Chlorproguanil?Dapsone?Artesunate versus Artemether?Lumefantrine: A Randomized,Double-Blind Phase III Trial in African Children and Adolescents with Uncomplicated Plasmodium falciparum Malaria

Background

Chlorproguanil−dapsone−artesunate (CDA) was developed as an affordable, simple, fixed-dose artemisinin-based combination therapy for use in Africa. This trial was a randomized parallel-group, double-blind, double-dummy study to compare CDA and artemether−lumefantrine (AL) efficacy in uncomplicated Plasmodium falciparum malaria and further define the CDA safety profile, particularly its hematological safety in glucose-6-phosphate dehydrogenase (G6PD) -deficient patients.

Methods and Findings

The trial was conducted at medical centers at 11 sites in five African countries between June 2006 and August 2007. 1372 patients (≥1 to <15 years old, median age 3 years) with acute uncomplicated P. falciparum malaria were randomized (2∶1) to receive CDA 2/2.5/4 mg/kg once daily for three days (N = 914) or six-doses of AL over three days (N = 458). Non-inferiority of CDA versus AL for efficacy was evaluated in the Day 28 per-protocol (PP) population using parasitological cure (polymerase chain reaction [PCR]-corrected). Cure rates were 94.1% (703/747) for CDA and 97.4% (369/379) for AL (treatment difference –3.3%, 95%CI –5.6, −0.9). CDA was non-inferior to AL, but there was simultaneous superiority of AL (upper 95%CI limit <0). Adequate clinical and parasitological response at Day 28 (uncorrected for reinfection) was 79% (604/765) with CDA and 83% (315/381) with AL. In patients with a G6PD-deficient genotype (94/603 [16%] hemizygous males, 22/598 [4%] homozygous females), CDA had the propensity to cause severe and clinically concerning hemoglobin decreases: the mean hemoglobin nadir was 75 g/L (95%CI 71, 79) at Day 7 versus 97 g/L (95%CI 91, 102) for AL. There were three deaths, unrelated to study medication (two with CDA, one with AL).

Conclusions

Although parasitologically effective at Day 28, the hemolytic potential of CDA in G6PD-deficient patients makes it unsuitable for use in a public health setting in Africa.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00344006","term_id":"NCT00344006"}}NCT00344006     

Signaling from an altered cell wall to the nucleus mediates sugar-responsive growth and development in Arabidopsis thaliana

Sugars such as glucose function as signal molecules that regulate gene expression, growth, and development in plants, animals, and yeast. To understand the molecular mechanisms of sugar responses, we isolated and characterized an Arabidopsis thaliana mutant, high sugar response8 (hsr8), which enhances sugar-responsive growth and gene expression. Light-grown hsr8 plants exhibited increased starch and anthocyanin and reduced chlorophyll content in response to glucose treatment. Dark-grown hsr8 seedlings showed glucose-hypersensitive hypocotyl elongation and development. The HSR8 gene, isolated using map-based cloning, was allelic to the MURUS4 (MUR4) gene involved in arabinose synthesis. Dark-grown mur1 and mur3 seedlings also exhibited similar sugar responses to hsr8/mur4. The sugar-hypersensitive phenotypes of hsr8/mur4, mur1, and mur3 were rescued by boric acid, suggesting that alterations in the cell wall cause hypersensitive sugar-responsive phenotypes. Genetic analysis showed that sugar-hypersensitive responses in hsr8 mutants were suppressed by pleiotropic regulatory locus1 (prl1), indicating that nucleus-localized PRL1 is required for enhanced sugar responses in hsr8 mutant plants. Microarray analysis revealed that the expression of many cell wall-related and sugar-responsive genes was altered in mur4-1, and the expression of a significant proportion of these genes was restored to wild-type levels in the mur4-1 prl1 double mutant. These findings reveal a pathway that signals changes in the cell wall through PRL1 to altered gene expression and sugar-responsive metabolic, growth, and developmental changes.     

