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G-quadruplex (G4) is a non-canonical nucleic acid structure formed in guanine-rich DNA or RNA. G4s are formed not only in vitro but also in vivo and are attracting considerable interest owing to their potential involvement in biological processes, including replication, transcription, mRNA splicing, translation and epigenetic regulation of the genome. In this review, we outline the functions of G4 in cellular biology and their implication in human pathogenesis, especially in cancer. Furthermore, we describe the properties of G4-stabilizing chemical compounds, G4 ligands, and their application for cancer therapeutics.

Liver metastasis (LM) is associated with poor prognosis in patients with metastatic colorectal cancer (mCRC). Here, we investigated the prognostic utility of several serum factors in mCRC patients with or without LM who were treated with anti-angiogenic agents in first-line (FL) or salvage-line (SL) settings. A combined cohort of 125 patients was analyzed in this single institute pooled analysis: FL cohort receiving bevacizumab (n = 71) and SL cohort receiving regorafenib (n = 54). Blood samples were obtained at baseline (BL) and during treatment, and serum factors were measured by ELISA. Overall survival (OS) was analyzed using Kaplan–Meier curves, the log-rank test, and Cox proportional hazard regression methods. In univariate analysis of the combined cohort, right-sided CRC, primary unresected tumor, wild-type KRAS, LM, ≥ 2 metastatic sites, and SL were associated with shorter OS; in multivariable analysis, LM and SL remained significant. Serum angiopoietin-2 (Ang-2) levels ≥ 2190.3 pg/ml and interleukin (IL)-8 levels ≥ 15.1 pg/ml at BL were significantly associated with LM. Using these cut-off values, patients with higher Ang-2 or IL-8 levels at BL had shorter OS than those with lower BL levels (Ang-2: hazard ratio [HR] 2.57, 95% confidence interval [CI] 1.47–4.51, P = 0.001; IL-8: HR 4.31, 95%CI 2.11–8.79, P < 0.001). High serum IL-8 level remained a significant predictor of shorter OS in multivariable analysis (HR 3.24, 95%CI 1.47–7.16, P = 0.004). Circulating IL-8 and Ang-2 levels are associated with LM in mCRC patients. IL-8 may be a prognostic marker of response to anti-angiogenic therapy, regardless of the treatment timing.

MICA/B proteins are expressed on the surface of various types of stressed cells, including cancer cells. Cytotoxic lymphocytes expressing natural killer group 2D (NKG2D) receptor recognize MICA/B and eliminate the cells. However, cancer cells evade such immune recognition by inducing proteolytic shedding of MICA/B proteins. Therefore, preventing the shedding of MICA/B proteins could enhance antitumor immunity. Here, by screening a protease inhibitor library, we found that the fatty-acid amide hydrolase (FAAH) inhibitor, URB597, suppresses the shedding of MICA/B. URB597 significantly reduced the soluble MICA level in culture medium and increased the MICA level on the surface of cancer cells. The effect was indirect, being mediated by increased expression of tissue inhibitor of metalloproteinases 3 (TIMP3). Knockdown of TIMP3 expression reversed the effect of URB597, confirming that TIMP3 is required for the MICA shedding inhibition by URB597. In contrast, FAAH overexpression reduced TIMP3 expression and the cell-surface MICA level and increased the soluble MICA level. These results suggest that inhibition of FAAH could prevent human cancer cell evasion of immune-mediated clearance.

Regorafenib has improved the survival of patients with refractory metastatic colorectal cancer (mCRC), yet the mechanisms of inherited or acquired resistance are not well understood. A total of 50 patients with refractory mCRC were enrolled. Circulating tumor cell (CTC) enumeration was carried out at baseline, day 21 after initiation of regorafenib, and at the time of progression of disease (PD) using the CellSearch System (Veridex LLC, NJ, USA). Poly(A) mRNA was extracted from CTCs, and gene expression of epithelial and epithelial‐mesenchymal transition markers was analyzed by a multiplex‐PCR based DNA Chip. Patients with fewer than 3 CTCs at baseline and day 21 had a longer progression‐free survival than those with 3 or more CTCs (3.3 vs 2.0 months, P = .008 and 3.3 vs 2.0 months, P = .004, respectively). Patients with fewer than 3 CTCs at baseline and day 21 had a longer overall survival (OS) than those with 3 or more CTCs (10.0 vs 4.6 months, P, We identified circulating tumor cell count as a prognostic marker in metastatic colorectal cancer patients treated with regorafenib. Circulating tumor cell EGFR expression was significantly increased with regorafenib at the time of disease progression, suggesting this to be a mode of resistance and lending further mechanistic evidence for the synergistic effects seen with regorafenib and anti‐EGFR mAbs.

Installation of a nitrogen at the C6 position of artemisinin facilitates the addition of a functional unit on the cyclohexane moiety (C-ring). In this study, conjugation of an amphiphilic chain, composed of sequentially connected hydrophilic oligoethylene glycol, hydrophobic alkyl chain, urea, and 4,4'-disubstituted biphenyl linker, imparted self-assembling properties. The fully synthetic mid-molecular weight 6-aza-artemisinin 6 bearing the amphiphilic moiety formed aggregates (approx. 200 nm) at ambient temperature and exhibited increased in vitro anti-cancer activities compared to the N-benzylated aza-artemisinin 5.

The let-7 complex in Drosophila encodes three evolutionarily conserved microRNAs: miR-100, let-7, and miR-125. These act as heterochronic genes in regulating developmental timing in response to the steroid hormone ecdysone and play important roles in cell differentiation. Here we identify two additional long non-coding RNAs in the let-7 complex, we named let-A and let-B. Both are transcribed in the large first intron of the primary RNA encoding the microRNAs. We show these RNAs to be sequentially expressed in early pupal stages in response to ecdysone signaling, albeit exhibiting a different expression pattern compared to the microRNA let-7. Surprisingly, ectopic expression of let-A in Drosophila cancer cells induces rapid cell death. Dead cells further release RNA molecules in the medium that is becoming toxic to other cancer cells. In vivo grown tumors lose their tumorigenicity after being incubated in the let-A induced medium. Moreover, feeding flies carrying transplanted tumor cells with such induced medium leads to reduced growth of tumors in a subset of hosts. Our results uncover a new lncRNA which can act as a potent and specific cell death effector for Drosophila tumor cells.

The vascular endothelial growth factor (VEGF)/VEGF receptor (VEGFR) axis is an essential regulator of angiogenesis and important therapeutic target in cancer. Ramucirumab is an anti-VEGFR2 monoclonal antibody used for the treatment of several cancers. Increased circulating VEGF-A levels after ramucirumab administration are associated with a worse prognosis, suggesting that excess VEGF-A induced by ramucirumab negatively affects treatment efficacy and that neutralizing VEGF-A may improve treatment outcomes. Here, we evaluated the effect of combination treatment with an anti-VEGFR2 antibody and anti-VEGF-A antibody on gastric tumor progression and normal tissues using a preclinical BALB/c-nu/nu mouse xenograft model. After anti-VEGFR2 antibody treatment in mice, a significant increase in plasma VEGF-A levels was observed, mirroring the clinical response. The elevated VEGF-A was host-derived. Anti-VEGF-A antibody co-administration enhanced the anti-tumor effect of the anti-VEGFR2-antibody without exacerbating the toxicity. Mechanistically, the combination treatment induced intra-tumor molecular changes closely related to angiogenesis inhibition and abolished the gene expression changes specifically induced by anti-VEGFR2 antibody treatment alone. We particularly identified the dual treatment-selective downregulation of ZEB1 expression, which was critical for gastric cancer cell proliferation. These data indicate that the dual blockade of VEGF-A and VEGFR2 is a rational strategy to ensure the anti-tumor effect of angiogenesis-targeting therapy.

