New Neplanocin Analogues. V. A Potent Adenosylhomocysteine Hydrolase Inhibitor Lacking Antiviral Activity. Synthesis And Antiviral Activity Of 6″-Carboxylic Acid Derivatives Of Neplanocin A 1

Abstract

The 6′-carboxylic acid derivative of neplanocin A 3 was synthesized from NPA, and was converted to the corresponding methyl ester 4 and amides 5 and 6. These were evaluated for their anti-RNA-virus activities. Of the derivatives synthesized, only 5 was active against RNA viruses within the concentration range of 0.14-4.88 μg/mL. Compounds 3 and 5 showed a potent inhibitory effect on S-adenosylhomocysteine (AdoHcy) hydrolase from rabbit erythrocytes. Although a close correlation between the inhibitory effect of adenosine analogues on AdoHcy hydrolase and their antiviral potency has been demonstrated, 3 did not show any anti-RNA-virus activities.

     

Structural Characterization of Cyclic ADP-Ribose by NMR Spectroscopy

Abstract

Structure of cyclic adenosine diphosphoribose (cADPR) was reinvestigated by using ¹H, ¹³C, and ³¹P NMR spectroscopy. The ¹H-¹H coupling constants and NOE data suggested that the adenosine and ribose moieties have a predominant C2′-endo conformation and an unusual flat conformation, respectively.     

Evaluation of high sensitive C-reactive protein and 5-10 methylenetetrahydrofolate reductase genotype in Japanese young adults

Primary objective: To carry out a preliminary evaluation of subclinical inflammation and its genetic background in young adults.

Research design: Fifty-five healthy Japanese young adults aged 19–27 years (37 males and 18 females, mean age: 22.3 years), and 58 healthy Japanese adults aged 40 to 60 years (21 males and 37 females, mean age: 51.5 years) were included in this study.

Methods and procedures: We measured plasma high-sensitive C-reactive protein (hs-CRP) levels and screened for the C677T polymorphism of the 5-10 methylenetetrahydrofolate reductase gene (MTHFR), which is considered a genetic risk factor for atherosclerosis, by HinfI digestion.

Main outcomes and results: Hs-CRP levels of the young adult group were significantly lower than the levels of the middle aged group (0.014±0.030 mg/dl vs. 0.031±0.040 mg/dl, p=0.005). The levels were significantly higher in males than in females (0.028±0.019 mg/dl vs. 0.013±0.010 mg/dl, p=0.008) among young adults. Furthermore, we evaluated the relationship of the C677T genotype and hs-CRP values, but found no association between them.

Conclusions: Although the sample size is limited, our preliminary study demonstrated the profiles of hs-CRP in Japanese young adults. Further investigation will be needed to establish the guidelines for customized school health education using sensitive laboratory and genetic markers.     

Rates of Serious Intracellular Infections in Autoimmune Disease Patients Receiving Initial Glucocorticoid Therapy

Background/Aims

The Japanese National Hospital Organization evidence-based medicine (EBM) Study group for Adverse effects of Corticosteroid therapy (J-NHOSAC) is a Japanese hospital-based cohort study investigating the safety of the initial use of glucocorticoids (GCs) in patients with newly diagnosed autoimmune diseases. Using the J-NHOSAC registry, the purpose of this observational study is to analyse the rates, characteristics and associated risk factors of intracellular infections in patients with newly diagnosed autoimmune diseases who were initially treated with GCs.

Methodology/Principal Findings

A total 604 patients with newly diagnosed autoimmune diseases treated with GCs were enrolled in this registry between April 2007 and March 2009. Cox proportional-hazards regression was used to determine independent risk factors for serious intracellular infections with covariates including sex, age, co-morbidity, laboratory data, use of immunosuppressants and dose of GCs. Survival was analysed according to the Kaplan-Meier method and was assessed by the log-rank test. There were 127 serious infections, including 43 intracellular infections, during 1105.8 patient-years of follow-up. The 43 serious intracellular infections resulted in 8 deaths. After adjustment for covariates, diabetes (Odds ratio [OR]: 2.5, 95% confidence interval [95% CI] 1.1–5.9), lymphocytopenia (≦1000/μl, OR: 2.5, 95% CI 1.2–5.2) and use of high-dose (≧30 mg/day) GCs (OR: 2.4, 95% CI 1.1–5.3) increased the risk of intracellular infections. Survival curves showed lower intracellular infection-free survival rate in patients with diabetes, lymphocytopaenia and high-dose GCs treatments.

Conclusions/Significance

Patients with newly diagnosed autoimmune diseases were at high risk of developing intracellular infection during initial treatment with GCs. Our findings provide background data on the risk of intracellular infections of patients with autoimmune diseases. Clinicians showed remain vigilant for intracellular infections in patients with autoimmune diseases who are treated with GCs.     

Quantitative Proteomic Profiling Identifies DPYSL3 as Pancreatic Ductal Adenocarcinoma-Associated Molecule That Regulates Cell Adhesion and Migration by Stabilization of Focal Adhesion Complex