CADMIUM ALTERS BEHAVIOUR AND THE BIOSYNTHETIC CAPACITY FOR CATECHOLAMINES AND SEROTONIN IN NEONATAL RAT BRAIN1

Abstract— Daily exposure to cadmium (10 μg/100g) for 30 days since birth significantly increased spontaneous locomotor activity as well as striatal tyrosine hydroxylase and mid-brain tryptophan hydroxylase. The endogenous levels of norepinephrine, dopamine and serotonin failed to change in various brain regions of cadmium-treated rats. In contrast, the concentration of 5-hydroxyindoleacetic acid tended to rise but was significantly different from controls only in the mid-brain region. The data suggest that cadmium treatment in early life increased the synthesis and physiological utilization of these putative transmitters which in turn probably altered locomotor performance. Increasing the dose of cadmium to 100 μg/100 g for 30 days decreased body weight (by 19%) and produced slight increases in the turnover of brain amines. However, the rise was not dose-dependent. Furthermore, the locomotor activity remained the same as that seen in rats treated with the low dose of cadmium. The levels of dopamine in hypothalamus and that of norepinephrine in several brain regions examined were enhanced. This could in part, be attributed to decreased (12%) activity of catechol-O-methyl transferase enzyme. Administration of the high dose of cadmium produced significant increases in 5-hydroxyindoleacetic acid level. Data suggest that cadmium acts at some step in the sequence of intracellular events that leads to increased synthesis and presumably turnover of brain catecholamines and serotonin. Since high dosage of this heavy metal failed to produce a dose-dependent change in locomotor activity, it is not possible to impute any casual role for these amines in the production of hyperactivity seen in cadmium-treated rats.     

Aspects of the biochemical toxicology of cadmium.

Cadmium, in addition to producing a variety of toxic manifestations, is known to accumulate in certain "target" organs which include liver and kidney where histological and functional damage becomes apparent. The daily intraperitoneal injection of cadmium chloride for 21 or 45 days stimulated the activities of hepatic pyruvate carboxylase, phosphoenolpyruvate carboxykinase, fructose-1, 6-diphosphatase and glucose-6-phosphatase elevated blood glucose and urea, and lowered hepatic glycogen in rats. Whereas chronic Cd treatment failed to alter adenosine-3', 5'-monophosphate phosphodiesterase (PDE) activity, cyclic AMP (cAMY and the activity of basal and fluoride-stimulated forms of hepatic adenylate cyclase (AC) were markedly increased. However, the cAMP binding to hepatic protein kinase was decreased as was the kinase activity ration. An acute dose of Cd decreased hepatic glycogen content and increased blood glucose, serum urea, and hepatic cAMP. Chronic exposure to Cd induced adrenal hypertrophy and augmented adrenal norepinephrine and epinephrine as well as the activity of adrenal tyrosine hydroxylase. This treatment decreased prostatic and testicular weights of mature rats. Although cAMP as well as AC activity of the prostate gland were reduced, cAMP binding to the prostatic protein kinase was increased as was the activity of the cAMP-dependent form of the enzyme. Testicular AC and PDE activities, however, were stimulated, although cAMP remained unaffected. Whereas the activities of the cAMP-dependent and the independent forms of testicular protein kinase were significantly depressed, the binding of cAMP to protein kinase from testes of Cd-treated rats was not affected. In most cases, the observed metabolic alterations persisted up to 28 days on cessation of Cd administration. Subacute Cd treatment suppressed pancreatic function as evidenced by lowered serum immunoreactive insulin (IRI) in presence of hyperglycemia, as well as by partial inhibition of phentolamine-stimulated increases in serum IRI. Although chronic Cd treatment failed to alter the concentration of brain stem norepinephrine and cerebrocortical acetylcholine esterase activity, serotonin levels of brain stem were depressed and the concentration of striatal dopamine and cerebrocortical acetylcholine were significantly elevated when compared with the values seen in control nonexposed animals.     

Catecholamines inEntamoebae: Recent (re)discoveries

Free-living and enteric amoebae have similar two-stage life cycles, and both organisms depend on being able to monitor environmental conditions to determine whether to continue multiplying as trophozoites, or to differentiate into dormant or transmissible cysts. Conditions that support high trophozoite densities might also be expected to select for mechanisms of information exchange between these cells. We recently determined that trophozoites of at least one species ofEntamoeba release and respond to catecholamine compounds during differentiation from the trophozoite stage into the cyst stage. It turns out that this is not an entirely novel finding, as a number of previous studies have demonstrated parts of this story in free-living or enteric amoebae. We briefly review here major points of the previous studies and describe some of our recent results that have extended them.     

