Novel Antiviral Agent DTriP-22 Targets RNA-Dependent RNA Polymerase of Enterovirus 71

Enterovirus 71 (EV71) has emerged as an important virulent neurotropic enterovirus in young children. DTriP-22 (4{4-[(2-bromo-phenyl)-(3-methyl-thiophen-2-yl)-methyl]-piperazin-1-yl}-1-pheny-1H-pyrazolo[3,4-d]pyrimidine) was found to be a novel and potent inhibitor of EV71. The molecular target of this compound was identified by analyzing DTriP-22-resistant viruses. A substitution of lysine for Arg163 in EV71 3D polymerase rendered the virus drug resistant. DTriP-22 exhibited the ability to inhibit viral replication by reducing viral RNA accumulation. The compound suppressed the accumulated levels of both positive- and negative-stranded viral RNA during virus infection. An in vitro polymerase assay indicated that DTriP-22 inhibited the poly(U) elongation activity, but not the VPg uridylylation activity, of EV71 polymerase. These findings demonstrate that the nonnucleoside analogue DTriP-22 acts as a novel inhibitor of EV71 polymerase. DTriP-22 also exhibited a broad spectrum of antiviral activity against other picornaviruses, which highlights its potential in the development of antiviral agents.Enterovirus 71 (EV71), a positive-stranded RNA virus, belongs to the genus Enterovirus in the family Picornaviridae. EV71 infection typically causes hand, foot, and mouth disease or herpangina, followed by severe central nervous system complications, including aseptic meningitis, encephalitis, poliomyelitis-like paralysis, neurogenic cardiopulmonary failure, and even death in some young children. Infants, following infection by EV71 with central nervous system complications, reportedly suffer from neurologic sequelae and delayed neurodevelopment (6). In 1998, a large outbreak of EV71 infection occurred in Taiwan, resulting in almost 80 fatalities and 405 severe cases (7, 21). Subsequently, many outbreaks of EV71 infection in Taiwan have been reported, with a total of 51 verified fatal cases in 2000 and 2001 (29). Severe EV71 infections continued to occur for several years thereafter. EV71 infection also has been reported to occur in many countries, such as Malaysia, Singapore, Australia, Japan, the United States, Germany, and mainland China (1-3, 5, 13, 22, 33, 34).The development of anti-EV71 agents is important because EV71 is regarded as the most important neurotropic enterovirus, after poliovirus control (34). A novel series of pyridyl imidazolidinones targeting VP1 protein, based on the skeletons of WIN compounds, has been developed using computer-assisted drug design (37). The new EV71 3C protease inhibitors, based on rhinovirus 3C protease inhibitor AG7088, exhibit inhibitory activities in both enzymatic and cell-based assays (25). A pharmacologically active drug library has been employed to identify anti-EV71 compounds (4). Ribavirin used in combination with interferon to treat patients with hepatitis C virus infection reduces the mortality rate of EV71-infected mice by reducing viral replication (26).We previously discovered piperazine-containing pyrazolo[3,4-d]pyrimidine derivatives as a novel class of anti-EV71 compounds (9). DTriP-22 (4{4-[(2-bromo-phenyl)-(3-methyl-thiophen-2-yl)-methyl]-piperazin-1-yl}-1-pheny-1H-pyr-azolo[3,4-d]pyrimidine) is one such compound, containing a diacrylmethyl group at the piperazine and a phenyl group at the pyrazolo[3,4-d]pyrimidine (9). Although DTriP-22 has a pyrazolo[3,4-d]pyrimidine structure and is thus similar to pyrazolo[3,4-d]pyrimidine nucleoside analogues, DTriP-22 differs from these analogues in that it lacks an appropriate carbocyclic ring, such as a ribose, for incorporation into the growing viral RNA chains (39). Moreover, the size of DTriP-22 differs markedly from those of the nucleoside analogues (39). In this study, we identified DTriP-22, a nonnucleoside analogue which targets EV71 3D polymerase, by analysis of DTriP-22-resistant viruses. Inhibition of 3D polymerase activity in vitro by DTriP-22 was also investigated.     

