Cenicriviroc,a Novel CCR5 (R5) and CCR2 Antagonist,Shows In Vitro Activity against R5 Tropic HIV-2 Clinical Isolates

Background

Maraviroc activity against HIV-2, a virus naturally resistant to different HIV-1 antiretroviral drugs, has been recently demonstrated. The aim of this study was to assess HIV-2 susceptibility to cenicriviroc, a novel, once-daily, dual CCR5 and CCR2 antagonist that has completed Phase 2b development in HIV-1 infection.

Methods

Cenicriviroc phenotypic activity has been tested using a PBMC phenotypic susceptibility assay against four R5-, one X4- and one dual-tropic HIV-2 clinical primary isolates. All isolates were obtained by co-cultivation of PHA-activated PBMC from distinct HIV-2-infected CCR5-antagonist-naïve patients included in the French HIV-2 cohort and were previously tested for maraviroc susceptibility using the same protocol. HIV-2 tropism was determined by phenotypic assay using Ghost(3) cell lines.

Results

Regarding the 4 R5 HIV-2 clinical isolates tested, effective concentration 50% EC₅₀ for cenicriviroc were 0.03, 0.33, 0.45 and 0.98 nM, similar to those observed with maraviroc: 1.13, 0.58, 0.48 and 0.68 nM, respectively. Maximum percentages of inhibition (MPI) of cenicriviroc were 94, 94, 93 and 98%, similar to those observed with maraviroc (93, 90, 82, 100%, respectively). The dual- and X4-tropic HIV-2 strains were resistant to cenicriviroc with EC50 >1000 nM and MPI at 33% and 4%, respectively.

Conclusions

In this first study assessing HIV-2 susceptibility to cenicriviroc, we observed an in vitro activity against HIV-2 R5-tropic strains similar to that observed with maraviroc. Thus, cenicriviroc may offer a once-daily treatment opportunity in the limited therapeutic arsenal for HIV-2. Clinical studies are warranted.     

An Extended,Boolean Model of the Septation Initiation Network in S.Pombe Provides Insights into Its Regulation

Cytokinesis in fission yeast is controlled by the Septation Initiation Network (SIN), a protein kinase signaling network using the spindle pole body as scaffold. In order to describe the qualitative behavior of the system and predict unknown mutant behaviors we decided to adopt a Boolean modeling approach. In this paper, we report the construction of an extended, Boolean model of the SIN, comprising most SIN components and regulators as individual, experimentally testable nodes. The model uses CDK activity levels as control nodes for the simulation of SIN related events in different stages of the cell cycle. The model was optimized using single knock-out experiments of known phenotypic effect as a training set, and was able to correctly predict a double knock-out test set. Moreover, the model has made in silico predictions that have been validated in vivo, providing new insights into the regulation and hierarchical organization of the SIN.     

Massive Diversification in Aging Colonies of Escherichia coli

The evolutionary success of bacteria depends greatly on their capacity to continually generate phenotypic diversity. Structured environments are particularly favorable for diversification because of attenuated clonal interference, which renders selective sweeps nearly impossible and enhances opportunities for adaptive radiation. We examined at the microscale level the emergence and the spatial and temporal dynamics of phenotypic diversity and their underlying causes in Escherichia coli colonies. An important dynamic heterogeneity in the growth, metabolic activity, morphology, gene expression patterns, stress response induction, and death patterns among cells within colonies was observed. Genetic analysis indicated that the phenotypic variation resulted mostly from mutations and that indole production, oxidative stress, and the RpoS-regulated general stress response played an important role in the generation of diversity. We observed the emergence and persistence of phenotypic variants within single colonies that exhibited variable fitness compared to the parental strain. Some variants showed improved capacity to produce biofilms, whereas others were able to use different nutrients or to tolerate antibiotics or oxidative stress. Taken together, our data show that bacterial colonies provide an ecological opportunity for the generation and maintenance of vast phenotypic diversity, which may increase the probability of population survival in unpredictable environments.     