Homocysteine modulates 5‐lipoxygenase expression level via DNA methylation

Elevated levels of homocysteinemia (Hcy), a risk factor for late‐onset Alzheimer's disease (AD), have been associated with changes in cell methylation. Alzheimer's disease is characterized by an upregulation of the 5‐lipoxygenase (5LO), whose promoter is regulated by methylation. However, whether Hcy activates 5LO enzymatic pathway by influencing the methylation status of its promoter remains unknown. Brains from mice with high Hcy were assessed for the 5LO pathway and neuronal cells exposed to Hcy implemented to study the mechanism(s) regulating 5LO expression levels and the effect on amyloid β formation. Diet‐ and genetically induced high Hcy resulted in 5LO protein and mRNA upregulation, which was associated with a significant increase of the S‐adenosylhomocysteine (SAH)/S‐adenosylmethionine ratio, and reduced DNA methyltrasferases and hypomethylation of 5‐lipoxygenase DNA. In vitro studies confirmed these results and demonstrated that the mechanism involved in the Hcy‐dependent 5LO activation and amyloid β formation is DNA hypomethylation secondary to the elevated levels of SAH. Taken together these findings represent the first demonstration that Hcy directly influences 5LO expression levels and establish a previously unknown cross talk between these two pathways, which is highly relevant for AD pathogenesis. The discovery of such a novel link not only provides new mechanistic insights in the neurobiology of Hcy, but most importantly new therapeutic opportunities for the individuals bearing this risk factor for the disease.     

Metabolic and antiproliferative consequences of activated polyamine catabolism in LNCaP prostate carcinoma cells

Depletion of intracellular polyamine pools invariably inhibits cell growth. Although this is usually accomplished by inhibiting polyamine biosynthesis, we reasoned that this might be more effectively achieved by activation of polyamine catabolism at the level of spermidine/spermine N(1)-acetyltransferase (SSAT); a strategy first validated in MCF-7 breast carcinoma cells. We now examine the possibility that, due to unique aspects of polyamine homeostasis in the prostate gland, tumor cells derived from it may be particularly sensitive to activated polyamine catabolism. Thus, SSAT was conditionally overexpressed in LNCaP prostate carcinoma cells via a tetracycline-regulatable (Tet-off) system. Tetracycline removal resulted in a rapid approximately 10-fold increase in SSAT mRNA and an increase of approximately 20-fold in enzyme activity. SSAT products N(1)-acetylspermidine, N(1)-acetylspermine, and N(1),N(12)-diacetylspermine accumulated intracellularly and extracellularly. SSAT induction also led to a growth inhibition that was not accompanied by polyamine pool depletion as it was in MCF-7 cells. Rather, intracellular spermidine and spermine pools were maintained at or above control levels by a robust compensatory increase in ornithine decarboxylase and S-adenosylmethionine decarboxylase activities. This, in turn, gave rise to a high rate of metabolic flux through both the biosynthetic and catabolic arms of polyamine metabolism. Treatment with the biosynthesis inhibitor alpha-difluoromethylornithine during tetracycline removal interrupted flux and prevented growth inhibition. Thus, flux-induced growth inhibition appears to derive from overaccumulation of metabolic products and/or from depletion of metabolic precursors. Metabolic effects that were not excluded as possible contributing factors include high levels of putrescine and acetylated polyamines, a 50% reduction in S-adenosylmethionine, and a 45% decline in the SSAT cofactor acetyl-CoA. Overall, the study demonstrates that activation of polyamine catabolism in LNCaP cells elicits a compensatory increase in polyamine biosynthesis and downstream metabolic events that culminate in growth inhibition.     

Expression of a bacterial, phenylpropanoid-metabolizing enzyme in tobacco reveals essential roles of phenolic precursors in normal leaf development and growth

Tobacco plants (Nicotiana tabacum cv XHFD 8) were genetically modified to express a bacterial 4-hydroxycinnamoyl-CoA hydratase/lyase (HCHL) enzyme which is active with intermediates of the phenylpropanoid pathway. We have previously shown that HCHL expression in tobacco stem resulted in various pleiotropic effects, indicative of a reduction in the carbon flux through the phenylpropanoid pathway, accompanied by an abnormal phenotype. Here, we report that in addition to the reduction in lignin and phenolic biosynthesis, HCHL expression also resulted in several gross morphological changes in poorly lignified tissue, such as abnormal mesophyll and palisade. The effect of HCHL expression was also noted in lignin-free single cells, with suspension cultures displaying an altered shape and different growth patterns. Poorly/non-lignified cell walls also exhibited a greater ease of alkaline extractability of simple phenolics and increased levels of incorporation of vanillin and vanillic acid. However, HCHL expression had no significant effect on the cell wall carbohydrate chemistry of these tissues. Evidence from this study suggests that changes in the transgenic lines result from a reduction in phenolic intermediates which have an essential role in maintaining structural integrity of low-lignin or lignin-deprived cell walls. These results emphasize the importance of the intermediates and products of phenylpropanoid pathway in modulating aspects of normal growth and development of tobacco. Analysis of these transgenic plants also shows the plasticity of the lignification process and reveals the potential to bioengineer plants with reduced phenolics (without deleterious effects) which could enhance the bioconversion of lignocellulose for industrial applications.