Phthalazinones 2: Optimisation and synthesis of novel potent inhibitors of poly(ADP-ribose)polymerase

We have previously described the discovery of poly(ADP-ribose)polymerase-1 (PARP-1) inhibitors based on a phthalazinone scaffold. Subsequent optimisation of inhibitory activity, metabolic stability and pharmacokinetic parameters has led to a novel series of meta-substituted 4-benzyl-2H-phthalazin-1-one PARP-1 inhibitors which retain low nM cellular activity and show good stability in vivo and efficacy in cell based models.     

Folding and misfolding mechanisms of the p53 DNA binding domain at physiological temperature

p53 modulates a large number of cellular response pathways and is critical for the prevention of cancer. Wild-type p53, as well as tumorigenic mutants, exhibits the singular property of spontaneously losing DNA binding activity at 37 degrees C. To understand the molecular basis for this effect, we examine the folding mechanism of the p53 DNA binding domain (DBD) at elevated temperatures. Folding kinetics do not change appreciably from 5 degrees C to 35 degrees C. DBD therefore folds by the same two-channel mechanism at physiological temperature as it does at 10 degrees C. Unfolding rates, however, accelerate by 10,000-fold. Elevated temperatures thus dramatically increase the frequency of cycling between folded and unfolded states. The results suggest that function is lost because a fraction of molecules become trapped in misfolded conformations with each folding-unfolding cycle. In addition, at 37 degrees C, the equilibrium stabilities of the off-pathway species are predicted to rival that of the native state, particularly in the case of destabilized mutants. We propose that it is the presence of these misfolded species, which can aggregate in vitro and may be degraded in the cell, that leads to p53 inactivation.     

Ecological risk and contamination history of heavy metals in the Andong tidal flat,Hangzhou Bay,China

In this study, we collected two sediment cores (C1 and C2) from the Andong tidal flat, Hangzhou Bay, and studied the temporal variations of heavy metals in the cores. Vertical distributions of heavy metals were almost unchanged in both the cores before 2000. After 2000, however, the heavy metal concentrations increased dramatically, suggesting that the sediments have been affected by enhanced human pollution in the recent decade. In the core C1, the sediments were severely polluted by Pb, moderately to considerably polluted by Cr and Zn, and low to moderately polluted by other heavy metals. The core C2 was relatively unpolluted before 2000 and low to moderately polluted after 2000. Multi-statistical analyses indicated that the core C1 was additionally contaminated by local human activities such as wastewater discharge and the Hangzhou Bay Bridge. The heavy metals in the core C2, however, were largely contributed by the Yangtze River and controlled by sedimentation process. The calculated sedimentary flux (4–8 g m^?2 a^?1) of heavy metals generally increased with time. It was closely related to the wastewater discharge in adjacent areas. This study reconstructed the local heavy metal pollution history and provides important information for environmental protection and policy making.     

Bayesian semiparametric intensity estimation for inhomogeneous spatial point processes

In this work we propose a fully Bayesian semiparametric method to estimate the intensity of an inhomogeneous spatial point process. The basic idea is to first convert intensity estimation into a Poisson regression setting via binning data points on a regular grid, and then model the log intensity semiparametrically using an adaptive version of Gaussian Markov random fields to smooth the corresponding counts. The inference is carried by an efficient Markov chain Monte Carlo simulation algorithm. Compared to existing methods for intensity estimation, for example, parametric modeling and kernel smoothing, the proposed estimator not only provides inference regarding the dependence of the intensity function on possible covariates, but also uses information from the data to adaptively determine the amount of smoothing at the local level. The effectiveness of using our method is demonstrated through simulation studies and an application to a rainforest dataset.     