G-quadruplexes (G4s) are higher-order structures formed by guanine-rich sequences of nucleic acids, such as the telomeric 5'-TTAGGG-3'/5'-UUAGGG-3' repeats and those in gene regulatory regions. G4s regulate various biological events, including replication, transcription, and translation. Imbalanced G4 dynamics is associated with diseases, such as cancer and neurodegenerative diseases. Telomestatin is a natural macrocyclic compound derived from Streptomyces anulatus 3533-SV4. It interacts with the guanine quartet via π-π stacking and potently stabilizes G4. Because G4 stabilization at the telomeric repeat inhibits the telomere-synthesizing enzyme telomerase, telomestatin was originally identified as a telomerase inhibitor. Whereas non-toxic doses of telomestatin induce gradual shortening of telomeres and eventual crisis in human cancer cells, higher doses trigger prompt replication stress and DNA damage responses, resulting in acute cell death. Suppression of the transcription and translation of G4-containing genes is also implicated in the anticancer effects of telomestatin. Because telomestatin is rare, labile, and insoluble, synthetic oxazole telomestatin derivatives have been developed and verified for their therapeutic efficacies in preclinical cancer models. Furthermore, a variety of G4-stabilizing compounds have been reported as promising seeds for molecular cancer therapeutics. To improve the design of future clinical studies, it will be important to identify predictive biomarkers of drug efficacy.

Background & Aims HBV causes hepatocellular carcinoma (HCC). While it was recently shown that the ability of HBV X protein (HBx) to impair the Smc5/6 (structural maintenance of chromosome 5/6) complex is important for viral transcription, HBx is also a potent driver of HCC. However, the mechanism by which HBx expression induces hepatocarcinogenesis is unclear. Methods Degradation of the Smc5/6 complex and accumulation of DNA damage were observed in both in vivo and in vitro HBV infection models. Rescue experiments were performed using nitazoxanide (NTZ), which inhibits degradation of the Smc5/6 complex by HBx. Results HBx-triggered degradation of the Smc5/6 complex causes impaired homologous recombination (HR) repair of DNA double-strand breaks (DSBs), leading to cellular transformation. We found that DNA damage accumulated in the liver tissue of HBV-infected humanized chimeric mice, HBx-transgenic mice, and human tissues. HBx suppressed the HR repair of DSBs, including that induced by the CRISPR-Cas9 system, in an Smc5/6-dependent manner, which was rescued by restoring the Smc5/6 complex. NTZ restored HR repair in, and colony formation by, HBx-expressing cells. Conclusions Degradation of the Smc5/6 complex by HBx increases viral transcription and promotes cellular transformation by impairing HR repair of DSBs. Lay summary The hepatitis B virus expresses a regulatory protein called HBV X protein (or HBx). This protein degrades the Smc5/6 complex in human hepatocytes, which is essential for viral replication. We found that this process also plays a key role in the accumulation of DNA damage, which contributes to HBx-mediated tumorigenesis.

Transactive response DNA-binding protein of 43 kDa (TDP-43) abnormally forms aggregates in certain subtypes of frontotemporal lobar degeneration (FTLD) and in amyotrophic lateral sclerosis (ALS). The pathological forms of TDP-43 have reported to be associated with poly(ADP-ribose) (PAR), which regulates the properties of these aggregates. A recent study has indicated that tankyrase, a member of the PAR polymerase (PARP) family, regulates pathological TDP-43 formation under conditions of stress, and tankyrase inhibitors suppress TDP-43 aggregate formation and cytotoxicity. Since we reported the development of tankyrase inhibitors that are more specific than conventional inhibitors, in this study, we examined their effects on the formation of TDP-43 aggregates in cultured cells. Time-lapse imaging showed that TDP-43 aggregates appeared in the nucleus within 30 min of treatment with sodium arsenite. Several tankyrase inhibitors suppressed the formation of aggregates and decreased the levels of the tankyrase protein. Immunohistochemical studies demonstrated that tankyrase was localized to neuronal cytoplasmic inclusions in the spinal cords of patients with ALS. Moreover, the tankyrase protein levels were significantly higher in the brains of patients with FTLD than in the brains of control subjects. These findings suggest that the inhibition of tankyrase activity protects against TDP-43 toxicity. Tankyrase inhibitors may be a potential treatment to suppress the progression of TDP-43 proteinopathies.

Macrocyclic hexaoxazoles (6OTDs) are G-quadruplex (G4) ligands, and some derivatives, such as L2H2-6OTD (1a) bearing two aminobutyl side chains, show cytotoxicity towards cancer cells. To identify the cellular target of 1a, we employed a post-target-binding strategy utilizing click reaction (Huisgen cyclization) between the azide-conjugated ligand L2H2-6OTD-Az (1b) and the cell-permeable dye CO-1 bearing a strained alkyne moiety and the BODIPY fluorophore under Cu-free conditions. We confirmed that introduction of the small azide group did not alter the physical or biological properties, including anti-cancer activity, of 1a, and we also demonstrated bias-free localization of CO-1. The post-binding visualization strategy suggested that L2H2-6OTD (1a) colocalized with RNA G4 in living cells.

Background and aims Polymerase chain reaction-based techniques require expensive equipment for fluorescence detection of the products. However, the measurement of inorganic pyrophosphate (PPi) released during DNA synthesis can be used to quantify target genes without such equipment. Here, we devised a high-sensitivity enzymatic assay for detection of PPi. Materials and methods In our assay method, PPi was converted to hypoxanthine by hypoxanthine phosphoribosyl transferase. Xanthine dehydrogenase converted the hypoxanthine to uric acid and yielded two molecules of NADH, which in turn reduced Fe3+ to Fe2+ (mediated by 1-methoxy-5-ethylphenazinium ethylsulfate). 2-Nitroso-5-(N-propyl-N-sulfopropylamino) phenol chelated the Fe2+, which resulted in an intensely colored product that could be measured using a biochemical automated analyzer. Results The assay was able to detect PPi within 10 min. It was linear between 0 and 10 µmol/L PPi, and intra-run and inter-run coefficients of variation were 1%–2%. Other validation tests with a biochemical automated analyzer were satisfactory. The assay could potentially be used to directly quantify samples after isothermal nucleic acid sequence-based amplification of a target gene. Conclusion The method developed here for detection of PPi can be used to measure nucleic acid biomarkers in biological samples in clinical practice using a high-throughput biochemical automated analyzer.

G-Quadruplex structures (G4s) in guanine-rich regions of DNA play critical roles in various biological phenomena, including replication, translation, and gene expression. The G4-forming DNAs can form three kinds of topologies, i.e., parallel, anti-parallel, and hybrid. In this paper, we present G4 ligands L2H2-2M2EA-6OTD (3) and L2G2-2M2EG-6OTD (4) bearing tetra-aminoalkyl and -guanidinylalkyl side chains, respectively, in a macrocyclic hexaoxazole structure. These ligands efficiently induce the parallel-type topology of telomeric G4 regardless of the effects of cations. Titration with 4 results in a drastic topology switch to the parallel topology from the anti-parallel structure induced by the structurally related ligand L2H2-6OTD (1).