Experimental evidence of the reciprocal oxidation of Bovine Serum Albumin and Linoleate in aqueous solution,initiated by HO free radicals

An investigation of radiation-induced oxidation of aqueous bovine serum albumin (BSA) in the presence of linoleate (LH) at pH 10.5 has been carried out in order to better understand the respective oxidative processes involved in both lipid and protein phases. Solutions containing BSA (15 μmol L⁻¹) and linoleate (15–600 μmol L⁻¹) below the critical micellar concentration (cmc = 2000 μmol L⁻¹), have been irradiated by γ-rays (¹³⁷Cs) at radiation doses ranging from 10 to 400 Gy (dose rate 9.5 Gy min⁻¹). It can be noticed that, in the absence of BSA, the main hydroperoxides formed from HO^•-induced linoleate oxidation below the cmc, do not exhibit a conjugated dienic structure. This was also verified in the presence of BSA. Selected chemical markers of oxidation have been monitored: non-conjugated dienic hydroperoxides and conjugated dienes (without hydroperoxide function) for linoleate oxidation, and carbonyl groups for BSA oxidation. We have shown that for the lowest linoleate concentration (15 μmol L⁻¹) in the presence of BSA (15 μmol L⁻¹), the formation of conjugated dienes was not observed, meaning that LH was not exposed to HO^• radicals attack. However, non-conjugated dienic lipid hydroperoxides were simultaneously detected, indicating that LH was secondarily oxidised by BSA oxidised species. Moreover, the oxidation of linoleate was found to be enhanced by the presence of BSA. For the highest linoleate concentration (600 μmol L⁻¹), the expected protection of BSA by LH was not observed, even if LH monomers were responsible for the total scavenging of HO^• radicals. In this latter case, the formation of non-conjugated dienic lipid hydroperoxides was lower than expected. Those results showed that BSA was not oxidised by the direct action of HO^• radicals but was undergoing a secondary oxidation by non-dienic lipid hydroperoxides and/or lipid radical intermediates, coming from the HO^•-induced linoleate oxidation.     

Mutations in IMPG2, Encoding Interphotoreceptor Matrix Proteoglycan 2, Cause Autosomal-Recessive Retinitis Pigmentosa

Retinitis pigmentosa (RP) is a heterogeneous group of inherited retinal diseases caused by progressive degeneration of the photoreceptor cells. Using autozygosity mapping, we identified two families, each with three affected siblings sharing large overlapping homozygous regions that harbored the IMPG2 gene on chromosome 3. Sequence analysis of IMPG2 in the two index cases revealed homozygous mutations cosegregating with the disease in the respective families: three affected siblings of Iraqi Jewish ancestry displayed a nonsense mutation, and a Dutch family displayed a 1.8 kb genomic deletion that removes exon 9 and results in the absence of seven amino acids in a conserved SEA domain of the IMPG2 protein. Transient transfection of COS-1 cells showed that a construct expressing the wild-type SEA domain is properly targeted to the plasma membrane, whereas the mutant lacking the seven amino acids appears to be retained in the endoplasmic reticulum. Mutation analysis in ten additional index cases that were of Dutch, Israeli, Italian, and Pakistani origin and had homozygous regions encompassing IMPG2 revealed five additional mutations; four nonsense mutations and one missense mutation affecting a highly conserved phenylalanine residue. Most patients with IMPG2 mutations showed an early-onset form of RP with progressive visual-field loss and deterioration of visual acuity. The patient with the missense mutation, however, was diagnosed with maculopathy. The IMPG2 gene encodes the interphotoreceptor matrix proteoglycan IMPG2, which is a constituent of the interphotoreceptor matrix. Our data therefore show that mutations in a structural component of the interphotoreceptor matrix can cause arRP.     

