER. In contrast, pretreatment of cells with the caspase-4 inhibitor had no effect. These results suggest that, in GCDCA-treated HepG2 cells, caspase-4 acts downstream of both the proapoptotic Bcl-2 protein Bax and caspase-9. Because the major mechanism underlying GCDCA-induced apoptosis involves a mitochondria-mediated pathway, it is unlikely that caspase-4 has a major role in the initiation and promotion of GCDCA-induced apoptosis.

ErbB2 is amplified in ~30% of breast cancer patients, and its amplification is associated with poor prognosis and worse survival outcome. We examined the synergistic effect of interferon-γ (IFN-γ) with anti-erbB2/neu mAb on erbB2-positive cancer model. IFN-γ showed marginal effects on its own, indicating that immune therapies mediated by this cytokine alone are unlikely to be beneficial. However, we discovered that treatment of the tumors with anti-erbB2/neu mAb concomitant with IFN-γ led to dramatic inhibition of in vivo tumor growth in the syngeneic tumor model and the preventional MMTV-neu transgenic mouse tumor model. We noted an increase of M1 macrophage accumulation in the tumor tissues as well as diminished myeloid derived suppressor cells, and increased CD8+ T cell cytotoxicity against tumor cells in the IFN-γ plus anti-erbB2/neu mAb treated mice. The tumor cells treated with both mAb and IFN-γ underwent changes in tumor stem cell marker ALDH-1, indicating a loss of stem cell-like properties, while mAb treatment alone did not accomplish this phenotypic change. We found that IFN-γ treatment significantly increase tumor PD-L1 expression, so we included anti-PD-1 or PD-L1 antibody with this combination therapy. Interestingly, anti-PD-L1 antibody further reduced tumor growth, while anti-PD-1 did not affect very much. These results indicate that the treatment of IFN-γ can improve targeted therapy, and further induce PD-L1 expression on tumor cells, creating a new target on the tumor cells for anti-PD-L1 therapy.

Phosphatase and tensin homolog deleted on chromosome 10 (PTEN) is a tumor suppressor gene that induces cell apoptosis by inhibiting the PI3K/Akt signaling pathway. Glioblastoma (GBM) is a brain tumor that is resistant to irradiation and chemotherapy and, thus, is difficult to cure. GBM stem-like cells (GSCs) have been implicated as a cause of this resistance. microRNA (miRNA/miR) inhibits the expression of proteins. The objective of the present study was to identify miRNAs that target PTEN, which induces apoptosis, in irradiation-resistant GSCs. When the expression of miRNAs was examined in GSCs irradiated at 60 Gy using the human GBM A172 cell line, the expression of PTEN-targeting miR-17-5p, -19a-3p, -19b-3p, -21-5p, -130b-3p, -221-3p and -222-3p was significantly higher in irradiated GSCs than in non-irradiated cells, and the PTEN expression levels, as revealed by immunostaining, were lower in the irradiated GSCs than in the non-irradiated cells. These results suggested that the expression of PTEN was suppressed through the overexpression of PTEN-targeting miRNAs in GSCs following irradiation.

No targeted-therapy has been established for triple-negative breast cancer accompanied by mutations in breast cancer susceptibility gene1 (BRCA1) mutation. In the present study, using BRCA1 wild-type cells (MDA-MB-231) and BRCA1-mutated cells (MDA-MB-436), we investigated miRNA expression and apoptosis on day 1 after addition of gemcitabine-alone and in combination with poly ADP-ribose polymerase-1 (PARP1) inhibitor. After drug treatment, there were significantly fewer apoptotic BRCA1 wild-type cells than BRCA1-mutated cells. Expression of miRNA-26a, -29b, -100, and -148a increased in BRCA1 wild-type cells exposed to gemcitabine-alone and in combination with the PARP1 inhibitor. The addition of PARP1 inhibitor reduced miR-206 expression in BRCA1 wild-type cells but increased it in BRCA1-mutated cells. It was suggested that miR-206 serves as a target molecule of PARP1 inhibitor combination therapy for BRCA1 wild-type triple-negative breast cancer cells.