Background The indocyanine green retention rate is important for assessing the severity of liver disorders. In the conventional method, blood needs to be collected twice. In the present study, we developed an automated indocyanine green method that does not require blood sampling before intravenous indocyanine green injections and is applicable to an automated biochemical analyser. Methods The serum samples of 471 patients collected before and after intravenous indocyanine green injections and submitted to the clinical laboratory of our hospital were used as samples. The standard procedure established by the Japan Society of Hepatology was used as the standard method. In the automated indocyanine green method, serum collected after an intravenous indocyanine green injection was mixed with the saline reagent containing a surfactant, and the indocyanine green concentration was measured at a dominant wavelength of 805 nm and a complementary wavelength of 884 nm. Results The coefficient of variations of the within- and between-run reproducibilities of this method were 2% or lower, and dilution linearity passing the origin was noted up to 10 mg/L indocyanine green. The reagent was stable for four weeks or longer. Haemoglobin, bilirubin and chyle had no impact on the results obtained. The correlation coefficient between the standard method (x) and this method (y) was r=0.995; however, slight divergence was noted in turbid samples. Conclusion Divergence in turbid samples may have corresponded to false negativity with the standard procedure. Our method may be highly practical because blood sampling before indocyanine green loading is unnecessary and measurements are simple.

G-quadruplex (G4) is a higher-order nucleic acid structure that is formed by guanine-rich sequences. G4 stabilization by small-molecule compounds called G4 ligands often causes cytotoxicity, although the potential medicinal impact of this effect has not been fully established. Here we demonstrate that a synthetic G4 ligand, Y2H2-6M(4)-oxazole telomestatin derivative (6OTD), limits the growth of intractable glioblastoma (grade IV glioma) and glioma stem cells (GSCs). Experiments involving a human cancer cell line panel and mouse xenografts revealed that 6OTD exhibits antitumor activity against glioblastoma. 6OTD inhibited the growth of GSCs more potently than it did the growth of differentiated non-stem glioma cells (NSGCs). 6OTD caused DNA damage, G1 cell cycle arrest, and apoptosis in GSCs but not in NSGCs. These DNA damage foci tended to colocalize with telomeres, which contain repetitive G4-forming sequences. Compared with temozolomide, a clinical DNA-alkylating agent against glioma, 6OTD required lower concentrations to exert anti-cancer effects and preferentially affected GSCs and telomeres. 6OTD suppressed the intracranial growth of GSC-derived tumors in a mouse xenograft model. These observations indicate that 6OTD targets GSCs through G4 stabilization and promotion of DNA damage responses. Therefore, G4s are promising therapeutic targets for glioblastoma.

// Tetsuo Mashima 1 , Yoko Taneda 1, 2 , Myung-Kyu Jang 1, 2 , Anna Mizutani 1 , Yukiko Muramatsu 1 , Haruka Yoshida 1 , Ayana Sato 1, 2 , Noritaka Tanaka 1, 3 , Yoshikazu Sugimoto 3 and Hiroyuki Seimiya 1, 2 1 Division of Molecular Biotherapy, Cancer Chemotherapy Center, Japanese Foundation for Cancer Research, Tokyo, Japan 2 Department of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of Tokyo, Tokyo, Japan 3 Division of Chemotherapy, Faculty of Pharmacy, Keio University, Tokyo, Japan Correspondence to: Hiroyuki Seimiya, email: hseimiya@jfcr.or.jp Keywords: tankyrase, Wnt, colorectal cancer, resistance, mTOR Received: August 26, 2016 Accepted: May 03, 2017 Published: May 24, 2017 ABSTRACT Activation of Wnt/β-catenin signaling is essential for colorectal carcinogenesis. Tankyrase, a member of the poly(ADP-ribose) polymerase (PARP) family, is a positive regulator of the Wnt/β-catenin signaling. Accordingly, tankyrase inhibitors are under preclinical development for colorectal cancer (CRC) therapy. However, Wnt-driven colorectal cancer cells are not equally sensitive to tankyrase inhibitors, and cellular factors that affect tankyrase inhibitor sensitivity remain elusive. Here, we established a tankyrase inhibitor-resistant cell line, 320-IWR, from Wnt/β-catenin-dependent CRC COLO-320DM cells. 320-IWR cells exhibited resistance to tankyrase inhibitors, IWR-1 and G007-LK, but remained sensitive to a PARP-1/2 inhibitor, olaparib, and several anti-CRC agents. In 320-IWR cells, nuclear localization of active β-catenin was decreased and expression of β-catenin target genes was constitutively repressed, suggesting that these cells repressed the Wnt/β-catenin signaling and were dependent on alternative proliferation pathways. 320-IWR cells exhibited upregulated mTOR signaling and were more sensitive to mTOR inhibition than the parental cells. Importantly, mTOR inhibition reversed resistance to tankyrase inhibitors and potentiated their anti-proliferative effects in 320-IWR cells as well as in CRC cell lines in which the mTOR pathway was intrinsically activated. These results indicate that mTOR signaling confers resistance to tankyrase inhibitors in CRC cells and suggest that the combination of tankyrase and mTOR inhibitors would be a useful therapeutic approach for a subset of CRCs.

Background The effectiveness of reintroducing oxaliplatin for metastatic colorectal cancer (mCRC) refractory to both oxaliplatin and irinotecan was previously reported in a phase II study (RE-OPEN). We conducted a phase I study to determine the maximum tolerated dose of oxaliplatin plus trifluridine/tipiracil (FTD/TPI) in patients with refractory mCRC. Patients and Methods Three dosages of intravenous oxaliplatin (50, 65 and 85 mg/m2) on days 1 and 15 and a fixed dose of FTD/TPI 35 mg/m2 twice daily (bid) on days 1–5 and 15–19 every 4 weeks were investigated in patients with refractory mCRC using a 3 + 3 design. Eligible patients had received prior oxaliplatin-based treatment that achieved a response or stable disease followed by confirmed disease progression at least 6 months before entering the study. Results Twelve patients were enrolled in the study. Three of six patients in the oxaliplatin 85 mg/m2 cohort had dose-limiting toxicities (DLTs) with treatment delays during the second cycle at ≥8 days due to grade ≥ 2 neutropenia or grade 2 AST/ALT increased. No DLTs were observed in the other cohorts. Grade ≥ 3 AEs were neutropenia (n = 3), thrombocytopenia (n = 1), anorexia (n = 1), and nausea (n = 1). There was no evidence of allergic reaction to oxaliplatin or severe peripheral sensory neuropathy. Conclusions A combination of FTD/TPI 35 mg/m2 bid on days 1–5 and 15–19 and oxaliplatin 85 mg/m2 on days 1 and 15 every 4 weeks could be a suitable regimen for the recommended dose of FTD/TPI plus oxaliplatin in patients with refractory mCRC.