Elucidation of how pancreatic cancer cells give rise to distant metastasis is urgently needed in order to provide not only a better understanding of the underlying molecular mechanisms, but also to identify novel targets for greatly improved molecular diagnosis and therapeutic intervention. We employed combined proteomic technologies including mass spectrometry and isobaric tags for relative and absolute quantification peptide tagging to analyze protein profiles of surgically resected human pancreatic ductal adenocarcinoma tissues. We identified a protein, dihydropyrimidinase-like 3, as highly expressed in human pancreatic ductal adenocarcinoma tissues as well as pancreatic cancer cell lines. Characterization of the roles of dihydropyrimidinase-like 3 in relation to cancer cell adhesion and migration in vitro, and metastasis in vivo was performed using a series of functional analyses, including those employing multiple reaction monitoring proteomic analysis. Furthermore, dihydropyrimidinase-like 3 was found to interact with Ezrin, which has important roles in cell adhesion, motility, and invasion, while that interaction promoted stabilization of an adhesion complex consisting of Ezrin, c-Src, focal adhesion kinase, and Talin1. We also found that exogenous expression of dihydropyrimidinase-like 3 induced activating phosphorylation of Ezrin and c-Src, leading to up-regulation of the signaling pathway. Taken together, the present results indicate successful application of combined proteomic approaches to identify a novel key player, dihydropyrimidinase-like 3, in pancreatic ductal adenocarcinoma tumorigenesis, which may serve as an important biomarker and/or drug target to improve therapeutic strategies.     

A Japanese child with geleophysic dysplasia caused by a novel mutation of FBN1

Geleophysic dysplasia (GD) is a rare disorder characterized by severe short stature, short hands and feet, limited joint mobility, skin thickening, characteristic facial features (e.g., a “happy” face), and cardiac valvular disorders that often result in an early death. The genes ADAMTSL2 (a disintegrin-like and metalloprotease with thrombospondin type 1 motif-like 2) and FBN1 (fibrillin 1) were recently identified as causative genes for GD. Here, we describe a 10-year-old Japanese female with GD who was born to non-consanguineous parents. At the age of 11 months, she was referred to our hospital because of very short stature for her age (− 4.4 standard deviations of the age-matched value) and a “happy” face with full cheeks, a shortened nose, hypertelorism, and a long and flat philtrum, characteristic of GD. Her hands and feet were small, her skin was thickened, and her joint mobility was generally limited. She had cardiac valvular disorders and history of recurrent respiratory failure. Mutation analysis revealed no abnormalities in ADAMTSL2. However, analysis of FBN1 revealed a novel heterozygous mutation (c.5161T > T/G) in exon 41, which encodes transforming growth factor-β-binding protein-like domain 5 (TB5). GD is an extremely rare disorder and, to our knowledge, only one case of GD with an FBN1 mutation has been reported in Japan. Similar to the previously reported cases of GD, the mutation in the current patient was located in the TB5 domain, which suggests that abnormalities in this domain of FBN1 are responsible for GD.     

miR‐197 induces epithelial–mesenchymal transition in pancreatic cancer cells by targeting p120 catenin

Invasive ductal adenocarcinoma (IDA) of the pancreas manifests poor prognosis due to the early invasion and distant metastasis. In contrast, intraductal papillary mucinous adenoma or carcinoma (IPMA or IPMC) reveals better clinical outcomes. Various molecular mechanisms contribute to these differences but entire picture is still unclear. Recent researches emphasized the important role of miRNA in biological processes including cancer invasion and metastasis. We previously described that miR‐126 is down‐regulated in IDA compared with IPMA or IPMC, and miR‐126 regulates the expression of invasion related molecule disintegrin and metalloproteinase domain‐containing protein 9 (ADAM9). Assessing the difference of miRNA expression profiles of IDA, IPMA, and IPMC, we newly identified miR‐197 as an up‐regulated miRNA specifically in IDA. Expression of miR‐197 in pancreatic cancer cells resulted in the induction of epithelial–mesenchymal transition (EMT) along with the down‐regulation of p120 catenin which is a putative target of miR‐197. Direct interaction between miR‐197 and p120 catenin mRNA sequence was confirmed by 3′UTR assay, and knockdown of p120 catenin recapitulated EMT induction in pancreatic cancer cells. In situ hybridization of miR‐197 and immunohistochemistry of p120 catenin showed mutually exclusive patterns suggesting pivotal role of miR‐197 in the regulation of p120 catenin. This miR‐197/p120 catenin axis could be a novel therapeutic target. J. Cell. Physiol. 228: 1255–1263, 2013. © 2012 Wiley Periodicals, Inc.     

Hyperthermia inhibits homologous recombination repair and sensitizes cells to ionizing radiation in a time‐ and temperature‐dependent manner

Hyperthermia has long been known as a radio‐sensitizing agent that displays anti‐tumor effects, and has been developed as a therapeutic application. The mechanisms of hyperthermia‐induced radio‐sensitization are highly associated with inhibition of DNA repair. Our investigations aimed to show how hyperthermia inactivate homologous recombination repair in the process of sensitizing cells to ionizing radiation by using a series of DNA repair deficient Chinese Hamster cells. Significant differences in cellular toxicity attributable to hyperthermia at and above 42.5°C were observed. In wild‐type and non‐homologous end joining repair mutants, cells in late S phase showed double the amount heat‐induced radio‐sensitization effects of G1‐phase cells. Both radiation‐induced DNA double strand breaks and chromatin damage resulting from hyperthermia exposure was measured to be approximately two times higher in G2‐phase cells than G0/G1 cells. Additionally, G2‐phase cells took approximately two times as long to repair DNA damage over time than G0/G1‐phase cells. To supplement these findings, radiation‐induced Rad51 foci formations at DNA double strand break sites were observed to gradually dissociate in response to the temperature and time of hyperthermia exposure. Dissociated Rad51 proteins subsequently re‐formed foci at damage sites with time, and occurred in a trend also related to temperature and time of hyperthermia exposure. These findings suggest Rad51's dissociation and subsequent reformation at DNA double strand break sites in response to varying hyperthermia conditions plays an important role in hyperthermia‐induced radio‐sensitization. J. Cell. Physiol. 228: 1473–1481, 2013. © 2012 Wiley Periodicals, Inc.