Crossing geographical, legal and moral boundaries: the Belgian cigarette black market

Objectives: To describe and analyse the cigarette smuggling trade in Belgium and its role in the international cigarette black market. Design: Analysis of Belgian customs and prosecution files concerning the cigarette smuggling trade in the period 2000 to 2006 and interviews with law enforcement authorities and private tobacco industry. Results: Analyses were made of the geographical aspects, the modus operandi and the participants of the cigarette smuggling trade in Belgium. Belgium is mainly a transit country. The cigarettes are transported via the fine-meshed Belgian highway network to the UK, which is often the destination country of the cigarettes. China is the most popular country of origin, especially for counterfeited cigarettes. In order to transport the cigarettes often use was made of legal transport companies and warehouses were frequently used to store the cigarettes. Many of the persons involved in the Belgian cigarette smuggling trade are strongly connected to legitimate business activities. Conclusions: Belgium is an important transit country for cigarette smuggling to the UK. This study pictures the illicit tobacco trade as a complex, ambiguous phenomenon involving several legal and illegal participants whereby the transit of cigarettes across the licit/illicit divide is paralleled by the moral careers of those who smuggle them, not to mention those who consume them. From the legal world to the illegal and back again, this trade and its practitioners and customers blur the line between criminality and non- criminality. Dealing with this phenomenon therefore requires more than a strategy focusing on these lawbreakers alone

Implementation of non-invasive prenatal testing by semiconductor sequencing in a genetic laboratory

Objectives: To implement non-invasive prenatal testing (NIPT) for fetal aneuploidies with semiconductor sequencing in an academic cytogenomic laboratory and to evaluate the first 15-month experience on clinical samples. Methods: We validated a NIPT protocol for cell-free fetal DNA sequencing from maternal plasma for the detection of trisomy 13, 18 and 21 on a semiconductor sequencing instrument. Fetal DNA fraction calculation for all samples and several quality parameters were implemented in the workflow. One thousand eighty-one clinical NIPT samples were analysed, following the described protocol. Results: Non-invasive prenatal testing was successfully implemented and validated on 201 normal and 74 aneuploid samples. From 1081 clinical samples, 17 samples showed an abnormal result: 14 trisomy 21 samples, one trisomy 18 and one trisomy 16 were detected. Also a maternal copy number variation on chromosome 13 was observed, which could potentially lead to a false positive trisomy 13 result. One sex discordant result was reported, possibly attributable to a vanishing twin. Moreover, our combined fetal fraction calculation enabled a more reliable risk estimate for trisomy 13, 18 and 21. Conclusions: Non-invasive prenatal testing for trisomy 21, 18 and 13 has a very high specificity and sensitivity. Because of several biological phenomena, diagnostic invasive confirmation of abnormal results remains required

In vitro activity profiling of Cumyl-PEGACLONE variants at the CB1 receptor : fluorination versus isomer exploration

Synthetic cannabinoid receptor agonists (SCRAs) are one of the largest groups of new psychoactive substances monitored in Europe. SCRAs are known to typically exert higher cannabinoid activity than tetrahydrocannabinol from cannabis, thereby entailing a greater health risk. Both Cumyl-PEGACLONE and 5F-Cumyl-PEGACLONE were not controlled by the national legislation upon their first detection in Germany in 2016 and 2017, respectively, and have been linked to several fatalities. In this study, the CB1 receptor activity of these compounds, together with two newly synthesized structural isomers (Cumyl-PEGACLONE ethylbenzyl isomer and npropylphenyl isomer), was assessed using two different in vitro receptor-proximal bioassays, monitoring the recruitment of either β-arrestin2 (β-arr2) or a modified G protein (mini-Gαi) to the activated CB1 receptor. In terms of both potency and relative efficacy, Cumyl-PEGACLONE and 5F-Cumyl-PEGACLONE were found to exert strong CB1 activation, with sub-nanomolar EC50 values and efficacy values exceeding those of the reference agonist JWH-018 threefold (β-arr2 assay) or almost twofold (mini-Gαi assay). The ethylbenzyl and n-propylphenyl isomers exhibited a strongly reduced CB1 activity (EC50 values >100 nM; efficacy <40% relative to JWH-018), which is hypothesized to originate from steric hindrance in the ligand-binding pocket. None of the evaluated compounds exhibited significant biased agonism. In conclusion, the functional assays applied here allowed us to demonstrate that 5-fluorination of Cumyl-PEGACLONE is not linked to an intrinsically higher CB1 activation potential and that the ethylbenzyl and n-propylphenyl isomers yield a strongly reduced CB1 activation