// Mitsukuni Suenaga 1 , Tetsuo Mashima 2 , Naomi Kawata 1, 2 , Takeru Wakatsuki 1 , Yuki Horiike 1 , Satoshi Matsusaka 1 , Shingo Dan 3 , Eiji Shinozaki 1 , Hiroyuki Seimiya 2 , Nobuyuki Mizunuma 1 , Kensei Yamaguchi 1 , Toshiharu Yamaguchi 4 1 Department of Gastroenterological and Chemotherapy Cancer Institute Hospital of Japanese Foundation for Cancer Research, Koto-ku, Tokyo 135–8550, Japan 2 Division of Molecular Biotherapy, Cancer Chemotherapy Center, Japanese Foundation for Cancer Research, Koto-ku, Tokyo 135–8550, Japan 3 Division of Molecular Pharmacology, Cancer Chemotherapy Center, Japanese Foundation for Cancer Research, Koto-ku, Tokyo 135–8550, Japan 4 Department of Gastroenterological and Surgery Cancer Institute Hospital of Japanese Foundation for Cancer Research, Koto-ku, Tokyo 135–8550, Japan Correspondence to: Mitsukuni Suenaga, email: m.suenaga@jfcr.or.jp Keywords: regorafenib, metastatic colorectal cancer, VEGF-A, CCL5, prognostic marker Received: December 27, 2015 Accepted: April 16, 2016 Published: May 05, 2016 ABSTRACT Regorafenib is an oral multi-kinase inhibitor used as salvage therapy for metastatic colorectal cancer (mCRC). We tested whether serum cytokine levels are associated with clinical outcome in the mCRC patients receiving regorafenib. Serum samples were collected before treatment start, day 21, and progressive disease, and eleven angiogenic and inflammatory cytokine serum levels were examined. Fifty-four patients of a total of 62 enrolled patients were eligible for the analyses. The chemokine ligand 5 (CCL5) levels ≤ cut-off value (59959 pg/ml) at baseline was associated with relative tumor shrinkage ( P = 0.021), better progression-free survival (PFS) ( P = 0.036) and overall survival (OS) ( P = 0.019). Vascular endothelial growth factor A (VEGF-A) levels showing a decrease on day 21 were significantly associated with a better PFS ( P = 0.021). CCL5 levels ≤ cut-off was associated with any grade hand-foot skin reaction (HFSR) ( P = 0.025) and thrombocytopenia ( P = 0.013). Low chemokine ligand 2 levels at baseline were associated with grade 2 ≤ HFSR. High angiopoietin-2 and basic fibroblast growth factor (bFGF) levels at baseline were associated with grade 3 ≤ total bilirubin increase and transaminases increase, respectively. Low bFGF levels at baseline were associated with grade 3 ≤ hypertension. No correlation with severe events was observed. Baseline serum CCL5 levels and decrease of the serum VEGF-A levels may serve as potential predictive markers for survival or treatment-specific toxicities in mCRC patients receiving regorafenib.

Background The aim of this study was to investigate cholesterol and triglyceride levels in the chylomicron fraction of preterm infants at birth and during the early postnatal period. Methods The subjects consisted of 133 infants (81 boys and 52 girls): 74 were term infants born at 37-41 weeks of gestation and 59 were preterm infants born at 29-36 weeks of gestation. Cholesterol and triglyceride in the chylomicron fraction were measured using high-performance liquid chromatography. Results Compared with term infants, preterm infants had higher cholesterol and lower triglyceride in the chylomicron fraction, both in cord blood and at 1 month after birth. Thus, the chylomicron triglyceride/cholesterol ratio was significantly lower in preterm infants than in term infants in cord blood and at 1 month of age. On single regression analysis the chylomicron triglyceride/cholesterol ratio correlated positively with gestational age at birth (r = 0.331, P = 0.0003) and at 1 month (r = 0.221, P = 0.0119). Conclusions Preterm infants have a less-lipidated chylomicron composition at birth and at 1 month of age. Some prenatal factors may persist to influence chylomicron lipidation during the early postnatal period.

Polycomb group proteins are epigenetic regulators maintaining transcriptional memory during cellular proliferation. In Drosophila larvae, malfunction of Polyhomeotic (Ph), a member of the PRC1 silencing complex, results in neoplastic growth. Here, we report an intrinsic tumour suppression mechanism mediated by the steroid hormone ecdysone during metamorphosis. Ecdysone alters neoplastic growth into a nontumorigenic state of the mutant ph cells which then become eliminated during adult stage. We demonstrate that ecdysone exerts this function by inducing a heterochronic network encompassing the activation of the microRNA lethal-7, which suppresses its target gene chronologically inappropriate morphogenesis. This pathway can also promote remission of brain tumours formed in brain tumour mutants, revealing a restraining of neoplastic growth in different tumour types. Given the conserved role of let-7, the identification and molecular characterization of this innate tumour eviction mechanism in flies might provide important clues towards the exploitation of related pathways for human tumour therapy., Nature Communications, 9 (1), ISSN:2041-1723

The role of the telomere repeat-binding factor 2 (TRF2) in telomere maintenance is well-established. However, recent findings suggest that TRF2 also functions outside telomeres, but relatively little is known about this function. Herein, using genome-wide ChIP-Seq assays of TRF2-bound chromatin from HT1080 fibrosarcoma cells, we identified thousands of TRF2-binding sites within the extra-telomeric genome. In light of this observation, we asked how TRF2 occupancy is organized within the genome. Interestingly, we found that extra-telomeric TRF2 sites throughout the genome are enriched in potential G-quadruplex–forming DNA sequences. Furthermore, we validated TRF2 occupancy at several promoter G-quadruplex motifs, which did adopt quadruplex forms in solution. TRF2 binding altered expression and the epigenetic state of several target promoters, indicated by histone modifications resulting in transcriptional repression of eight of nine genes investigated here. Furthermore, TRF2 occupancy and target gene expression were also sensitive to the well-known intracellular G-quadruplex–binding ligand 360A. Together, these results reveal an extensive genome-wide association of TRF2 outside telomeres and that it regulates gene expression in a G-quadruplex–dependent fashion.

Glioblastoma (GBM) is an invariably fatal brain tumor in which a small subpopulation of self-renewable glioma stem cells (GSCs) contributes to tumor propagation and relapse. Targeting GSCs could therefore have a significant clinical impact for GBM. Telomestatin is a naturally-occurring compound that preferentially impairs GSC growth by perturbing transcription and inducing a DNA damage response. Telomestatin stabilizes G-quadruplexes (G4s), which are guanine-rich four-strand nucleic acid structures observed in vitro and in vivo . However, the mechanism underlying the GSC-selective nature of the DNA damage response remains unknown. Here we demonstrate that GSCs are more susceptible to telomestatin-induced telomere dysfunction and replication stress when compared with GSC-derived non-stem glioma cells (NSGCs). Telomestatin induced dissociation of the telomere-capping protein TRF2 from telomeres, leading to telomeric DNA damage in GSCs—but not in NSGCs. BIBR1532, a telomerase catalytic inhibitor, did not preferentially inhibit GSC growth, suggesting that telomestatin promotes telomere dysfunction in a telomerase-independent manner. GSCs and NSGCs had comparable levels of G4s in their nuclei, and both responded to telomestatin with phosphorylation of RPA2 at Ser33—a hallmark of replication stress. However, activation of the checkpoint kinase Chk1, induction of a DNA damage response, and subsequent growth inhibition occurred only in telomestatin-treated GSCs. These observations suggest that telomestatin impairs GSC growth through removal of TRF2 from telomeres and potent activation of the replication stress response pathway. Therefore, a novel G4-directed therapeutic strategy could specifically target cancer stem cells in GBM.

Background Proteomic approaches may provide new insights into pathological conditions associated with alcoholism. The aim of this study was to conduct a proteomic analysis of liver tissue and serum in chronically alcohol-fed rats using agarose 2-dimensional gel electrophoresis (2-DE) and 3-step serum proteome analysis. Methods A total of 12 rats were pair-fed nutritionally adequate liquid diet containing ethanol as 36% of the total energy or an isocaloric control diet for 2 months. Rat liver homogenates and cytosol fractions were subjected to agarose 2-DE. Serum samples were subjected to 3-step serum proteome analysis involving immunodepletion of abundant proteins followed by fractionation using reverse-phase high-performance liquid chromatography and 1-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Candidate proteins were digested with trypsin and identified using mass spectrometry. Observed differences in protein expression levels were confirmed using Western blotting. Results A total of 46 protein spots were found to be differentially expressed in the liver homogenates and cytosol fractions of alcohol-fed rats relative to pair-fed controls. The most notable change was down-regulation of a 29-kDa protein, which was subsequently identified as carbonic anhydrase III (CA III). Down-regulation of this protein in alcohol-fed rats was confirmed by Western blotting. The messenger RNA level of CA III was decreased as well. In rat serum, a total of 41 proteins were differentially expressed. Of these proteins, only betaine–homocysteine methyltransferase (BHMT) was also found to be differentially expressed in the liver. Conclusions A combined proteomic analysis of liver tissue and serum in chronically alcohol-fed rats revealed that the expression of CA III is significantly down-regulated in the liver of alcohol-fed rats. Our results also showed that BHMT expression is up-regulated in both the liver and serum of alcohol-fed rats.

