The design and discovery of novel amide CCR5 antagonists

The synthesis of a range of novel amine-containing structures and their primary potency as inhibitors of HIV-1 fusion via blocking of the CCR5 receptor is described. The development of the medicinal chemistry strategy and SAR’s which led to the identification of the piperidine amide compounds 33 and 36 as excellent leads for further evaluation is described, along with key physicochemical data which highlighted their lead potential.     

Evaluation of creatinine phosphokinase in screening patients for malignant hyperpyrexia.

Evidence is presented that serum creatinine phosphokinase (CPK) activity is of no direct value in screening patients for susceptibility to malignant hyperpyrexia and does not correlate with halothane-induced muscle contracture or the presence of myopathy. Widely differing CPK values were found at different times in the same people. In most "malignant hyperpyrexia" families the susceptible patients had either normal or inconsistently raised CPK values.     

Evolution of ruminant Sarcocystis (Sporozoa) parasites based on small subunit rDNA sequences

We present an evolutionary analysis of 13 species of Sarcocystis, including 4 newly sequenced species with ruminants as their intermediate host, based on complete small subunit rDNA sequences. Those species with ruminants as their intermediate host form a well-supported clade, and there are at least two major clades within this group, one containing those species forming microcysts and with dogs as their definitive host and the other containing those species forming macrocysts and with cats as their definitive host. Those species with nonruminants as their intermediate host form the paraphyletic sister group to these clades. Most of the species have considerable genotypic differences (differing in more than 100 nucleotide positions), except for S. buffalonis and S. hirsuta. There is a large suite of genotypic differences indicating that those species infecting ruminant and nonruminant hosts have had very different evolutionary histories, and similarly for the felid- and canid-infecting species. Furthermore, the rDNA sequences that represent the different structural regions of the rRNA molecule have very different genotypic behavior within Sarcocystis. The evolution of these regions should be functionally constrained, and their differences can be explained in terms of the importance of the nucleotide sequences to their functions.     

Stimulation of prostaglandin-dependent macrophage suppressor cells by the subcutaneous injection of polyunsaturated fatty acids

Polyunsaturated fatty acids (PUFAs), in the form of pure linoleic, linolenic, or arachidonic acid, were injected subcutaneously into male C57Bl/6 mice daily for 10 days. Injection of 3.6 mg/day of PUFA resulted in up to a two- to threefold increase in spleen weight. Spleen cell response to mitogens was reduced by about 70%; mixed lymphocyte responses were reduced by about 90% when compared to normal values. In admixture experiments, spleen cells from PUFA treated mice suppressed the mitogen induced blastogenic response of control spleen cells by up to 90%. Fractionation of spleen cells from PUFA treated mice by G-10 adherence resulted in an enrichment of suppressive activity in the adherent cells. The suppressive effect of G-10 adherent cells was abolished by the addition of indomethacin as well as by depletion of macrophages by treatments with agents such as carbonyl iron and leucine methyl ester. These studies indicate that the administration of PUFA has marked immunosuppressive effects in mice. These effects may be related to increased prostaglandin production and appear to be mediated by a macrophage type cell.     

Helicobacter Hypothesis for Idiopathic Parkinsonism: Before and Beyond

We challenge the concept of idiopathic parkinsonism (IP) as inevitably progressive neurodegeneration, proposing a natural history of sequential microbial insults with predisposing host response. Proof-of-principle that infection can contribute to IP was provided by case studies and a placebo-controlled efficacy study of Helicobacter eradication. "Malignant" IP appears converted to "benign", but marked deterioration accompanies failure. Similar benefit on brady/hypokinesia from eradicating "low-density" infection favors autoimmunity. Although a minority of UK probands are urea breath test positive for Helicobacter , the predicted probability of having the parkinsonian label depends on the serum H. pylori antibody profile, with clinically relevant gradients between this "discriminant index" and disease burden and progression. In IP, H. pylori antibodies discriminate for persistently abnormal bowel function, and specific abnormal duodenal enterocyte mitochondrial morphology is described in relation to H. pylori infection. Slow intestinal transit manifests as constipation from the prodrome. Diarrhea may flag secondary small-intestinal bacterial overgrowth. This, coupled with genetically determined intense inflammatory response, might explain evolution from brady/hypokinetic to rigidity-predominant parkinsonism.     