Mechanism of Arachidonic Acid Modulation of the T-type Ca2+ Channel α1G

Arachidonic acid (AA) modulates T-type Ca2+ channels and is therefore a potential regulator of diverse cell functions, including neuronal and cardiac excitability. The underlying mechanism of modulation is unknown. Here we analyze the effects of AA on the T-type Ca2+ channel α1G heterologously expressed in HEK-293 cells. AA inhibited α1G currents within a few minutes, regardless of preceding exposure to inhibitors of AA metabolism (ETYA and 17-ODYA). Current inhibition was also observed in cell-free inside-out patches, indicating a membrane-delimited interaction of AA with the channel. AA action was consistent with a decrease of the open probability without changes in the size of unitary currents. AA shifted the inactivation curve to more negative potentials, increased the speed of macroscopic inactivation, and decreased the extent of recovery from inactivation at −80 mV but not at −110 mV. AA induced a slight increase of activation near the threshold and did not significantly change the deactivation kinetics or the rectification pattern. We observed a tonic current inhibition, regardless of whether the channels were held in resting or inactivated states during AA perfusion, suggesting a state-independent interaction with the channel. Model simulations indicate that AA inhibits T-type currents by switching the channels into a nonavailable conformation and by affecting transitions between inactivated states, which results in the negative shift of the inactivation curve. Slow-inactivating α1G mutants showed an increased affinity for AA with respect to the wild type, indicating that the structural determinants of fast inactivation are involved in the AA–channel interaction

Extracellular Ca2+ Modulates the Effects of Protons on Gating and Conduction Properties of the T-type Ca2+ Channel α1G (CaV3.1)

Since Ca2+ is a major competitor of protons for the modulation of high voltage–activated Ca2+ channels, we have studied the modulation by extracellular Ca2+ of the effects of proton on the T-type Ca2+ channel α1G (CaV3.1) expressed in HEK293 cells. At 2 mM extracellular Ca2+ concentration, extracellular acidification in the pH range from 9.1 to 6.2 induced a positive shift of the activation curve and increased its slope factor. Both effects were significantly reduced if the concentration was increased to 20 mM or enhanced in the absence of Ca2+. Extracellular protons shifted the voltage dependence of the time constant of activation and decreased its voltage sensitivity, which excludes a voltage-dependent open pore block by protons as the mechanism modifying the activation curve. Changes in the extracellular pH altered the voltage dependence of steady-state inactivation and deactivation kinetics in a Ca2+-dependent manner, but these effects were not strictly correlated with those on activation. Model simulations suggest that protons interact with intermediate closed states in the activation pathway, decreasing the gating charge and shifting the equilibrium between these states to less negative potentials, with these effects being inhibited by extracellular Ca2+. Extracellular acidification also induced an open pore block and a shift in selectivity toward monovalent cations, which were both modulated by extracellular Ca2+ and Na+. Mutation of the EEDD pore locus altered the Ca2+-dependent proton effects on channel selectivity and permeation. We conclude that Ca2+ modulates T-type channel function by competing with protons for binding to surface charges, by counteracting a proton-induced modification of channel activation and by competing with protons for binding to the selectivity filter of the channel

Mg2+-dependent Gating and Strong Inward Rectification of the Cation Channel TRPV6