Japanese barnyard millet is an important food source in East Asian countries. However its crumbly texture limits desirability and consumption. Controlling amylose level in the endosperm is important to improve the eating quality of the millet. Because it is well known that the waxy gene determines the amylose level in the endosperm, we conducted a molecular analysis of the gene. Segregation analysis revealed that wild-type cultivars had three functional genes while low-amylose cultivars had one. We determined complete sequences of the three homoeologous waxy structural genes, EeWx1, EeWx2 and EeWx3, in a wild-type cultivar. These sequences showed high homology in the exon regions (97 %), and lower homology in the introns (82 %). Two spontaneous mutations were characterized in the low-amylose cultivars. In addition, one induced mutation was found in the fully waxy cultivar, Chojuromochi. Spontaneous mutations are deletions of whole and terminal regions in the EeWx2 and EeWx3 alleles, respectively. The induced mutation is a single-base deletion that led to a premature termination codon in EeWx1. These findings led us to develop useful markers for selecting low-amylose and waxy lines in millet.

Purpose: Glioma stem cells (GSC) are a critical therapeutic target of glioblastoma multiforme (GBM). Experimental Design: The effects of a G-quadruplex ligand, telomestatin, were evaluated using patient-derived GSCs, non-stem tumor cells (non-GSC), and normal fetal neural precursors in vitro and in vivo. The molecular targets of telomestatin were determined by immunofluorescence in situ hybridization (iFISH) and cDNA microarray. The data were then validated by in vitro and in vivo functional assays, as well as by immunohistochemistry against 90 clinical samples. Results: Telomestatin impaired the maintenance of GSC stem cell state by inducing apoptosis in vitro and in vivo. The migration potential of GSCs was also impaired by telomestatin treatment. In contrast, both normal neural precursors and non-GSCs were relatively resistant to telomestatin. Treatment of GSC-derived mouse intracranial tumors reduced tumor sizes in vivo without a noticeable cell death in normal brains. iFISH revealed both telomeric and non-telomeric DNA damage by telomestatin in GSCs but not in non-GSCs. cDNA microarray identified a proto-oncogene, c-Myb, as a novel molecular target of telomestatin in GSCs, and pharmacodynamic analysis in telomestatin-treated tumor-bearing mouse brains showed a reduction of c-Myb in tumors in vivo. Knockdown of c-Myb phenocopied telomestatin-treated GSCs both in vitro and in vivo, and restoring c-Myb by overexpression partially rescued the phenotype. Finally, c-Myb expression was markedly elevated in surgical specimens of GBMs compared with normal tissues. Conclusions: These data indicate that telomestatin potently eradicates GSCs through telomere disruption and c-Myb inhibition, and this study suggests a novel GSC-directed therapeutic strategy for GBMs. Clin Cancer Res; 18(5); 1268–80. ©2012 AACR.

The effects of certain disinfectants on the stability of a polymethyl methacrylate denture base resin were investigated, including those of a novel disinfection method using reactive oxygen species (ROS). The surface roughness and flexural strength were analyzed to assess the effects of the disinfectants on material properties. The following disinfectants were tested: 5% sodium hypochlorite, 70% alcohol, and ROS. Furthermore, the attachment of Candida albicans to the resin surface was investigated. The disinfection method using sodium hypochlorite significantly increased the surface roughness and decreased flexural strength. The surface roughness and flexural strength of the ROS-treated specimens did not significantly differ from those of the control specimens, and the ROS-treated specimens exhibited diminished Candida attachment. These results demonstrate that the ROS disinfection method preserves acceptable material stability levels in polymethyl methacrylate resins.

The fruit of Illicium verum Hook. f. (star anise) is used by many as a spice. The fragrance of I. verum fruit is characteristically anise-like. In this study, hexane-extracted I. verum fruit oil (IVO), trans-anethole as the main component, and related compounds (propiophenone, 4′-methoxy-propiophenone, trans-β-methylstyrene) were analyzed in order to clarify the emotional effect of inhaling the fragrance of I. verum fruit. As a result, although 4 μL/L air IVO did not exhibit an anxiolytic-like effect, 1 μL/L air trans-anethole exhibited a significant effect (p

Al-3.7 pct Si-0.18 pct Mg foams strengthened by AlN particle dispersion were prepared by a melt foaming method, and the effect of foaming temperature on the foaming behavior was investigated. Al-3.7 pct Si-0.18 pct Mg alloy containing AlN particles was prepared by noncompressive infiltration of Al powder compacts with molten Al alloy in nitrogen atmosphere, and it was foamed at different foaming temperatures ranging from 1023 to 1173 K. The porosity of prepared foam decreases and the pore structure becomes homogeneous with increasing foaming temperature. When the foaming temperature is higher than 1123 K, homogeneous pores are formed in the prepared ingot without using oxide particles and metallic calcium granules, which are usually used for stabilizing a foaming process. This stabilization of the foaming at high temperatures is possibly caused by Al3Ti intermetallic compounds formed at high temperature and AlN particles. Compression tests for the prepared foams revealed that the absorbed energy per unit mass of prepared Al-3.7 pct Si-0.18 pct Mg foam is higher than those of aluminum foams strengthened by alloying or dispersion of reinforcements. It is remarkable that the oscillation in stress, which usually appears in strengthened aluminum foams, does not appear in the plateau stress region of the present Al-3.7 pct Si-0.18 pct Mg foam. The homogeneity in cell walls and pore morphology due to the stabilization of pore formation and growth by AlN and Al3Ti particles is a possible cause of this smooth plateau stress region.

Insulin-like growth factor (IGF) signaling plays a pivotal role in cell proliferation and mitogenesis. Secreted IGF-binding proteins (IGFBPs) are important modulators of IGF bioavailability; however, their intracellular functions remain elusive. We sought to assess the antiapoptotic properties of intracellular IGFBP-2 in lung adenocarcinomas. IGFBP-2 overexpression resulted in a decrease in procaspase-3 expression; however, it did not influence the phosphorylation status of either IGF receptor or its downstream targets, including Akt and extracellular signal-regulated kinase. Apoptosis induced by camptothecin was significantly inhibited by IGFBP-2 overexpression in NCI-H522 cells. Conversely, selective knockdown of IGFBP-2 using small-interfering RNA resulted in an increase in procaspase-3 expression and sensitization to camptothecin-induced apoptosis in NCI-H522 cells. LY294002, an inhibitor of phosphatidyl-inositol 3-kinase, caused a decrease in IGFBP-2 levels and enhanced apoptosis in combination with camptothecin. Immunohistochemistry demonstrated that intracellular IGFBP-2 was highly expressed in lung adenocarcinomas compared with normal epithelium. Intracellular IGFBP-2 and procaspase-3 were expressed in a mutually exclusive manner. These findings suggest that intracellular IGFBP-2 regulates caspase-3 expression and contributes to the inhibitory effect on apoptosis independent of IGF. IGFBP-2, therefore, may offer a novel therapeutic target and serve as an antiapoptotic biomarker for lung adenocarcinoma.