Unidirectional cross-activation of GRPR by MOR1D uncouples itch and analgesia induced by opioids

Spinal opioid-induced itch, a prevalent side effect of pain management, has been proposed to result from pain inhibition. We now report that the μ-opioid receptor (MOR) isoform MOR1D is essential for morphine-induced scratching (MIS), whereas the isoform MOR1 is required only for morphine-induced analgesia (MIA). MOR1D heterodimerizes with gastrin-releasing peptide receptor (GRPR) in the spinal cord, relaying itch information. We show that morphine triggers internalization of both GRPR and MOR1D, whereas GRP specifically triggers GRPR internalization and morphine-independent scratching. Providing potential insight into opioid-induced itch prevention, we demonstrate that molecular and pharmacologic inhibition of PLCβ3 and IP3R3, downstream effectors of GRPR, specifically block MIS but not MIA. In addition, blocking MOR1D-GRPR association attenuates MIS but not MIA. Together, these data suggest that opioid-induced itch is an active process concomitant with but independent of opioid analgesia, occurring via the unidirectional cross-activation of GRPR signaling by MOR1D heterodimerization.     

The polypyrimidine/polypurine motif in the mouse mu opioid receptor gene promoter is a supercoiling-regulatory element

The mu opioid receptor (MOR) is the principle molecular target of opioid analgesics. The polypyrimidine/polypurine (PPy/u) motif enhances the activity of the MOR gene promoter by adopting a non-B DNA conformation. Here, we report that the PPy/u motif regulates the processivity of torsional stress, which is important for endogenous MOR gene expression. Analysis by topoisomerase assays, S1 nuclease digests, and atomic force microscopy showed that, unlike homologous PPy/u motifs, the position- and orientation-induced structural strains to the mouse PPy/u element affect its ability to perturb the relaxation activity of topoisomerase, resulting in polypurine strand-nicked and catenated DNA conformations. Raman spectrum microscopy confirmed that mouse PPy/u containing-plasmid DNA molecules under the different structural strains have a different configuration of ring bases as well as altered Hoogsteen hydrogen bonds. The mouse MOR PPy/u motif drives reporter gene expression fortyfold more effectively in the sense orientation than in the antisense orientation. Furthermore, mouse neuronal cells activate MOR gene expression in response to the perturbations of topology by topoisomerase inhibitors, whereas human cells do not. These results suggest that, interestingly among homologous PPy/u motifs, the mouse MOR PPy/u motif dynamically responds to torsional stress and consequently regulates MOR gene expression in vivo.     

Length–weight relationships for 39 species of moray eel from the waters around Taiwan

Length–weight relationships were determined in 39 muraenid species belonging to seven genera collected during 2003–2008 from the waters around Taiwan. Various fishing methods were used to collect both the adult and pre‐adult moray eel specimens. Length–weight relationship records of 28 species are provided for the first time. New maximum size records of six species were also established: 61.2 cm for Gymnothorax monostigma, 88.2 cm for G. niphostigmus, 68.6 cm for G. shaoi, 53.9 cm for G. taiwanensis, 73.2 cm for G. thyrsoideus, and 37.3 cm for U. micropterus.     

Molecular and morphological characterization of neural tube defects in embryos of diabetic Swiss Albino mice

BACKGROUND AND RESULTS: Embryos from diabetic mice exhibit several forms of neural tube defects, including non-closure of the neural tube. In the present study, embryos collected at embryonic day 11.5 from diabetic pregnancies displayed open neural tube with architectural disruption of the surrounding tissues. The percentage of proliferating cells was found to be increased in the dorsal and ventral domains of the spinal neural tube of embryos from diabetic mice, indicating a defect in the proliferation index. We have analyzed the development of various cell types, including motoneurons, interneurons, oligodendrocytes and migrating neurons, as well as radial glial cells in the open neural tube using specific molecular markers. Immunofluorescence results revealed a significantly reduced number of Pax2+ interneurons and increased number of Isl-1+ motoneurons, as well as Olig2+ oligodendrocytes in the neural tube of embryos from diabetic mice as compared to controls. In addition, these embryos exhibited a decreased number of doublecortin positive migrating neurons and Glast/Blbp positive radial glial cells with shortened processes in the neural tube. Expression levels of several developmental control genes involved in the generation of different neuronal cell types (such as Shh, Ngn, Ngn2, Ascl1) were also found to be altered in the neural tube of embryos from diabetic mice. CONCLUSIONS: Overall, the open neural tube in embryos of diabetic mice exhibits defects in the specification of different cell types, including motoneurons and interneurons, as well as glial cells along the dorsoventral axis of the developing spinal cord. Although these defects are associated with altered expression of several development control genes, the exact mechanisms by which maternal diabetes contributes to these changes remain to be investigated.