Aeromonas (A) gum and Erwinia (E) gum are acidic heteropolysaccharides, which exist in water as expanded chains caused by the electrostatic repulsion between the charged groups. A modified approach, which was suggested to estimate miscibility of stiff polymer in our previous study, was used to determine miscibility parameter μ for blends between A gum or E gum with other polysaccharides, by using viscometry in aqueous 0.1M NaCl solution. When weight ratios of E gum/pectin were in the range of 3: 7 ∼ 7: 3, the blends in aqueous solution are miscible. The blends of A gum/xanthan, A gum/pectin, and E gum/xanthan in aqueous solution are obviously immiscible. When the weight fraction (ω 2 ) of A gum or E gum in the mixture solution is 0.7, the μ predicts that the miscibility of blend films was on the order of E gum/pectin > E gum/xanthan > A gum/pectin > A gum/xanthan. These results are in good agreement with those observed by scanning electron microscopy and Fourier transform IR.

Aeromonas (A) gum, a stabilizer, and thickening agent of food, is composed of xylose, mannose, galactose, glucose, and mannuronic acid. Weight-average molecular weights Mw and intrinsic viscosities [η] of the unfractionated A gum and three fractions in water. 0.5 M NaCl aqueous solution and cadoxen were studied by light scattering and viscometry. Experimental results showed that A gum exists as an aggregate in 0.5 M NaCl aqueous solution and mainly as a single chain in cadoxen. The apparent aggregation number (Nap) of A gum in 0.5 M NaCl aqueous solution was ca. 22, and decreased with increasing sample molecular weight. Change of aggregation and disaggregation of A gum in 0.5 M NaCl aqueous/cadoxen mixtures occurred from 0 to 0.4 of vcad (volume fraction of cadoxen), and was reversible. The stable aggregation of A gum in 0.5 M NaCl aqueous/3% cadoxen mixture was achieved after 150 min.

The foundation for targeted therapy of cancers driven by members of the ErbB oncoprotein family was established initially by the demonstration that ectodomain binding monoclonal antibodies (mAb) could disable the protein kinase encoded by the HER2/neu oncogene. Homomeric and heteromeric erbB kinases play critical roles in the development of cancer and in the spread of early lesions. In particular, antibodies targeting the p185erbB2/neu receptor provide major clinical benefits in the treatment of breast cancer and also stomach cancer. As suggested by our study with oncogenic neu transgenic mice, anti-p185erbB2/neu antibodies are also effective in preventing the tissue hyperplasia that precedes tumorigenesis, tumor growth and the dissemination of ErbB2/neu kinase-positive cells into other tissues. As a therapeutic principle, “reversion of phenotype” for established tumors and “prevention” of tumorigenesis and spread can explain the basis for the benefits invoked by therapeutic and adjuvant therapies for breast cancer patients after cancers are surgically removed. These emerging principles being enlightened by ongoing studies of monoclonal antibody therapy will continue to provide guidance for the development of new targeted therapies for resistant tumors that arise after treatment.

SummaryReversion of the malignant phenotype of erbB2-transformed cells can be driven by anti-erbB2/neu monoclonal antibodies (mAbs), which disrupt the receptor’s kinase activity. We examined the biologic effects of IFN-γ alone or after anti-erbB2/neu mAb treatment of erbB2-positive cells. IFN-γ had no effect on its own. Treatment of the tumors with anti-erbB2/neu mAbs followed by IFN-γ led to dramatic inhibition of tumor growth in vitro and in vivo with minimal mAb dosing. Sequential therapy enhanced the effects of chemotherapy. Moreover, IFN-γ with mAb treatment of mice with IFNγR knockdown tumors did not demonstrate marked synergistic eradication effects, indicating an unexpected role of IFN-γ on the tumor itself. Additionally, mAb and IFN-γ treatment also induced immune host responses that enhanced tumor eradication. Biochemical analyses identified loss of Snail expression in tumor cells, reflecting diminution of tumor-stem-cell-like properties as a consequence of altered activity of GSK3-β and KLF molecules.