79 Background: Ramucirumab (RAM) showed significantly improved survivals of gastric cancer in the second line with paclitaxel or single use. It is known that RAM causes pharmacodynamic changes of plasma VEGF-A family levels during treatment. However, its clinical significance is still unclear. The aim of this study is to reveal the changes of plasma biomarker levels until disease progression, and to examine associations between plasma biomarker levels at disease progression and survival after RAM treatment failure. Methods: Plasma samples were collected at three points: base line, day 8, and disease progression. Nine kinds of plasma biomarker related to angiogenesis: VEGF-A, C, D, PlGF, sVEGFR-1, 2, Neuropilin-1 (NRP1), Angiopietin-1, and SDF-1α, were measured by means of ELISA. The dynamics of plasma biomarkers were compared using the ANOVA. Patients were dichotomized by optimal cut-off value in each biomarker. Survival after RAM treatment failure was estimated by Kaplan-Maier methods and compared by log-rank test. Results: Plasma samples were collected from 21 patients. Median age was 67 y.o and female was dominant (57%). Plasma VEGF-A and PlGF levels sharply increased at day 8 and these higher plasma levels were sustained until disease progression: the median VEGF-A and PlGF levels at baseline, day 8, and disease progression were 20.2, 350.2, and 596.7 pg/ml for VEGF-A (p

Hepatocellular carcinoma (HCC) is the third-leading cause of cancer-related mortality worldwide (1). Although its early diagnosis can lead to curative treatment by operation or radiofrequency ablation (2), sorafenib, a multikinase inhibitor, has long been used for patients with advanced tumors and is the only drug available for treating advanced HCC (3). However, the clinical benefit of sorafenib is limited and almost all patients become rapidly refractory to therapy. Recently, two other multikinase inhibitors, regorafenib and lenvatinib, have become available (4), but sorafenib is still widely used worldwide. Thus, the development of novel therapeutic options against HCC, particularly against sorafenib-resistant HCC, is urgently needed.

Telomeres are the capping structures of the eukaryotic chromosome ends. Tankyrase 1 is a poly(ADP-ribose) polymerase that elongates telomeres in a telomerase-dependent manner. This function of tankyrase 1 is mediated by down-regulation of TRF1, a negative regulator of telomere access to telomerase. Namely, tankyrase 1 poly(ADP-ribosyl)ates (PARsylates) TRF1, which in turn dissociates TRF1 from telomeres. The resulting telomeres become better substrates for telomerase-mediated DNA extension. Tankyrase 1 has five independent TRF1 binding sites, ARC (ANK repeat cluster) I to V. Among them, the most C-terminal ARC V is required for TRF1 PARsylation and its release from telomeres. By contrast, functional significance of other four ARCs remains elusive. In this study, we generated a mutant tankyrase 1 that had inactive ARC IV and lacked ARC V but elongated telomeres without TRF1 PARsylation. Consistent with the failure in PARsylation, this mutant only marginally released TRF1 from telomeres. Still, it decreased telomere binding of POT1, a downstream effector of TRF1-mediated telomere length control, and elongated the telomeric 3'-overhang as the wild-type tankyrase 1 did. Thus even without TRF1 PARsylation, this mutant tankyrase 1 seemed to loosen the closed structure of the telomeric heterochromatin. These findings suggest a new role for multiple ARCs in telomere extension by tankyrase 1.

The biological function of hnRNP family proteins is widely diverse and involved in pre-mRNA processing, transcriptional regulation, recombination, and telomere maintenance. In the course of our study on the elucidation of biological functions of minisatellite DNA, we isolated several nuclear proteins that bind to the mouse minisatellite Pc-1, which consists of a tandem array of d(GGCAG) repeats, from NIH3T3 cells. One of the minisatellite binding proteins, MNBP-A, which binds to a single-stranded G-rich strand of the Pc-1 repeat, was proven identical to the hnRNP A3. Recombinant hnRNP A3 was demonstrated to bind to the single-stranded telomeric d(TTAGGG) repeat with much higher affinity than the d(GGCAG) repeat. Binding of hnRNP A3 to the single-stranded telomeric repeat protected the repeat from nuclease attack, and inhibited both telomerase reaction and DNA synthesis in vitro. These results suggest a possible biological role of hnRNP A3 in the stable maintenance of telomere repeats.

G-quadruplexes (G4) are thought to be important factors for telomerase inhibition and transcriptional/translational modulations. Bioinformatic analyses imply that the human genome and mRNA contain a multitude of G4-forming sequences; however, their analysis requires selective and detectable ligands. Given that two molecules of fluorescent berberine (BBR) coordinate to telomeric G4 in their co-crystals, we designed hydrocarbon-linked BBR-analogue dimers because we expected the alignment of two BBR chromophores would avoid Watson-Crick base pair intercalation, which should result in high selectivity towards G4. An alkene-cis-C2 BBR dimer showed the highest affinity (Kd ≤2.6 nM) and selectivity (ca. 900-fold vs. duplex) towards G4. The intrinsic "light-up" fluorescence properties of this BBR dimer, derived from its conformational switching by G4, allowed a selective visualization of various G4 in the gel without using additional bulky fluorescence dyes, which, combined with the observed lack of conformational change of the ligand, suggested future applications in in vitro detection systems.

BACKGROUND Although serum albumin levels (sALB) have been quantified by the dye-binding method with bro- mocresol green (BCG) or bromocresol purple (BCP) on a routine basis, accurate measurement of sALB with these methods is difficult. The modified BCP method is highly specific to albumin without being affected by sample preservation to enable stable and accurate quantification of albumin concentrations. A change in the albumin measurement method may alter the diagnosis of nephrotic syndrome. METHODS sALB was measured in 295 patients including 98 patients with chronic renal disease by the modified BCP method, BCG method, and immunonephelometry as the gold standard. RESULTS sALB measured by the modified BCP method was well correlated with levels measured by immunonephelometry. sALB obtained by the BCG method was significantly higher than the levels measured by the modified BCP method (p < 0.001, Student's t-test). This tendency was more evident in patients with chronic renal disease than other patients. When the threshold value of sALB for the diagnosis criteria of nephrotic syndrome (≤ 25 g/L) and a high risk of thrombosis (≤ 20 g/L) in nephrotic syndrome was based on the BCG method, the revised criteria in the modified BCP method would be ≤ 20.5 and ≤ 14.9 g/L, respectively. CONCLUSIONS Overestimation of sALB by the BCG method affected diagnosis of nephrotic syndrome. The method by which sALB is measured should be specified in both clinical and research settings in nephrology.

We have further developed the clip-type strain gauge dilatometer in order to measure an angular deformation by applying the cosine theorem. In this research, not only a linear thermal expansion along three crystallographic axes but also a monoclinic angle β of ferroelectric TGS in the monoclinic system were measured below 380 K down to the cryogenic temperature region. Each of these components showed a clear bend and a typical feature of the second order transition at the Curie temperature. On the other hand, the volume thermal expansion did not show a big change at the Curie temperature.

Inhibition of telomerase activity by telomerase inhibitors induces a gradual loss of telomeres, and this in turn causes cancer cells to enter to a crisis stage. Here, we report the telomerase inhibitor telomestatin, which is known to stabilize G-quadruplex structures at 3' single-stranded telomeric overhangs (G-tails), rapidly dissociates TRF2 from telomeres in cancer cells within a week, when given at a concentration that does not cause normal cells to die. The G-tails were dramatically reduced upon short-term treatment with the drug in cancer cell lines, but not in normal fibroblasts and epithelial cells. In addition, telomestatin also induced anaphase bridge formation in cancer cell lines. These effects of telomestatin were similar to those of dominant negative TRF2, which also causes a prompt loss of the telomeric G-tails and induces an anaphase bridge. These results indicate that telomestatin exerts its anticancer effect not only through inhibiting telomere elongation, but also by rapidly disrupting the capping function at the very ends of telomeres. Unlike conventional telomerase inhibitors that require long-term treatments, the G-quadruplex stabilizer telomestatin induced prompt cell death, and it was selectively effective in cancer cells. This study also identifies the TRF2 protein as a therapeutic target for treating many types of cancer which have the TRF2 protein at caps of the telomere DNA of each chromosome.