Associations between physical activity, sedentary behavior, and glycemic control in a large cohort of adolescents with type 1 diabetes: the Hvidoere Study Group on Childhood Diabetes

Background: The Hvidoere Study Group on Childhood Diabetes has demonstrated persistent differences in metabolic outcomes between pediatric diabetes centers. These differences cannot be accounted for by differences in demographic, medical, or treatment variables. Therefore, we sought to explore whether differences in physical activity or sedentary behavior could explain the variation in metabolic outcomes between centers. Methods: An observational cross‐sectional international study in 21 centers, with demographic and clinical data obtained by questionnaire from participants. Hemoglobin A1c (HbA1c) levels were assayed in one central laboratory. All individuals with diabetes aged 11–18 yr (49.4% female), with duration of diabetes of at least 1 yr, were invited to participate. Individuals completed a self‐reported measure of quality of life (Diabetes Quality of Life ‐ Short Form [DQOL‐SF]), with well‐being and leisure time activity assessed using measures developed by Health Behaviour in School Children WHO Project. Results: Older participants (p < 0.001) and females (p < 0.001) reported less physical activity. Physical activity was associated with positive health perception (p < 0.001) but not with glycemic control, body mass index, frequency of hypoglycemia, or diabetic ketoacidosis. The more time spent on the computer (r = 0.06; p < 0.05) and less time spent doing school homework (r = ?0.09; p < 0.001) were associated with higher HbA1c. Between centers, there were significant differences in reported physical activity (p < 0.001) and sedentary behavior (p < 0.001), but these differences did not account for center differences in metabolic control. Conclusions: Physical activity is strongly associated with psychological well‐being but has weak associations with metabolic control. Leisure time activity is associated with individual differences in HbA1c but not with intercenter differences.     

Transmission electron microscopic study of early stage gamma(1) (Ag(2)Hg(3)) and beta(1) (Ag-Hg) phases: technical note.

Incomplete reaction, residual free mercury, and high volatility of mercury are often major causes of the discrepancy in chemical composition of gamma(1) measured by different methods. The high mercury volatility also proves to be a major difficulty in transmission electron microscopy (TEM) study of high mercury materials. Provided in this report is a preparation method for TEM thin foils that can largely eliminate the mercury loss problem. The results show that the lattice constant of early stage gamma(1) crystals is 10.05 Delta with a chemical composition of Ag(41.08)Hg(58.92) (or Ag(2)Hg(2.87)). The chemical composition data shows that little or no mercury is lost during specimen preparation. The lattice parameters, a and c, of beta(1) crystals are 3.01 and 4.88 Delta, respectively with a chemical composition of Ag(1.1)Hg(0.94). It seems that the present specimen preparation method allows for accurate TEM microanalysis of high mercury amalgam phases such as gamma(1) and beta(1), which has been extremely difficult to do in the past.     

Role of p21 as a determinant of 1,6-Bis[4-(4-amino-3-hydroxyphenoxy)phenyl] diamantane response in human HCT-116 colon carcinoma cells

1,6-Bis[4-(4-amino-3-hydroxyphenoxy)phenyl]diamantane (DPD) induces growth inhibition in human cancer cells. In our previous study, we discovered that DPD irreversibly inhibits the growth of Colo 205 colon cancer cells at the G0/G1 phase and induces cell differentiation. However, the detailed mechanism is still unknown. In this study, we examined the functional importance of p21 and p53 in DPD-induced anticancer effects. We used three isogenic cell lines, HCT-116, HCT-116 p53-/- and HCT-116 p21-/-, to evaluate the roles of p21 and p53 in the in?vitro anticancer effects of DPD. The in?vivo anti-proliferative effect of DPD was demonstrated by HCT-116 and HCT-116 p21-/- xenograft models. DPD significantly inhibited the growth as well as increased the number of HCT-116 cells in the G0/G1 phase, but not in HCT-116 p53-/- and HCT-116 p21-/- cells examined by flow cytometry. Additionally, western blot analysis showed that DPD treatment induced p21, but not p53 protein expression in HCT-116 cells. The p21-associated cell cycle regulated proteins, such as cyclin D, CDK4 and pRb were decreased after DPD treatment in HCT-116 cells. The DPD-increased G0/G1 phase and induced cell cycle regulated protein expression were not observed in HCT-116 p21-/- and HCT-116 p53-/- cells. DPD decreased cell migration in HCT-116 and HCT-116 p53-/- but not in HCT-116 p21-/- cells. p21 was required for the DPD-induced in?vitro anti-colon cancer effect. The in?vivo study also showed that DPD significantly inhibited tumor growth through p21 signaling. Our results clearly demonstrate that DPD-induced in?vitro and in?vivo anticancer effects through the activation of p21 in HCT-116 cells.     

Critical incidents: the respiratory system

Respiratory complications are expensive, not just in terms of the overall litigation burden faced by anaesthetists but also, and more importantly, the mortality and morbidity burden faced by our patients. Critical incidents arising in the respiratory system can cause rapid deterioration if left unchecked: trauma to airway structures can be debilitating or even life threatening; hypoxaemia may result in damage to other organ systems, most notably the brain. Each patient carries their own risk profile, as well as unique ideas, concerns and expectations of their anaesthetist. An understanding of the potential critical incidents that may befall the respiratory system, a patient-centred approach to discussing these risks, and familiarity with the procedures for mitigating harm are all necessary components of safe, effective practice in anaesthesia.