Next-Generation Sequencing of a 40 Mb Linkage Interval Reveals TSPAN12 Mutations in Patients with Familial Exudative Vitreoretinopathy

Familial exudative vitreoretinopathy (FEVR) is a genetically heterogeneous retinal disorder characterized by abnormal vascularisation of the peripheral retina, often accompanied by retinal detachment. To date, mutations in three genes (FZD4, LRP5, and NDP) have been shown to be causative for FEVR. In two large Dutch pedigrees segregating autosomal-dominant FEVR, genome-wide SNP analysis identified an FEVR locus of ∼40 Mb on chromosome 7. Microsatellite marker analysis suggested similar at risk haplotypes in patients of both families. To identify the causative gene, we applied next-generation sequencing in the proband of one of the families, by analyzing all exons and intron-exon boundaries of 338 genes, in addition to microRNAs, noncoding RNAs, and other highly conserved genomic regions in the 40 Mb linkage interval. After detailed bioinformatic analysis of the sequence data, prioritization of all detected sequence variants led to three candidates to be considered as the causative genetic defect in this family. One of these variants was an alanine-to-proline substitution in the transmembrane 4 superfamily member 12 protein, encoded by TSPAN12. This protein has very recently been implicated in regulating the development of retinal vasculature, together with the proteins encoded by FZD4, LRP5, and NDP. Sequence analysis of TSPAN12 revealed two mutations segregating in five of 11 FEVR families, indicating that mutations in TSPAN12 are a relatively frequent cause of FEVR. Furthermore, we demonstrate the power of targeted next-generation sequencing technology to identify disease genes in linkage intervals.     

“Working the System”—British American Tobacco's Influence on the European Union Treaty and Its Implications for Policy: An Analysis of Internal Tobacco Industry Documents

Background

Impact assessment (IA) of all major European Union (EU) policies is now mandatory. The form of IA used has been criticised for favouring corporate interests by overemphasising economic impacts and failing to adequately assess health impacts. Our study sought to assess how, why, and in what ways corporations, and particularly the tobacco industry, influenced the EU''s approach to IA.

Methods and Findings

In order to identify whether industry played a role in promoting this system of IA within the EU, we analysed internal documents from British American Tobacco (BAT) that were disclosed following a series of litigation cases in the United States. We combined this analysis with one of related literature and interviews with key informants. Our analysis demonstrates that from 1995 onwards BAT actively worked with other corporate actors to successfully promote a business-oriented form of IA that favoured large corporations. It appears that BAT favoured this form of IA because it could advance the company''s European interests by establishing ground rules for policymaking that would: (i) provide an economic framework for evaluating all policy decisions, implicitly prioritising costs to businesses; (ii) secure early corporate involvement in policy discussions; (iii) bestow the corporate sector with a long-term advantage over other actors by increasing policymakers'' dependence on information they supplied; and (iv) provide businesses with a persuasive means of challenging potential and existing legislation. The data reveal that an ensuing lobbying campaign, largely driven by BAT, helped secure binding changes to the EU Treaty via the Treaty of Amsterdam that required EU policymakers to minimise legislative burdens on businesses. Efforts subsequently focused on ensuring that these Treaty changes were translated into the application of a business orientated form of IA (cost–benefit analysis [CBA]) within EU policymaking procedures. Both the tobacco and chemical industries have since employed IA in apparent attempts to undermine key aspects of European policies designed to protect public health.

Conclusions

Our findings suggest that BAT and its corporate allies have fundamentally altered the way in which all EU policy is made by making a business-oriented form of IA mandatory. This increases the likelihood that the EU will produce policies that advance the interests of major corporations, including those that produce products damaging to health, rather than in the interests of its citizens. Given that the public health community, focusing on health IA, has largely welcomed the increasing policy interest in IA, this suggests that urgent consideration is required of the ways in which IA can be employed to undermine, as well as support, effective public health policies. Please see later in the article for the Editors'' Summary