With a purpose of obtaining materials of high specific strength at room and high temperatures, Al-8at%Mg mixed powder was mechanically alloyed with addition of oxide powders (GeO2, SnO2, PbO) of the Group IV elements of the periodic table. Solid state reactions during mechanical alloying (MA) and subsequent thermomechanical processing were studied. The MA powders were consolidated to the P/M materials by vacuum hot pressing and hot extrusion, and their solid state reaction and mechanical properties were studied. It was observed by X-ray diffraction that PbO was decomposed during MA, while GeO2 and SnO2 were decomposed during hot pressing. Reduced Ge and Sn were combined with Mg to form Mg2Ge and Mg2Sn, while reduced Pb remained as metallic Pb. Preferential oxidation of solute Mg occurred resulting in formation of MgO and MgAl2O4. The P/M material of Al-Mg-GeO2 system is harder and more ductile than those of Al-Mg-SnO2 and Al-Mg-PbO systems. The P/M material of Al-Mg-GeO2 showed the highest tensile strength of 629 MPa at RT and 361 MPa at 473K.

Clast5/Stra13/DEC1 is a member of the helix-loop-helix family of transcriptional repressors. We have previously shown that Clast5 is rapidly down-regulated upon B cell activation and its overexpression inhibits cell cycle progression in B lymphoma cells. In the present study, we show that Clast5 expression is developmentally regulated during B cell differentiation, being expressed at the progenitor B cells, down-regulated at the precursor B cells, elevated in immature and mature resting B lymphocytes, and down-regulated again in germinal center B cells. To investigate the function of Clast5 in regulating lymphocyte development, we have generated transgenic mice expressing Clast5 in B- and T-lineage cells (Clast5-Tg). Clast5-Tg mice grew and bred normally but their spleen and thymus cellularity was reduced compared with control littermates. The development of B cells in the bone marrow and T cells in the thymus was impaired, with the expansion of progenitor B and T cells most strongly affected. The frequency of IL-7-responsive cells in the bone marrow of Clast5-Tg mice was reduced by >80% and their proliferative response to IL-7 was also compromised. Mature B cells from Clast5-Tg mice were hyporesponsive to antigen receptor cross-linking and exhibited mild reduction in the proliferative response to CD40 ligation or lipopolysaccharide stimulation. Moreover, the development of germinal center B cells and antibody production against a T-dependent antigen were reduced in Clast5-Tg mice. These results reveal a critical role for Clast5/Stra13/DEC1 in negatively regulating lymphocyte development and function in vivo.

We prepared Bi-2223 sintered bulk composed with silver alloy wires to improve simultaneously both the superconductivity and the mechanical properties. When 24 Ag alloy-wires of 0.4 mm in diameter were composed, the critical current density ( J c ) at 77.3 K in the self-field was obtained 700 A/cm 2 . Scanning electron microscope observation results showed that dense structure of Bi-2223 plate-like grains was formed around the interfacial region between the Bi-2223 oxide and metal Ag. The mechanical property was estimated by three point bending test. The bending stress of 80 MPa at the rupture was obtained for Bi-2223 AgMg alloy composite bulk irrespective of the number of the wires. In the AgZr alloy-wires composite bulks, the bending stress decreased to the half value of that in Bi-2223 bulk without the wires. After the maximum bending stress the sample fractured separately in Bi-2223 bulk without alloy-wires, while the composite did not fracture but only fine cracks were induced.

The enthalpy of hydrophobic hydration, Δ H HH , was separately estimated for each apolar segment of amphiphiles and of related compounds. The specimens analyzed in this study were the homologs and analogs of sodium alkyl sulfate, alkyltrimethylammonium bromide, ammonium perfluoroalkylcarboxylate, and polyethylene glycols. The Δ H HH was calculated from the adiabatic compressibility β of aqueous solution systematically measured for each homologs at varying concentrations and temperatures. Those data reported in the previous paper were utilized. The Δ H HH estimated for each CH 2 , CF 2 , and OCH 2 CH 2  segments in both hydrophobic and hydrophilic parts of the amphiphiles was found to change depending on its locations in the molecule. The Δ H HH for the first CH 2  unit which is adjacent to the head sulfate group of H(CH 2 ) n SO 4 Na was found to be negligibly small; however, it increases gradually as the position of the unit moves apart from the anionic head group and attains as large as −26.5 kcal/mol for the eighth CH 2 . The change of Δ H HH of CF 2  unit in F(CF 2 ) n COONH 4 shows the similar tendency to H(CH 2 ) n SO 4 Na. The value of Δ H HH is as small as −8 kcal/mol for the first CF 2 , however, it increases to −16 kcal/mol for the fifth CF 2 , which is 1.5 times larger than that of the fifth CH 2  of alkyl sulfate. Contrary to this, all the Δ H HH of the methylenes of cationic surfactant ranging from the second to the eighth CH 2  units of H(CH 2 ) n N(CH 3 ) 3 Br take a similar value around −8 kcal/mol of CH 2  irrespective of their positions. The first CH 2  unit which is equivalent to that of quaternary ammonium [HCH 2 ] 4 NBr shows slightly higher value of −9.0 kcal/mol of the first CH 2  than those for the second to eighth CH 2 . This seems to suggest that the cationic core nitrogen has an enhancing effect on hydrophobic hydration. The hydration enthalpy Δ H OEH of each oxyethylene (OE) unit of polyethylene glycol was also estimated for H(OCH 2 CH 2 ) p OH ( p =1–4) homologues to see the roles of ether oxygen and hydroxyl groups in hydrophobic hydration. The value of Δ H OEH showed its maximum −13.1 kcal/mol for the OE unit located at the middle of three. The other two units at both ends were found to take the value of −9.9 kcal/mol, smaller than that at the center. This could suggest that the hydrophobic hydration of the ethylene segment is largely enhanced by the neighboring ether oxygens.

72 Background: Ramucirumab, anti-VEGFR2 receptor antibody, showed significantly improved survivals of gastric cancer in the second line with paclitaxel or single use; however, no validated biomarker has been available until today. The aim of this study is 1) to reveal the dynamics of plasma biomarkers, 2) to assess the associations between these baseline levels or dynamics and efficacy. Methods: Plasma samples were collected at three points: base line, day 8, and after disease progression. Nine kinds of plasma biomarker involved in angiogenesis were measured by means of ELISA. Progression-free survival (PFS) was estimated using Kaplan-Meier methods and compared using the log-rank test. Comparisons of plasma biomarker levels were performed using Mann-Whitney-U test. Results: Samples were collected from 35 patients. Among them, 25 patients were evaluated when analyzed and 6 patients showed disease progression at the first evaluation. In VEGF-A and PlGF, plasma biomarker levels of day 8 were dramatically elevated in all patients; Day8/ baseline ratios were 18.1 folds in VEGF-A and 19.3 folds in PlGF. On the other hand, Day8/ baseline ratio of VEGFR1 and NRP1 were 0.7 and 0.8, respectively. The others appeared to show variable change at day 8. In survival analysis, lower than median Day8/ baseline ratios of VEGF-A were significantly associated with longer PFS with 6.3 months in < median vs. 2.4 months in ≥ median (p = 0.004). In addition, patients with disease progression at the first evaluation by CT showed higher Day8/ baseline ratios of VEGF-C, Angiopoietin 1, and lower baseline NRP1 levels than those of patients with non-disease progression. Conclusions: Dramatic changes of biomarker levels at day 8 were observed among patients in each biomarker. These changes in VEGF-A, C, and Angiopoietin-1 may be candidate predictive marker of ramucirumab in Japanese patients with gastric cancer.