TRPV6 (CaT1/ECaC2), a highly Ca2+-selective member of the TRP superfamily of cation channels, becomes permeable to monovalent cations in the absence of extracellular divalent cations. The monovalent currents display characteristic voltage-dependent gating and almost absolute inward rectification. Here, we show that these two features are dependent on the voltage-dependent block/unblock of the channel by intracellular Mg2+. Mg2+ blocks the channel by binding to a site within the transmembrane electrical field where it interacts with permeant cations. The block is relieved at positive potentials, indicating that under these conditions Mg2+ is able to permeate the selectivity filter of the channel. Although sizeable outward monovalent currents were recorded in the absence of intracellular Mg2+, outward conductance is still ∼10 times lower than inward conductance under symmetric, divalent-free ionic conditions. This Mg2+-independent rectification was preserved in inside-out patches and not altered by high intracellular concentrations of spermine, indicating that TRPV6 displays intrinsic rectification. Neutralization of a single aspartate residue within the putative pore loop abolished the Mg2+ sensitivity of the channel, yielding voltage-independent, moderately inwardly rectifying monovalent currents in the presence of intracellular Mg2+. The effects of intracellular Mg2+ on TRPV6 are partially reminiscent of the gating mechanism of inwardly rectifying K+ channels and may represent a novel regulatory mechanism for TRPV6 function in vivo

Pore Structure Influences Gating Properties of the T-type Ca2+ Channel α1G

The selectivity filter of all known T-type Ca2+ channels is built by an arrangement of two glutamate and two aspartate residues, each one located in the P-loops of domains I–IV of the α1 subunit (EEDD locus). The mutations of the aspartate residues to glutamate induce changes in the conduction properties, enhance Cd2+ and proton affinities, and modify the activation curve of the channel. Here we further analyze the role of the selectivity filter in the gating mechanisms of T-type channels by comparing the kinetic properties of the α1G subunit (CaV3.1) to those of pore mutants containing aspartate-to-glutamate substitution in domains III (EEED) or IV (EEDE). The change of the extracellular pH induced similar effects on the activation properties of α1G and both pore mutants, indicating that the larger affinity of the mutant channels for protons is not the cause of the gating modifications. Both mutants showed alterations in several gating properties with respect to α1G, i.e., faster macroscopic inactivation in the voltage range from −10 to 50 mV, positive voltage shift and decrease in the voltage sensitivity of the time constants of activation and deactivation, decrease of the voltage sensitivity of the steady-state inactivation, and faster recovery from inactivation for long repolarization periods. Kinetic modeling suggests that aspartate-to-glutamate mutations in the EEDD locus of α1G modify the movement of the gating charges and alter the rate of several gating transitions. These changes are independent of the alterations of the selectivity properties and channel protonation

Recent developments in the treatment of small cell lung cancer

Small cell lung cancer (SCLC) comprises about 15% of all lung cancers. It is an aggressive disease, with early metastasis and a poor prognosis. Until recently, SCLC treatment remained relatively unchanged, with chemotherapy remaining the cornerstone of treatment. In this overview we will highlight the recent advances in the field of staging, surgery, radiotherapy and systemic treatment. Nevertheless, the prognosis remains dismal and there is a pressing need for new treatment options. We describe the progress that has been made in systemic treatment by repurposing existing drugs and the addition of targeted treatment. In recent years, immunotherapy entered the clinic with high expectations of its role in the treatment of SCLC. Unravelling of the genomic sequence revealed new possible targets that may act as biomarkers in future treatment of patients with SCLC. Hopefully, in the near future, we will be able to identify patients who may benefit from targeted therapy or immunotherapy to improve prognoses

Endobronchial Lipomas: Rare Benign Lung Tumors, Two Case Reports

Abstract:Endobronchial lipoma is a rare benign lung tumor. Here we present two cases. One case is the first report of the association of and endobronchial lipoma with a hilar lipoma. We discuss the epidemiology, difficulties in establishing the diagnosis, and the management of this rare condition