Oncogenic erbB2/HER2 is a major transforming determinant of human breast cancer and its amplification is prognostic of poor survival. Monoclonal antibody (mAb)-based therapy to specifically target erbB2/HER2 has been effective in patients with HER2-positive breast cancer; however, some patients do not respond and those who do frequently relapse despite ongoing therapy. The evolution of tumors that resist targeted therapy is complex and involves the emergence of complex transcriptional functions. Snail is overexpressed in recurrent tumors of transgenic HER2/neu mice and snail expression participates in tumor recurrence. Previous studies from our laboratory and others have revealed that targeted therapy requires cooperation with elements of the endogenous immune system for optimal tumor regression activity. The cytokine interferon-gamma (IFN-γ) is relevant to the regression process although the mechanism by which IFN operates on targeted cells remains undefined and the effects of IFN on the transformed cell have not been studied during phenotypic reversion. In the present study, we utilized cell biological and biochemical-based assays to define if targeted therapy could be improved through the addition of IFN-γ and unexpectedly defined a unique mechanism operative to achieve this end. Cell culture model systems were used to examine the co-treatment of erbB2/HER2 targeting with IFN-γ to eliminate issues such as aberrant immune responses, which arise as a consequence of adding IFN-γ to cells. We defined that while mAb targeting of erbB2 reversed the transformed phenotype of HER2-positive breast cancer cells reversion of phenotype was increased upon inclusion of IFN-γ. Co-treatment with the erbB2 mAb and IFN-γ resulted in a greater reduction of snail protein levels compared to either modality alone. Snail degradation was dependent on GSK3-β and active proteasome dependent elimination processes. Furthermore, IFN-γ activated GSK3-β in mAb treated cells to the extent of that observed in cells treated with specific small molecule inhibitors to erbB receptors. Our findings provide a mechanistic explanation as to how to use IFN-γ to optimize mAb-mediated tumor regression. The implication of this study is improvement of targeted efficacy could lead to more comprehensive therapy options that may eliminate tumor recurrence. Citation Format: Edwin Aaron Runkle, Peter Young, Yasuhiro Nagai, Hiromichi Tsuchiya, Hongtao Zhang, Mark I. Greene. Interferon-gamma potentiates the targeted phenotypic reversion of erbb2/her2/neu transformed human tumor cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1390. doi:10.1158/1538-7445.AM2014-1390

2'- or 3'-O-(4-Methoxybenzyl)nucleoside derivatives were synthesized by treatment of uridine, N4-benzoylcytidine, N6-benzoyladenosine, and N2-benzoylguanosine with 4-methoxy-phenyldiazomethane. The separation of 2'- or 3'-isomers became possible on a silica gel column chromatography by using N-acylated nucleosides and these compounds could be used as useful starting materials for the synthesis of oligoribonucleotides by the phosphotriester method. The trimer blocks, U-U-Cp, A-C-Cp, and A-C-A were synthesized by the rapid method. The 3'-terminal nonanucleotide, U-U-C-A-C-C-A-C-A, of Rous Sarcoma Virus 35S RNA was obtained by the block condensation of trimers.

A highly sensitive homogeneous enzyme immunoassay has been developed for the determination of antigens with high molecular weight. The method presented is based on steric hindrance of the enzyme against an insoluble substrate by antigen binding to antibody: enzyme, conjugate, and consists of α-amylase from Bacillus subtilis as label with antibody, human α-fetoprotein (AFP) or human ferritin as ligands, and insoluble substrate. The assay procedure is as follows: sample is mixed and incubated with the conjugate at 37° C for 20 min. Thereafter, the mixture is incubated at 37° C for 20 min following addition of substrate, and the enzyme reaction is terminated by adding the stopper. Then the mixture is centrifuged at 3,000 rpm for 1 minute, and the absorbance of the supernatant is measured at 620 nm. The measurable range for AFP was 5-200 ng/ml for ferritin, it was 10-800 ng/ml. In addition, the method was well correlated with radioimmunoassay; i.e., for AFP r = 0.892 and for ferritin r = 0.951. Furthermore, there was no interference during the assay of endogenous a-amylase in human serum as a result of the use of the inhibitor specific to mammalian amylase which was purified from bacteria.

5-Chloro-8-quinolyl protecting group of internucleotidic bonds was efficiently removed by treatment with zinc acetate or 2-pyridine-carboxaldehyde oxime at room temperature. On the other hand, alkaline hydrolysis of phosphotriester intermediates with free 3'-hydroxyl function (2a) leads to the formation of 3'-3'in addition to 3'-5'-inter-nucleotidic bonds.

A new phosphorylating agent, di(2,2,2-trifluoroethyl) phosphonate (1) was found to be a useful agent for the phosphorylation of 3′-hydroxyl group of deoxyribonucleosides. When 5′-O-protected deoxyribonucleosides are treated with 1 in the absence of coupling agents and the products subjected to oxidation, the corresponding 3′-phosphodiesters are obtained in good yields. They are key intermediates for the synthesis of oligodeoxyribonucleotides by the phosphotriester approach.

The mononucleotide units (4) was prepared by the action of 2,6-dichlorophenyl 5-chloro-8-quinolyl phosphorochloridate on nucleosides. 2,6-Dichlorophenyl group in the mononucleotide units (4) have been employed as a protecting group for 3′-terminal phosphodiester functions; it is stable under standard operations required for the deoxyribooligonucleotide synthesis and readily removed by the action of t-BuNH2 in pyridine-H2O within 90 min at room temperature.