Predicting in vitro drug sensitivity using Random Forests

MOTIVATION: Panels of cell lines such as the NCI-60 have long been used to test drug candidates for their ability to inhibit proliferation. Predictive models of in vitro drug sensitivity have previously been constructed using gene expression signatures generated from gene expression microarrays. These statistical models allow the prediction of drug response for cell lines not in the original NCI-60. We improve on existing techniques by developing a novel multistep algorithm that builds regression models of drug response using Random Forest, an ensemble approach based on classification and regression trees (CART). RESULTS: This method proved successful in predicting drug response for both a panel of 19 Breast Cancer and 7 Glioma cell lines, outperformed other methods based on differential gene expression, and has general utility for any application that seeks to relate gene expression data to a continuous output variable. Implementation: Software was written in the R language and will be available together with associated gene expression and drug response data as the package ivDrug at http://r-forge.r-project.org.     

Toluene inhibits voltage-sensitive calcium channels expressed in pheochromocytoma cells

Commercial solvents such as toluene are commonly used as drugs of abuse by children and adolescents. The cellular and molecular sites and mechanisms of actions of these compounds are not well studied but their effects on behavior resemble those of central nervous system depressants such as alcohol, barbiturates and benzodiazepines. In this study, the effects of toluene on voltage-sensitive calcium channels (VSCCs) were measured in pheochromocytoma cells. The KCl-induced rise in intracellular calcium as measured by calcium imaging was almost completely blocked by the dihydropyridine calcium channel antagonist nifedipine verifying that undifferentiated pheochromocytoma cells express mainly the L-type of calcium channel. Toluene (0.3–3000 μM) by itself did not affect intracellular calcium levels in resting cells but dose-dependently inhibited the KCl-induced rise in calcium. This inhibition was substantially reversed upon washout of the toluene-containing solution. KCl-dependent increases in intracellular calcium in cells differentiated with nerve growth factor (NGF) were largely insensitive to nifedipine. Toluene produced a greater inhibition of the KCl response in NGF treated cells as compared with undifferentiated cells. A similar finding was obtained when whole-cell patch-clamp-electrophysiology was used to directly monitor the effects of toluene on voltage-activated calcium currents in undifferentiated and differentiated cells. These results show that dihydropyridine sensitive and insensitive calcium channels are inhibited by toluene and may represent important sites of action for this compound.     

The chemical ecology of <Emphasis Type="Italic">Harmonia axyridis</Emphasis>

We review the chemical ecology of the ladybird beetle Harmonia axyridis from the perspective of its invasiveness and the deleterious effects it exerts in the regions it has colonised. We outline the nature and quantification of its chemical defence, and discuss the protection this provides against natural enemies, particularly intraguild predators. We consider the role of infochemicals in location of prey, intraspecific communication and intraguild interactions. We also discuss the role of prey allelochemicals in relation to H. axyridis extreme dietary generalism. Harmonia axyridis poses a number of practical problems for human health and well-being, including “ladybug taint” wine contamination and problems resulting from large aggregations overwintering in buildings. We consider chemical insights into these issues and, in particular, how attractants and repellents might help manage H. axyridis populations through a push–pull strategy. We conclude by discussing future perspectives for research.     

Synergy of fatty acid and reactive alkenal activation of proton conductance through uncoupling protein 1 in mitochondria

The kinetics of proton transport through mammalian UCP1 (uncoupling protein 1) expressed in yeast mitochondria were measured. There was little or no UCP1 activity in the absence of added palmitate, but significant activity in its presence. The activator 4-HNE (4-hydroxy-2-nonenal) had little effect when added alone, but significantly enhanced proton conductance in the presence of added palmitate. Activation of the proton conductance of UCP1 was synergistic: proton conductance in the presence of both palmitate and 4-HNE was significantly greater than the sum of the individual effects. Mitochondria from control yeast transformed with empty vector showed no such synergy, showing that synergy is a property of UCP1. Activation by the 4-HNE analogue trans-cinnamate showed essentially the same characteristics as activation by 4-HNE. Mitochondria from brown adipose tissue also showed synergistic activation of GDP-sensitive proton conductance by palmitate and 4-HNE. These results show that reactive alkenals activate the proton conductance of UCP1 more strongly when fatty acids are also added, with implications for both mechanistic and physiological models of UCP1 activation.     

Dynamic regulation of uncoupling protein 2 content in INS-1E insulinoma cells

Uncoupling protein 2 (UCP2) regulates glucose-stimulated insulin secretion in pancreatic beta-cells. UCP2 content, measured by calibrated immunoblot in INS-1E insulinoma cells (a pancreatic beta-cell model) grown in RPMI medium, and INS-1E mitochondria, was 2.0 ng/million cells (7.9 ng/mg mitochondrial protein). UCP2 content was lower in cells incubated without glutamine and higher in cells incubated with 20 mM glucose, and varied from 1.0-4.4 ng/million cells (2.7-14.5 ng/mg mitochondrial protein). This dynamic response to nutrients was achieved by varied expression rates against a background of a very short UCP2 protein half-life of about 1 h.     

Activated lymphocytes increase expression of 5-lipoxygenase and its activating protein in THP-1 cells

The aim of this study was to investigate the regulation of the5-lipoxygenase pathway of arachidonic acid metabolism by lymphocytes using the monocyte-like cell line, THP-1. When THP-1 cells were incubated over 4-7 days in 10% supernatant from lectin-activated human lymphocytes, their capacity to synthesize 5-lipoxygenase productswas significantly increased. In contrast, the supernatant fromnonactivated lymphocytes had no effect. The increase in capacity tosynthesize 5-lipoxygenase products was mimicked by the addition ofeither granulocyte macrophage colony-stimulating factor (GM-CSF) orinterleukin-3. These increases in synthetic capacity reflected increased enzymatic activity. Increased immunoreactive protein and mRNAfor the enzymes 5-lipoxygenase and 5-lipoxygenase-activating protein were also found in cells conditioned with activated lymphocyte supernatants. Furthermore, the increase in mRNA for both enzymes wasnot blocked by cycloheximide, suggesting that the effect on steady-state mRNA levels does not require the synthesis of new protein.The increase in mRNA could be reproduced by GM-CSF. We conclude thatlymphocytes can regulate the expression of 5-lipoxygenase in THP-1cells over a period of days via the release of soluble factors.