Star anise is an important fragrance material that has a characteristic anise-like odor. Although the main component of star anise is ( E)-anethole, which accounts for over 90% of the constituents, the odor of ( E)-anethole is different from that of the material itself. Here, we examined the aroma profile of star anise. GC-MS analysis of star anise extracts showed that it contains many compounds with structures similar to ( E)-anethole. Our results indicate that ( E)-anethole is the key compound in the odor of star anise, but structurally similar compounds play an important role in creating its odor. We examined the structure-odor relationship of ( E)-anethole, focusing on the methoxy and 1-propenyl substituents. Altering the 1-propenyl group changed the odors of all the anethole derivatives. Replacing the methoxy group with a hydrogen atom created compounds with similar fatty odors. This shows that the methoxy group is important for the characteristic odor of anethole. We synthesized anethole derivatives where the methoxy group was replaced with a methyl group. In both methoxy- and methyl-substituted anethole derivatives, altering the 1-propenyl group changed the odors of the derivatives. Therefore, the methoxy and methyl benzene moieties are important structural features for the odor of star anise. The structural characteristics of anethole are closely related to its odor expression.

The hydrophilicity of polar and apolar domains of various amphiphiles was systematically estimated for their homologues and analogues by measuring the molar adiabatic compressibility of an aqueous solution at infinite dilution. The homologues of protic alkyl H(CH(2))(n)-, perfluoroalkyl F(CF(2))(n)-, and alkylphenyl H(CH(2))(n)(C(6)H(5))- groups (n=0-10) were chosen to represent apolar hydrophobic domains. The polar hydrophilic domains tested were -SO(4)Na, -SO(3)Na, -COONH(4), -N(CH(3))(3)Br, N(C(m)H(2m+1))(4)Br (m=1-5), and -NH(CH(2))(n)SO(3) (n=3, 4) groups. Also tested were the tetraphenyl ionic compounds (C(6)H(5))(4)MX (M=B/X=Na, M=P/X=Cl, M=As/X=Cl) to study the effect of the ionic sign of the core atom across the tetraphenyl apolar shell, the polyethylene glycols H(OCH(2)CH(2))(m)OH (m=1-4) to study the role of apolar -CH(2)- units in the hydrophilic oxyethylene group, and the zwitterionic dimethylaminoalkylsulfonate (CH(3))(2)NH(CH(2))(n)SO(3) homologues to study the effect of intramolecular salt formation on the hydrophilicity of the zwitterion. The adiabatic compressibility of the solution was calculated from measurement of the sound velocity and density of solutions. The introduction of laboratory automation and the numerical control of the system improved the accuracies and efficiencies of the measurements a great deal. The range of the temperature scan was 0-40 degrees C with an effective accuracy of +/-0.001 degrees C and the concentration was automatically scanned down to far below the cmc of the surfactant. The hydrophilicity of various polar and apolar substances was estimated as the decrease of molar adiabatic compressibility of the aqueous solution with increased concentration of their homologues and analogues. The hydrophobic hydration of nonpolar substances was found to be very small at room temperature and was barely detected above 40 degrees C; however, it became large as the temperature was lowered and attained a maximum at 0 degrees C. The cationic charge of quaternary ammonium N(+)(C(n)H(2n+1))(4) was found to enhance the hydrophobic hydration of methylene groups located at a distance of 4 to 6 Å from the core nitrogen atom, while the terminal negative charge of the anionic surfactant R-SO(4)(-), R-SO(3)(-), or R-COO(-) was found to decrease the hydrophobic hydration of -CH(2)- units within the same range. The hydrophilicity of quaternary ammonium and the tetraphenyl ions should be synergistically given by both hydrophobic and ionic hydrations. The hydrophilicity of the perfluoromethylene unit -CF(2)- was found to have a value comparable to that of the protic methylene unit -CH(2)-. The hydrophobic hydration seems to offer a good measure of the hydrophilicity of apolar substances; however, it does not necessarily represent the "hydrophobicity" of the apolar segment when the "surface activity" of the amphiphile is concerned. Copyright 2000 Academic Press.

The apparent molar volume and the apparent molar adiabatic compressibility of the sugar-based surfactants methyl 6-O-octanoyl-α-d-glucopyranoside (α-MOnG), methyl 6-O-octanoyl-β-d-glucopyranoside (β-MOnG), and octyl β-d-glucoside were measured over a wide concentration range. Also, the aggregation number of their micelles was determined from the Debye plot using static light scattering data. It was found that the micellar aggregation number for α-MOnG is 179 at 35 °C, which is 1.5 times larger than that for β-MOnG, suggesting that the anomerism of the head group influences the packing of the monomers during micelle formation.

Velocity distributions of desorbing product CO2 were studied in the CO oxidation on Pt(557) over a wide range of surface temperatures, in both the active and inhibited regions at the steady state. The translational energy of CO2 in the active region, where the reaction is in a first order with respect to CO, is very high and it is insensitive to the surface temperature. It decreases sharply with increasing CO partial pressure in the boundary region. In the inhibited region, where the reaction is retarded by CO, the translational energy is relatively low and increases with increasing surface temperature.

The interaction between sodium hyaluronate (NaHy) and decyltrimethylammonium bromide (C 10 TAB) has been investigated by phase behavior, viscosity, and dynamic shear moduli studies. When the concentration of C 10 TAB, C a , was between 0.03 and 0.35 M and the polymer concentration was above 0.05 g L -1 , the system separated into two phases. For the isotropic solutions containing a large excess of surfactant (C s ≥ 0.35 M), the boundary between the dilute and semidilute regime was determined by using the zero-shear specific viscosity data. Further, the intrinsic viscosity and the dynamic shear moduli of NaHy in C 10 TAB solutions were compared with those in NaCl solutions. The viscosity results suggested that NaHy and C 10 TAB formed compact complexes in 0.35 M C 10 TAB to give a small intrinsic viscosity and the size of the complexes increased with the increase of C 10 TAB concentration. When the added salt concentration was 1.00 M and the polymer concentration was in semidilute regime, it was suggested from the data on dynamic shear moduli that the entanglement density of Hy chains in C 10 TAB was higher than that in NaCl.

Angular and velocity distributions of desorbing CO 2 molecules formed by the CO oxidation on a Pt(113) surface were studied. Measurements were performed by means of angle-resolved thermal desorption combined with time-of-flight techniques. At initial reactant coverages of 0.10≤ Θ O ≤0.30 and 0.08≤ Θ CO ≤0.15, the CO 2 formation mostly occurs around 300–400 K during heating procedures. The working reaction sites are found to be both on the (111) terrace and the (100) step where the preferential occurrence of the reaction depends significantly on the reactant coverages. The translational temperature T 〈 E 〉 of these CO 2 molecules reaches around 1300 K. No significant difference in T 〈 E 〉 was found between the terrace and the step sites. On the other hand, at a high coverage of Θ O =0.35 and Θ CO =0.54, five CO 2 species are produced over a wide temperature range of 150–400 K. The reaction takes place both on the terrace and the step sites. T 〈 E 〉 for these CO 2 molecules is higher than or around 1300 K on the terrace sites. A decrease in T 〈 E 〉 of 150–270 K was found on the step sites in comparison with that on the terrace sites. The change of the reactant coverage affects to some extent the translational energy of the product CO 2 . Such a difference in the translational energy is discussed through the energy transfer mechanism between the product molecule and the substrate surface.