Expression of engrailed proteins in arthropods, annelids, and chordates

engrailed is a homeobox gene that has an important role in Drosophila segmentation. Genes homologous to engrailed have been identified in several other organisms. Here we describe a monoclonal antibody that recognizes a conserved epitope in the homeodomain of engrailed proteins of a number of different arthropods, annelids, and chordates; we use this antibody to isolate the grasshopper engrailed gene. In Drosophila embryos, the antibody reveals engrailed protein in the posterior portion of each segment during segmentation, and in a segmentally reiterated subset of neuronal cells during neurogenesis. Other arthropods, including grasshopper and two crustaceans, have similar patterns of engrailed expression. However, these patterns of expression are not shared by the annelids or chordates we examined. Our results provide the most comprehensive view that has been obtained of how expression patterns of a regulatory gene vary during evolution. On the basis of these patterns, we suggest that engrailed is a gene whose ancestral function was in neurogenesis and whose function was co-opted during the evolution of segmentation in the arthropods, but not in the annelids and chordates.     

Nifs and Sufs in malaria

This review assembles data from three bodies of literature (bacterial genetics, plastid biogenesis and parasitology) that seldom have much direct cross-talk. After overcoming terminological complications to sort out microbial nifS from sufS genes, we connect a bacterial operon, recently found to be involved in iron metabolism, the formation of [Fe-S] clusters and oxidative stress to a potentially important gene (sufB) carried on the degenerate plastid genome of malaria and related parasites.     

Classifying aquatic macrophytes as indicators of eutrophication in European lakes

Aquatic macrophytes are one of the biological quality elements in the Water Framework Directive (WFD) for which status assessments must be defined. We tested two methods to classify macrophyte species and their response to eutrophication pressure: one based on percentiles of occurrence along a phosphorous gradient and another based on trophic ranking of species using Canonical Correspondence Analyses in the ranking procedure. The methods were tested at Europe-wide, regional and national scale as well as by alkalinity category, using 1,147 lakes from 12 European states. The grouping of species as sensitive, tolerant or indifferent to eutrophication was evaluated for some taxa, such as the sensitive Chara spp. and the large isoetids, by analysing the (non-linear) response curve along a phosphorous gradient. These thresholds revealed in these response curves can be used to set boundaries among different ecological status classes. In total 48 taxa out of 114 taxa were classified identically regardless of dataset or classification method. These taxa can be considered the most consistent and reliable indicators of sensitivity or tolerance to eutrophication at European scale. Although the general response of well known indicator species seems to hold, there are many species that were evaluated differently according to the database selection and classification methods. This hampers a Europe-wide comparison of classified species lists as used for the status assessment within the WFD implementation process.     

RNF4 is a poly-SUMO-specific E3 ubiquitin ligase required for arsenic-induced PML degradation

In acute promyelocytic leukaemia (APL), the promyelocytic leukaemia (PML) protein is fused to the retinoic acid receptor alpha (RAR). This disease can be treated effectively with arsenic, which induces PML modification by small ubiquitin-like modifiers (SUMO) and proteasomal degradation. Here we demonstrate that the RING-domain-containing ubiquitin E3 ligase, RNF4 (also known as SNURF), targets poly-SUMO-modified proteins for degradation mediated by ubiquitin. RNF4 depletion or proteasome inhibition led to accumulation of mixed, polyubiquitinated, poly-SUMO chains. PML protein accumulated in RNF4-depleted cells and was ubiquitinated by RNF4 in a SUMO-dependent fashion in vitro. In the absence of RNF4, arsenic failed to induce degradation of PML and SUMO-modified PML accumulated in the nucleus. These results demonstrate that poly-SUMO chains can act as discrete signals from mono-SUMOylation, in this case targeting a poly-SUMOylated substrate for ubiquitin-mediated proteolysis.